1997年, 井上財団奨励賞　　　　　　　　　　　　　　　

1996年, FIMSA Young Investigators Award　　　　　　　　　　　　　　　

Tumor cell-induced macrophage senescence plays a pivotal role in tumor initiation followed by stable growth in immunocompetent condition.
Haruka Wada, Ryo Otsuka, Wilfred T V Germeraad, Tomoki Murata, Toru Kondo, Ken-Ichiro Seino
Journal for immunotherapy of cancer, 11, 11, 2023年11月, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: The cancer stem cell theory proposes that tumor formation in vivo is driven only by specific tumor-initiating cells having stemness; however, clinical trials conducted to test drugs that target the tumor stemness provided unsatisfactory results thus far. Recent studies showed clear involvement of immunity in tumors; however, the requirements of tumor-initiation followed by stable growth in immunocompetent individuals remain largely unknown. METHODS: To clarify this, we used two similarly induced glioblastoma lines, 8B and 9G. They were both established by overexpression of an oncogenic H-RasL61 in p53-deficient neural stem cells. In immunocompromised animals in an orthotopic transplantation model using 1000 cells, both show tumor-forming potential. On the other hand, although in immunocompetent animals, 8B shows similar tumor-forming potential but that of 9G's are very poor. This suggests that 8B cells are tumor-initiating cells in immunocompetent animals. Therefore, we hypothesized that the differences in the interaction properties of 8B and 9G with immune cells could be used to identify the factors responsible for its tumor forming potential in immunocompetent animals and performed analysis. RESULTS: Different from 9G, 8B cells induced senescence-like state of macrophages around tumors. We investigated the senescence-inducing factor of macrophages by 8B cells and found that it was interleukin 6. Such senescence-like macrophages produced Arginase-1, an immunosuppressive molecule known to contribute to T-cell hyporesponsiveness. The senescence-like macrophages highly expressed CD38, a nicotinamide adenine dinucleotide (NAD) glycohydrolase associated with NAD shortage in senescent cells. The addition of nicotinamide mononucleotide (NMN), an NAD precursor, in vitro inhibited to the induction of macrophage senescence-like phenotype and inhibited Arginase-1 expression resulting in retaining T-cell function. Moreover, exogenous in vivo administration of NMN after tumor inoculation inhibited tumor-initiation followed by stable growth in the immunocompetent mouse tumor model. CONCLUSIONS: We identified one of the requirements for tumor-initiating cells in immunocompetent animals. In addition, we have shown that tumor growth can be inhibited by externally administered NMN against macrophage senescence-like state that occurs in the very early stages of tumor-initiating cell development. This therapy targeting the immunosuppressive environment formed by macrophage senescence-like state is expected to be a novel promising cancer therapeutic strategy.

Induced pluripotent stem cell-derived hematopoietic stem and progenitor cells induce mixed chimerism and donor-specific allograft tolerance.
Tomoki Murata, Naoki Hama, Tomoki Kamatani, Akihiro Mori, Ryo Otsuka, Haruka Wada, Ken-Ichiro Seino
American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons, 23, 9, 1331, 1344, 2023年09月, ［国際誌］
英語, 研究論文（学術雑誌）, In transplantation using allogeneic induced pluripotent stem cells (iPSCs), strategies focused on major histocompatibility complexes were adopted to avoid immune rejection. We showed that minor antigen mismatches are a risk factor for graft rejection, indicating that immune regulation remains one of the most important issues. In organ transplantation, it has been known that mixed chimerism using donor-derived hematopoietic stem/progenitor cells (HSPCs) can induce donor-specific tolerance. However, it is unclear whether iPSC-derived HSPCs (iHSPCs) can induce allograft tolerance. We showed that 2 hematopoietic transcription factors, Hoxb4 and Lhx2, can efficiently expand iHSPCs with a c-Kit+Sca-1+Lineage- phenotype, which possesses long-term hematopoietic repopulating potential. We also demonstrated that these iHSPCs can form hematopoietic chimeras in allogeneic recipients and induce allograft tolerance in murine skin and iPSC transplantation. With mechanistic analyses, both central and peripheral mechanisms were suggested. We demonstrated the basic concept of tolerance induction using iHSPCs in allogeneic iPSC-based transplantation.

Augmented interferon regulatory factor 7 axis in whole tumor cell vaccines prevents tumor recurrence by inducing interferon gamma-secreting B cells
Nabeel Kajihara, Yoshino Tanaka, Riko Takeuchi, Takuto Kobayashi, Masafumi Tanji, Tsukasa Ataka, Shiho Nakano, Taisho Yamada, Akinori Takaoka, Yoshinori Hasegawa, Ken-Ichiro Seino, Haruka Wada
OncoImmunology, 12, 1, Informa UK Limited, 2023年05月22日
研究論文（学術雑誌）

Induction of allograft tolerance by adoptive transfer of donor B cells: an immune regulatory strategy for transplantation using MHC-matched iPS cells.
Tomoki Murata, Ryo Otsuka, Airi Sasaki, Tomoki Kamatani, Haruka Wada, Hisashi Yamakawa, Yoshinori Hasegawa, Ken-Ichiro Seino
International immunology, 2023年04月13日, ［国際誌］
英語, 研究論文（学術雑誌）, For cellular or tissue transplantation using iPS cells (iPSCs), from the viewpoint of time and economic cost, an use of allogeneic ones is being considered. Immune regulation is one of the key issues in successful allogeneic transplantation. To reduce the risk of rejection, several attempts have been reported to eliminate effects of major histocompatibility complex (MHC) on the iPSCs-derived grafts. On the other hand, we have shown that minor antigen-induced rejection is not negligible even when the MHC's impact is mitigated. In organ transplantation, it is known that donor-specific blood transfusion (DST) can specifically control immune responses to the donor. However, whether DST could control the immune response in iPSC-based transplantation was not clarified. In this study, using a mouse skin transplantation model, we demonstrate that infusion of donor splenocytes can promote allograft tolerance in the MHC-matched but minor antigen-mismatched condition. When narrowing down the cell types, we found that infusion of isolated splenic B cells was sufficient to control rejection. As a mechanism, the administration of donor B cells induced unresponsiveness but not deletion in recipient T cells, suggesting that the tolerance was induced in the periphery. The donor B cell transfusion induced allogeneic iPSCs engraftment. These results suggest for the first time a possibility that DST using donor B cells could induce tolerance against allogeneic iPSC-derived grafts.

Interleukin-34 cancels anti-tumor immunity by PARP inhibitor.
Takayoshi Nakamura, Nabeel Kajihara, Naoki Hama, Takuto Kobayashi, Ryo Otsuka, Nanumi Han, Haruka Wada, Yoshinori Hasegawa, Nao Suzuki, Ken-Ichiro Seino
Journal of gynecologic oncology, 2022年12月21日, ［国際誌］
英語, 研究論文（学術雑誌）, OBJECTIVE: Breast cancer susceptibility gene 1 (BRCA1)-associated ovarian cancer patients have been treated with A poly (ADP-ribose) polymerase (PARP) inhibitor, extending the progression-free survival; however, they finally acquire therapeutic resistance. Interleukin (IL)-34 has been reported as a poor prognostic factor in several cancers, including ovarian cancer, and it contributes to the therapeutic resistance of chemotherapies. IL-34 may affect the therapeutic effect of PARP inhibitor through the regulation of tumor microenvironment (TME). METHODS: In this study, The Cancer Genome Atlas (TCGA) data set was used to evaluate the prognosis of IL-34 and human ovarian serous carcinoma. We also used CRISPR-Cas9 genome editing technology in a mouse model to evaluate the efficacy of PARP inhibitor therapy in the presence or absence of IL-34. RESULTS: We found that IL34 was an independent poor prognostic factor in ovarian serous carcinoma, and its high expression significantly shortens overall survival. Furthermore, in BRCA1-associated ovarian cancer, PARP inhibitor therapy contributes to anti-tumor immunity via the XCR1+ DC-CD8+ T cell axis, however, it is canceled by the presence of IL-34. CONCLUSION: These results suggest that tumor-derived IL-34 benefits tumors by creating an immunosuppressive TME and conferring PARP inhibitor therapeutic resistance. Thus, we showed the pathological effect of IL-34 and the need for it as a therapeutic target in ovarian cancer.

Tumor-derived interleukin-34 creates an immunosuppressive and chemoresistant tumor microenvironment by modulating myeloid-derived suppressor cells in triple-negative breast cancer.
Nabeel Kajihara, Takuto Kobayashi, Ryo Otsuka, Junko Nio-Kobayashi, Tomohiro Oshino, Masato Takahashi, Seiichi Imanishi, Ari Hashimoto, Haruka Wada, Ken-Ichiro Seino
Cancer immunology, immunotherapy : CII, 2022年09月14日, ［国際誌］
英語, 研究論文（学術雑誌）, Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype characterized by a lack of therapeutic targets. The paucity of effective treatment options motivated a number of studies to tackle this problem. Immunosuppressive cells infiltrated into the tumor microenvironment (TME) of TNBC are currently considered as candidates for new therapeutic targets. Myeloid-derived suppressor cells (MDSCs) have been reported to populate in the TME of TNBC, but their roles in the clinical and biological features of TNBC have not been clarified. This study identified that interleukin-34 (IL-34) released by TNBC cells is a crucial immunomodulator to regulate MDSCs accumulation in the TME. We provide evidence that IL-34 induces a differentiation of myeloid stem cells into monocytic MDSCs (M-MDSCs) that recruits regulatory T (Treg) cells, while suppressing a differentiation into polymorphonuclear MDSCs (PMN-MDSCs). As a result, the increase in M-MDSCs contributes to the creation of an immunosuppressive TME, and the decrease in PMN-MDSCs suppresses angiogenesis, leading to an acquisition of resistance to chemotherapy. Accordingly, blockade of M-MDSC differentiation with an estrogen receptor inhibitor or anti-IL-34 monoclonal antibody suppressed M-MDSCs accumulation causing retardation of tumor growth and restores chemosensitivity of the tumor by promoting PMN-MDSCs accumulation. This study demonstrates previously poorly understood mechanisms of MDSCs-mediated chemoresistance in the TME of TNBC, which is originated from the existence of IL-34, suggesting a new rationale for TNBC treatment.

Evaluation of immunosuppression protocols for MHC-matched allogeneic iPS cell-based transplantation using a mouse skin transplantation model.
Tomoki Kamatani, Ryo Otsuka, Tomoki Murata, Haruka Wada, Takeshi Takahashi, Akihiro Mori, Soichiro Murata, Hideki Taniguchi, Ken-Ichiro Seino
Inflammation and regeneration, 42, 1, 4, 4, 2022年02月02日, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Off-the-shelf major histocompatibility complex (MHC)-matched iPS cells (iPSC) can potentially initiate host immune responses because of the existence of numerous minor antigens. To suppress allo-immune responses, combination of immunosuppressants is usually used, but its efficacy to the allogeneic iPSC-based transplantation has not been precisely evaluated. METHODS: Three transplantation models were used in this study; MHC-matched, minor antigen-mismatched mouse skin or iPSC-graft transplantation, and fully allogeneic human iPSC-derived liver organoid transplantation in immune-humanized mice. The recipients were treated with triple drugs combination (TDC; tacrolimus, methylprednisolone, and mycophenolate mofetil) or co-stimulatory molecule blockade (CB) therapy with some modifications. Graft survival as well as anti-donor T and B cell responses was analyzed. RESULTS: In the mouse skin transplantation model, immunological rejection caused by the minor antigen-mismatch ranged from mild to severe according to the donor-recipient combination. The TDC treatment could apparently control the mild skin graft rejection when combined with a transient T cell depletion, but unexpected anti-donor T or B cell response was observed. On the other hand, CB therapy, particularly when combined with rapamycin treatment, was capable of attenuating both mild and severe skin graft rejection and allowing them to survive long-term without any unfavorable anti-donor immune responses. The efficacy of the CB therapy was confirmed in both mouse and human iPSC-derived graft transplantation. CONCLUSIONS: The findings suggest that the CB-based treatment seems suitable to well manage the MHC-matched allogeneic iPSC-based transplantation. The TDC-based treatment may be also used to suppress the rejection, but screening of its severity prior to the transplantation seems to be needed.

Erratum: Interleukin-34 Limits the Therapeutic Effects of Immune Checkpoint Blockade.
Naoki Hama, Takuto Kobayashi, Nanumi Han, Fumihito Kitagawa, Nabeel Kajihara, Ryo Otsuka, Haruka Wada, Hee-Kyung Lee, Hwanseok Rhee, Yoshinori Hasegawa, Hideo Yagita, Muhammad Baghdadi, Ken-Ichiro Seino
iScience, 25, 1, 103713, 103713, 2022年01月21日, ［国際誌］
英語, [This corrects the article DOI: 10.1016/j.isci.2020.101584.].

Macrophage-like iPS-derived Suppressor Cells Reduce Th1-mediated Immune Response to a Retinal Antigen.
Keitaro Hase, Kenichi Namba, Haruka Wada, Hyuma Tsuji, Aoi Maeda, Tomoki Murata, Ryo Otsuka, Daiju Iwata, Atsuhiro Kanda, Kousuke Noda, Nobuyoshi Kitaichi, Ken-Ichiro Seino, Susumu Ishida
Current eye research, 46, 12, 1908, 1916, 2021年12月, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: To investigate the immunotherapeutic effects of macrophage-like induced pluripotent stem (iPS) cell-derived suppressor cells (SCs) in ocular immune response and experimental autoimmune uveoretinitis (EAU). METHODS: The genes of Oct3/4, Sox2, Klf4, and c-Myc were transferred to B cells enriched from the spleen cells of C57BL/6 mice by using retrovirus vectors. Transferred B cells were cultured for 17 days to obtain colonies of iPS cells. Through additional steps, iPS-SCs were induced. An antigen-specific T cell proliferation assay was performed with CD4+ T cells collected from draining lymph nodes of the mice immunized with human interphotoreceptor retinoid-binding protein (hIRBP) peptide and co-cultured with iPS-SCs. Cytokine concentrations in the culture supernatant were examined. Mice were immunized with hIRBP peptide to induce EAU. The iPS-SCs were administered into the mice one day before the induction of EAU. RESULTS: The iPS-SCs decreased hIRBP-specific T cell proliferation depending on the number of cells. Productions of tumor necrosis factor-α and interferon-γ were significantly decreased; however, transforming growth factor-β1, nitric oxide, interleukin (IL)-13, IL-17A, and IL-17 F levels were elevated in the supernatant when the collected T cells were co-cultured with iPS-SCs. The iPS-SCs had immunosuppressant effects even without cell-to-cell contact, and their effects were non-specific to the antigen preloaded on iPS-SCs. EAU was significantly milder in the mice administered iPS-SCs prior to immunization. CONCLUSIONS: Macrophage-like iPS-SCs reduced Th1 immune response to a retinal antigen and Th1-mediated EAU in mice. These results showed the possibility of the application of iPS technology to the treatment of noninfectious ocular inflammation, endogenous uveitis, in the future.

細胞・臓器の移植に伴う免疫反応とその制御-最新の研究- MHC型一致他家iPS細胞由来組織移植に有効な免疫抑制剤併用プロトコールの構築
鎌谷 智紀, 大塚 亮, 村田 智己, 和田 はるか, 高橋 武司, 森 淳祐, 村田 聡一郎, 谷口 英樹, 清野 研一郎
Organ Biology, 28, 3, 75, 75, (一社)日本臓器保存生物医学会, 2021年10月
日本語

死にゆくがん細胞ががん細胞ワクチン効果を発揮する　　　　　　　　　　　　　　　
和田 はるか, 梶原 ナビール, 清野 研一郎
日本癌学会総会記事, 80回, [P12, 4], (一社)日本癌学会, 2021年09月
英語

An optimized protocol for patient-derived xenograft in humanized mice to evaluate the role of IL-34 in immunotherapeutic resistance.
Nanumi Han, Hye Yoon Jang, Naoki Hama, Takuto Kobayashi, Ryo Otsuka, Haruka Wada, Ken-Ichiro Seino
STAR protocols, 2, 2, 100460, 100460, 2021年06月18日, ［国際誌］
英語, 研究論文（学術雑誌）, Previously, we identified a therapy-resistant role of IL-34 in an immune checkpoint blockade in murine models. To investigate whether a similar mechanism is applicable in human tumors as well, we used this protocol for the selection of IL-34-neutralizing antibody and transplanting human tumor tissue expressing both IL-34 and PD-L1 as a patient-derived xenograft in immunologically humanized mice. This model helps to determine the effect of IL-34 neutralization along with the immune checkpoint blockade in human tumors. For complete details on the use and execution of this protocol, please refer to Hama et al. (2020).

Author Correction: Efficient generation of thymic epithelium from induced pluripotent stem cells that prolongs allograft survival.
Ryo Otsuka, Haruka Wada, Hyuma Tsuji, Airi Sasaki, Tomoki Murata, Mizuho Itoh, Muhammad Baghdadi, Ken-Ichiro Seino
Scientific reports, 11, 1, 12857, 12857, 2021年06月14日, ［国際誌］
英語

IL-34, the rationale for its expression in physiological and pathological conditions.
Ryo Otsuka, Haruka Wada, Ken-Ichiro Seino
Seminars in immunology, 54, 101517, 101517, 2021年04月, ［国際誌］
英語, 研究論文（学術雑誌）, IL-34 is a cytokine that shares one of its receptors with CSF-1. It has long been thought that CSF-1 receptor (CSF-1R) receives signals only from CSF-1, but the identification of IL-34 reversed this stereotype. Regardless of low structural homology, IL-34 and CSF-1 emanate similar downstream signaling through binding to CSF-1R and provoke similar but different physiological events afterward. In addition to CSF-1R, protein-tyrosine phosphatase (PTP)-ζ and Syndecan-1 were also identified as IL-34 receptors and shown to be at play. Although IL-34 expression is limited to particular tissues in physiological conditions, previous studies have revealed that it is upregulated in several diseases. In cancer, IL-34 is produced by several types of tumor cells and contributes to therapy resistance and disease progression. A recent study has demonstrated that tumor cell-derived IL-34 abrogates immunotherapy efficacy through myeloid cell remodeling. On the other hand, IL-34 expression is downregulated in some brain and dermal disorders. Despite accumulating insights, our understanding of IL-34 may not be even close to its nature. This review aims to comprehensively describe the physiological and pathological roles of IL-34 based on its similarity and differences to CSF-1 and discuss the rationale for its disease-dependent expression pattern.

Establishment of Human Leukocyte Antigen-Mismatched Immune Responses after Transplantation of Human Liver Bud in Humanized Mouse Models.
Akihiro Mori, Soichiro Murata, Nao Tashiro, Tomomi Tadokoro, Satoshi Okamoto, Ryo Otsuka, Haruka Wada, Tomoki Murata, Takeshi Takahashi, Ken-Ichiro Seino, Hideki Taniguchi
Cells, 10, 2, 2021年02月23日, ［国際誌］
英語, 研究論文（学術雑誌）, Humanized mouse models have contributed significantly to human immunology research. In transplant immunity, human immune cell responses to donor grafts have not been reproduced in a humanized animal model. To elicit human T-cell immune responses, we generated immune-compromised nonobese diabetic/Shi-scid, IL-2RγKO Jic (NOG) with a homozygous expression of human leukocyte antigen (HLA) class I heavy chain (NOG-HLA-A2Tg) mice. After the transplantation of HLA-A2 human hematopoietic stem cells into NOG-HLA-A2Tg, we succeeded in achieving alloimmune responses after the HLA-mismatched human-induced pluripotent stem cell (hiPSC)-derived liver-like tissue transplantation. This immune response was inhibited by administering tacrolimus. In this model, we reproduced allograft rejection after the human iPSC-derived liver-like tissue transplantation. Human tissue transplantation on the humanized mouse liver surface is a good model that can predict T-cell-mediated cellular rejection that may occur when organ transplantation is performed.

Bromodomain-containing protein 4 regulates interleukin-34 expression in mouse ovarian cancer cells
Nanumi Han, Delnur Anwar, Naoki Hama, Takuto Kobayashi, Hidefumi Suzuki, Hidehisa Takahashi, Haruka Wada, Ryo Otsuka, Muhammad Baghdadi, Ken-ichiro Seino
Inflammation and Regeneration, 40, 1, Springer Science and Business Media LLC, 2020年12月
研究論文（学術雑誌）,


Interleukin (IL)-34 acts as an alternative ligand for the colony-stimulating factor-1 receptor and controls the biology of myeloid cells, including survival, proliferation, and differentiation. IL-34 has been reported to be expressed in cancer cells and to promote tumor progression and metastasis of certain cancers via the promotion of angiogenesis and immunosuppressive macrophage differentiation. We have shown in our previous reports that targeting IL-34 in chemo-resistant tumors in vitro resulted in a remarkable inhibition of tumor growth. Also, we reported poor prognosis in patients with IL-34-expressing tumor. Therefore, blocking of IL-34 is considered as a promising therapeutic strategy to suppress tumor progression. However, the molecular mechanisms that control IL-34 production are still largely unknown.





IL-34 producing ovarian cancer cell line HM-1 was treated by bromodomain and extra terminal inhibitor JQ1. The mRNA and protein expression of IL-34 was evaluated after JQ1 treatment. Chromatin immunoprecipitation was performed to confirm the involvement of bromodomain-containing protein 4 (Brd4) in the regulation of the Il34 gene. Anti-tumor effect of JQ1 was evaluated in mouse tumor model.





We identified Brd4 as one of the critical molecules that regulate Il34 expression in cancer cells. Consistent with this, we found that JQ1 is capable of efficiently suppressing the recruitment of Brd4 to the promotor region of Il34 gene. Additionally, JQ1 treatment of mice bearing IL-34-producing tumor inhibited the tumor growth along with decreasing Il34 expression in the tumor.





The results unveiled for the first time the responsible molecule Brd4 that regulates Il34 expression in cancer cells and suggested its possibility as a treatment target.

A Clinical Trial With Adoptive Transfer of Ex Vivo-induced, Donor-specific Immune-regulatory Cells in Kidney Transplantation-A Second Report.
Ichiro Koyama, Hisashi Bashuda, Koichiro Uchida, Ken-Ichiro Seino, Sonoko Habu, Ichiro Nakajima, Shohei Fuchinoue, Ko Okumura, Satoshi Teraoka
Transplantation, 104, 11, 2415, 2423, 2020年11月, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Although the outcome of kidney transplantation (KTx) has improved, various adverse effects of immunosuppressants and chronic rejection aggravate the long-term prognosis of patients. Therefore, the induction of immune tolerance may be an effective therapeutic strategy. METHODS: A clinical trial aiming at immune tolerance induction was conducted in kidney transplant recipients from HLA mismatched living donors by infusing autologous donor-specific regulatory T cells (Treg). To obtain Treg, recipient's peripheral blood mononuclear cells were cocultured with irradiated donor cells in the presence of anti-CD80/CD86 monoclonal antibody for 2 weeks. For preconditioning, splenectomy + cyclophosphamide (CP) was employed in the first series (group A; n = 9). In group B, splenectomy was substituted by preadministration of rituximab (group B; n = 3). In the latest cases, rituximab + rabbit antithymocyte globulin was administered instead of cyclophosphamide (group C; n = 4). Twelve days after KTx, the cultured cells were intravenously infused, and immunosuppressants were gradually tapered thereafter. RESULTS: Although mixed lymphocyte reaction was remarkably suppressed in a donor-specific fashion, 6 out of 9 patients from group A, 1 out of 3 from group B, and 1 out of 4 from group C developed acute rejection within 1 year after KTx. Complete cessation of immunosuppression was not achieved, and a small dose of immunosuppressants was continued. CONCLUSIONS: The adoptive transfer of autologous ex vivo-expanded Treg is 1 of the options to possibly induce alloimmune hyporesponsiveness. However, in the present study, further regimen optimization is still required and should be the focus of future investigations.

Interleukin-34 contributes to poor prognosis in triple-negative breast cancer.
Nabeel Kajihara, Fumihito Kitagawa, Takuto Kobayashi, Haruka Wada, Ryo Otsuka, Ken-Ichiro Seino
Breast cancer (Tokyo, Japan), 27, 6, 1198, 1204, 2020年11月, ［国内誌］
英語, 研究論文（学術雑誌）, Triple-negative breast cancer (TNBC) is a subtype characterized by the absence of therapeutic targets. It shows rapid progression, higher relapse, and poor prognosis, so the establishment of an effective therapeutic target is required. We focused on interleukin-34 (IL-34) that is a novel cytokine relating to inflammation and tumorigenesis. It has been reported that IL-34 correlates with poor prognosis of various cancers. In this study, we evaluated the relationship of IL-34 and prognosis in TNBC using human clinical information and mice model. We found that IL-34 was highly expressed in TNBC, and the survival rate in TNBC was significantly lower in patients with high IL-34 expression. Furthermore, multivariate analysis revealed that IL-34 independently affects prognosis. In murine TNBC model, IL-34 deficiency in tumor cells decreased in vivo tumor growth and increased inflammatory cytokine production from macrophages. These results suggest that tumor-derived IL-34 creates a favorable environment for TNBC cells. Thus, we showed a novel pathological role of IL-34 in TNBC and the potential of IL-34 as a therapeutic target for it.

腫瘍開始細胞は炎症環境を形成し免疫細胞を老化させて抗腫瘍応答の低下を導くことで免疫健常個体における腫瘍発生を許容させる　　　　　　　　　　　　　　　
和田 はるか, 近藤 亨, 清野 研一郎
日本癌学会総会記事, 79回, OJ12, 8, (一社)日本癌学会, 2020年10月
英語

Interleukin-34 Limits the Therapeutic Effects of Immune Checkpoint Blockade
Naoki Hama, Takuto Kobayashi, Nanumi Han, Fumihito Kitagawa, Nabeel Kajihara, Ryo Otsuka, Haruka Wada, Hee-kyung Lee, Hwanseok Rhee, Yoshinori Hasegawa, Hideo Yagita, Muhammad Baghdadi, Ken-ichiro Seino
iScience, 23, 10, 101584, 101584, Elsevier BV, 2020年10月
研究論文（学術雑誌）

腫瘍由来Interleukin-34を標的とした新規がん治療法の確立　　　　　　　　　　　　　　　
羽馬 直希, 小林 拓斗, 韓 ナヌミ, 北川 郁人, 梶原 ナビール, 大塚 亮, 和田 はるか, Lee Hee-kyung, Rhee Hwanseok, 長谷川 嘉則, 八木田 秀雄, Baghdadi Muhammad, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 24回, 102, 102, 日本がん免疫学会, 2020年09月
日本語

IL-34が大腸がん患者の予後に与える影響の解析　　　　　　　　　　　　　　　
小林 拓斗, バグダーディ・ムハンマド, 韓 ナヌミ, 村田 智己, 大塚 亮, 和田 はるか, 宮城 洋平, 醍醐 弥太郎, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 24回, 104, 104, 日本がん免疫学会, 2020年09月
日本語

Establishment of an experimental model for MHC homo-to-hetero transplantation.
Tomoki Murata, Haruka Wada, Ryo Otsuka, Airi Sasaki, Hyuma Tsuji, Mizuho Itoh, Nanami Eguchi, Tatsuo Kawai, Ken-Ichiro Seino
Scientific reports, 10, 1, 13560, 13560, 2020年08月11日, ［国際誌］
英語, 研究論文（学術雑誌）, Preventing rejection is a major challenge in transplantation medicine, even when using pluripotent stem cell-derived grafts. In iPS cell (iPSC)-based transplantation, to reduce the risk of rejection, it is thought to be optimal that preparing the cells from donors whose human leukocyte antigen-haplotype are homozygous. Generally, this approach is referred to as major histocompatibility complex (MHC) homo-to-hetero transplantation, which is MHC-matched but minor antigen-mismatched. To investigate the immune response in the MHC homo-to-hetero transplantation, we established a murine experimental system in which MHC-matched but minor antigen-mismatched tissue (skin) grafts were transplanted into MHC-heterozygous recipients. Unexpectedly, only minor antigen-mismatched grafts were rejected at the same time points as rejection of fully allogeneic grafts. A vigorous anti-donor type T cell response was detected in vitro and conventional immunosuppressants targeting T cell activation had limited effects on controlling rejection. However, anti-donor antibodies were not detected only in the minor antigen-mismatched transplantation. This murine transplantation model can be used to further analyze immunological subjects for MHC homo-to-hetero iPSC-based transplantation.

Induction of macrophage-like immunosuppressive cells from common marmoset ES cells by stepwise differentiation with DZNep.
Hyuma Tsuji, Ryo Otsuka, Haruka Wada, Tomoki Murata, Airi Sasaki, Mizuho Itoh, Muhammad Baghdadi, Erika Sasaki, Ken-Ichiro Seino
Scientific reports, 10, 1, 12625, 12625, 2020年07月28日, ［国際誌］
英語, 研究論文（学術雑誌）, Recent progress in regenerative medicine has enabled the utilization of pluripotent stem cells (PSCs) as the resource of therapeutic cells/tissue. However, immune suppression is still needed when the donor-recipient combination is allogeneic. We have reported previously that mouse PSCs-derived immunosuppressive cells contribute to prolonged survival of grafts derived from the same mouse PSCs in allogeneic recipients. For its clinical application, a preclinical study using non-human primates such as common marmoset must be performed. In this study, we established the induction protocol of immunosuppressive cells from common marmoset ES cells. Although similar immunosuppressive macrophages could not be induced by same protocol as that for mouse PSCs, we employed an inhibitor for histone methyltransferase, DZNep, and succeeded to induce them. The DZNep-treated macrophage-like cells expressed several immunosuppressive molecules and significantly inhibited allogeneic mixed lymphocyte reaction. The immunosuppressive cells from non-human primate ESCs will help to establish an immunoregulating strategy in regenerative medicine using PSCs.

Steps towards COVID-19 suppression
Hideyuki Okano, Ken-Ichiro Seino
Inflammation and Regeneration, 40, 1, BioMed Central, 2020年06月22日
英語, 研究論文（学術雑誌）

【マクロファージの功罪-疾患病態誘導と制御におけるマクロファージの役割】がん・腫瘍 IL-34とマクロファージ
小林 拓斗, 大塚 亮, 清野 研一郎
医学のあゆみ, 273, 10, 993, 998, 医歯薬出版(株), 2020年06月
日本語, インターロイキン-34(IL-34)は、マクロファージコロニー刺激因子(M-CSF)に続いて単球およびマクロファージに発現するコロニー刺激因子1受容体(CSF-1R)と結合する第2のリガンドとして近年発見されたサイトカインである。IL-34は皮膚や神経系、赤脾髄に発現し、骨髄系細胞の生存、分化に関与し、また炎症性腸疾患や関節リウマチなどの疾患、および多数のがん種においては局所で発現が上昇することが知られている。興味深いことに、がん患者の腫瘍微小環境で産生されるIL-34はがん細胞の生存、増殖などに関与し、免疫抑制型マクロファージを誘導しがんに有利な腫瘍微小環境を構築するなど、腫瘍の悪性度に強く関わる可能性が示唆されている。その機能から、IL-34は新規治療標的、およびがん患者の予後を予測するためのバイオマーカーとして有効であると考えられ、研究が行われている。(著者抄録)

Interleukin-34 expression in ovarian cancer: a possible correlation with disease progression.
Hiraku Endo, Naoki Hama, Muhammad Baghdadi, Kozo Ishikawa, Ryo Otsuka, Haruka Wada, Hiroshi Asano, Daisuke Endo, Yosuke Konno, Tatsuya Kato, Hidemichi Watari, Akiko Tozawa, Nao Suzuki, Tomoyuki Yokose, Atsushi Takano, Hisamori Kato, Yohei Miyagi, Yataro Daigo, Ken-Ichiro Seino
International immunology, 32, 3, 175, 186, 2020年03月07日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Ovarian cancer is the second-most lethal gynecological malignancy and the seventh-commonest cause of cancer-related death in women around the world. Most of the ovarian cancer patients are diagnosed at advanced stages and suffer from recurrence after primary cytoreductive surgery and standard first-line chemotherapy. Thus, the successful management of ovarian cancer patients requires the identification of factors that contribute to progression and relapse. Interleukin-34 (IL-34) is a novel cytokine that acts as a tissue-specific ligand of colony-stimulating factor-1 receptor (CSF-1R). In cancer, IL-34 exerts pro-tumorigenic functions that promote tumor growth, metastasis, angiogenesis, immune suppression and therapeutic resistance. In this study, we evaluate the impact of IL-34 on progression and survival of ovarian cancer patients. First, IL-34 was found to be expressed in several human ovarian cancer cell lines and cancer tissues from patients. The expression of IL-34 was enhanced by cytotoxic chemotherapy in ovarian cancer cell lines and cancer tissues from chemotherapy-treated ovarian cancer patients. Importantly, high IL-34 expression correlated with worse progression-free survival (PFS) and overall survival in different cohorts. The assessment of PFS based on a combination between IL34 expression and other related genes such as CSF1R and CD163 helped further to reach more statistical significance compared with IL34 alone. Furthermore, in the murine ovarian cancer cell HM-1 in vivo model, it was suggested that IL-34-derived tumor cells was correlated with tumor progression and survival by modulating the immune environment. Collectively, these findings indicate a possible correlation between IL-34 expression and disease progression in ovarian cancer patients and the mouse model.

Efficient generation of thymic epithelium from induced pluripotent stem cells that prolongs allograft survival.
Ryo Otsuka, Haruka Wada, Hyuma Tsuji, Airi Sasaki, Tomoki Murata, Mizuho Itoh, Muhammad Baghdadi, Ken-Ichiro Seino
Scientific reports, 10, 1, 224, 224, 2020年01月14日, ［国際誌］
英語, 研究論文（学術雑誌）, The thymus plays a significant role in establishing immunological self-tolerance. Previous studies have revealed that host immune reaction to allogeneic transplants could be regulated by thymus transplantation. However, physiological thymus involution hinders the clinical application of these insights. Here, we report an efficient generation of thymic epithelial-like tissue derived from induced pluripotent stem cells (iPSCs) and its potential to regulate immune reaction in allogeneic transplantation. We established an iPSC line which constitutively expresses mouse Foxn1 gene and examined the effect of its expression during in vitro differentiation of thymic epithelial cells (TECs). We found that Foxn1 expression enhances the differentiation induction of cells expressing TEC-related cell surface molecules along with upregulation of endogenous Foxn1. iPSC-derived TECs (iPSC-TECs) generated T cells in nude recipient mice after renal subcapsular transplantation. Moreover, iPSC-TEC transplantation to immuno-competent recipients significantly prolonged the survival of allogeneic skin. Our study provides a novel concept for allogeneic transplantation in the setting of regenerative medicine.

Transcriptomic Features of T Cell-Barren Tumors Are Conserved Across Diverse Tumor Types.
Eric D Routh, Ashok K Pullikuth, Guangxu Jin, Julia Chifman, Jeff W Chou, Ralph B D'Agostino Jr, Ken-Ichiro Seino, Haruka Wada, Cristin G Print, Wei Zhang, Yong Lu, Lance D Miller
Frontiers in immunology, 11, 57, 57, 2020年, ［国際誌］
英語, 研究論文（学術雑誌）, Background: Understanding how tumors subvert immune destruction is essential to the development of cancer immunotherapies. New evidence suggests that tumors limit anti-tumor immunity by exploiting transcriptional programs that regulate intratumoral trafficking and accumulation of effector cells. Here, we investigated the gene expression profiles that distinguish immunologically "cold" and "hot" tumors across diverse tumor types. Methods: RNAseq profiles of tumors (n = 8,920) representing 23 solid tumor types were analyzed using immune gene signatures that quantify CD8+ T cell abundance. Genes and pathways associated with a low CD8+ T cell infiltration profile (CD8-Low) were identified by correlation, differential expression, and statistical ranking methods. Gene subsets were evaluated in immunotherapy treatment cohorts and functionally characterized in cell lines and mouse tumor models. Results: Among different cancer types, we observed highly significant overlap of genes enriched in CD8-Low tumors, which included known immunomodulatory genes (e.g., BMP7, CMTM4, KDM5B, RCOR2) and exhibited significant associations with Wnt signaling, neurogenesis, cell-cell junctions, lipid biosynthesis, epidermal development, and cancer-testis antigens. Analysis of mutually exclusive gene clusters demonstrated that different transcriptional programs may converge on the T cell-cold phenotype as well as predict for response and survival of patients to Nivo treatment. Furthermore, we confirmed that a top-ranking candidate belonging to the TGF-β superfamily, BMP7, negatively regulates CD8+ T cell abundance in immunocompetent murine tumor models, with and without anti-PD-L1 treatment. Conclusions: This study presents the first evidence that solid tumors of diverse anatomical origin acquire conserved transcriptional alterations that may be operative in the T cell-cold state. Our findings demonstrate the potential clinical utility of CD8-Low tumor-associated genes for predicting patient immunotherapy outcomes and point to novel mechanisms with potential for broad therapeutic exploitation.

Immune reaction and regulation in transplantation based on pluripotent stem cell technology.
Ryo Otsuka, Haruka Wada, Tomoki Murata, Ken-Ichiro Seino
Inflammation and regeneration, 40, 12, 12, 2020年, ［国際誌］
英語, 研究論文（学術雑誌）, The development of pluripotent stem cell (PSC)-based technologies provides us a new therapeutic approach that generates grafts for transplantation. In order to minimize the risk of immune reaction, the banking of induced pluripotent stem cells (iPSCs) from donors with homozygous human leukocyte antigen (HLA) haplotype is planned in Japan. Even though pre-stocked and safety validated HLA-homozygous iPSCs are selected, immunological rejection may potentially occur because the causes of rejection are not always due to HLA mismatches. A couple of studies concerning such immunological issues have reported that genetic ablation of HLA molecules from PSC combined with gene transduction of several immunoregulatory molecules may be effective in avoiding immunological rejection. Also, our research group has recently proposed a concept that attempts to regulate recipient immune system by PSC-derived immunoregulatory cells, which results in prolonged survival of the same PSC-derived allografts. PSC-based technologies enable us to choose a new therapeutic option; however, considering its safety from an immunological point of view should be of great importance for safe clinical translation of this technology.

Macrophage activation syndrome and COVID-19.
Ryo Otsuka, Ken-Ichiro Seino
Inflammation and regeneration, 40, 19, 19, 2020年, ［国際誌］
英語, 研究論文（学術雑誌）, An emerging, rapidly spreading coronavirus SARS-CoV-2 is causing a devastating pandemic. As we have not developed curative medicine and effective vaccine, the end of this life-threatening infectious disease is still unclear. Severe COVID-19 is often associated with hypercytokinemia, which is typically found in macrophage activation syndrome. SARS-CoV-2 infection causes this strong inflammation within the lung and propagates to respiratory and, ultimately, systemic organ malfunction. Although we have not fully understood the physiological and pathological aspects of COVID-19, current research progress indicates the effectiveness of anti-cytokine therapy. Here, we summarize macrophage activation syndrome and its possible contribution to COVID-19, and cytokine targeted attempts in severe COVID-19 cases.

Prognostic value of IL-34 in colorectal cancer patients.
Takuto Kobayashi, Muhammad Baghdadi, Nanumi Han, Tomoki Murata, Naoki Hama, Ryo Otsuka, Haruka Wada, Manabu Shiozawa, Tomoyuki Yokose, Yohei Miyagi, Atsushi Takano, Yataro Daigo, Ken-Ichiro Seino
Immunological medicine, 42, 4, 169, 175, 2019年12月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The mortality of colorectal cancer is expected to increase in some countries including the United States, which necessitates the identification of new molecules that help in prognosis assessment and survival improvement. In this brief report, we evaluated the potential of interleukin-34 (IL-34) as a prognostic factor in colorectal cancer. IL-34 was reported for the first time in 2008 as a novel cytokine that controls the biology of the myeloid cell lineage. Accumulating evidence suggests important roles for IL-34 in modifying the tumor microenvironment and enhancing therapeutic resistance of cancer. In this study, we found that IL-34 expression was detectable in various colorectal cancer cell lines in addition to primary cancer tissues from a cohort of Japanese colorectal cancer patients, ranging from high to absent. A Kaplan-Meier analysis showed that high expression of IL-34 correlated with poor survival of colorectal cancer patients. Importantly, in both univariate and multivariate analysis, high IL-34 expression correlated with unfavorable prognosis. A similar relationship between IL-34 expression and the poorer prognosis was also observed in a cohort of colorectal cancer patients registered at The Cancer Genome Atlas. Together, these findings indicate a potential role for IL-34 as a prognostic factor in colorectal cancer.

Flow cytometric identification and cell-line establishment of macrophages in naked mole-rats.
Wada H, Shibata Y, Abe Y, Otsuka R, Eguchi N, Kawamura Y, Oka K, Baghdadi M, Atsumi T, Miura K, Seino KI
Scientific reports, 9, 1, 17981, 17981, 2019年11月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Naked mole rats (NMRs) have extraordinarily long lifespans and anti-tumorigenic capability. Recent studies of humans and mice have shown that many age-related diseases, including cancer, are strongly correlated with immunity, and macrophages play particularly important roles in immune regulation. Therefore, NMR macrophages may contribute to their unique phenotypes. However, studies of the roles of macrophages are limited by material restrictions and the lack of an established experimental strategy. In this study, we developed a flow cytometric strategy to identify NMR macrophages. The NMR macrophages were extractable using an off-the-shelf anti-CD11b antibody, M1/70, and forward/side scatter data obtained by flow cytometry. NMR macrophages proliferated in response to human/mouse recombinant M-CSF and engulfed Escherichia coli particles. Interestingly, the majority of NMR macrophages exhibited co-staining with an anti-NK1.1 antibody, PK136. NK1.1 antigen crosslinking with PK136 results in mouse NK cell stimulation; similarly, NMR macrophages proliferated in response to NK1.1 antibody treatment. Furthermore, we successfully established an NMR macrophage cell line, NPM1, by transduction of Simian virus 40 early region that proliferated indefinitely without cytokines and retained its phagocytotic capacity. The NPM1 would contribute to further studies on the immunity of NMRs.

がん幹細胞はそれ自身が原炎症性細胞であり微小環境に免疫老化及び免疫抑制をもたらすことで健常動物における造腫瘍能を担保する(Tumor-initiating cell induce immuno-hyporesponsiveness following cellular senescence to Mφs guarantee its tumorigenesis)　　　　　　　　　　　　　　　
和田 はるか, バグダーディー・ムハンマド, 近藤 亨, 清野 研一郎
日本癌学会総会記事, 78回, J, 1042, (一社)日本癌学会, 2019年09月
英語

ATLLにおけるM-CSFR発現とM-CSFR阻害剤の有効性(Potential anti-lymphoma effect of M-CSFR inhibitor in adult T-cell leukemia/lymphoma)　　　　　　　　　　　　　　　
菰原 義弘, Muhammad Baghdadi, 清野 研一郎, 奥野 豊, 野坂 生郷, 松岡 雅雄
日本癌学会総会記事, 78回, P, 1014, 日本癌学会, 2019年09月
英語

がん幹細胞はそれ自身が原炎症性細胞であり微小環境に免疫老化及び免疫抑制をもたらすことで健常動物における造腫瘍能を担保する　　　　　　　　　　　　　　　
和田 はるか, 村田 智己, 大塚 亮, 森口 徹夫, バグダーディー・ムハンマド, 近藤 亨, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 23回, 131, 131, 日本がん免疫学会, 2019年07月
日本語

IL-34は免疫チェックポイント阻害剤の治療効果を抑制する(IL-34 limits the therapeutic effects of immune checkpoints blockade)　　　　　　　　　　　　　　　
バグダーディー・ムハンマド, 小林 拓斗, 羽馬 直希, 韓 ナヌミ, 和田 はるか, 八木田 秀雄, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 23回, 132, 132, 日本がん免疫学会, 2019年07月
英語

人工多能性幹細胞由来マクロファージ様免疫抑制細胞を用いたぶどう膜炎モデルの軽症化　　　　　　　　　　　　　　　
長谷 敬太郎, 南場 研一, 北市 伸義, 岩田 大樹, 辻 飛雄馬, 和田 はるか, 清野 研一郎, 石田 晋
日本眼科学会雑誌, 123, 臨増, 226, 226, (公財)日本眼科学会, 2019年03月
日本語

A role for IL-34 in osteolytic disease of multiple myeloma.
Baghdadi M, Ishikawa K, Nakanishi S, Murata T, Umeyama Y, Kobayashi T, Kameda Y, Endo H, Wada H, Bogen B, Yamamoto S, Yamaguchi K, Kasahara I, Iwasaki H, Takahata M, Ibata M, Takahashi S, Goto H, Teshima T, Seino KI
Blood advances, 3, 4, 541, 551, 2019年02月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Multiple myeloma (MM) is a hematological malignancy that grows in multiple sites of the axial skeleton and causes debilitating osteolytic disease. Interleukin-34 (IL-34) is a newly discovered cytokine that acts as a ligand of colony-stimulating factor-1 (CSF-1) receptor and can replace CSF-1 for osteoclast differentiation. In this study, we identify IL-34 as an osteoclastogenic cytokine that accelerates osteolytic disease in MM. IL-34 was found to be expressed in the murine MM cell line MOPC315.BM, and the expression of IL-34 was enhanced by stimulation with proinflammatory cytokines or by bone marrow (BM) stromal cells. MM-cell-derived IL-34 promoted osteoclast formation from mouse BM cells in vitro. Targeting Il34 by specific small interfering RNA impaired osteoclast formation in vitro and attenuated osteolytic disease in vivo. In BM aspirates from MM patients, the expression levels of IL-34 in CD138+ populations vary among patients from high to weak to absent. MM cell-derived IL-34 promoted osteoclast formation from human CD14+ monocytes, which was reduced by a neutralizing antibody against IL-34. Taken together, this study describes for the first time the expression of IL-34 in MM cells, indicating that it may enhance osteolysis and suggesting IL-34 as a potential therapeutic target to control pathological osteoclastogenesis in MM patients.

High co-expression of IL-34 and M-CSF correlates with tumor progression and poor survival in lung cancers
Muhammad Baghdadi, Hiraku Endo, Atsushi Takano, Kozo Ishikawa, Yosuke Kameda, Haruka Wada, Yohei Miyagi, Tomoyuki Yokose, Hiroyuki Ito, Haruhiko Nakayama, Yataro Daigo, Nao Suzuki, Ken-Ichiro Seino
Scientific Reports, 8, 1, Nature Publishing Group, 2018年12月01日, ［査読有り］
英語, 研究論文（学術雑誌）, Despite recent advances in diagnosis and treatment of lung cancers, the 5-year survival rate remains unsatisfactory, which necessitates the identification of novel factors that associates with disease progression and malignant degree for improving diagnostic and therapeutic strategies. Recent progress in cancer immunology research has unveiled critical roles for colony stimulating factor 1 receptor (CSF1R) in multiple aspects of the tumor microenvironment. CSF1R is expressed on tumor-associated macrophages (TAMs), and mediates important pro-tumorigenic functions. CSF1R also provides critical autocrine signals that promote cancer cell survival and proliferation. Activation of CSF1R can be achieved by two independent ligands
macrophage colony-stimulating factor (M-CSF) and interleukin 34 (IL-34). Accordingly, the expression of these ligands in cancer is expected to result in poor prognosis. In this study, we show that IL-34 and M-CSF expression correlates with poor survival in a cohort of lung cancer patients. Importantly, high co-expression of IL-34 and M-CSF associates with the poorest survival compared to cancers that show weak or absent expression of the two ligands. Furthermore, high expression of IL-34 and M-CSF associates with advanced stages of lung cancers. Together, these results indicate a correlation between IL-34/M-CSF expression with poor survival and disease progression in lung cancer patients.

iPS細胞由来組織移植に想定される拒絶反応の精査と制御法の検討
村田 智己, 和田 はるか, 大塚 亮, 辻 飛雄馬, 佐々木 愛里, 柴田 悠平, 安宅 司, 小林 拓斗, ムハンマド・バグダーディー, 清野 研一郎
移植, 53, 総会臨時, 485, 485, (一社)日本移植学会, 2018年09月
日本語

免疫健常個体において造腫瘍能を発揮する腫瘍開始細胞の免疫学的因子の同定(Tumor initiating cell in immunocompetent animal defined by immunological features)　　　　　　　　　　　　　　　
和田 はるか, Baghdadi Muhammad, 森口 徹生, 近藤 亨, 清野 研一郎
日本癌学会総会記事, 77回, 2327, 2327, (一社)日本癌学会, 2018年09月
英語

Interleukin-34, a comprehensive review.
Baghdadi M, Umeyama Y, Hama N, Kobayashi T, Han N, Wada H, Seino KI
Journal of leukocyte biology, 104, 5, 931, 951, 2018年08月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, IL-34 is a novel cytokine that was identified in 2008 in a comprehensive proteomic analysis as a tissue-specific ligand of CSF-1 receptor (CSF-1R). IL-34 exists in all vertebrates including fish, amphibians, birds, and mammals, showing high conservation among species. Structurally, IL-34 belongs to the short-chain helical hematopoietic cytokine family but shows no apparent consensus structural domains, motifs, or sequence homology with other cytokines. IL-34 is synthesized as a secreted homodimeric glycoprotein that binds to the extracellular domains of CSF-1R and receptor-type protein-tyrosine phosphatase-zeta (PTP-ζ) in addition to the chondroitin sulfate chains of syndecan-1. These interactions result in activating several signaling pathways that regulate major cellular functions, including proliferation, differentiation, survival, metabolism, and cytokine/chemokine expression in addition to cellular adhesion and migration. In the steady state, IL-34 contributes to the development and maintenance of specific myeloid cell subsets in a tissue-specific manner: Langerhans cells in the skin and microglia in the brain. In pathological conditions, changes in IL-34 expression-increased or decreased-are involved in disease pathogenesis and correlate with progression, severity, and chronicity. One decade after its discovery, IL-34 has been introduced as a newcomer to the big family of interleukins with specific physiological functions, critical pathological roles, and promising clinical applications in disease diagnosis and treatment. In this review, we celebrate the 10th anniversary of IL-34 discovery, introducing its biological characteristics, and discussing the importance of IL-34 signaling network in health and disease.

肺癌におけるIL-34と治療耐性、疾患進行、予後不良との関連(IL-34 correlates with therapeutic resistance, disease progression, and poor prognosis in lung cancers)　　　　　　　　　　　　　　　
バグダーディー・ムハンマド, 和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 22回, 111, 111, 日本がん免疫学会, 2018年07月
英語

がん細胞ワクチンの効果発揮には死細胞でなく「死にゆく」細胞が重要である　　　　　　　　　　　　　　　
和田 はるか, 安宅 司, 田中 睦乃, バグダーディー・ムハンマド, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 22回, 151, 151, 日本がん免疫学会, 2018年07月
日本語

Enhanced expression of IL-34 in an inflammatory cyst of the submandibular gland: a case report.
Baghdadi M, Ishikawa K, Endo H, Umeyama Y, Ataka T, Wada H, Oyamada Y, Hyakushima N, Seino KI
Inflammation and regeneration, 38, 12, 12, 12, 2018年07月, ［査読有り］, ［国際誌］
英語, Background: Cysts of the salivary glands are common lesions that occur in the context of various etiologies. Although the diagnostic importance of cysts in salivary gland diseases has been well studied, molecular mechanisms that control the related pathological process remain largely unknown. IL-34 is a novel cytokine that was discovered recently as a tissue-specific ligand of colony stimulating factor-1 receptor. Since its discovery, accumulating evidence has revealed emerging roles of IL-34 in various pathological conditions and has been suggested to correlate remarkably with inflammation. In this study, we report a medical case of an inflammatory cyst within the submandibular gland, through which evaluating the possible involvement of IL-34 in salivary gland disorders. Case presentation: A 37-year-old male patient suffered from a sudden swelling in the right submandibular region, started initially small and had gradually increased in size to reach 3-4 cm in 1 week, accompanied by pain and local fever. Ultrasonography and MRI imaging revealed the existence of a well-defined cystic lesion with sharp borders measuring 39.8 mm × 19.7 mm within the right submandibular gland. The cyst was removed surgically, and the diagnostic decision was determined based on histopathological observations as an inflammatory cyst in the submandibular gland. Sections were generated from different regions of the surgically resected inflammatory cyst and used to examine IL-34 expression by immunohistochemistry compared to normal salivary gland tissues. Immunohistochemical staining showed enhanced expression of IL-34 in the ductal epithelial cells and endothelial cells of blood vessels, with a tendency to be accompanied with high infiltration of immune cells, which suggests a possible involvement of IL-34 in the pathogenesis of salivary gland inflammation. Conclusions: In this report, we introduce interesting findings of enhanced IL-34 expression in a case of an inflamed submandibular gland. Our findings emphasize the pathological roles of IL-34 as an inflammation amplifier and angiogenic enhancer in inflammatory conditions, such as in salivary gland disorders.

免疫療法の新たなステージへ ～Interleukin-34標的薬の可能性～　　　　　　　　　　　　　　　
羽馬直希, 韓ナヌミ、ムハンマド・バグダーディー, 清野研一郎
日本免疫治療学会誌, 6, 3, 6, 2018年06月, ［査読有り］, ［招待有り］
日本語

Enhanced IL-34 expression in Nivolumab-resistant metastatic melanoma.
Han N, Baghdadi M, Ishikawa K, Endo H, Kobayashi T, Wada H, Imafuku K, Hata H, Seino KI
Inflammation and regeneration, 38, 3, 3, 3, 2018年03月, ［査読有り］, ［国際誌］
英語, BACKGROUND: Immunotherapies that target immune-checkpoint molecules such PD-1 have helped to achieve durable responses in melanoma treatment. However, 25% of melanoma patients who showed objective responses to PD-1 blockade develop resistance and suffer from disease progression and ultimately death, which necessitates the identification of related resistance mechanisms.IL-34 is a cytokine that controls the biology of myeloid cell lineage through binding to CSF-1R. IL-34 is importantly involved in the pathogenesis of various diseases. In cancer, the expression of IL-34 has been suggested to associate with tumor growth, metastasis, angiogenesis, and therapeutic resistance such as in lung cancers and malignant pleural mesotheliomas. In this study, we evaluate the possible involvement of IL-34 in immunotherapeutic resistance. CASE PRESENTATION: Melanoma resection species were obtained from a patient who developed a refractory melanoma against immunotherapy with Nivolumab, and stained with anti-IL-34, anti-melanoma antigens and anti-CD163 antibody. Staining of these markers was compared between primary or metastatic refractory melanoma tissues. Immunohistochemistry staining of melanoma tissues showed an enhanced expression of IL-34 in metastatic refractory melanoma compared to primary melanoma tissues, which correlates with increased frequencies of CD163+ macrophages. CONCLUSION: We introduce for the first time a clinical case of a patient with metastatic refractory melanoma that acquired resistance to anti-PD-1 immunotherapy, showing an enhanced expression of IL-34 in refractory melanoma tissues.

Potential anti-lymphoma effect of M-CSFR inhibitor in adult T-cell leukemia/lymphoma.
Yoshihiro Komohara, Osamu Noyori, Yoichi Saito, Hiroto Takeya, Muhammad Baghdadi, Fumihito Kitagawa, Naoki Hama, Kozo Ishikawa, Yutaka Okuno, Kisato Nosaka, Ken-Ichiro Seino, Masao Matsuoka, Shinya Suzu
Journal of clinical and experimental hematopathology : JCEH, 58, 4, 152, 160, 2018年, ［査読有り］, ［国内誌］
英語, 研究論文（学術雑誌）, The c-fms proto-oncogene is also known as macrophage colony stimulating factor receptor (M-CSFR) or colony-stimulating factor-1 receptor (CSF-1R), and is expressed on several types of malignant tumor cells and myeloid cells. In the present study, we found that overexpression of M-CSFR was present in adult T-cell leukemia/lymphoma (ATLL) cases. M-CSFR signaling was associated with lymphoma cell proliferation, and M-CSFR inhibition induced apoptosis in lymphoma cells. The ATLL cell line ATL-T expressed M-CSF/CSF-1 and interleukin (IL)-34, which are both M-CSFR ligands. M-CSF and IL-34 expression was seen in ATLL cases, and co-expression of these ligands was detected in 11 of 13 ATLL cases. M-CSFR inhibition suppressed programmed death-1 and -2 ligand in ATL-T cells and macrophages stimulated with conditioned medium from ATL-T cells. Thus, an M-CSFR inhibitor may be useful as additional therapy against ATLL due to direct and indirect mechanisms.

Interleukin 34, from pathogenesis to clinical applications
Muhammad Baghdadi, Hiraku Endo, Yoshino Tanaka, Haruka Wada, Ken-ichiro Seino
CYTOKINE, 99, 139, 147, ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 2017年11月, ［査読有り］
英語, Interleukin-34 (IL-34) is a hematopoietic cytokine that was described for the first time in 2008 as a second ligand of CSF1R in addition to M-CSF. IL-34 and M-CSF share no sequence homology, but have similar functions, affecting the biology of myeloid cell lineage. In contrast to M-CSF, IL-34 shows unique signaling and expression patterns. Physiologically, IL-34 expression is restricted to epidermis and CNS, acting as a regulator of Langerhans cells and microglia, respectively. However, IL-34 expression can be induced and regulated by NF-kappa B under pathological conditions. Importantly, growing evidence indicates a correlation between IL-34 and disease severity, chronicity and progression. In addition to its promising roles as a novel diagnostic and prognostic bio-marker of disease, IL-34 may also serve as a powerful target for therapeutic intervention. Here, we review the current knowledge regarding the emerging roles of IL-34 in disease, and focus on the clinical applications of IL-34 in medicine.

Intravenous dendritic cell administration enhances suppression of lung metastasis induced by carbon-ion irradiation
Ken Ando, Hidetoshi Fujita, Akihiro Hosoi, Liqiu Ma, Masaru Wakatsuki, Ken-ichiro Seino, Kazuhiro Kakimi, Takashi Imai, Takashi Shimokawa, Takashi Nakano
JOURNAL OF RADIATION RESEARCH, 58, 4, 446, 455, OXFORD UNIV PRESS, 2017年07月, ［査読有り］
英語, 研究論文（学術雑誌）, Carbon-ion radiotherapy (CIRT) is an advanced radiotherapy and has achieved good local control, even in tumors that are resistant to conventional photon beam radiotherapy (PBRT). However, distant metastasis control is an important issue. Recently, the combination of radiotherapy and immunotherapy has attracted the attention. In immunotherapy, dendritic cells (DCs) play a pivotal role in the anti-tumor immune system. However, the mechanisms underlying the combination therapy of DCs and radiotherapy have been unclear. In the present study, we evaluated anti-metastatic effects of this combination therapy, focused on the irradiation type and the route of DC administration, using a mouse model. C3H/He mice bearing NR-S1 cells were treated with CIRT or PBRT, using biologically equivalent doses. Subsequently, DCs were administered intratumorally (IT) or intravenously (IV). IV and IT DC administrations combined with CIRT to the local tumor, but not alone, significantly suppressed pulmonary metastasis, whereas the combination of DCs with PBRT suppressed metastasis at a relatively higher dose. Additionally, the anti-metastatic effect was greater in IV DC administration compared with in IT DC administration in both CIRT and PBRT. The expression levels of CD40 and IL-12 in DCs were significantly increased after co-culturing with CIRT-treated NR-S1 cells. In addition, IV administration of those co-cultured DCs significantly suppressed pulmonary metastasis. Furthermore, ecto-calreticulin levels from CIRT-treated NR-S1 cells significantly increased compared with those of a PBRT-treated tumor. Taken together, these results suggest that local CIRT combined with IV DCs augments an immunogenicity of the tumor cells by ecto-calreticulin expression and the maturation of DCs to stimulate anti-tumor immunity to decrease lung metastases.

マウスiPS細胞から誘導した制御性マクロファージ様細胞によるアログラフト生着延長の検討
佐々木 元, 和田 はるか, 森田 研, 清野 研一郎, 篠原 信雄
泌尿器外科, 30, 6, 1067, 1067, 医学図書出版(株), 2017年06月
日本語

化学療法誘発性のIL-34はオートクラインの機序により化学療法耐性腫瘍細胞の生存を促進する(Chemotherapy-induced IL-34 promotes survival of chemoresistant cancer cells via autocrine mechanism)　　　　　　　　　　　　　　　
Putra Wira Eka, Baghdadi Muhammad, 和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 21回, 115, 115, 日本がん免疫学会, 2017年06月
英語

腫瘍始原細胞の免疫学的な特徴(Immunological features of tumor initiating cells)　　　　　　　　　　　　　　　
和田 はるか, Baghdadi Muhammad, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 21回, 121, 121, 日本がん免疫学会, 2017年06月
英語

多発性骨髄腫においてIL-34は破骨細胞の形成を促進し骨疾患を増悪化させる(IL-34 promotes ostelcclast formation and enhances bone disease in multiple myeloma)　　　　　　　　　　　　　　　
Baghdadi Muhammad, 石川 浩三, 和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 21回, 145, 145, 日本がん免疫学会, 2017年06月
英語

Chemotherapy-Induced IL34 Enhances Immunosuppression by Tumor-Associated Macrophages and Mediates Survival of Chemoresistant Lung Cancer Cells
Muhammad Baghdadi, Haruka Wada, Sayaka Nakanishi, Hirotake Abe, Nanumi Han, Wira Eka Putra, Daisuke Endo, Hidemichi Watari, Noriaki Sakuragi, Yasuhiro Hida, Kichizo Kaga, Yohei Miyagi, Tomoyuki Yokose, Atsushi Takano, Yataro Daigo, Ken-ichiro Seino
CANCER RESEARCH, 76, 20, 6030, 6042, AMER ASSOC CANCER RESEARCH, 2016年10月, ［査読有り］
英語, 研究論文（学術雑誌）, The ability of tumor cells to escape immune destruction and their acquired resistance to chemotherapy are major obstacles to effective cancer therapy. Although immune checkpoint therapies such as anti-PD-1 address these issues in part, clinical responses remain limited to a subpopulation of patients. In this report, we identified IL34 produced by cancer cells as a driver of chemoresistance. In particular, we found that IL34 modulated the functions of tumor-associated macrophages to enhance local immunosuppression and to promote the survival of chemoresistant cancer cells by activating AKT signaling. Targeting IL34 in chemoresistant tumors resulted in a remarkable inhibition of tumor growth when accompanied with chemotherapy. Our results define a pathogenic role for IL34 in mediating immunosuppression and chemoresistance and identify it as a tractable target for anticancer therapy. (C) 2016 AACR.

Transcriptional regulator Bhlhe40 works as a cofactor of T-bet in the regulation of IFN-γ production in iNKT cells.
Kanda M, Yamanaka H, Kojo S, Usui Y, Honda H, Sotomaru Y, Harada M, Taniguchi M, Suzuki N, Atsumi T, Wada H, Baghdadi M, Seino K
Proceedings of the National Academy of Sciences of the United States of America, 113, 24, E3394, E3402, NATL ACAD SCIENCES, 2016年06月, ［査読有り］
英語, 研究論文（学術雑誌）, Invariant natural killer T (iNKT) cells are a subset of innate-like T cells that act as important mediators of immune responses. In particular, iNKT cells have the ability to immediately produce large amounts of IFN-gamma upon activation and thus initiate immune responses in various pathological conditions. However, molecular mechanisms that control IFN-gamma production in iNKT cells are not fully understood. Here, we report that basic helix-loop-helix transcription factor family, member e40 (Bhlhe40), is an important regulator for IFN-gamma production in iNKT cells. Bhlhe40 is highly expressed in stage 3 thymic iNKT cells and iNKT1 subsets, and the level of Bhlhe40 mRNA expression is correlated with Ifng mRNA expression in the resting state. Although Bhlhe40-deficient mice show normal iNKT cell development, Bhlhe40-deficient iNKT cells show significant impairment of IFN-gamma production and antitumor effects. Bhlhe40 alone shows no significant effects on Ifng promoter activities but contributes to enhance T-box transcription factor Tbx21 (T-bet)-mediated Ifng promoter activation. Chromatin immunoprecipitation analysis revealed that Bhlhe40 accumulates in the T-box region of the Ifng locus and contributes to histone H3-lysine 9 acetylation of the Ifng locus, which is impaired without T-bet conditions. These results indicate that Bhlhe40 works as a cofactor of T-bet for enhancing IFN-gamma production in iNKT cells.

Tumour resistance in induced pluripotent stem cells derived from naked mole-rats
Shingo Miyawaki, Yoshimi Kawamura, Yuki Oiwa, Atsushi Shimizu, Tsuyoshi Hachiya, Hidemasa Bono, Ikuko Koya, Yohei Okada, Tokuhiro Kimura, Yoshihiro Tsuchiya, Sadafumi Suzuki, Nobuyuki Onishi, Naoko Kuzumaki, Yumi Matsuzaki, Minoru Narita, Eiji Ikeda, Kazuo Okanoya, Ken-ichiro Seino, Hideyuki Saya, Hideyuki Okano, Kyoko Miura
NATURE COMMUNICATIONS, 7, 11471, NATURE PUBLISHING GROUP, 2016年05月, ［査読有り］
英語, 研究論文（学術雑誌）, The naked mole-rat (NMR, Heterocephalus glaber), which is the longest-lived rodent species, exhibits extraordinary resistance to cancer. Here we report that NMR somatic cells exhibit a unique tumour-suppressor response to reprogramming induction. In this study, we generate NMR-induced pluripotent stem cells (NMR-iPSCs) and find that NMR-iPSCs do not exhibit teratoma-forming tumorigenicity due to the species-specific activation of tumour-suppressor alternative reading frame (ARF) and a disruption mutation of the oncogene ES cell-expressed Ras (ERAS). The forced expression of Arf in mouse iPSCs markedly reduces tumorigenicity. Furthermore, we identify an NMR-specific tumour-suppression phenotype-ARF suppression-induced senescence (ASIS)-that may protect iPSCs and somatic cells from ARF suppression and, as a consequence, tumorigenicity. Thus, NMR-specific ARF regulation and the disruption of ERAS regulate tumour resistance in NMR-iPSCs. Our findings obtained from studies of NMR-iPSCs provide new insight into the mechanisms of tumorigenicity in iPSCs and cancer resistance in the NMR.

多発性骨髄腫由来IL-34による破骨細胞誘導と骨破壊性疾患に対する影響
バグダーディ・ムハンマド, 中西 沙耶香, 和田 はるか, 清野 研一郎
International Journal of Myeloma, 6, 2, 125, 125, (一社)日本骨髄腫学会, 2016年04月
日本語

Identification of a highly immunogenic mouse breast cancer sub cell line, 4T1-S
Hirotake Abe, Haruka Wada, Muhammad Baghdadi, Sayaka Nakanishi, Yuu Usui, Takahiro Tsuchikawa, Toshiaki Shichinohe, Satoshi Hirano, Ken-ichiro Seino
HUMAN CELL, 29, 2, 58, 66, SPRINGER JAPAN KK, 2016年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Cancer vaccines serve as a promising clinical immunotherapeutic strategy that help to trigger an effective and specific antitumor immune response compared to conventional therapies. However, poor immunogenicity of tumor cells remains a major obstacle for clinical application, and developing new methods to modify the immunogenicity of tumor cells may help to improve the clinical outcome of cancer vaccines. 4T1 mouse breast cancer cell line has been known as poorly immunogenic and highly metastatic cell line. Using this model, we identified a sub cell line of 4T1-designated as 4T1-Sapporo (4T1-S)-which shows immunogenic properties when used as a vaccine against the same line. In 4T1-S-vaccinated mice, subcutaneous injection of 4T1-S resulted in an antitumor inflammatory response represented by significant enlargement of draining lymph nodes, accompanied with increased frequencies of activated CD8 T cells and a subpopulation of myeloid cells. Additionally, 4T1-S vaccine was ineffective to induce tumor rejection in nude mice, which importantly indicate that 4T1-S vaccine rely on T cell response to induce tumor rejection. Further analysis to identify mechanisms that control tumor immunogenicity in this model may help to develop new methods for improving the efficacies of clinical cancer vaccines.

Identification of a highly immunogenic mouse breast cancer sub cell line, 4t1-s
Hirotake Abe, Haruka Wada, Muhammad Baghdadi, Sayaka Nakanishi, Yuu Usui, Takahiro Tsuchikawa, Toshiaki Shichinohe, Satoshi Hirano, Ken-Ichiro Seino
Human Cell, 29, 2, 58, 66, Springer Tokyo, 2016年, ［査読有り］
英語, 研究論文（学術雑誌）, Cancer vaccines serve as a promising clinical immunotherapeutic strategy that help to trigger an effective and specific antitumor immune response compared to conventional therapies. However, poor immunogenicity of tumor cells remains a major obstacle for clinical application, and developing new methods to modify the immunogenicity of tumor cells may help to improve the clinical outcome of cancer vaccines. 4T1 mouse breast cancer cell line has been known as poorly immunogenic and highly metastatic cell line. Using this model, we identified a sub cell line of 4T1—designated as 4T1-Sapporo (4T1-S)—which shows immunogenic properties when used as a vaccine against the same line. In 4T1-S-vaccinated mice, subcutaneous injection of 4T1-S resulted in an antitumor inflammatory response represented by significant enlargement of draining lymph nodes, accompanied with increased frequencies of activated CD8 T cells and a sub-population of myeloid cells. Additionally, 4T1-S vaccine was ineffective to induce tumor rejection in nude mice, which importantly indicate that 4T1-S vaccine rely on T cell response to induce tumor rejection. Further analysis to identify mechanisms that control tumor immunogenicity in this model may help to develop new methods for improving the efficacies of clinical cancer vaccines.

Invariant natural killer T cell deficiency leads to the development of spontaneous liver inflammation dependent on γδT cells in mice.
Nishio K, Miyagi T, Tatsumi T, Mukai K, Yokoyama Y, Yoshioka T, Sakamori R, Hikita H, Kodama T, Shimizu S, Shigekawa M, Nawa T, Yoshihara H, Hiramatsu N, Yamanaka H, Seino K, Takehara T
Journal of gastroenterology, 50, 11, 1124, 1133, SPRINGER JAPAN KK, 2015年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Immune tolerance is maintained in the liver, and perturbation of tolerance can lead to immune-mediated liver diseases such as autoimmune hepatitis (AIH). Invariant natural killer T (iNKT) cells and gamma delta T cells have been shown to maintain immune homeostasis as regulatory cells and to play pathogenic roles in immune-mediated diseases as effector cells. We hypothesized that iNKT cells and gamma delta T cells are involved in the maintenance of hepatic immune tolerance and immune-mediated liver disease.
We measured liver inflammation and the cytokine profiles of liver mononuclear cells in BALB/c wild-type (WT) mice and BALB/c J alpha 18-deficient (KO) mice lacking iNKT cells. We also examined the role of gamma delta T cells in AIH using liver tissue from AIH patients and control subjects.
Spontaneous liver inflammation, hepatocyte damage, and anti-nuclear-antibody production occurred in J alpha 18 KO mice but not in WT mice. Furthermore, liver mononuclear cells from J alpha 18 KO mice, but not those from WT mice, produced interleukin-17 (IL-17). gamma delta T cells were the primary producers of the cytokine, and they were more abundant in the livers of J alpha 18 KO mice than in those of WT mice. In J alpha 18 KO mice, the administration of anti-gamma delta T-cell-receptor antibody abolished liver inflammation, hepatocyte damage, and IL-17 production. gamma delta T cells accumulated in the livers of AIH patients but not in those of the control subjects.
Our results suggest a protective role for iNKT cells, a pathologic role for gamma delta T cells, and an association between these cells in the pathogenesis of AIH.

New Immunosuppressive Cell Therapy to Prolong Survival of Induced Pluripotent Stem Cell-Derived Allografts
Hajime Sasaki, Haruka Wada, Muhammad Baghdadi, Hyuma Tsuji, Ryo Otsuka, Ken Morita, Nobuo Shinohara, Ken-ichiro Seino
TRANSPLANTATION, 99, 11, 2301, 2310, LIPPINCOTT WILLIAMS & WILKINS, 2015年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. Induced pluripotent stem cell (iPSC) technology provides new opportunities in regenerative medicine to generate grafts from donors for transplantation. However, particularly when allogeneic iPSCs are used, immune suppression is required to avoid rejection of iPSC-derived grafts. In this study, we examine a concept that protection of iPSCs-derived allografts can be achieved when transplantation is accompanied with the administration of immunosuppressive cells generated from the same iPSCs resource. Methods. Mouse iPSCs were differentiated into immunosuppressive cells by a culture protocol using granulocyte macrophage-colony-stimulating factor, macrophage-colony-stimulating factor, IL-4, and lipopolysaccharide. Adherent clusters were collected and examined for the ability to suppress allogeneic T-and B-cell responses, as well as for the contribution to prolonged allogeneic graft survival in transplantation models. Results. Myeloid cells with immunosuppressive features were successfully induced from iPSCs, and thus referred to as iPSC-derived suppressor cells (iPS-SCs). The iPS-SCs resemble macrophages in terms of cell surface molecules and gene expressions. Furthermore, iPS-SCs efficiently suppressed allogeneic T-and B-cell proliferation in a nitric oxide-dependent manner, and iPS-SCs were found to suppress alloantibody production and prolong substantially the survival of iPSC-derived grafts, such as embryoid bodies and cardiomyocytes, in in vivo allogeneic transplantation models. Conclusions. A certain fraction of macrophage-like cells with immunosuppressive functions can be generated from donor iPSCs, which contribute to the prolonged survival of grafts derived from the same iPSCs in allogeneic recipients. These results suggest a new immunosuppressive strategy of combined donor iPSC-derived graft and immunosuppressive cell transplantation in regenerative medicine using iPSCs.

Generation of T-lineage cells from iPS cells and its application
Haruka Wada, Muhammad Baghdadi, Ken-Ichiro Seino
Hematopoietic Differentiation of Human Pluripotent Stem Cells, 81, 90, Springer Netherlands, 2015年08月25日, ［査読有り］
英語, 論文集(書籍)内論文, The development of cell reprogramming technology will alter many clinical situations, including immunotherapy. One instance is donor lymphocyte infusion (DLI) therapy. DLI is an effective therapy for lymphoma or leukemia patient for the inhibition of relapses after bone marrow transplantation (BMT). However, DLI treatment depends on the availability of T lymphocytes isolated from a donor for the generation of lymphocytes. The application of cell reprogramming technology facilitates the generation of lymphocytes without donor apheresis, offering a sustainable and repeatable DLI therapy. In this chapter, we show an instance of this new age immunotherapy that utilizes cell reprogramming technology.

Myeloid molecular characteristics of human γδ T cells support their acquisition of tumor antigen-presenting capacity.
Muto M, Baghdadi M, Maekawa R, Wada H, Seino K
Cancer immunology, immunotherapy : CII, 64, 8, 941, 949, SPRINGER, 2015年08月, ［査読有り］
英語, 研究論文（学術雑誌）, Human T cells expressing gamma delta T cell receptor have a potential to show antigen-presenting cell-like phenotype and function upon their activation. However, the mechanisms that underlie the alterations in human gamma delta T cells remain largely unclear. In this study, we have investigated the molecular characteristics of human gamma delta T cells related to their acquisition of antigen-presenting capacity in comparison with activated alpha beta T cells. We found that activated gamma delta but not alpha beta T cells upregulated cell surface expression of a scavenger receptor, CD36, which seemed to be mediated by signaling through mitogen-activated protein kinase and/or NF-kappa B pathways. Confocal microscopical analysis revealed that activated gamma delta T cells can phagocytose protein antigens. Activated gamma delta T cells could induce tumor antigen-specific CD8(+) T cells using both apoptotic and live tumor cells as antigen resources. Furthermore, we detected that C/EBP alpha, a critical transcription factor for the development of myeloid-lineage cells, is expressed much higher in gamma delta T cells than in alpha beta T cells. These results unveiled the molecular mechanisms for the elicitation of antigen-presenting functions in gamma delta T cells and would also help designing new approaches for gamma delta T cell-mediated human cancer immunotherapy.

Chemotherapy-Derived Inflammatory Responses Accelerate the Formation of Immunosuppressive Myeloid Cells in the Tissue Microenvironment of Human Pancreatic Cancer
Shintaro Takeuchi, Muhammad Baghdadi, Takahiro Tsuchikawa, Haruka Wada, Toru Nakamura, Hirotake Abe, Sayaka Nakanishi, Yuu Usui, Kohtaro Higuchi, Mizuna Takahashi, Kazuho Inoko, Syoki Sato, Hironobu Takano, Toshiaki Shichinohe, Ken-ichiro Seino, Satoshi Hirano
CANCER RESEARCH, 75, 13, 2629, 2640, AMER ASSOC CANCER RESEARCH, 2015年07月, ［査読有り］
英語, 研究論文（学術雑誌）, Pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic malignancies. PDAC builds a tumor microenvironment that plays critical roles in tumor progression and metastasis. However, the relationship between chemotherapy and modulation of PDAC-induced tumor microenvironment remains poorly understood. In this study, we report a role of chemotherapy-derived inflammatory response in the enrichment of PDAC microenvironment with immunosuppressive myeloid cells. Granulocyte macrophage colony-stimulating factor (GM-CSF) is a major cytokine associated with oncogenic KRAS in PDAC cells. GM-CSF production was significantly enhanced in various PDAC cell lines or PDAC tumor tissues from patients after treatment with chemotherapy, which induced the differentiation of monocytes into myeloid-derived suppressor cells (MDSC). Furthermore, blockade of GM-CSF with monoclonal antibodies helped to restore T-cell proliferation when cocultured with monocytes stimulated with tumor supernatants. GM-CSF expression was also observed in primary tumors and correlated with poor prognosis in PDAC patients. Together, these results describe a role of GM-CSF in the modification of chemotherapy-treated PDAC microenvironment and suggest that the targeting of GM-CSF may benefit PDAC patients' refractory to current anticancer regimens by defeating MDSC-mediated immune escape. (C)2015 AACR.

多能性幹細胞を用いた再生医療における新しい免疫制御法
和田はるか, 工藤浩也, 佐々木元, Muhammad Baghdadi, 清野研一郎
日本臨床, 73, 5, 90, 95, 2015年06月, ［査読有り］, ［招待有り］
日本語

マウスiPS細胞から誘導した制御性マクロファージ様細胞によるiPS細胞由来アログラフトの生着延長効果
佐々木 元, 和田 はるか, 森田 研, 清野 研一郎, 篠原 信雄
泌尿器外科, 28, 5, 1012, 1012, 医学図書出版(株), 2015年05月
日本語

マウスiPS細胞から誘導した制御性マクロファージ様細胞によるアログラフト生着延長の検討　　　　　　　　　　　　　　　
佐々木 元, 和田 はるか, 森田 研, 清野 研一郎, 篠原 信雄
日本泌尿器科学会総会, 103回, 518, 518, (一社)日本泌尿器科学会総会事務局, 2015年04月
日本語

New immunosuppressive strategies for transplantation based on pluripotent stem cell (PSC)-derived immunoregulatory cells
Stem Cell and Translational Investigation, 2015年02月, ［査読有り］

α-MSH stimulation contributes to TGF-β1 production via MC1R-MITF signaling pathway in melanoma cell
Hayashi Erika, Hachiya Kaori, Kojo Satoshi, Baghdadi Muhammad, Takeuchi Shintaro, Yamanaka Hiroyuki, Abe Hirotak, Wada Haruka, Seino Ken-ichiro
Inflammation and Regeneration, 35, 5, 244, 254, 日本炎症・再生医学会, 2015年
日本語, Transforming growth factor-β (TGF-β) is a multifunctional cytokine that play critical roles in melanoma progression. Although the impact of TGF-β signaling on melanoma progression has been well characterized, little is known about the molecular mechanisms that control TGF-β production in melanoma cells. In this study, we describe a novel role for Melanocortin Receptor 1 (MC1R) in the regulation of TGF-β production. MC1R is a cell surface endocytic receptor expressed in melanoma cells and serves as a receptor for α-Melanocyte Stimulating Hormone (α-MSH). The activation of MC1R with α-MSH resulted in increased levels of TGF-β, which was mediated by ERK1/2 and p38 signaling pathways. Furthermore, Microphthalmia Transcription Factor (MITF), the master regulator of melanocytes, was found to act downstream of MC1R to regulate TGF-β production. Targeting of MC1R-MITF axis was effective to decrease TGF-β production, and resulted in delayed tumor growth of B16 melanoma in vivo. Collectively, these results give new insight into the molecular mechanisms that control TGF-β production in melanoma cells.

New immune regulation strategy in the age of regenerative medicine using pluripotent stem cells
Wada Haruka, Kudo Hiroya, Sasaki Hajime, Baghdadi Muhammad, Seino Ken-ichir
Inflammation and Regeneration, 35, 5, 238, 243, 日本炎症・再生医学会, 2015年
日本語, Recent progress of manipulating pluripotent stem cells expands possibilities of regenerative medicine and opens novel transplantation medicine. However, in many cases of these medicines, the relationship between therapeutic cells and recipients would be allogeneic. In this context, we proposed new concept of immune regulation therapy in new-age medicine using pluripotent stem cells. In our concept, not only grafts but also immune regulating cells are generated from pluripotent stem cells by exertion of its pluripotency. We have recently developed immune suppressive macrophage-like cells from pluripotent stem cells. These cells suppressed allo-antigen stimulated T cell proliferation in an iNOS dependent manner. Furthermore, these immune suppressive macrophage-like cells derived from pluripotent stem cells prolonged survival of grafts derived from same pluripotent stem cells in allogeneic recipients. Thus, series of our study proved the efficacy of our new immune regulating strategy in the age of regenerative medicine which utilize pluripotent stem cells as a therapeutic cell source.

Induction of Macrophage-Like Immunosuppressive Cells from Mouse ES Cells That Contribute to Prolong Allogeneic Graft Survival
Hiroya Kudo, Haruka Wada, Hajime Sasaki, Hyuma Tsuji, Ryo Otsuka, Muhammad Baghdadi, Satoshi Kojo, Tatsuya Chikaraishi, Ken-ichiro Seino
PLOS ONE, 9, 10, e111826, PUBLIC LIBRARY SCIENCE, 2014年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Recent progress in regenerative medicine has enabled the utilization of pluripotent stem cells (PSCs) such as embryonic stem cells (ESCs) as a donor resource for transplantation. However, immune suppression is still needed when the donor-recipient combination is allogeneic. Protection of ESCs-derived grafts from host immune response might be achieved thought the utilization of immunosuppressive cells generated from ESCs. In the present study, we show that a certain fraction of immunosuppressive cells can be generated from ESCs and help to suppress immune response against allogeneic grafts. ESCs-derived suppressor cells (ES-SCs) resembled macrophages in terms of cell surface molecule and gene expressions. Furthermore, gene expression analysis including microarray showed that ES-SCs have M1/M2 hybrid phenotype with high expression of genes correlated to immunosuppression of T cell response. Indeed, ES-SCs were effective to block allogeneic T cell proliferation in a nitric oxide-dependent manner, and prolonged the survival of ESCs-derived embryoid bodies or cardiomyocytes grafts transplanted into mouse kidney capsule. Thus, we consider the potential use of these ESCs-derived macrophage-like immunosuppressive cells as cellular therapies to promote long-term graft survival in future therapies.

膵癌は化学療法投与環境下でMyeloid derived suppressor cellの形成を促進する(Pancreatic cancer facilitates myeloid derived suppressor cell formation under chemotherapy treated conditions)　　　　　　　　　　　　　　　
武内 慎太郎, 土川 貴裕, 和田 はるか, 中村 透, 七戸 俊明, 清野 研一郎, 平野 聡
日本癌学会総会記事, 73回, E, 2042, (一社)日本癌学会, 2014年09月
英語

Blocking monoclonal antibodies of TIM proteins as orchestrators of anti-tumor immune response
Muhammad Baghdadi, Shintaro Takeuchi, Haruka Wada, Ken-ichiro Seino
MABS, 6, 5, 1124, 1132, TAYLOR & FRANCIS INC, 2014年09月, ［査読有り］
英語, Monoclonal antibody (mAb)-based treatment of cancer has a significant effect on current practice in medical oncology, and is considered now as one of the most successful therapeutic strategies for cancer treatment. MAbs are designed to initiate or enhance anti-tumor immune responses, which can be achieved by either blocking inhibitory immune checkpoint molecules or triggering activating receptors. TIM gene family members are type-I surface molecules expressed in immune cells, and play important roles in the regulation of both innate and adaptive arms of the immune system. Therapeutic strategies based on anti-TIMs mAbs have shown promising results in experimental tumor models, and synergistic combinations of anti-TIMs mAbs with cancer vaccines, adoptive T-cell therapy, radiotherapy and chemotherapy will have great impact on cancer treatment in future clinical development.

マウスiPS細胞から誘導した制御性マクロファージ様細胞によるMLRアロT細胞増殖抑制の検討　　　　　　　　　　　　　　　
佐々木 元, 和田 はるか, 清野 研一郎
日本移植学会総会プログラム抄録集, 50回, 359, 359, (一社)日本移植学会, 2014年08月
日本語

コモンマーモセットES細胞由来免疫制御細胞による免疫寛容誘導の試み　　　　　　　　　　　　　　　
辻 飛雄馬, 和田 はるか, 佐々木 えりか, 清野 研一郎
日本移植学会総会プログラム抄録集, 50回, 359, 359, (一社)日本移植学会, 2014年08月
日本語

多能性幹細胞由来胸腺組織による中枢性寛容誘導の試み　　　　　　　　　　　　　　　
大塚 亮, 和田 はるか, 清野 研一郎
日本移植学会総会プログラム抄録集, 50回, 446, 446, (一社)日本移植学会, 2014年08月
日本語

小型霊長類コモンマーモセットの多能性幹細胞を用いた免疫抑制細胞の誘導
辻 飛雄馬, 和田 はるか, 清野 研一郎, 佐々木 えりか
北海道外科雑誌, 59, 1, 91, 91, 北海道外科学会, 2014年06月
日本語

ワクチン効果の異なるマウス乳がん細胞株の比較・検討によるワクチン効果増強因子の検索　　　　　　　　　　　　　　　
阿部 紘丈, 和田 はるか, 林 えりか, 山中 弘之, 武内 慎太郎, 平野 聡, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 18回, 155, 155, 日本がん免疫学会, 2014年06月
日本語

多能性幹細胞を用いた免疫抑制療法　　　　　　　　　　　　　　　
工藤 浩也, 和田 はるか, 佐々木 元, 辻 飛雄馬, 香城 諭, 清野 研一郎, 力石 辰也
日本泌尿器科学会総会, 102回, 429, 429, (一社)日本泌尿器科学会総会事務局, 2014年04月
日本語

マウスiPS細胞から誘導した制御性マクロファージ様細胞によるMLRアロT細胞増殖抑制の検討　　　　　　　　　　　　　　　
佐々木 元, 和田 はるか, 香城 論, 清野 研一郎, 野々村 克也
日本泌尿器科学会総会, 102回, 621, 621, (一社)日本泌尿器科学会総会事務局, 2014年04月
日本語

Treg-enriched CD4+T cells attenuate collagen synthesis in keloid fibroblasts
Naoki Murao, Ken-ichiro Seino, Toshihiko Hayashi, Masaki Ikeda, Emi Funayama, Hiroshi Furukawa, Yuhei Yamamoto, Akihiko Oyama
EXPERIMENTAL DERMATOLOGY, 23, 4, 266, 271, WILEY-BLACKWELL, 2014年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Keloid is an inflammatory and fibrotic disease with an unknown pathogenesis. Regulatory T cells (Tregs) of CD4+ lineage can suppress other effector CD4+ T cells and modulate the immune response. A relative decrease in the number of Tregs may be involved in the pathogenesis of inflammatory and fibrotic diseases. We therefore investigated the number of Tregs in keloids using immunohistochemistry and examined the interaction between Tregs and keloid fibroblasts (KFs) using a coculture system. It was found that the ratio of Tregs/CD4+ T cells was lower compared with that in other common inflammatory skin conditions. In addition, Treg-enriched CD4+ T cells reduced collagen synthesis by KFs. Our findings suggest that a local imbalance of Tregs contributes to the development of keloids and that correction of this imbalance might represent a novel therapeutic approach to keloid fibrosis.

【再生医療の最新の進歩(前篇) 次世代再生医療に向けた基盤研究】再生医療普及のための基盤技術 再生医療における免疫制御
佐々木 元, 和田 はるか, 清野 研一郎
最新医学, 69, 3月増刊, 707, 713, (株)最新医学社, 2014年03月
日本語, 再生医療として多能性幹細胞から誘導した細胞・組織を移植する際,多くの場合同種異系(アロ)移植となることが予想され,免疫抑制が必要である.臓器移植の発展とともに進化してきた免疫抑制療法を再生医療特有の状況に適合させていく作業や,ES細胞やiPS細胞から免疫制御に役立つような新しい細胞を作製するといった,再生医療時代にも通用する新しいコンセプトに基づいた免疫制御研究が必要である.(著者抄録)

Donor specific hyporesponsiveness in kidney transplantation using adoptive transfer of ex vivo induced tolerogenic cells　　　　　　　　　　　　　　　
Koyama Ichiro, Bashuda Hisashi, Seino Ken-Ichiro, Kai Kotaro, Sannomiya Akihito, Murakami Toru, Nakajima Ichiro, Nakajima Ichiro, Fuchinoue Shohei, Okumura Ko, Teraoka Satoshi
XENOTRANSPLANTATION, 20, 5, 375, 376, 2013年09月, ［査読有り］

Mouse models of human INAD by Pla2g6 deficiency
Haruka Wada, Satoshi Kojo, Ken-ichiro Seino
HISTOLOGY AND HISTOPATHOLOGY, 28, 8, 965, 969, F HERNANDEZ, 2013年08月, ［査読有り］
英語, Infantile neuroaxonal dystrophy (INAD) is a severe neurodegenerative disease characterized by its early onset. PLA2G6, which encodes a phospholipase A2, iPLA(2)beta, has been identified as a causative gene of INAD. iPLA(2)beta has been shown to be involved in various physiological and pathological processes, including immunity, cell death, and cell membrane homeostasis. Gene targeted mice with a null mutation of Pla2g6 develop the INAD phenotype as late as approximately 1 to 2 years after birth. Recently, another INAD mouse model, Pla2g6-INAD mice line, has been established. The Pla2g6-INAD mice bear a point mutation in the ankyrin repeat domain of Pla2g6 generated by N-ethyl-N-nitrosourea mutagenesis. These mutant mice develop severe motor dysfunction and hematopoietic abnormality in a manner following Mendelian law. The mice showed the abnormal gait and poor performance as early as 7 to 8 weeks of age, detected by hanging grip test. Neuropathological examination revealed widespread formation of spheroids containing tubulovesicular membranes similar to human INAD. Molecular and biochemical analysis revealed that the mutant mice expressed Pla2g6 mRNA and protein, but the mutated Pla2g6 protein had no glycerophospholipid-catalyzing enzyme activity. When analyzed the offspring which bear Pla2g6 knockout allele and Pla2g6-INAD allele, abnormal gait appeared slightly later than Pla2g6-INAD homozygotes but with earlier onset than the Pla2g6 knockout homozygotes. This result suggests that mutant Pla2g6 protein contributes to early onset of INAD symptoms in the absence of intact Pla2g6 protein. The analysis of various INAD mouse models may help to understand the pathogenesis of neurodegenerative diseases, including INAD.

【免疫反応と疾患】免疫が関与する病態と免疫応答 移植と免疫
工藤 浩也, 和田 はるか, 清野 研一郎
Medicina, 50, 3, 416, 418, (株)医学書院, 2013年03月
日本語, ＜ポイント＞拒絶反応の種類をメカニズムの違いを踏まえて理解することが重要である.免疫抑制剤の作用機序を理解し,各拒絶反応の治療に用いる.生体内には免疫を抑制する機能をもつ細胞が存在する.(著者抄録)

多能性幹細胞を用いた免疫制御法の研究
工藤 浩也, 和田 はるか, 香城 諭, 力石 辰也, 清野 研一郎
移植, 47, 総会臨時, 343, 343, (一社)日本移植学会, 2012年09月
日本語

【臓器移植臨床における免疫寛容導入の試みと現状】細胞治療による免疫寛容の誘導
工藤 浩也, 和田 はるか, 清野 研一郎
今日の移植, 25, 4, 293, 300, (株)日本医学館, 2012年08月
日本語, 臓器移植においてドナー特異的免疫寛容の誘導は非常に重要な課題となっている。これまで免疫寛容の機序が明らかになるに従って、それに準じたさまざまな寛容の誘導法が試みられてきた。そして、生体内に免疫を抑制する細胞が存在することが明らかになってきており、その誘導法や免疫抑制の機序もしだいに解明されつつある。そのため免疫抑制性細胞を用いた細胞治療は一部で臨床応用の段階まできており、将来的に移植医療の標準治療になる日がやってくるのかもしれない。今後、より詳細な機序の解明と安全性の確立が必要である。また多能性幹細胞による再生医療の研究が進むなか、併行して免疫抑制性細胞による多能性幹細胞特異的な免疫寛容の誘導法の確立にも注目していく必要がある。(著者抄録)

Role of gamma delta T cells in the regulation of HTLV-1 infection
Tomoo Sato, Masato Muto, Natsumi Araya, Satoshi Kojo, Ariyoshi Naoko, Ryuji Maekawa, Atae Utsunomiya, Ken-ichiro Seino, Yoshihisa Yamano
JOURNAL OF NEUROVIROLOGY, 18, 100, 101, SPRINGER, 2012年05月, ［査読有り］
英語

次世代の移植医療に向けて 先端技術を応用した移植医療の開発 多能性幹細胞を用いた新しい免疫制御法に関する研究
工藤 浩也, 和田 はるか, 香城 諭, 力石 辰也, 清野 研一郎
移植, 46, 総会臨時, 126, 126, (一社)日本移植学会, 2011年10月
日本語

iPSテクノロジーを導入した新しいがん免疫細胞療法の確立に向けた取り組みと現状　　　　　　　　　　　　　　　
和田 はるか, 細井 亮宏, 垣見 和宏, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 15回, 104, 104, 日本がん免疫学会, 2011年06月
日本語

Side population is increased in paclitaxel-resistant ovarian cancer cell lines regardless of resistance to cisplatin
Yoichi Kobayashi, Ken-ichiro Seino, Shinji Hosonuma, Tatsuru Ohara, Hiroaki Itamochi, Seiji Isonishi, Tsunekazu Kita, Haruka Wada, Satoshi Kojo, Kazushige Kiguchi
GYNECOLOGIC ONCOLOGY, 121, 2, 390, 394, ACADEMIC PRESS INC ELSEVIER SCIENCE, 2011年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Objectives. In recent years, cancer stem cells (CSCs) have been reported to be correlated with chemoresistance and may also be enriched in side populations (SPs). In this study, the relationship between resistance to paclitaxel (PTX) and cisplatin (CDDP) and side populations was examined in three parental PTX- and CDDP-sensitive ovarian cancer cell lines (2008, KF28, and TLI-OM-1) and several other cell lines derived from these as well as the additional effects of interferon-alpha (INF-alpha).
Methods. SP of three different parental cell lines and PTX- and/or CDDP-resistant cell lines derived from these was analyzed with flow cytometry. The expression of ABCB1 and ABCG2 in KF28 and its derived cell lines was examined. Additional cell-death effect of INF-alpha with PTX was also examined.
Results. In the three parental cell lines and the FIX-sensitive cell lines derived from these lines, SP was very low. Conversely, in PTX-resistant cell lines, regardless of CDDP resistance, SP increased. ABCB1 was strongly expressed in the PTX-resistant cells, but not in their parental lines, which are sensitive to PTX. While INF-alpha showed only slight enhancement of the cell-death effect of FIX in PTX-sensitive cells, INF-alpha itself strongly induced apoptosis in PTX-resistant cells regardless of PTX concentration.
Conclusions. The SP could be correlated with resistance to PTX. SP could be a target of INF-alpha, and resistance to PTX might be overcome by INF-alpha. (C) 2011 Elsevier Inc. All rights reserved.

Clinical significance of side population in ovarian cancer cells
Shinji Hosonuma, Yoichi Kobayashi, Satoshi Kojo, Haruka Wada, Ken-ichiro Seino, Kazushige Kiguchi, Bunpei Ishizuka
Human Cell, 24, 1, 9, 12, 2011年03月, ［査読有り］
英語, 研究論文（学術雑誌）, Recently, accumulating evidence has suggested that tumors, including ovarian cancer, are composed of a heterogeneous cell population with a small subset of cancer stem cells (CSCs) that sustain tumor formation and growth. The emergence of drug resistance is one of the most difficult problems in the treatment of ovarian cancer, which has been explained recently by the potential of CSCs to have superior resistance against anti-cancer drugs than conventional cancer cells. In this study, we expanded this line of study to examine whether this phenomenon is also observed in clinical specimens of ovarian cancer cells. In total we could analyze 28 samples out of 60 obtained from ovarian cancer patients. The clinical samples were subjected to testing of the expression of side population (SP) as a CSC marker, and according to the presence of SP (SP+) or absence of SP (SP-), clinicopathological significances were analyzed. Although there was no statistical significance, there were more SP+s in recurrent cases as well as in ascitic and peritoneal dissemination than in primary tumor of the ovary. There was no correlation between SP status and FIGO staging. In 19 cases of those who could be followed more than 6 months from initial therapy, there were 8 cases of recurrence or death from disease, and all of these were SP+. On the other hand, in 11 cases of disease-free survivors, 6 were SP+. There was a significant difference in prognosis between SP+ and SP- (p = 0.017). Although this study was limited, it revealed that SP could be contained more in recurrent or metastatic tumors than in primary tumors, and also that the presence of SP could be a risk factor of recurrence in ovarian cancer. Therefore, a novel therapeutic strategy targeting SP could improve the prognosis of ovarian cancer. © 2010 The Author(s).

Clinical significance of side population in ovarian cancer cells
Shinji Hosonuma, Yoichi Kobayashi, Satoshi Kojo, Haruka Wada, Ken-ichiro Seino, Kazushige Kiguchi, Bunpei Ishizuka
HUMAN CELL, 24, 1, 9, 12, SPRINGER TOKYO, 2011年, ［査読有り］
英語, 研究論文（学術雑誌）, Recently, accumulating evidence has suggested that tumors, including ovarian cancer, are composed of a heterogeneous cell population with a small subset of cancer stem cells (CSCs) that sustain tumor formation and growth. The emergence of drug resistance is one of the most difficult problems in the treatment of ovarian cancer, which has been explained recently by the potential of CSCs to have superior resistance against anti-cancer drugs than conventional cancer cells. In this study, we expanded this line of study to examine whether this phenomenon is also observed in clinical specimens of ovarian cancer cells. In total we could analyze 28 samples out of 60 obtained from ovarian cancer patients. The clinical samples were subjected to testing of the expression of side population (SP) as a CSC marker, and according to the presence of SP (SP+) or absence of SP (SP-), clinicopathological significances were analyzed. Although there was no statistical significance, there were more SP+s in recurrent cases as well as in ascitic and peritoneal dissemination than in primary tumor of the ovary. There was no correlation between SP status and FIGO staging. In 19 cases of those who could be followed more than 6 months from initial therapy, there were 8 cases of recurrence or death from disease, and all of these were SP+. On the other hand, in 11 cases of disease-free survivors, 6 were SP+. There was a significant difference in prognosis between SP+ and SP- (p = 0.017). Although this study was limited, it revealed that SP could be contained more in recurrent or metastatic tumors than in primary tumors, and also that the presence of SP could be a risk factor of recurrence in ovarian cancer. Therefore, a novel therapeutic strategy targeting SP could improve the prognosis of ovarian cancer.

Successful differentiation to T cells, but unsuccessful B-cell generation, from B-cell-derived induced pluripotent stem cells
Haruka Wada, Satoshi Kojo, Chie Kusama, Naoki Okamoto, Yorino Sato, Bunpei Ishizuka, Ken-ichiro Seino
INTERNATIONAL IMMUNOLOGY, 23, 1, 65, 74, OXFORD UNIV PRESS, 2011年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Forced expression of certain transcription factors in somatic cells results in generation of induced pluripotent stem (iPS) cells, which differentiate into various cell types. We investigated T-cell and B-cell lineage differentiation from iPS cells in vitro. To evaluate the impact of iPS cell source, murine splenic B-cell-derived iPS (B-iPS) cells were generated after retroviral transduction of four transcription factors (Oct4, Sox2, Klf4 and c-Myc). B-iPS cells were identical to embryonic stem (ES) cells and mouse embryonic fibroblast (MEF)-derived iPS cells in morphology, ES cell marker expression as well as teratoma and chimera mouse formation. Both B-iPS and MEF-derived iPS cells differentiated into lymphocytes in OP9 co-culture systems. Both efficiently differentiated into T-cell lineage that produced IFN-gamma on T-cell receptor stimulation. However, iPS cells including B-iPS cells were relatively resistant to B-cell lineage differentiation. One of the reasons of the failure of B-cell lineage differentiation seemed due to a defect of Pax5 expression in the differentiated cells. Therefore, current in vitro differentiation systems using iPS cells are sufficient for inducing T-cell but not B-cell lineage.

Combining carbon ion radiotherapy and local injection of α-galactosylceramide-pulsed dendritic cells inhibits lung metastases in an in vivo murine model.
Ohkubo Y, Iwakawa M, Seino K, Nakawatari M, Wada H, Kamijuku H, Nakamura E, Nakano T, Imai T
International journal of radiation oncology, biology, physics, 78, 5, 1524, 1531, ELSEVIER SCIENCE INC, 2010年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose Our previous report indicated that carbon ion beam irradiation upregulated membrane-associated immunogenic molecules, underlining the potential clinical application of radioimmunotherapy The antimetastatic efficacy of local combination therapy of carbon ion radiotherapy and immunotherapy was examined by use of an en vivo murine model
Methods and Materials Tumors of mouse squamous cell carcinoma (NR-S1) cells inoculated in the legs of C3H/HeSlc mice were locally irradiated with a single 6 Gy dose of carbon ions (290 MeV/nucleon, 6 cm spread out Bragg peak) Thirty-six hours after irradiation, a galactosylceramide pulsed dendritic cells (DCs) were injected Into the leg tumor We investigated the effects on distant lung metastases by counting the numbers of lung tumor colonies, making pathologic observations, and assessing immunohistochemistry
Results The mice with no treatment (control) presented with 168 53 8 metastatic nodules in the lungs, whereas the mice that received the combination therapy of carbon ion irradiation and DCs presented with 2 6 1 9 (P = 0 009) at 2 weeks after irradiation Immunohistochemistry showed that intracellular adhesion molecule 1, which activates DCs, increased from 6 h to 36 h after irradiation in the local tumors of the carbon ion irradiated group The expression of S100A8 in lung tissue, a marker of the lung pre metastatic phase, was decreased only in the group with a combination of carbon ions and DCs
Conclusions The combination of carbon ion radiotherapy with the injection of a galactosylceramide pulsed DCs into the primary tumor effectively inhibited distant lung metastases (C) 2010 Elsevier Inc

iPS細胞を用いた新しい免疫細胞再生医療の可能性(DLIモデルによる検討)　　　　　　　　　　　　　　　
和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 14回, 55, 55, 日本がん免疫学会, 2010年07月
日本語

インフルエンザ感染におけるCTLA4-Igの効果　　　　　　　　　　　　　　　
神宿 元, 香城 諭, 和田 はるか, 清野 研一郎
リウマチ科, 43, 3, 291, 297, (有)科学評論社, 2010年03月
日本語

Establishment of an Improved Mouse Model for Infantile Neuroaxonal Dystrophy That Shows Early Disease Onset and Bears a Point Mutation in Pla296
Haruka Wada, Takuwa Yasuda, Ikuo Miura, Kazuhiko Watabe, Chika Sawa, Hajime Kamijuku, Satoshi Kojo, Masaru Taniguchi, Ichizo Nishino, Shigeharu Wakana, Hisahiro Yoshida, Ken-ichiro Seino
AMERICAN JOURNAL OF PATHOLOGY, 175, 6, 2257, 2263, ELSEVIER SCIENCE INC, 2009年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Calcium-independent group VIA phospholipase A, (iPLA,P), encoded by PLA2G6, has been shown to be involved in various physiological and pathological processes, including immunity, cell death, and cell membrane homeostasis. Mutations in the PLA2G6 gene have been recently identified in patients with infantile neuroaxonal dystrophy (INAD). Subsequently, it was reported that similar neurological impairment occurs in gene-targeted mice with a null mutation of iPLA(2)beta, whose disease onset became apparent approximately 1 to 2 years after birth. Here, we report the establishment of an improved mouse model for INAD that bears a point mutation in the ankyrin repeat domain of Pla2g6 generated by N-ethyl-N-nitrosourea mutagenesis. These mutant mice developed severe motor dysfunction, including abnormal gait and poor performance in the hanging grip test, as early as 7 to 8 weeks of age, in a manner following Mendelian law. Neuropathological examination revealed widespread formation of spheroids containing tubulovesicular membranes similar to human INAD. Molecular and biochemical analysis revealed that the mutant mice expressed Pla2g6 mRNA and protein, but the mutated Pla2g6 protein had no glycerophospholipid-catalyzing enzyme activity. Because of the significantly early onset of the disease, this mouse mutant (Pla2g6-inad) could be highly useful for further studies of pathogenesis and experimental interventions in INAD and neurodegeneration. (Am j Pathol 2009, 175:2257-226, DOI: 10.2353/ajpath.2009.090343)

免疫制御分子とがん免疫療法 Zoledronateにより活性化させたγδT細胞の抗原提示能に関する検討　　　　　　　　　　　　　　　
武藤 真人, 富山 舞, 贄田 美江, 前川 隆司, 香城 諭, 和田 はるか, 清野 研一郎
日本免疫学会総会・学術集会記録, 39, 199, 199, (NPO)日本免疫学会, 2009年11月
日本語

Severe loss of invariant NKT cells exhibiting anti-HTLV-1 activity in patients with HTLV-1-associated disorders
Kazuko Azakami, Tomoo Sato, Natsumi Araya, Atae Utsunomiya, Ryuji Kubota, Kenshi Suzuki, Daisuke Hasegawa, Toshihiko Izumi, Hidetoshi Fujita, Satoko Aratani, Ryoji Fujii, Naoko Yagishita, Hajime Kamijuku, Takuro Kanekura, Ken-ichiro Seino, Kusuki Nishioka, Toshihiro Nakajima, Yoshihisa Yamano
BLOOD, 114, 15, 3208, 3215, AMER SOC HEMATOLOGY, 2009年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Invariant natural killer T (iNKT) cells are unique T cells that regulate the immune response to microbes, cancers, and autoimmunity. We assessed the characteristics of iNKT cells from persons infected with human T-lymphotropic virus type 1 (HTLV-1). Whereas most infected persons remain asymptomatic carriers (ACs) throughout their lives, a small proportion, usually with high equilibrium proviral loads, develop 2 diseases: HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T-cell leukemia (ATL). We demonstrated that the frequency of iNKT, NK, and dendritic cells in the peripheral blood of HAM/TSP and ATL patients is decreased. We also observed an inverse correlation between the iNKT cell frequency and the HTLV-1 proviral load in the peripheral blood of infected persons. Notably, in vitro stimulation of peripheral blood cells with alpha-galactosylceramide led to an increase in the iNKT cell number and a subsequent decrease in the HTLV-1-infected T-cell number in samples from ACs but not HAM/TSP or ATL patients. Our results suggest that iNKT cells contribute to the immune defense against HTLV-1, and iNKT-cell depletion plays an important role in the pathogenesis of HAM/TSP and ATL. Therefore, iNKT cell-based immunotherapy may be an effective strategy for preventing these HTLV-1-associated disorders. (Blood. 2009; 114: 3208-3215)

The study of regulatory T cells and NKT cells in Japan: a historical perspective
Hisashi Arase, Ken-ichiro Seino
INTERNATIONAL IMMUNOLOGY, 21, 10, 1101, 1103, OXFORD UNIV PRESS, 2009年10月, ［査読有り］
英語, Immune regulation plays an important role in maintaining homeostasis of the immune system. A number of Japanese immunologists have made significant contributions to the elucidation of the mechanisms of immune regulation. In particular, lymphocyte populations that could regulate immune responses-for example regulatory T cells and NKT cells-have been extensively analyzed. Here, we present an overview of research on immune regulation by highlighting the work of several Japanese contributors.

多能性幹細胞からの機能的T細胞の生成
和田 はるか, 清野 研一郎
移植, 44, 総会臨時, 255, 255, (一社)日本移植学会, 2009年09月
日本語

Research highlights: Immunomodulation.
Wada H, Kojo S, Seino K
Immunotherapy, 1, 5, 737, 739, 2009年09月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）

Cell therapy(細胞療法)の新たなアプローチ ES細胞と人工多能性幹細胞(iPS)細胞由来の機能的T細胞のin vitro作製(Generation of functional T cells from ES cells and induced pluripotent stem (iPS) cells in vitro)　　　　　　　　　　　　　　　
和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 13回, 36, 36, 日本がん免疫学会, 2009年06月
英語

樹状細胞 マウスモデルを用いた炭素イオン線照射・樹状細胞療法併用による肺転移抑制効果　　　　　　　　　　　　　　　
大久保 悠, 岩川 眞由美, 神宿 元, 和田 はるか, 清野 研一郎
日本がん免疫学会総会プログラム・抄録集, 13回, 46, 46, 日本がん免疫学会, 2009年06月
日本語

Cancellation of NKT cell immunosuppression targeting myeloid suppressor cells
Wada Haruka, Yanagisawa Kazuhiko, Seino Ken-ichiro
炎症・再生 : 日本炎症・再生医学会雑誌 = Inflammation and regeneration, 29, 2, 128, 130, 日本炎症・再生医学会, 2009年03月25日
英語, CD1d-restricted natural killer T (NKT) cells are one of immunoregulatory cells. NKT cells can be specifically activated by a synthetic glycolipid, &alpha;-galactosylceramide (&alpha;-GalCer). Using some glycolipids such as &alpha;-GalCer, it is expected to develop a new NKT cell-mediated therapeutic strategy against cancer. However, it is known that, in human cancer patients, NKT cells express a degree of hyporesponsiveness to &alpha;-GalCer. For example, we have reported that, in gastrointestinal cancer patients, NKT cell proliferation and cytokine production were impaired. We have further examined the mechanism by which hyporesponsiveness to &alpha;-GalCer can be induced using cancer-bearing mice. In the animal study, &alpha;-GalCer-induced NKT cell expansion, cytokine production, cytotoxicity, and anti-metastatic effect in vivo were all significantly impaired. In fact, &alpha;-GalCer could eliminate metastatic disease in naive animals, but failed to protect cancer-bearing mice. We found that CD11b<SUP>+</SUP> Gr-1<SUP>+</SUP> cells were particularly increased in cancer-bearing mice and were necessary and sufficient for the suppression of NKT cells to &alpha;-GalCer. We also found that the increased CD11b<SUP>+</SUP> Gr-1<SUP>+</SUP> cells suppressed NKT cell function in a nitric oxide-mediated fashion. To reduce the population of CD11b<SUP>+</SUP> Gr-1<SUP>+</SUP> cells, we administered a retinoic acid to cancer-bearing mice. This treatment significantly reduced the population of CD11b<SUP>+</SUP> Gr-1<SUP>+</SUP> cells and effectively restored &alpha;-GalCer-induced NKT cell responses. These results demonstrate a novel feature of NKT cell function in cancer, and suggest a new strategy to enhance NKT cell-mediated anti-cancer immune responses by suppressing CD11b<SUP>+</SUP> Gr-1<SUP>+</SUP> cell functions.

ENU誘発PLA2G6変異マウス ヒト乳児型軸索ジストロフィーモデルの樹立　　　　　　　　　　　　　　　
渡部 和彦, 和田 はるか, 高澤 隆紀, 澤 智華, 秋山 けい子, 安田 琢和, 吉田 尚弘, 西野 一三, 清野 研一郎
臨床神経学, 48, 12, 1116, 1116, (一社)日本神経学会, 2008年12月, ［査読有り］
日本語

RCAI-17, 22, 24-26, 29, 31, 34-36, 38-40, and 88, the analogs of KRN7000 with a sulfonamide linkage: Their synthesis and bioactivity for mouse natural killer T cells to produce Th2-biased cytokines
Takuya Tashiro, Naomi Hongo, Ryusuke Nakagawa, Ken-ichiro Seino, Hiroshi Watarai, Yasuyuki Ishii, Masaru Taniguchi, Kenji Mori
BIOORGANIC & MEDICINAL CHEMISTRY, 16, 19, 8896, 8906, PERGAMON-ELSEVIER SCIENCE LTD, 2008年10月, ［査読有り］
英語, 研究論文（学術雑誌）, RCAI-17, 22, 24-26, 29, 31, 34-36, 38-40, and 88, the analogs of KRN7000 (1) with a sulfonamide linkage instead of an amide bond, were synthesized to examine their bioactivity for mouse natural killer (NK) T cells. RCAI-17, 22, 24-26, 29, 31, 34-36, and 88 are the aromatic sulfonamide analogs, while RCAI-39 and 40 are the aliphatic ones. RCAI-38 is a C-galactoside analog of RCAI-26, which is the p-toluenesulfonamide analog of KRN7000. According to their bioassay, these sulfonamide analogs were shown to be the stimulants of mouse NKT cells to induce the production of Th2-biased cytokines in vitro, while RCAI-38 did not induce any cytokine production. (C) 2008 Elsevier Ltd. All rights reserved.

Mechanism of NKT cell activation by intranasal coadministration of alpha-galactosylceramide, which can induce cross-protection against influenza viruses
H. Kamijuku, Y. Nagata, X. Jiang, T. Ichinohe, T. Tashiro, K. Mori, M. Taniguchi, K. Hase, H. Ohno, T. Shimaoka, S. Yonehara, T. Odagiri, M. Tashiro, T. Sata, H. Hasegawa, K-i Seino
MUCOSAL IMMUNOLOGY, 1, 3, 208, 218, NATURE PUBLISHING GROUP, 2008年05月, ［査読有り］
英語, 研究論文（学術雑誌）, In a nasal vaccine against influenza, the activation of natural killer T (NKT) cells by intranasal coadministration of alpha-galactosylceramide (alpha-GalCer) can potently enhance protective immune responses. The results of this study show that the NKT cell-activated nasal vaccine can induce an effective cross-protection against different strains of influenza virus, including H5 type. To analyze the mechanism of NKT cell activation by this nasal vaccine, we prepared fluorescence-labeled alpha-GalCer by which we detect a direct interaction between NKT cells and alpha-GalCer-stored dendritic cells in nasal mucosa-associated tissues. Accordingly, although very few NKT cells exist at mucosa, the nasal vaccination induced a localized increase in NKT cell population, which is partly dependent on CXCL16/CXCR6. Furthermore, we found that NKT cell activation stimulates mucosal IgA production by a mechanism that is dependent on interleukin (IL)-4 production. These results strengthen the basis of nasal vaccination via NKT cell activation, which can induce immune cross-protection.

Distinct regulatory functions of SLP-76 and MIST in NK cell cytotoxicity and IFN-gamma production
Shinya Hidano, Hiroki Sasanuma, Keiko Ohshima, Ken-Ichiro Seino, Lalit Kumar, Katsuhiko Hayashi, Masaki Hikida, Tomohiro Kurosaki, Masaru Taniguchi, Raif S. Geha, Daisuke Kitamura, Ryo Goitsuka
INTERNATIONAL IMMUNOLOGY, 20, 3, 345, 352, OXFORD UNIV PRESS, 2008年03月, ［査読有り］
英語, 研究論文（学術雑誌）, Activation of NK cells is triggered by multiple receptors. We demonstrate here that SLP-76 is required for CD16- and NKG2D-mediated NK cell cytotoxicity, while MIST negatively regulates these responses in an SLP-76-dependent manner. Exceptionally, MIST acts as a positive regulator of cytotoxicity against YAC-1 cells, although SLP-76 plays a more key role. SLP-76 acts as a dominant positive regulator for both NKG2D-mediated and YAC-1 cell-triggered IFN-gamma production. Although NKG2D-mediated IFN-gamma production depends on phospholipase C (PLC) gamma 2, YAC-1 cell-triggered IFN-gamma production is PLC gamma 2- and Syk/ZAP-70 independent and nuclear factor-kappa B mediated. SLP-76 is required for this process in the presence of MIST but is dispensable in the absence of MIST. Thus, YAC-1 cell-triggered NKG2D-independent IFN-gamma production appears to be regulated by SLP-76-dependent and -independent pathways, in which the latter is negatively regulated by MIST. Taken together, these results suggest that SLP-76 and MIST distinctly but interactively regulate NK cell cytotoxicity and IFN-gamma production.

RCAI-8, 9, 18, 19, and 49-52, conformationally restricted analogues of KRN7000 with an azetidine or a pyrrolidine ring: Their synthesis and bioactivity for mouse natural killer T cells to produce cytokines
Ken-ichi Fuhshuku, Naomi Hongo, Takuya Tashiro, Yui Masuda, Ryusuke Nakagawa, Ken-ichiro Seino, Masaru Taniguchi, Kenji Mori
BIOORGANIC & MEDICINAL CHEMISTRY, 16, 2, 950, 964, PERGAMON-ELSEVIER SCIENCE LTD, 2008年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Conformationally restricted analogues of KRN7000, an alpha-D-galactosyl ceramide, were synthesized to examine their bioactivity for mouse natural killer (NK) T cells to produce cytokines. RCAI-8, 9, 5 1, and 52 are the analogues with a pyrrolidine ring, and RCAI-18, 19, 49, and 50 are those with an azetidine ring. RCAI-18 was shown to be a potent inducer of cytokine production by mouse NKT cells, while RCAI-51 was a moderately active inducer. (C) 2007 Elsevier Ltd. All rights reserved.

Critical role for CXC chemokine ligand 16 (SR-PSOX) in Th1 response mediated by NKT cells
Takeshi Shimaoka, Ken-ichiro Seino, Noriaki Kume, Manabu Minami, Chiyoko Nishime, Makoto Suematsu, Toru Kita, Masaru Taniguchi, Kouji Matsushima, Shin Yonehara
JOURNAL OF IMMUNOLOGY, 179, 12, 8172, 8179, AMER ASSOC IMMUNOLOGISTS, 2007年12月, ［査読有り］
英語, 研究論文（学術雑誌）, The transmembrane chemokine CXCL 16 (CXCL16), which is the same molecule as the scavenger receptor that binds phosphatidylserine and oxidized lipoprotein (SR-PSOX), has been shown to mediate chemotaxis and adhesion of CXC chemokine receptor 6-expressing cells such as NKT and activated Th1 cells. We generated SR-PSOX/CXCL16-deficient mice and examined the role of this chemokine in vivo. The mutant mice showed a reduced number of liver NKT cells, and decreased production of IFN-gamma and IL-4 by administration of alpha-galactosylceramide (alpha GalCer). Of noted the alpha GalCer-induced production of IFN-gamma was more severely impaired than the production of IL-4 in SR-PSOX-deficient mice. In this context, SR-PSOX-deficient mice showed impaired sensitivity to aGalCer-induced anti-tumor effect mediated by IFN-gamma from NKT cells. NKT cells from wild-type mice showed impaired production of IFN-gamma, but not IL-4, after their culture with alpha GalCer and APCs from mutant mice. Moreover, Propionibacterium acnes-induced in vivo Th1 responses were severely impaired in SR-PSOX-deficient as well as NKT KO mice. Taken together, SR-PSOX/CXCL16 plays an important role in not only the production of IFN-gamma by NKT cells, but also promotion of Th1-inclined immune responses mediated by NKT cells.

Construction of an open-access database that integrates cross-reference information from the transcriptome and proteome of immune cells
Atsushi Hijikata, Hiroshi Kitamura, Yayoi Kimura, Ryo Yokoyama, Yuichi Aiba, Yanyuan Bao, Shigeharu Fujita, Koji Hase, Shohei Hori, Yasuyuki Ishii, Osami Kanagawa, Hiroshi Kawamoto, Kazuya Kawano, Haruhiko Koseki, Masato Kubo, Ai Kurita-Miki, Tomohiro Kurosaki, Kyoko Masuda, Mitsumasa Nakata, Keisuke Oboki, Hiroshi Ohno, Mariko Okamoto, Yoshimichi Okayama, Jiyang O-Wang, Hirohisa Saito, Takashi Saito, Machie Sakuma, Katsuaki Sato, Kaori Sato, Ken-Ichiro Seino, Ruka Setoguchi, Yuki Tamura, Masato Tanaka, Masaru Taniguchi, Ichiro Taniuchi, Annabelle Teng, Takeshi Watanabe, Hiroshi Watarai, Sho Yamasaki, Osamu Ohara
BIOINFORMATICS, 23, 21, 2934, 2941, OXFORD UNIV PRESS, 2007年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Motivation: Although a huge amount of mammalian genomic data does become publicly available, there are still hurdles for biologists to overcome before such data can be fully exploited. One of the challenges for gaining biological insight from genomic data has been the inability to cross-reference transcriptomic and proteomic data using a single informational platform. To address this, we constructed an open-access database that enabled us to cross-reference transcriptomic and proteomic data obtained from immune cells.
Results: The database, named RefDIC (Reference genomics Data-base of Immune Cells), currently contains: (i) quantitative mRNA profiles for human and mouse immune cells/tissues obtained using Affymetrix GeneChip technology; (ii) quantitative protein profiles for mouse immune cells obtained using two-dimensional gel electrophoresis (2-DE) followed by image analysis and mass spectrometry and (iii) various visualization tools to cross-reference the mRNA and protein profiles of immune cells. RefDIC is the first open-access database for immunogenomics and serves as an important information-sharing platform, enabling a focused genomic approach in immunology.

Mechanism of NKT cell-mediated transplant tolerance
X. Jiang, S. Kojo, M. Harada, N. Ohkohchi, M. Taniguchi, K.-I. Seino
AMERICAN JOURNAL OF TRANSPLANTATION, 7, 6, 1482, 1490, BLACKWELL PUBLISHING, 2007年06月, ［査読有り］
英語, 研究論文（学術雑誌）, The mechanism by which CD1d-restricted V alpha 14 natural killer T (NKT) cells participate in transplant tolerance has yet to be completely clarified. Recently, we showed that repeated activation of NKT cells by their specific glycolipid ligand, alpha-galactosylceramide, leads to a change in function to an immune regulatory role with IL-10 production. Moreover, these cells were shown to be able to induce regulatory dendritic cells (DCs). In this study, we showed that NKT cells from transplant tolerant recipients of cardiac allograft produced higher levels of IL-10, which is required for the maintenance of tolerance; this was proved by adoptive transfer experiments. In addition, DCs from wild-type (WT) tolerant recipients but not NKT cell-deficient recipients showed a higher IL-10-producing profile, a more immature phenotype, and tolerogenic capability. CD4 T cells from WT tolerant recipients but not NKT cell-deficient recipients also produced higher levels of IL-10 upon alloantigen stimulation and showed lower proliferative activity that was reversed by blocking the IL-10 receptor. These data indicate the existence of IL-10-dependent immune regulatory interplay among NKT cells, DCs, and CD4 T cells, even in the absence of artificial stimulation of NKT cells with synthetic glicolipids, which is required for the maintenance of transplant tolerance.

Innate Immunity: NKT Cells
K. I. Seino, M. Taniguchi
Comprehensive Glycoscience: From Chemistry to Systems Biology, 4-4, 9, 16, Elsevier, 2007年01月01日, ［査読有り］
英語, 論文集(書籍)内論文

Differential role of thymic stromal lymphopoietin in the induction of airway hyperreactivity and Th2 immune response in antigen-induced asthma with respect to natural killer T cell function
Yuko Nagata, Hajime Kamijuku, Masaru Taniguchi, Steven Ziegler, Ken-ichiro Seino
INTERNATIONAL ARCHIVES OF ALLERGY AND IMMUNOLOGY, 144, 4, 305, 314, KARGER, 2007年, ［査読有り］
英語, 研究論文（学術雑誌）, Asthma is an inflammatory lung disease, in which CD1d-restricted natural killer T (NKT) cells play an important pathogenic role. Also, recent reports indicated that a cytokine, thymic stromal lymphopoietin ( TSLP), is essential for the development of antigen-induced asthma. Here we examined the relationship between NKT cells and TSLP in a mouse model of asthma. NKT cells express TSLP receptor as well as IL-7 receptor alpha-chain. TSLP acts on NKT cells to preferentially increase their IL-13 production but not IFN-gamma and IL- 4. In an allergen-induced asthma model, the development of airway hyperreactivity, a cardinal feature of asthma, was increased in TSLP transgenic mice, whereas this effect was not observed in TSLP transgenic mice lacking NKT cells. Interestingly, in the NKT cell-lacking TSLP transgenic mice, pulmonary eosinophilia and increase in lgE did not improve. Pulmonary lymphocytes from the NKT cell-lacking TSLP transgenic mice produced much less IL-13 upon CD3 stimulation than those from NKT cell-competent TSLP transgenic mice. These results suggest that, in allergen-induced asthma, TSLP acts on NKT cells to enhance airway hyperreactivity by upregulating their IL-13 production, whereas eosinophilia and lgE production are not influenced. Copyright (C) 2007 S. Karger AG, Basel

Hyporesponsiveness to natural killer T-cell ligand alpha-galactosylceramide in cancer-bearing state mediated by CD11b(+) Gr-1(+) cells producing nitric oxide
Kazuhiko Yanagisawa, Mark A. Exley, Xiaofeng Jiang, Nobuhiro Ohkochi, Masaru Taniguchi, Ken-ichiro Seino
CANCER RESEARCH, 66, 23, 11441, 11446, AMER ASSOC CANCER RESEARCH, 2006年12月, ［査読有り］
英語, 研究論文（学術雑誌）, CD1d-restricted natural killer T (NKT) cells are a potential therapeutic target for cancer, for which several clinical trials have already been reported. NKT cells are specifically activated by a synthetic glycolipid, alpha-galactosylceramide (alpha-GalCer). However, it is known that, in human cancer patients, NKT cells express a degree of hyporesponsiveness to alpha-GalCer. In this study, we have examined the mechanism by which hyporesponsiveness to alpha-GalCer can be induce. In cancer-bearing mice, alpha-GalCer-induced NKT cell expansion, cytokine production, cytotoxicity, and antimetastatic effect in vivo were all significantly impaired. In fact, alpha-GalCer could eliminate metastatic disease in naive animals but failed to protect cancer-bearing mice. CD11b(+) Gr-1(+) cells were particularly increased in cancer-bearing mice and were necessary and sufficient for the suppression of the alpha-GalCer response in a nitric oxide-mediated fashion. Administration of a retinoic acid to cancer-bearing mice reduced the population of CD11b(+) Gr-1(+) cells and effectively restored alpha-GalCer-induced protection. These results show a novel feature of NKT cell function in cancer. Furthermore, our data suggest a new strategy to enhance NKT cell-mediated anticancer immune responses by suppressing CD11b(+) Gr-1(+) cell functions.

IL-21-induced B epsilon cell apoptosis mediated by natural killer T cells suppresses IgE responses
Michishige Harada, Kumiko Magara-Koyanagi, Hiroshi Watarai, Yuko Nagata, Yasuyuki Ishii, Satoshi Kojo, Shigetoshi Horiguchi, Yoshitaka Okamoto, Toshinori Nakayama, Nobutaka Suzuki, Wen-Chen Yeh, Shizuo Akira, Hiroshi Kitamura, Osamu Ohara, Ken-ichiro Seino, Masaru Taniguchi
JOURNAL OF EXPERIMENTAL MEDICINE, 203, 13, 2929, 2937, ROCKEFELLER UNIV PRESS, 2006年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Epidemiological studies have suggested that the recent increase in the incidence and severity of immunoglobulin (Ig) E-mediated allergic disorders is inversely correlated with Mycobacterium bovis bacillus Calmette Guerin ( BCG) vaccination; however, the underlying mechanisms remain uncertain. Here, we demonstrate that natural killer T (NKT) cells in mice and humans play a crucial role in the BCG-induced suppression of IgE responses. BCG-activated murine V alpha 14 NKT cells, but not conventional CD4 T cells, selectively express high levels of interleukin (IL)-21, which preferentially induces apoptosis in B epsilon cells. Signaling from the IL-21 receptor increases the formation of a complex between Bcl-2 and the proapoptotic molecule Bcl-2-modifying factor, resulting in B epsilon cell apoptosis. Similarly, BCG vaccination induces IL-21 expression by human peripheral blood mononuclear cells (PBMCs) in a partially NKT cell-dependent fashion. BCG-activated PBMCs significantly reduce IgE production by human B cells. These findings provide new insight into the therapeutic effect of BCG in allergic diseases.

Natural killer T cell-mediated antitumor immune responses and their clinical applications
Ken-ichiro Seino, Shinichiro Motohashi, Takehiko Fujisawa, Toshinori Nakayama, Masaru Taniguchi
CANCER SCIENCE, 97, 9, 807, 812, BLACKWELL PUBLISHING, 2006年09月, ［査読有り］
英語, A unique lymphocyte population, CD1d-restricted NKT cells, has been revealed to be a key player in both the innate and acquired immune responses, including antitumor effects. Recent studies revealed that at least two subsets of CD1d-restricted NKT cells exist: type I, having invariant V alpha 14 receptor; and type II, having heterogeneous non-V alpha 14 receptor. The specific glycolipid ligand, alpha-GalCer, effectively stimulates mouse and human type I NKT cells. The activation of type I NKT cells substantially influences function of other various cell types, particularly DC, NK cells, CD4 Th1 cells, and CD8 cytotoxic T cells, all contributing to the antitumor immune responses. Recent studies also indicated that, unlike type I NKT cells, type II NKT cells have a potential to repress antitumor immune responses. In this review, we summarize the characteristics of the antitumor immune responses mediated by both mouse and human CD1d-restricted NKT cells and discuss their potential in clinical applications against cancer.

CD1d-restricted NKT cell activation enhanced homeostatic proliferation of CD8(+) T cells in a manner dependent on IL-4
Naoko Ueda, Hiroko Kuki, Daisuke Kamimura, Shinichiro Sawa, Kenichiro Seino, Takuya Tashiro, Ken-ichi Fushuku, Masaru Taniguchi, Toshio Hirano, Masaaki Murakami
INTERNATIONAL IMMUNOLOGY, 18, 9, 1397, 1404, OXFORD UNIV PRESS, 2006年09月, ［査読有り］
英語, 研究論文（学術雑誌）, CD1d-restricted NKT cells are activated by TCR-mediated stimulation via CD1d plus lipid antigens such as alpha-galactosylceramide (alpha-GaICer). These cells suppressed autoimmunity and graft rejection, but sometimes enhanced resistance to infection and tumor immunity. This double-action phenomenon of NKT cells is partly explained by cytokines produced by NKT cells. Therefore, roles of cytokines from activated NKT cells have been extensively examined; however, their roles on T cell homeostatic proliferation in lymphopenic condition have not been investigated. Here, we showed that alpha-GaICer enhanced homeostatic proliferation of CD8(+) but not CD4(+) T cells and this effect of alpha-GaICer was required for NKT cells. IL-4 was essential and sufficient for this NKT cell action on CD8(+) T cell homeostatic proliferation. Importantly, the expression of IL-4R alpha and STAT6 in CD8(+) T cells was essential for the NKT activity, indicating a direct action of IL-4 on CD8(+) T cells. Consistent with this, the level of IL-4R alpha expression on memory phenotype CD8(+) T cells was higher than that on naive phenotype one and CD4(+) T cells. Thus, these results showed the 'involvement' of IL-4 that is produced from activated NKT cells for CD8(+) T cell homeostatic proliferation in vivo.

The importance of CD25(+)CD4(+) regulatory T cells in mouse hepatic allograft tolerance
Jiang, X, M Morita, A Sugioka, M Harada, S Kojo, H Wakao, H Watarai, N Ohkohchi, M Taniguchi, KI Seino
LIVER TRANSPLANTATION, 12, 7, 1112, 1118, JOHN WILEY & SONS INC, 2006年07月, ［査読有り］
英語, 研究論文（学術雑誌）, In mouse liver transplantation, tolerance is readily inducible. Recent studies have revealed that CD25(+)CD4(+) regulatory T cells play an important role in regulating various immune responses, including transplant tolerance. However, the contribution of these cells to tolerance in mouse liver transplantation has not been elucidated. We showed here that depletion of CD25(+)CD4(+) T cells increased proliferative response of CD4(+) T cells and cytotoxic T lymphocyte induction of CD8(+) T cells. Depletion of these cells in the recipient but not in the donor before liver transplantation caused rejection. Furthermore, the number of CD25(+)CD4(+) population and forkhead/winged helix transcription factor expression in liver mononuclear lymphocytes derived from tolerant mice were higher than those from grafts undergoing rejection. In conclusion, these results indicate that CD25(+)CD4(+) regulatory T cells in the recipient but not in the donor of liver transplantation are important for the tolerance induction.

Regulatory dendritic cells act as regulators of acute lethal systemic inflammatory response
S Fujita, K Seino, K Sato, Y Sato, K Eizumi, N Yamashita, M Taniguchi, K Sato
BLOOD, 107, 9, 3656, 3664, AMER SOC HEMATOLOGY, 2006年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Bacterial infection triggers host inflammation through the activation of immune cells, leading to the elimination of bacteria. However, the regulatory mechanisms of the host inflammatory response remain unknown. Here we report that a subset of potent tolerogenic dendritic cells (DCs), regulatory DCs (DCregs), control the systemic inflammatory response. Unlike normal DCs, which produced proinflammatory cytokines in response to bacterial lipopolysaccharide (LPS), DCregs produced fewer proinflammatory cytokines and instead preferentially produced interleukin-10 (IL-10), and these events involved the expression Of I kappa BNS and Bcl-3 as well as cyclic AMP (cAMP)mediated activation of protein kinase A (PKA). In addition, DCregs not only suppressed LPS-induced production of proinflammatory cytokines in macrophages, but also reduced their serum levels in mice. Furthermore, DCregs protected mice against the lethality induced by experimental endotoxemia and bacterial peritonitis. The inhibitory effect of DCregs against inflammatory responses involved the production of IL-10. On the other hand, naturally existing tolerogenic DC subsets producing IL-10, CD11c(low)CD45RB(high) DCs, also suppressed LPS-induced host inflammatory responses. Thus, a subset of tolerogenic DCs act as potential regulators of the host inflammatory response, and they might have preventive and therapeutic potential for the treatment of systemic as well as local inflammatory diseases.

Indications for surgery in intestinal Behçet's disease.
Kurata M, Nozue M, Seino K, Murata H, Sumita T, Katashi F
Hepato-gastroenterology, 53, 67, 60, 63, H G E UPDATE MEDICAL PUBLISHING S A, 2006年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Background/Aims: As advances in steroids and immunosuppressants made medical treatment first-line therapy for Behcet's disease, the criteria for elective surgery became unclear. The aim of this paper is to establish surgical criteria for Behcet's.
Methodology: We reviewed the medical records of 8 consecutive Japanese patients who were diagnosed with intestinal Behcet's at Tsukuba University Hospital between 1976 and 2001. Records were examined for medical treatment including dose of steroids, and surgical treatment including length of resected intestine, incidence of recurrence, and the form and site of recurrence.
Results: Elective surgery was performed in cases where the preoperative steroid administration could not be reduced to under 40mg/day, once steroids exceeded 40mg/day to control symptoms and inflammation. Cases with emergency operations (perforated cases), the preoperative doses of steroids were not so high (0, 0, 10, 20mg/day). In all cases, the post-surgical steroid dose was reduced under 20mg/day. Three cases showed recurrence after surgery within 2 years. These cases also had an increase in steroid dosage at the time of recurrence, but have been controlled medically so far.
Conclusions: Surgery should be strongly considered when the steroid dose exceeds 40mg/day.

Functionally distinct NKT cell subsets and subtypes
K Seino, M Taniguchi
JOURNAL OF EXPERIMENTAL MEDICINE, 202, 12, 1623, 1626, ROCKEFELLER UNIV PRESS, 2005年12月, ［査読有り］
英語, Natural killer T (NKT) cells are a population of autoreactive cells that mediate both protective and regulatory immune functions. NKT cells comprise several subsets of cells, but it has been unclear whether these different NKT cell subsets possess distinct functions in vivo. New studies now demonstrate that subsets of NKT cells are indeed functionally distinct and that the specific functions of these cells may be dictated in part by organ-specific mechanisms.

Suppression of IgE antibody responses by NKT cells-mechanisms of NKT cell-mediated regulatory function
Satoshi Kojo, Michishige Harada, Ken-ichiro Seino, Masaru Taniguchi
International Congress Series, 1285, 179, 183, 2005年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Vα14 natural killer T (NKT) cells exhibit various immune-regulatory properties in vivo. A repeated injection of α-galactosylceramide (α-GalCer) into mice induced suppression of antigen-specific IgE, IgG1 and IgG2a production in vivo. The suppression of antigen specific Ig responses seems to be due to the generation of regulatory DCs producing high IL-10 and low IL-12 by IL-10 derived from Vα14 NKT cells, because repeated α-GalCer stimulation changed cytokine profiles, such as high IL-10 and low IFN-γ production in Vα14 NKT cells. The unique cytokine profile (high IL-10 and low IL-12) in the regulatory DCs appeared to be regulated by upregulation of phosphorylation of extracellular signal-regulated kinase-1/2 and augmented production of IκBNS. The regulatory DCs thus induced in turn generated antigen specific Tr1 type regulatory T cells producing IL-10 suppressing Ig responses in an antigen specific fashion. © 2005 Elsevier B.V. All rights reserved.

Induction of regulatory properties in dendritic cells by V alpha 14 NKT cells
S Kojo, K Seino, M Harada, H Watarai, H Wakao, T Uchida, T Nakayama, M Taniguchi
JOURNAL OF IMMUNOLOGY, 175, 6, 3648, 3655, AMER ASSOC IMMUNOLOGISTS, 2005年09月, ［査読有り］
英語, 研究論文（学術雑誌）, V alpha 14 NKT cells exhibit various immune regulatory properties in vivo, but their precise mechanisms remain to be solved. In this study, we demonstrate the mechanisms of generation of regulatory dendritic cells (DCs) by stimulation of V alpha 14 NKT cells in vivo. After repeated injection of a-galactosylceramide (alpha-GalCer) into mice, splenic DCs acquired properties of regulatory DCs in IL-10-dependent fashion, such as nonmatured phenotypes and increased IL-10 but reduced IL-12 production. The unique cytokine profile in these DCs appears to be regulated by ERK1/2 and I kappa B-NS. These DCs also showed an ability to suppress the development of experimental allergic encephalomyelitis by generating IL-10-producing regulatory CD4 T cells in vivo. These findings contribute to explaining how V alpha 14 NKT cells regulate the immune responses in vivo.

Cutting edge: Critical role of CXCL16/CXCR6 in NKT cell trafficking in allograft tolerance
XF Jiang, T Shimaoka, S Kojo, M Harada, H Watarai, H Wakao, N Ohkohchi, S Yonehara, M Taniguchi, K Seino
JOURNAL OF IMMUNOLOGY, 175, 4, 2051, 2055, AMER ASSOC IMMUNOLOGISTS, 2005年08月, ［査読有り］
英語, 研究論文（学術雑誌）, It is well-documented that certain chemokines or their receptors play important roles in the graft rejection. However, the roles of chemokines and their receptors in the maintenance of transplantation tolerance remain unclear. In this study, we demonstrate that blocking of the interaction between the chemokine receptor, CXCR6, highly expressed on V alpha 14(+) NKT cells and its ligand, CXCL16, resulted in the failure to maintain graft tolerance and thus in the induction of acceleration of graft rejection. In a mouse transplant tolerance model, the expression of CXCL16 was up-regulated in the tolerated allografts, and anti-CXCL16 mAb inhibited intragraft accumulation of NKT cells. In vitro experiments further showed that blocking of CXCL16/CXCR6 interaction significantly affected not only chemotaxis but also cell adhesion of NKT cells. These results demonstrate the unique role of CYCL16 and CXCR6 molecules in the maintenance of cardiac allograft tolerance mediated by NKT cells.

Renal allograft rejection is prevented by adoptive transfer of anergic T cells in nonhuman primates
H Bashuda, M Kimikawa, K Seino, Y Kato, F Ono, A Shimizu, H Yagita, S Teraoka, K Okumura
JOURNAL OF CLINICAL INVESTIGATION, 115, 7, 1896, 1902, AMER SOC CLINICAL INVESTIGATION INC, 2005年07月, ［査読有り］
英語, 研究論文（学術雑誌）, Anergic T cells generated ex vivo are reported to have immunosuppressive effects in vitro and in vivo. Here, we tested this concept in nonhuman primates. Alloreactive T cells were rendered anergic ex vivo by coculture with donor alloantigen in the presence of anti-CD80/CD86 mAbs before adoptive transfer via renal allograft to rhesus monkey recipients. The recipients were briefly treated with cyclophosphamide and cyclosporine A during the preparation of the anergic cells. Thirteen days after renal transplantation, the anergic T cells were transferred to the recipient, after which no further immunosuppressive agents were administered. Rejection-free survival was prolonged in all treated recipients, and 3 of 6 animals survived long term (410-880 days at study's end). In the long-surviving recipients, proliferative responses against alloantigen were inhibited in a donor-specific manner, and donor-type, but not third-party skin allografts were also accepted, which demonstrated that antigen-specific tolerance had been induced. We conclude that anergic T cells generated ex vivo by blocking CD28/B7 costimulation can suppress renal allograft rejection after adoptive transfer in nonhuman primates. This strategy may be applicable to the design of safe clinical trials in humans.

V alpha 14 NKT cell-mediated anti-tumor responses and their clinical application
K Seino, S Fujii, M Harada, S Motohashi, T Nakayama, T Fujisawa, M Taniguchi
SPRINGER SEMINARS IN IMMUNOPATHOLOGY, 27, 1, 65, 74, SPRINGER, 2005年06月, ［査読有り］
英語, 研究論文（学術雑誌）, A unique lymphocyte population, V alpha 14 NKT cells, has recently been revealed to be a key player in the immune responses against tumors. Activation of V alpha 14 NKT cells affects various cell types, particularly dendritic cells (DCs), NK cells, CD4 Th1 cells, and CD8 cytotoxic T cells in the innate and acquired immune systems, eventually resulting in the enhanced activation of NKT cell-mediated cellular cascade in the anti-tumor responses. The specific ligand, alpha-galactosylceramide (alpha-GalCer), effectively stimulates mouse and human NKT cells, making NKT cells an ideal target for the development of cancer immunotherapy. Clinical trials using alpha-GalCer have actually started in several centers in the world. In this review, we summarize the Va14 NKT cell-mediated cellular cascade in the anti-tumor response in mice and discuss potential clinical applications of alpha-GalCer-pulsed DC therapy.

Generation of cloned mice by direct nuclear transfer from natural killer T cells
K Inoue, H Wakao, N Ogonuki, H Miki, KI Seino, R Nambu-Wakao, S Noda, H Miyoshi, H Koseki, M Taniguchi, A Ogura
CURRENT BIOLOGY, 15, 12, 1114, 1118, CELL PRESS, 2005年06月, ［査読有り］
英語, 研究論文（学術雑誌）, Cloning mammals by nuclear transfer (NT) remains inefficient. One fundamental question is whether clones have really been derived from differentiated cells rather than from rare stem cells present in donor-cell samples. To date, cells, such as mature lymphocytes, with genetic differentiation markers have been cloned to generate mice only via a two-step NT involving embryonic stem (ES) cell generation and tetraploid complementation [1-3]. Here, we show that the genome of a unique T-cell population, natural killer T (NKT) cells, can be fully reprogrammed by a single-step NT. The pups and their placentas possessed the rearranged TCR loci specific for NKT cells. The NKT-cell-cloned embryos had a high developmental potential in vitro: Most (71%) developed to the morula/blastocyst stage, in marked contrast to embryos from peripheral blood T cells (12%; p < 1 x 10(-25)). Furthermore, ES cell lines were efficiently established from these NKT-cell blastocysts. These findings clearly indicate a high level of plasticity in the NKT-cell genome. Thus, differentiation of the genome is not always a barrier to NT cloning for either reproductive or therapeutic purposes, so we can now postulate that at least some mammals cloned to date have indeed been derived from differentiated donor cells.

Effect of E6060 [4-{5-[7-Fluoro-4-(trifluoromethyl)benzo[b]furan-2-yl]-1H-2-pyrrolyl}benzoic acid], a novel subtype-selective retinoid, on lupus-like nephritis in female (NZBxNZW)F1 mice
T Yamauchi, A Ishibashi, K Shikata, N Tokuhara, K Seino, S Kobayashi, M Nagai
JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, 312, 3, 938, 944, AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS, 2005年03月, ［査読有り］
英語, 研究論文（学術雑誌）, Disease amelioration by retinoids in various nephritic models has been reported from either immunological or pathophysiologic viewpoints. It has also been reported that retinoids exert immunosuppressive effects in a retinoic acid receptor (RAR)-alpha-dependent manner. In particular, synthetic retinoid agonists with selectivity to RAR-alpha have been reported to have a remarkable disease-ameliorating effect in some immune disease models via their potent immunosuppressive activities; however, there has been no report in which the effect of RAR-alpha-selective agonists in the nephritic models was examined. In this report, we investigated the effect of a newly synthesized RAR-alpha-selective retinoid agonist, E6060 [4-{5-[7-fluoro-4-(trifluoromethyl)benzo[b]furan-2-yl]-1H-2-pyrrolyl}benzoic acid], on the disease progression in a murine lupus nephritis model. Female (NZBxNZW)F1 mice were prophylactically treated with E6060 from 5 months of age, and their nephritic (proteinuria, blood urea nitrogen) and immunological parameters (serum anti-DNA autoantibodies and total serum immunoglobulins) were monitored with age up to 10 months old. E6060 at 0.03 and 0.1 mg/kg (once daily, p.o.) significantly improved survival rate and prevented the development of proteinuria in (NZBxNZW) F1 mice. Anti-DNA autoantibodies and total serum IgG were also significantly reduced in the E6060-treated mice. Among IgG isotypes, IgG2a was substantially reduced by E6060 treatment, indicating reduced T helper 1 responses in E6060-treated mice. In accordance with this possibility, elevation of serum interleukin-12 (p40) in old female (NZBxNZW) F1 mice was significantly inhibited by E6060 treatment. Our data suggest that the RAR-alpha-selective retinoid E6060 is a promising candidate of new remedy for lupus nephritis in systemic lupus erythematosus patients.

NKT cells: A regulator in both innate and acquired immunity
Ken-Ichiro Seino, Masaru Taniguchi
Current Medicinal Chemistry: Anti-Inflammatory and Anti-Allergy Agents, 4, 1, 59, 64, 2005年02月, ［査読有り］
英語, CDd-restricted NKT cells are a unique subset of lymphocytes bridging innate and acquired immunity and mediating both effector and regulatory functions in immune responses. NKT cells are essential for the protection against pathogens or tumors, and also play a regulatory role in transplantation tolerance and autoimmune disease development. This review focuses on the various functions of NKT cells and discusses fundamental mechanisms in NKT cell biology. © 2005 Bentham Science Publishers Ltd.

Functional roles of NKT cell in the immune system
K Seino, M Taniguchi
FRONTIERS IN BIOSCIENCE, 9, 2577, 2587, FRONTIERS IN BIOSCIENCE INC, 2004年09月
英語, 研究論文（学術雑誌）, CD1d-restricted Natural Killer T cells (NKT cells), a novel lymphocyte lineage, are considered to play an intermediary role bridging innate and acquired immunity. This review discusses the characteristics of NKT cells and their biological significance in the immune system, and summarizes their in vivo functions observed in a number of pathological settings, including infectious diseases, cancer, autoimmunity, and transplantation. Further, we discuss recent data that have generated considerable interest in utilizing NKT cells as targets of new therapeutic interventions in various human diseases.

Role of a NK receptor, KLRE-1, in bone marrow allograft rejection: analysis with KLRE-1-deficient mice
E Shimizu, J Koike, H Wakao, K Seino, H Koseki, T Kakiuchi, T Nakayama, M Taniguchi
BLOOD, 104, 3, 781, 783, AMER SOC HEMATOLOGY, 2004年08月, ［査読有り］
英語, 研究論文（学術雑誌）, Natural killer (NK) cells play a pivotal role in the immune reaction during the bone marrow allograft. rejection. Little is known, however, about the molecular mechanisms underlying the NK cell-mediated allograft recognition and rejection. In this report, we assessed the role of a recently identified NK receptor, killer cell lectinlike receptor 1 (KLRE-1), by generating knockout mice. KLRE-1-deficient mice were born at an expected frequency and showed no aberrant phenotype on growth and lymphoid development. Nevertheless, KLRE-1 -deficient cells showed a severely compromised allogeneic cytotoxic activity compared with the wild-type cells. Furthermore, allogeneic bone marrow transfer culminated in colony formation in the spleen of KLRE-1-deficient mice, whereas no colony formation was born at an expected frequency and showed no aberrant phenotype on growth and lymphold development. Nevertheless, KLRE-1 -deficient cells showed a severely compromised allogeneic cytotoxic activity compared with the wild-type cells. Furthermore, allogeneic bone marrow transfer culminated in colony formation in the spleen of KLRE-1-deficient mice, whereas no colony formation was observed in wild-type recipient mice. These results demonstrate that KLRE-1 is a receptor mediating recognition and rejection of allogeneic target cells in the host immune system.

Prevention of acute and chronic allograft rejection by a novel retinoic acid receptor-alpha-selective agonist
K Seino, T Yamauchi, K Shikata, S Kobayashi, M Nagai, M Taniguchi, K Fukao
INTERNATIONAL IMMUNOLOGY, 16, 5, 665, 673, OXFORD UNIV PRESS, 2004年05月, ［査読有り］
英語, 研究論文（学術雑誌）, To investigate the involvement of retinoic acid receptor (RAR)-alpha in allograft rejection, we investigated the effect of a novel selective agonist to the receptor, ER-38925, in a mouse cardiac allograft model. Prophylactic treatment with ER-38925 inhibited the acute rejection of the mouse cardiac allograft (BALB/c --> C3H/HeN) at 0.3 and 3 mg/kg, and its effect was enhanced in combination with tacrolimus. In this model, ER-38925 remarkably inhibited cytotoxic T lymphocyte induction and alloantigen-stimulated production of cytokines, i.e. IL-2, IL-12 and IFN-gamma. In the chronic rejection model, combined treatment with tacrolimus and ER-38925 reduced the grade and incidence of arteriosclerosis in the cardiac allografts significantly more potently than tacrolimus monotherapy. ER-38925 inhibited the proliferation of rat aortic smooth muscle cells stimulated in vitro, presumably through the induction of a cyclin-dependent kinase inhibitor, p27(kip-1). Those results provide a rationale for using RAR-alpha agonists as immunosuppressants in human organ transplantation.

NKT cells are relatively resistant to apoptosis
K Seino, M Harada, M Taniguchi
TRENDS IN IMMUNOLOGY, 25, 5, 219, 221, ELSEVIER SCI LTD, 2004年05月, ［査読有り］
英語, 研究論文（学術雑誌）, CD1d-restricted natural killer T (NKT) cells are relatively resistant to activation-induced apoptosis, a characteristic that seems to be acquired in the late stage of intrathymic development. On strong antigen stimulation, peripheral NKT cells respond by downregulating their antigen receptor, while preserving their functional activities. Receptor downregulation might contribute to the homeostasis of the peripheral NKT-cell population and thus has an important regulatory role in the immune system.

Impaired IFN-γ production of V_α24 NKT cells in non-remitting sarcoidosis
KOBAYASHI Seiichiro, KANEKO Yoshikatsu, SEINO Ken-ichiro, YAMADA Yoshihito, MOTOHASHI Shinichiro, KOIKE Junzo, SUGAYA Kaoru, KURIYAMA Takayuki, ASANO Shigetaka, TSUDA Tomiyasu, WAKAO Hiroshi, HARADA Michishige, KOJO Satoshi, NAKAYAMA Toshinori, TANIGUCHI Masaru
International immunology, 16, 2, 215, 222, OXFORD UNIV PRESS, 2004年02月
英語, 研究論文（学術雑誌）, Sarcoidosis is a systemic disorder associated with granuloma characterized by an abnormal T(h)1-type cytokine production and accumulation of T(h)1 CD4 T cells in the granuloma lesions, suggesting an importance of T(h)1 responses in sarcoidosis. However, the pathogenesis of sarcoidosis remains to be solved. Here, we investigated the nature of V(alpha)24 NKT cells with immunoregulatory functions in sarcoidosis. Patients with non-remitting sarcoidosis displayed a decrease in the number of V(alpha)24 NKT cells in peripheral blood, but an accumulation of these cells in granulomatous lesions. When stimulated with the specific glycolipid ligand, alpha-galactosylceramide, peripheral blood V(alpha)24 NKT cells from patients with non-remitting disease produced significantly less IFN-gamma than those from healthy volunteers, but normal levels of IL-4. The reduced IFN-gamma production was observed only in V(alpha)24 NKT cells and not conventional CD4 T cells, but was normal in patients with remitting disease, suggesting that non-remitting sarcoidosis involves an insufficient IFN-gamma production of V(alpha)24 NKT cells which is well correlated with disease activity. Thus, these results suggest that V(alpha)24 NKT cells play a crucial role in the disease status of sarcoidosis.

Down-regulation of the invariant Vα14 antigen receptor in NKT cells upon activation
HARADA Michishige, SEINO Ken-ichiro, WAKAO Hiroshi, SAKATA Sakura, ISHIZUKA Yuko, ITO Toshihiro, KOJO Satoshi, NAKAYAMA Toshinori, TANIGUCHI Masaru
International immunology, 16, 2, 241, 247, OXFORD UNIV PRESS, 2004年02月
英語, 研究論文（学術雑誌）, NKT cells expressing the invariant Valpha14 antigen receptor constitute a novel lymphocyte subpopulation with immunoregulatory functions. Stimulation via their invariant Valpha14 receptor with anti-CD3 or a ligand, alpha-galactosylceramide (alpha-GalCer), triggers activation of Valpha14 NKT cells, resulting in a rapid cytokine production such as IFN-gamma and IL-4. Soon after their receptor activation, Valpha14 NKT cells disappeared as judged by staining with CD1d tetramer loaded with alpha-GalCer (alpha-GalCer/CD1d tetramer), which has been believed to be due to apoptotic cell death. Here we show that such a disappearance was largely attributed to down-regulation of the Valpha14 receptor. In fact, Valpha14 NKT cells were relatively resistant to apoptosis compared to the conventional T cells as evidenced by less staining with Annexin-V, a limited DNA fragmentation, and their preferential expression of anti-apoptotic genes such as NAIP and MyD118. Furthermore, they did not become tolerant, and maintained their proliferative capacity and cytokine production even after their receptor down-regulation. These as yet unrecognized facets of Valpha14 NKT cells are discussed in relation to their regulatory functions.

Down-regulation of the invariant V alpha 14 antigen receptor in NKT cells upon activation
M Harada, K Seino, H Wakao, S Sakata, Y Ishizuka, T Ito, S Kojo, T Nakayama, M Taniguchi
INTERNATIONAL IMMUNOLOGY, 16, 2, 241, 247, OXFORD UNIV PRESS, 2004年02月
英語, 研究論文（学術雑誌）, NKT cells expressing the invariant Valpha14 antigen receptor constitute a novel lymphocyte subpopulation with immunoregulatory functions. Stimulation via their invariant Valpha14 receptor with anti-CD3 or a ligand, alpha-galactosylceramide (alpha-GalCer), triggers activation of Valpha14 NKT cells, resulting in a rapid cytokine production such as IFN-gamma and IL-4. Soon after their receptor activation, Valpha14 NKT cells disappeared as judged by staining with CD1d tetramer loaded with alpha-GalCer (alpha-GalCer/CD1d tetramer), which has been believed to be due to apoptotic cell death. Here we show that such a disappearance was largely attributed to down-regulation of the Valpha14 receptor. In fact, Valpha14 NKT cells were relatively resistant to apoptosis compared to the conventional T cells as evidenced by less staining with Annexin-V, a limited DNA fragmentation, and their preferential expression of anti-apoptotic genes such as NAIP and MyD118. Furthermore, they did not become tolerant, and maintained their proliferative capacity and cytokine production even after their receptor down-regulation. These as yet unrecognized facets of Valpha14 NKT cells are discussed in relation to their regulatory functions.

Impaired IFN-gamma production of V(alpha)24NKT cells in non-remitting sarcoidosis
S Kobayashi, Y Kaneko, K Seino, Y Yamada, S Motohashi, J Koike, K Sugaya, T Kuriyama, S Asano, T Tsuda, H Wakao, M Harada, S Kojo, T Nakayama, M Taniguchi
INTERNATIONAL IMMUNOLOGY, 16, 2, 215, 222, OXFORD UNIV PRESS, 2004年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Sarcoidosis is a systemic disorder associated with granuloma characterized by an abnormal T(h)1-type cytokine production and accumulation of T(h)1 CD4 T cells in the granuloma lesions, suggesting an importance of T(h)1 responses in sarcoidosis. However, the pathogenesis of sarcoidosis remains to be solved. Here, we investigated the nature of V(alpha)24 NKT cells with immunoregulatory functions in sarcoidosis. Patients with non-remitting sarcoidosis displayed a decrease in the number of V(alpha)24 NKT cells in peripheral blood, but an accumulation of these cells in granulomatous lesions. When stimulated with the specific glycolipid ligand, alpha-galactosylceramide, peripheral blood V(alpha)24 NKT cells from patients with non-remitting disease produced significantly less IFN-gamma than those from healthy volunteers, but normal levels of IL-4. The reduced IFN-gamma production was observed only in V(alpha)24 NKT cells and not conventional CD4 T cells, but was normal in patients with remitting disease, suggesting that non-remitting sarcoidosis involves an insufficient IFN-gamma production of V(alpha)24 NKT cells which is well correlated with disease activity. Thus, these results suggest that V(alpha)24 NKT cells play a crucial role in the disease status of sarcoidosis.

Bone marrow allograft rejection mediated by a novel murine NK receptor, NKG2I
J Koike, H Wakao, Y Ishizuka, T Sato, M Hamaoki, K Seino, H Koseki, T Nakayama, M Taniguchi
JOURNAL OF EXPERIMENTAL MEDICINE, 199, 1, 137, 143, ROCKEFELLER UNIV PRESS, 2004年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Natural killer (NK) cells mediate bone marrow allograft rejection. However, the molecular mechanisms underlying such a rejection remain elusive. In previous analyses, it has been shown that NK cells recognize allogeneic target cells through Ly-49s and CD94/NKG2 heterodimers. Here, we describe identification and characterization of a novel murine NK receptor, NKG2I, belonging to the NKG2 family. NKG2I, which was composed of 226 amino acids, showed similar to40% homology to the murine NKG2D and CD94 in the C-type lectin domain. Flow cytometric analysis with anti-NKG2I monoclonal antibody (mAb) revealed that expression of NKG2I was largely confined to NK and NKT cells, but was not seen in T cells. Furthermore, anti-NKG2I mAb inhibited NK cell-mediated cytotoxicity, whereas cross-linking of NKG2I enhanced interleukin 2- and interleukin 12-dependent interferon-gamma production. Similarly, the injection of anti-NKG2I mAb before the allogeneic bone marrow transfer in vivo impinged on the function of NKG2I, resulting in the enhanced colony formation in the spleen. NKG2I is a novel activating receptor mediating recognition and rejection of allogeneic target cells.

The NKT cell system: bridging innate and acquired immunity
M Taniguchi, K Seino, T Nakayama
NATURE IMMUNOLOGY, 4, 12, 1164, 1165, NATURE PUBLISHING GROUP, 2003年12月, ［査読有り］
英語

Auxiliary partial orthotopic liver transplantation for fulminant hepatitis: Regeneration of the diseased native liver in a pig model
S Ishiguro, Y Takada, M Gu, K Fukunaga, H Taniguchi, K Seino, T Kawamoto, K Yuzawa, M Otsuka, T Todoroki, K Fukao
TRANSPLANTATION, 75, 11, 1901, 1904, LIPPINCOTT WILLIAMS & WILKINS, 2003年06月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. This study was, performed to develop a clinically relevant porcine model of auxiliary partial orthotopic liver transplantation (APOLT) for fulminant hepatic failure.
Methods. FHF was induced by intraportal administration of alpha-amanitin and lipopolysaccharide. Thereafter, pigs were divided into four groups. Group 1 was an untreated, control group. In group 2, pigs underwent only a left hemihepatectomy. Pigs in groups 3 and 4 received APOLT after hemihepatectomy. Tacrolimus was administered to pigs in group 4, but not to those in group 3.
Results. Two-week survival rates were 0%, 20%,40%, and 100% for groups 1, 2, 3, and 4, respectively. In group 4, after abolishing the graft function at 1 week, pigs survived for more than 2 weeks, and regeneration of the native liver was confirmed histologically.
Conclusions. Pigs suffering from fulminant hepatic failure could achieve long survival and liver regeneration with a temporary support of the auxiliary graft.

Impaired proliferative response of V alpha 24 NKT cells from cancer patients against alpha-galactosylceramide
K Yanagisawa, K Seino, Y Ishikawa, M Nozue, T Todoroki, K Fukao
JOURNAL OF IMMUNOLOGY, 168, 12, 6494, 6499, AMER ASSOC IMMUNOLOGISTS, 2002年06月, ［査読有り］
英語, 研究論文（学術雑誌）, Human invariant Valpha24(+) NKT cells are a relatively new subpopulation of lymphocytes. It has been reported that Valpha24 NKT cells are significantly involved in some human diseases. We have evaluated the number and function of Valpha24 NKT cells in both healthy volunteers and cancer patients. In this study we found that Valpha24 NKT cells in unfractionated PBMCs obtained from cancer patients did not respond efficiently to alpha-galactosylceramide (alpha-GalCer) in vitro. Thus, their proportion after stimulation with alpha-GalCer was smaller than that found in healthy volunteers. However, the cancer patients' Valpha24 NKT cells retained cytotoxic activity against malignant target cells, and they could efficiently proliferate to alpha-GalCer when fractionated by sorting. Furthermore, we found that addition of G-CSF to the culture could restore the low proliferative response of Valpha24 NKT cells from cancer patients. These results suggest that some functions of NKT cells in cancer patients are impaired, and this observation carries significant implications for immunotherapy-based cancer treatments.

Role of Fas/FasL pathway in the activation of infiltrating cells in murine acute myocarditis caused by Coxsackievirus B3
Y Seko, N Kayagaki, K Seino, H Yagita, K Okumura, R Nagai
JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY, 39, 8, 1399, 1403, ELSEVIER SCIENCE INC, 2002年04月, ［査読有り］
英語, 研究論文（学術雑誌）, OBJECTIVES This study was designed to investigate the roles of Fas/FasL pathway in myocardial damage in murine acute myocarditis caused by Coxsackie virus B3 (CVB3).
BACKGROUND Cardiac myocyte apoptosis rarely occurs in murine acute myocarditis caused by CVB3. Fas/FasL belong to the tumor necrosis factor receptor/ligand superfamily of costimulatory molecules and are known to play a critical role in the induction of apoptosis, as well as in the cytotoxicty mediated by T-cells and natural killer cells.
METHODS We first analyzed the expression of Fas on cardiac myoctyes in vivo and in vitro. Second, we examined the development of myocardial damage, in C3H/He mice treated with an anti-FasL monoclonal antibody (mAb), and in C3H/He-lpr/lpr mice and C3H/He-gld/gld mice infected with CVB3. Third, to investigate the effects of anti-FasL mAb treatment on the activation of the infiltrating cells, we examined the expression of interferon (IFN)-gamma and interleukin (IL)-2 as activation markets in the heart of mice by semiquantitative polymerase chain reaction.
RESULTS Fas was markedly, induced on cardiac myocytes with acute myocarditis. Myocardial inflammation was decreased in mice treated with anti-Fas L mAb, C3H/He-lpr/lpr mice and C3H/He-gld/gld mice. Anti-FasL mAb-treatment also decreased the expression of IFNgamma, IL-2, inducible nitric oxide synthase and CVB3 genomes in myocardial tissue.
CONCLUSIONS Our findings strongly suggested that the Fas/FasL pathway played a critical role in the development of massive myocardial necrosis through activation of infiltrating cells, and raise the possibility of immunotherapy by blocking the Fas/FasL pathway to prevent myocardial damage and improve the prognosis of patients with viral myocarditis. (J Am Coll Cardiol 2002;39:1399 - 403) (C) 2002 by the American College of Cardiology Foundation.

Effect of a novel potent cytokine suppressor on toxic liver syndrome in the pig treated by auxiliary partial orthotopic liver transplantation for fatal fulminant hepatic failure
S Ishiguro, Y Takada, K Fukunaga, Jiang, X, H Taniguchi, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Fukao
PROCEEDINGS OF THE 37TH CONGRESS OF THE EUROPEAN SOCIETY FOR SURGICAL RESEARCH, 235, 237, MEDIMOND S R L, 2002年, ［査読有り］
英語, 研究論文（国際会議プロシーディングス）, 'Toxic liver syndrome' can lead to considerable morbidity and mortality in some clinical cases of auxiliary partial orthotopic liver transplantation (APOLT) against fulminant hepatic failure (FHF). After inducing FHF to male pigs by the administration of alpha-amanitin and lipopolysaccharide, all of the pigs died within 5 days with clinically relevant features of FHF. Pigs under the same condition were rescued by APOLT 8 hours after induction of FHF. When the timing of APOLT was 24 hours after toxin injection, none of the pigs could survive more than 5 days. Additional treatment with a novel potent cytokine suppressor 'ONO-SM362' to APOLT prolonged the survival time of these pigs more than 2 weeks. Controlling cytokines being released from necrotic native liver is important to obtain good results of APOLT against FHF.

Treatment of advanced colorectal adenocarcinoma with weekly high-dose l-leucovorin and 5-fluorouracil
M Nozue, N Isaka, T Maruyama, T Kawamoto, K Seino, H Tanagichi, K Fukao
ONCOLOGY REPORTS, 9, 1, 93, 96, PROFESSOR D A SPANDIDOS, 2002年01月, ［査読有り］
英語, 研究論文（学術雑誌）, We report the results of 5-fluorouracil (5-FU) combined with high-dose l-folinic acid (leucovorin) therapy for patients with advanced colorectal carcinoma. In each treatment course, the patients weekly received both 5-FU (600 mg/m(2) by intravenous 15 min infusion) and l-folinic acid (250 mg/m(2) by intravenous infusion over a period of 2 h). A total of six treatments were administered with a 14-day interval to the next course of six treatments. Forty-eight patients were evaluated for toxicity and 32 for response. The combined complete and partial response rate was 25% in 32 patients. Toxicity was within acceptable limits without grade 4 toxicity. Although the response rate was slightly lower than those reported in phase II trials in Japan, the result was satisfactory. This therapy can be the standard chemotherapy for colorectal cancer patients, even in Japan.

Treatment and prognosis in colorectal cancer patients with bone metastasis
M Nozue, Y Oshiro, M Kurata, K Seino, N Koike, T Kawamoto, H Taniguchi, T Todoroki, K Fukao
ONCOLOGY REPORTS, 9, 1, 109, 112, PROFESSOR D A SPANDIDOS, 2002年01月, ［査読有り］
英語, 研究論文（学術雑誌）, We have reviewed the cases of every patient presenting with bone metastasis after colorectal surgery and tried to establish the features of this clinical entity and generate basic strategies to the therapeutic management of this condition. Of 928 primary tumor resected colorectal cancer patients, 12 (1.3%) were identified with bone metastasis and included in this study. The majority of primary tumors were located at the rectosigmoid portion of the colon. All cases were highly advanced at the time of diagnosis, including 8 cases of stage IV by TNM classification. Sites of metastatic tumors were concentrated in lumber or pelvic bones. At the onset of bone metastasis, 9 of the 12 cases had other metastatic sites. i.e., only 3 patients had bone metastasis alone. Survival after onset of bone metastasis was very poor, with a median survival of approximately 5 months and a 20% survival rate at I year. With regard to cause of death, seven patients died of pulmonary failure, one of liver failure, and one of DIC. Only 2 cases of solitary osseous metastasis have survived more than 1 year. In order to significantly improve prognosis, the early detection of bone metastases is important.

Transition and improvement in surgical treatment for rectal cancer during the last 21 years in our department
Y Oshima, M Nozue, H Taniguchi, KI Seino, N Koike, T Kawamoto, T Todoroki, K Fukao
INTERNATIONAL JOURNAL OF ONCOLOGY, 19, 6, 1283, 1286, PROFESSOR D A SPANDIDOS, 2001年12月, ［査読有り］
英語, 研究論文（学術雑誌）, The subject of this study was to examine the net effect of numerous changes in basic strategies, personnel and devices, upon the clinical courses and outcomes of rectal cancer patients. A total of 151 rectal cancer patients who underwent low anterior resection were divided into 4 groups (period I to 4) based upon the time period of the operation. They were compared among groups based upon the following para-meters: blood loss, operation time, incidence of leakage and urinary dysfunction, incidence of ileus, duration of nasogastric tube insertion, timing of initial oral feeding and survival. The blood loss during the operations, urinary dysfunction and duration of naso-gastric tube insertion tended to decrease in every period. Timing of initial oral feeding became faster. The operation times, incidence of leakage and ileus were nearly the same in each period. The 5-year survival rates on Dukes' C cases were 100% in period 4, 82.4% in period 3 and 50% in period 2. Survival rates became better. Our net outcome for rectal cancer treatment was satisfactory, because the survival rates became better under function preserving strategies.

Protection against Fas-mediated and tumor necrosis factor receptor 1-mediated liver injury by blockade of FADD without loss of nuclear factor-kappa B activation
K Seino, Y Setoguchi, T Ogino, N Kayagaki, H Akiba, H Nakano, H Taniguchi, Y Takada, K Yuzawa, T Todoroki, Y Fukuchi, H Yagita, K Okumura, K Fukao
ANNALS OF SURGERY, 234, 5, 681, 688, LIPPINCOTT WILLIAMS & WILKINS, 2001年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Objective To investigate the role of FADD (Fas-associated protein with death domain) in Fas and tumor necrosis factor receptor I (TNFR1)-mediated hepatic injury and inflammatory response in vivo.
Summary Background Data Fas and TNFR1 are cell surface molecules that trigger apoptosis or inflammation on engagement by a specific ligand or antibody. FADD is recruited to the cytoplasmic domain of these receptors on their activation and works as a common mediator to induce apoptosis. It is known that a blockade of FADD can inhibit apoptosis mediated by Fas or TNFR1 in vitro. However, it is not known whether the blockade can prevent organ injury and whether the inflammatory cascade is affected in vivo.
Methods A FADD deletion mutant lacking the death effector domain was introduced into mice by transduction with an adenovirus vector, and the effect of this FADD dominant negative mutant was examined in several liver injury models.
Results Hepatic injury induced by anti-Fas monoclonal antibody or tumor necrosis factor (TNF)-alpha plus D-galactosamine was markedly ameliorated by the FADD dominant negative transduction, which abrogated the death rate. Further, the FADD dominant negative transduction efficiently blocked T cell-mediated concanavalin A-induced hepatitis while not affecting TNF-alpha production or TNF-alpha -induced nuclear factor-kappaB activation in the liver.
Conclusions These results provide the basis for a novel therapeutic modality in which an unfavorable apoptotic process can be inhibited without affecting a favorable response for liver regeneration; this would be relevant to the clinical treatment of acute and chronic liver diseases as well as to some inflammatory disorders with hypercytokinemia, such as sepsis.

Living donor with type IV-A choledochal cyst in liver transplantation
Y Takada, T Hori, K Yuzawa, K Seino, H Taniguchi, M Otsuka, M Kaneko, K Fukao
TRANSPLANTATION, 72, 3, 551, 552, LIPPINCOTT WILLIAMS & WILKINS, 2001年08月, ［査読有り］
英語

Increased intracranial pressure in a porcine model of fulminant hepatic failure using amatoxin and endotoxin
Y Takada, S Ishiguro, K Fukunaga, M Gu, H Taniguchi, KI Seino, K Yuzawa, M Otsuka, T Todoroki, K Fukao
JOURNAL OF HEPATOLOGY, 34, 6, 825, 831, ELSEVIER SCIENCE BV, 2001年06月, ［査読有り］
英語, 研究論文（学術雑誌）, Background/Aims: The purpose of this study was to develop a clinically relevant porcine model of fulminant hepatic failure (FHF) by means of administration of amatoxin and endotoxin.
Methods: Pigs were intraportally administered only saline in group 1 (n = 3), 1 mug/kg of Lipopolysaccharide (LPS) in group 2 (n = 4), 0.1 mg/kg of alpha -amanitin in group 3 (n = 5), and amanitin plus LPS in group 4 (n = 9).
Results: All the pigs in groups 1 and 2 survived with minimal changes in liver function tests, In contrast to the 60% mortality in group,3, all the pigs in group 4 died within 96 h, with a significant increase in aspartate transaminase at 24 h (9757 +/- 2167 IU/I). In addition, they demonstrated severe metabolic disorders, such as serum lactate accumulation, hypoglycemia, coagulopathy, plasma amino acid imbalance, and hyperammonemia. The intracranial pressure significantly increased to 17.8 +/- 2.5 mmHg immediately before death. Reversal of FHF in these pigs following orthotopic liver transplantation confirmed that the toxicity is liver-specific and that the graft liver is unaffected.
Conclusions: This porcine model of FHF induced by a combination of amanitin and LPS will be of much use in the development of new therapies for human FHF. (C) 2001 European Association for the Study of the Liver. Published by Elsevier Science B.V. All rights reserved.

Role of platelet-activating factor in hepatectomy with Pringle's maneuver
M Gu, Y Takada, K Fukunaga, S Ishiguro, H Taniguchi, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Fukao
JOURNAL OF SURGICAL RESEARCH, 96, 2, 233, 238, ACADEMIC PRESS INC, 2001年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. Interruption of hepatic inflow is commonly used to reduce blood loss during extensive liver resection, but may cause liver dysfunction, The present study investigated the effects of platelet-activating factor (PAF) antagonist E5880 on total liver warm ischemia and 70% hepatectomy.
Methods. Rabbits were used in this study and were divided into four groups: group 1, those treated with only 70% hepatectomy; group 2, those treated with only 20 min Pringle's maneuver; group 3, those treated with both Pringle's maneuver and 70% hepatectomy without pretreatment; and group 4, those pretreated with PAF antagonist E5880 (0.3 mg/kg) followed by Pringle's maneuver and 70% hepatectomy. The remnant liver function was then evaluated after reperfusion.
Results. Seven-day survival rates in both groups 1 and 2 were 100%, E5880 treatment significantly increased 7-day survival rate (group 4: 38% vs group 3: 0%, P < 0.05) after a combination of Pringle's maneuver and 10% hepatectomy. The elevations of serum liver enzymes (GOT, GPT, mGOT, and LDH) were significantly inhibited in group 4 at 1 and 4 h after reperfusion. Portal venous pressure and the energy charge of liver were also significantly improved in group 4, compared with those in group 3. Endothelin-1 levels of arterial and portal venous blood progressively increased after reperfusion; however, there were no significant differences between the two groups. Leukocyte infiltration into the liver was significantly inhibited in group 4.
Conclusion. E5880 pretreatment has protective effects on liver function after 70% hepatectomy with Pringle's maneuver in rabbits. (C) 2001 Academic Press.

Requirement for natural killer T (NKT) cells in the induction of allograft tolerance
K Seino, K Fukao, K Muramoto, K Yanagisawa, Y Takada, S Kakuta, Y Iwakura, L Van Kaer, K Takeda, T Nakayama, M Taniguchi, H Bashuda, H Yagita, K Okumura
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 98, 5, 2577, 2581, NATL ACAD SCIENCES, 2001年02月
英語, 研究論文（学術雑誌）, In this study, we investigated the role of V alpha 14 natural killer T(NKT) cells in transplant immunity. The ability to reject allografts was not significantly different between wild-type (WT) and V alpha 14 NKT cell-deficient mice. However, in models in which tolerance was induced against cardiac allografts by blockade of lymphocyte function-associated antigen-1/intercellular adhesion molecule-1 or CD28/B7 interactions, long-term acceptance of the grafts was observed only in WT but not V alpha 14 NKT cell-deficient mice. Adoptive transfer with V alpha 14 NKT cells restored long-term acceptance of allografts in V alpha 14 NKT cell-deficient mice. The critical role of V alpha 14 NKT cells to mediate immunosuppression was also observed in vitro in mixed lymphocyte cultures in which lymphocyte function-associated antigen-1/intercellular adhesion molecule-1 or CD28/B7 interactions were blocked. Experiments using IL-4- or IFN-gamma -deficient mice suggested a critical contribution of IFN-gamma to the V alpha 14 NKT cell-mediated allograft acceptance in vivo. These results indicate a critical contribution of Va14 NKT cells to the induction of allograft tolerance and provide a useful model to investigate the regulatory role of V alpha 14 NKT cells in various immune responses.

The effect of alpha-galactosylceramide upon allogenic rejection
K Seino, K Yanagisawa, H Taniguchi, Y Takada, K Yuzawa, M Otsuka, K Fukao
TRANSPLANTATION PROCEEDINGS, 33, 1-2, 437, 438, ELSEVIER SCIENCE INC, 2001年02月, ［査読有り］
英語, 研究論文（学術雑誌）

Improvement of graft function without donor pretreatment in liver transplantation from non-heart-heating donors
M Gu, Y Takada, K Fukunaga, S Ishiguro, K Seino, H Taniguchi, K Yuzawa, M Otsuka, T Todoroki, K Fukao
TRANSPLANTATION PROCEEDINGS, 33, 1-2, 837, 838, ELSEVIER SCIENCE INC, 2001年02月, ［査読有り］
英語, 研究論文（学術雑誌）

Hepatic resection for metastatic tumors from noncolorectal carcinoma
Y Takada, M Otsuka, K Seino, H Taniguchi, N Koike, T Kawamoto, K Koda, S Adachi, K Yuzawa, M Nozue, T Todoroki, K Fukao
HEPATO-GASTROENTEROLOGY, 48, 37, 83, 86, H G E UPDATE MEDICAL PUBL LTD., 2001年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Background/Aims: The role of liver resection for hepatic metastases from noncolorectal carcinomas has yet to be clarified. The present study examines a single institutional experience of hepatic resection for noncolorectal metastases.
Methodology: From January 1987 to March 1999, 14 patients underwent curative resection for liver metastases from noncolorectal carcinomas. Records of these patients were reviewed.
Results: Resections were performed for liver metastases from gastric cancers (n = 8), pancreatic cancers (n = 2), and cancers of bile duct, the papilla of Vater, kidney, and breast (n = 1, each). Six patients (5 with gastric cancers and 1 with pancreas cancer) pre- sented with synchronous disease and 8 with metachronous disease. In the gastric cancer patients, there are 2 disease-free survivors (26 and 53 months) in the metachronous group, though all of the 5 patients with synchronous disease died within 29 months. All of the 4 patients with pancreatobiliary carcinomas died within 2 years. One case of breast cancer and another of renal cell cancer are alive without disease at 49 and 9 months, respectively.
Conclusions: For metastases from gastric cancers, better survival after hepatic resection is expected in metachronous cases than in synchronous cases. Hepatic resection may afford little benefit for patients with liver metastases from pancretobiliary cancers.

Idiopathic calcifying pancreatitis in a Japanese pediatric patient
K Seino, Nishimori, I, Y Nagai, H Inoue, Y Takada, S Adachi, T Todoroki, K Fukao
JOURNAL OF GASTROENTEROLOGY, 35, 12, 941, 944, SPRINGER-VERLAG, 2000年12月, ［査読有り］
英語, 研究論文（学術雑誌）, We recently experienced a rare case of chronic pancreatitis in a 13-year-old Japanese boy. Recently, in hereditary pancreatitis patients, some mutations have been identified in the trypsinogen gene. The purpose of this study was to investigate whether the same mutations could also be found in this patient. Polymerase chain reaction (PCR)-amplified products of his cationic and anionic trypsinogen genes were examined by direct sequence analysis. The gene analysis failed to show any mutation in any exons and their flanking intronic sequences of his trypsinogen genes. These findings indicate that the chronic calcifying pancreatitis in the present patient is "idiopathic", and thus a rare case of juvenile pancreatitis.

Peri- and postoperative kinetics of endothelin-1/big endothelin-1 and effects of endothelin antagonist in porcine liver transplantation from non-heart-beating donors
K Fukunaga, Y Takada, M Gu, S Ishiguro, K Seino, H Taniguchi, K Yuzawa, M Otsuka, K Goto, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 7, 1647, 1649, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Long-term graft survival of living-related kidneys after donor-specific transfusion
M Otsuka, K Yuzawa, Y Takada, H Taniguchi, K Fukunaga, K Seino, T Todoroki, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 7, 1741, 1742, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Inhibition of CD95 ligand-mediated inflammation
K Seino, T Tun, N Ohshima, H Hamada, K Yoshino, S Ikeda, K Fukunaga, H Taniguchi, Y Takada, K Yuzawa, M Otsuka, T Todoroki, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 7, 2038, 2039, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Porcine model of fulminant hepatic failure treated by liver transplantation
Y Takada, K Fukunaga, G Gei, S Ishiguro, H Taniguchi, K Seino, Y Yuzawa, M Otsuka, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 7, 2243, 2244, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Clonal colony formation of hepatic stem/progenitor cells enhanced by embryonic fibroblast conditioning medium
A Suzuki, H Taniguchi, YW Zheng, Y Takada, K Fukunaga, K Seino, K Yazawa, M Otsuka, K Fukao, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 32, 7, 2328, 2330, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Proliferative and functional ability of transplanted murine neonatal hepatocytes in adult livers
A Suzuki, H Taniguchi, YW Zheng, Y Takada, K Fukunaga, K Seino, K Yazawa, M Otsuka, A Yoshiki, M Kusakabe, K Fukao, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 32, 7, 2370, 2371, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Effects of combined growth factors on clonal growth and albumin secretion of murine fetal hepatocytes in low density culture
YW Zheng, H Taniguchi, A Suzuki, Y Takada, K Fukunaga, K Seino, K Yuzawa, M Otsuka, K Fukao, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 32, 7, 2372, 2373, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Effects of four extracellular matrices associated with growth factors on clonal culture and proliferation of murine fetal hepatocytes
YW Zheng, H Taniguchi, A Suzuki, Y Takada, K Fukunaga, K Seino, K Yuzawa, M Otsuka, K Fukao, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 32, 7, 2498, 2499, ELSEVIER SCIENCE INC, 2000年11月, ［査読有り］
英語, 研究論文（学術雑誌）

Pharmacologic graft protection without donor pretreatment in liver transplantation from non-heart-beating donors
M Gu, Y Takada, K Fukunaga, S Ishiguro, H Taniguchi, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Fukao
TRANSPLANTATION, 70, 7, 1021, 1025, LIPPINCOTT WILLIAMS & WILKINS, 2000年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. Non-heart-beating donors (NHBDs) are considered potential sources of transplant organs in an effort to alleviate the problem of donor shortage in clinical liver transplantation. We investigated the possibility of pharmacologic protection of hepatic allo-graft function from NHBDs without donor pretreatment.
Methods. Orthotopic liver transplantation was performed using pigs. In donors, cardiac arrest was induced by stopping the respirator. Forty-five minutes after cessation of the respirator, the liver was flushed with cold lactated Ringer's solution including heparin and with the University of Wisconsin (UW) solution, and then preserved for 8 hr at 4 degreesC in the UW solution. The pigs were divided into two groups: a control group and a treated group. In the treated group, an endothelin antagonist TAR-044 was added to the UW solutions (10 mg/L), and TAK-044 (10 mg/kg body weight) and a platelet activating factor antagonist E5880 (0.3 mg/kg body weight) were also administered to the recipients.
Results. TAK-044 and E5880 treatment significantly increased the 7-day survival rate of the recipients (100% vs. 17%, P<0.05), In the treated group, portal venous pressure immediately after reperfusion of the graft was significantly lower than in the control group, and postoperative increase in serum concentrations of glutamic oxaloacetic transaminase and total bilirubin was attenuated. Moreover, the energy charge and adenosine triphosphate concentration of the liver were rapidly restored after reperfusion.
Conclusions. Pharmacologic modulation with TAK-044 and E5880 avoiding donor pretreatment can improve the viability of hepatic allografts procured from NHBDs.

Clonogenic colony-forming ability of flow cytometrically isolated hepatic progenitor cells in the murine fetal liver
H Taniguchi, R Kondo, A Suzuki, YW Zheng, Y Takada, K Fukunaga, K Seino, K Yuzawa, M Otsuka, K Fukao, H Nakauchi
CELL TRANSPLANTATION, 9, 5, 697, 700, COGNIZANT COMMUNICATION CORP, 2000年09月, ［査読有り］
英語, 研究論文（学術雑誌）, Stem cells are defined as cells having multilineage differentiation potential and self-renewal capability. Hepatic stem cells have aroused considerable interest not only because of their developmental importance but also for their therapeutic potential. However, their presence in the liver has not yet been demonstrated. With the use of a fluorescence-activated cell sorter (FACS) and monoclonal antibodies, we attempted to ascertain whether hepatic stem cells are present in the murine fetal liver. For this purpose, we optimized a cell isolation technique for FAGS sorting of fetal liver cells. When isolated CD45(-)TER119(-) cells (the non-blood cell fraction in the fetal liver) were tested for their clonogenic colony-forming ability, mechanical dissociation (pipetting) was the most suitable cell isolation technique for FAGS sorting. We confirmed that these colonies contained not only cells expressing hepatocyte markers but also cells expressing cholangiocyte markers. To identify hepatic stem cells, studies must focus on CD45(-)TER119(-) cells in the murine fetal liver.

Improvement of allograft viability with organs procured from non-heart-beating donors in porcine liver transplantation
Y Takada, K Fukunaga, M Gu, S Ishiguro, H Taniguchi, K Seino, K Yuzawa, M Otsuka, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 2, 277, 278, ELSEVIER SCIENCE INC, 2000年03月, ［査読有り］
英語, 研究論文（学術雑誌）

Prolongation of mouse skin allograft survival by novel agonists selective for retinoic acid receptor-alpha
K Seino, T Yamauchi, A Ishibashi, N Tokuhara, S Kobayashi, K Fukunaga, H Taniguchi, Y Takada, K Yuzawa, M Otsuka, T Todoroki, K Fukao
TRANSPLANTATION PROCEEDINGS, 32, 2, 257, 258, ELSEVIER SCIENCE INC, 2000年03月, ［査読有り］
英語, 研究論文（学術雑誌）

Usefulness of flow-cytometric cell sorting for enrichment of hepatic stem and progenitor cells in the liver
H Taniguchi, A Suzuki, Y Zheng, R Kondo, Y Takada, K Fukunaga, K Seino, K Yuzawa, M Otsuka, K Fukao, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 32, 2, 249, 251, ELSEVIER SCIENCE INC, 2000年03月, ［査読有り］
英語, 研究論文（学術雑誌）

Attempts to reveal the mechanism of CD95-ligand-mediated inflammation
K Seino, T Ogino, K Fukunaga, H Taniguchi, Y Takada, K Yuzawa, M Otsuka, H Yagita, K Okumura, K Fukao
TRANSPLANTATION PROCEEDINGS, 31, 5, 1942, 1943, ELSEVIER SCIENCE INC, 1999年08月, ［査読有り］
英語, 研究論文（学術雑誌）

An endothelin receptor antagonist ameliorates injuries of sinusoid lining cells in porcine liver transplantation
K Fukunaga, Y Takada, G Mei, H Taniguchi, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Goto, K Fukao
AMERICAN JOURNAL OF SURGERY, 178, 1, 64, 68, CAHNERS PUBL CO, 1999年07月, ［査読有り］
英語, 研究論文（学術雑誌）, BACKGROUND: TAK-044 is an endothelin receptor antagonist. Whether the agent has protective effects on liver graft injuries from non-heart-beating donors is unknown.
METHODS: In donor pigs, cardiac arrest was induced by stopping the respirator. Forty-five minutes after cessation of the respirator, the liver was flushed with University of Wisconsin (UW) solution, preserved for 8 hours at 4 degrees C, and transplanted orthotopically. The pigs were divided into two groups: a control group and a drug-treated group in which TAK-044 was given in the UW solutions (10 mg/L) and was administered to recipients (10 mg/kg body weight).
RESULTS: TAK-044 treatment significantly increased recipient survival rate. After reperfusion of the graft, portal venous pressure and 15-minute retention rate of indocyanine green were significantly reduced in the drug-treated group. Electron microscopic findings indicated that TAK-044 attenuated endothelial cell injuries.
CONCLUSION: TAK-044 treatment improves the viability of livers harvested from non-heart-beating donors. The main effect of the agent is protection of endothelial cells from ischemia/reperfusion injuries. Am J Surg. 1999;178:64-68. (C) 1999 by Excerpta Medica, Inc.

Soluble forms of CD95 and CD95 ligand after living related liver transplantation
K Seino, N Kayagaki, N Yamaguchi, Y Takada, S Uyama, T Kiuchi, K Tanaka, H Yagita, K Okumura, K Fukao
TRANSPLANTATION, 67, 4, 634, 636, LIPPINCOTT WILLIAMS & WILKINS, 1999年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. Soluble forms of CD95 and CD95 ligand (sCD95 and sCD95L, respectively) can increase in the serum of patients with some inflammatory disease. In this study, we investigated the serum levels of sCD95 and sCD95L in liver transplantation recipients.
Methods. Serum levels of sCD95 and sCD95L in living related liver transplant recipients were analyzed by ELISA and their relation to the clinical findings estimated.
Results. Serum samples from the recipients did not show detectable levels of sCD95L but showed significantly increased levels of sCD95. The increase of sCD95 was positively associated with that of total-bilirubin and incidence of rejection, infection, and graft ischemia.
Conclusions. The present results indicate an existence of sCD95 in the recipients of living related liver transplants. The increased serum levels of sCD95 may modify the immunological situation of the recipients after transplantation or represent the ongoing graft damage.

Evidence for the presence of hepatic stem cells in the murine fetal liver
H Taniguchi, R Kondo, A Suzuki, Y Zheng, S Ito, Y Takada, K Fukunaga, K Seino, K Yuzawa, M Otsuka, K Fukao, A Yoshiki, M Kusakabe, H Nakauchi
TRANSPLANTATION PROCEEDINGS, 31, 1-2, 454, 454, ELSEVIER SCIENCE INC, 1999年02月, ［査読有り］
英語, 研究論文（学術雑誌）

Biological factors that affect CD95 ligand-mediated inflammation
K Seino, T Ogino, ST Ju, H Hamada, H Yagita, K Okumura, K Fukao
TRANSPLANTATION PROCEEDINGS, 31, 1-2, 893, 895, ELSEVIER SCIENCE INC, 1999年02月, ［査読有り］
英語, 研究論文（学術雑誌）

Endothelin antagonist treatment for successful liver transplantation from non-heart-beating donors
K Fukunaga, Y Takada, H Taniguchi, G Mei, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Goto, K Fukao
TRANSPLANTATION, 67, 2, 328, 332, LIPPINCOTT WILLIAMS & WILKINS, 1999年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Background. With the shortage of cadaveric donors, non-heart-beating donors (NHBDs) are a potential source of liver allografts, However, warm ischemic injury in NHBDs seriously affects the viability of graft liver. Endothelin (ET)-1 has been reported to be involved in the hepatic microcirculatory disturbances after ischemia-reperfusion.
Methods. In a porcine orthotopic liver transplantation model, changes in the serum and liver tissue ET-1 concentration were measured and the effects of an ET receptor antagonist, TAK-044, were evaluated. After cardiac arrest of the donors, liver allografts were subjected to 90 min of warm ischemia, flushed, and preserved for 4 hr at 4 degrees C. The pigs were divided into two groups: a control group (no drug treatment) and a drug-treated group, in which donors and recipients were treated with TAK-044 (10 mg/kg body, drip intravenous injection). Both groups had six donor/recipient pairs.
Results. The ET-1 concentration in the hepatic venous blood increased after reperfusion of the graft in the control group recipients. ET-1 in the graft, liver significantly increased during the cold preservation period. TBK-044 treatment significantly increased recipient 7-day survival rate. After reperfusion of the graft, the concentrations of serum liver enzymes and arterial lactate in the drug-treated group were significantly lower than in the control group. The postoperative increase in portal venous pressure was significantly reduced in the drug-treated group. Measurements of liver enzymes in the washed-out preservation fluid at the time of graft rinsing indicated that TAK-044 treatment of the donors significantly suppressed liver enzyme release during ischemia.
Conclusions. These findings indicate TAK-044 treatment has protective effects on postoperative function of hepatic allografts procured from NHBDs.

Cutting edge: Chemotactic activity of soluble Fas ligand against phagocytes
K Seino, K Iwabuchi, N Kayagaki, R Miyata, Nagaoka, I, A Matsuzawa, K Fukao, H Yagita, K Okumura
JOURNAL OF IMMUNOLOGY, 161, 9, 4484, 4488, AMER ASSOC IMMUNOLOGISTS, 1998年11月, ［査読有り］
英語, 研究論文（学術雑誌）, A recombinant soluble form of human Fas ligand (sFasL) was tested for its chemotactic activity against human and mouse polymorphonuclear neutrophils (PMN) by the Boyden chamber method, sFasL exhibited a potent chemotactic activity against both human and mouse PMN and HL-60 cells when differentiated into neutrophils or monocytes. A neutralizing anti-FasL mAb abolished the chemotactic activity, while control mAb did not, Ligation of Fas by either IgM- or IgG-type anti-Fas mAb also induced PMN migration, PMN derived from lpr mice that express few Fas molecules did not respond, to sFasL. In contrast, those derived from lpr(cg) mice that express Fas molecules with a mutated death domain normally responded to sFasL chemotaxis. These results directly indicated a chemotactic activity of sFasL against PMN and suggest a novel signaling function of Fas, which appears to be independent of the death domain-mediated apoptosis.

Can expression of CD95 (Fas/APO-1) ligand on grafts or tumor cells prevent their rejection?
J Allison, K Seino, H Yagita
SPRINGER SEMINARS IN IMMUNOPATHOLOGY, 19, 3, 311, 322, SPRINGER VERLAG, 1998年, ［査読有り］
英語, 研究論文（学術雑誌）

Contribution of Fas ligand to T cell-mediated hepatic injury in mice
KI Seino, N Kayagaki, K Takeda, K Fukao, K Okumura, H Yagita
GASTROENTEROLOGY, 113, 4, 1315, 1322, W B SAUNDERS CO, 1997年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Background & Aims: Fas has been implicated in liver damage, The aim of this study was to investigate the role of its ligand to induce hepatocyte death and liver damage in T cell-dependent hepatitis, Methods: Fas ligand-mediated lysis of primary hepatocytes from C57BL/6 wild-type, Fas ligand-deficient gld, and Fas-deficient lpr mice and concanavalin A-induced hepatitis in these mice were assessed. Results: Freshly isolated hepatocytes from wild-type or gld mice, but not those from lpr mice, were susceptible to Fas ligand-mediated lysis, When concanavalin A was intravenously administered into wild-type mice, they developed acute hepatic injury with massive degenerative changes in hepatocytes, In contrast, both gld and lpr mice had lower aminotransferase levels with milder histological changes, Reverse-transcription polymerase chain reaction and flow cytometric analysis showed that Fas ligand was induced in the liver shortly after the concanavalin A injection and was predominantly expressed on intrahepatic T cells, Administration of monoclonal antibody neutralizing mouse Fas ligand could reduce the aminotransferase increase, Conclusions: The results indicate that Fas ligand plays a role in the T cell-dependent hepatitis induced by concanavalin A administration.

Transplantation of CD95 ligand-expressing grafts - Influence of transplantation site and difficulty in protecting allo- and xenografts
K Seino, N Kayagaki, N Tsukada, K Fukao, H Yagita, K Okumura
TRANSPLANTATION, 64, 7, 1050, 1054, WILLIAMS & WILKINS, 1997年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Background, CD95 and its ligand (CD95L) have been implicated in the regulation of immune responses. Recently, it was reported that CD95L expression prevented rejection of allogeneic grafts transplanted under the kidney capsule. In contrast, we reported that enforced CD95L expression in subcutaneously grafted cells induced acute rejection even in the syngeneic or immunodeficient hosts, In this study, we investigated, whether the CD95L-expressing cells could be protected from rejection when transplanted under the kidney capsule,
Methods, CD95-negative cells (baby hamster kidney and L5178Y lymphoma cells) were transfected with CD95L cDNA to express functional CD95L, The cells were transplanted into skin or renal subcapsular space of immunocompetent or T cell-deficient nu/nu mice.
Results, The parental cells grew well in nu/nu or syngeneic mice but were rejected in allogeneic or xenogeneic immunocompetent mice, The CD95L transfectants; were rejected when transplanted subcutaneously in all types of nice studied, However, when transplanted under the kidney capsule, they survived in nu/nu or syngeneic nice but were rejected in allogeneic or xenogeneic immunocompetent mice.
Conclusions. These results imply that CD95L expression may not be sufficient to protect the grafts from rejection, and the survival of CD95L-bearing grafts is substantially influenced by the site of transplantation.

Immunoregulation via adhesion molecules in allogenic and xenogenic hepatocyte transplantation to Nagase's analbuminemic rats
H Horimoto, M Nozawa, N Kokudo, M Nakao, S Takahashi, M Miyasaka, K Seino, H Yagita, K Okumura
CELL TRANSPLANTATION, 6, 5, 535, 536, PERGAMON-ELSEVIER SCIENCE LTD, 1997年09月, ［査読有り］
英語, 研究論文（学術雑誌）, We investigated the effects of monoclonal antibodies (mAbs) against lymphocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) on intrasplenic allogenic and xenogenic hepatocyte transplantation (HCTx) to analbuminemic rats, Ten to 12-wk-old male Nagase's analbuminemic rats (RT1(l)) were used as recipients, Wistar/Shi rats (RT1(k)) were used as donors for allografts and BALB/C mice were used as donors for xenografts. The experimental groups were as follows: group 1, allo-HCTx (n = 7); group 2, allo-HCTx + antirat ICAM-1/antirat LFA-1 mAbs (1.0 mg/kg/day, for 7 days, respectively) (n = 6); group 3, xeno-HCTx (n = 5); group 4, xeno-HCTx + mAbs (antimouse LFA-1/antirat ICAM-1) (n = 5), group 5, xeno-HCTx + mAbs (antirat LFA-1/antimouse ICAM-1) (n = 5), Serum rat albumin levels were measured in groups 1 and 2, and serum mouse albumin levels were measured in groups 3, 4, and 5, as indicators of the function of grafted hepatocytes, In allotransplantation groups, the serum rat albumin levels in the mAbs-treated group were significantly higher than those in the control group for 6 wk after transplantation. In xenotransplantation groups, no increase in the serum mouse albumin levels was detected in any group. (C) 1997 Elsevier Science Inc.

Antitumor effect of locally produced CD95 ligand
KI Seino, N Kayagaki, K Okumura, H Yagita
NATURE MEDICINE, 3, 2, 165, 170, NATURE PUBLISHING CO, 1997年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Activation of the cell-surface antigen CD95 induces apoptosis of CD95-bearing tumor cells. In this study, we investigated the antitumor effect of locally produced CD95 ligand (CD95L) on CD95-negative tumor cells in vivo. Introduction of CD95L cDNA into murine tumor cells did not affect growth in vitro but caused rejection in vivo. Neutrophils were primarily responsible for this rejection. A CD8(+) T cell-mediated protective immunity against subsequent challenge with parental tumor cells was also elicited. These results provide evidence for the potential utility of CD95L in tumor eradication and also reveal a proinflammatory function of CD95L.

Rejection of Fas ligand-expressing grafts
K Seino, N Kayagaki, K Fukao, K Okumura, H Yagita
TRANSPLANTATION PROCEEDINGS, 29, 1-2, 1092, 1093, ELSEVIER SCIENCE INC, 1997年02月, ［査読有り］
英語, 研究論文（学術雑誌）

Lack of cognate help by CD4(+) T cells and anergy of CD8(+) T cells are the principal mechanisms for anti-leukocyte function-associated antigen-1 intercellular adhesion molecule-1-induced cardiac allograft tolerance
H Bashuda, K Seino, C Ra, H Yagita, K Okumura
TRANSPLANTATION, 63, 1, 113, 118, WILLIAMS & WILKINS, 1997年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Combined treatment with anti-leukocyte function-associated antigen-1 (LFA-1) and anti-intercellular adhesion molecule-1 (ICAM-1) monoclonal antibodies leads to allograft tolerance in murine cardiac transplantation. In the present study, we analyzed the mechanisms for this tolerance induction. In the tolerant mice, proliferative response of splenic T cells against donor-type cardiac myocytes and of CD8(+) T cells against donor-type alloantigens was impaired as compared with responses in naive or rejected mice, but was completely restored with exogenous interleukin 2. This suggests that class I-restricted CD8(+) T cells of tolerant mice were rendered anergic against donor-type alloantigens in the periphery. In contrast, proliferative response of CD4(+) T cells against donor-type alloantigens in vitro was comparable between tolerant and naive mice. When heart and skin grafts from the same donor (BALB/c [H2(d)]) were simultaneously transplanted to C3H mice (H2(k)), both were rejected within 29 days, even though the mice were similarly treated with anti-LFA-1 and anti-ICAM-1 monoclonal antibodies, In contrast, when heart graft from BALB/c and skin graft hom third-party donor (C57BL/6 [H2(b)]) were simultaneously transplanted to C3H mice under the same condition, the heart graft was accepted indefinitely and the skin graft was rejected. These findings suggest that the peripheral tolerance against cardiac allografts could be induced by selective inactivation of alloreactive CD8(+) T cells resulting from the lack of cognate help by CD4(+) T cells.

Contribution of Fas ligand to cardiac allograft rejection
K Seino, N Kayagaki, H Bashuda, K Okumura, H Yagita
INTERNATIONAL IMMUNOLOGY, 8, 9, 1347, 1354, OXFORD UNIV PRESS UNITED KINGDOM, 1996年09月, ［査読有り］
英語, 研究論文（学術雑誌）, Effector mechanisms for allograft injury remain unclear. In the present study, we verified the contribution of Fas and Fas ligand (Fast) to cardiac allograft rejection by utilizing the Fas-deficient lpr or FasL-deficient gld mice as the donor or recipient. Cardiac myocytes prepared from normal mice, but not those from lpr mice, constitutively expressed Fas and were susceptible to FasL-mediated lysis. Survival of cardiac allografts was substantially prolonged when gld or lpr mice were used as the recipient. In contrast, cardiac allografts from lpr mice were normally rejected without a delay. Histological examination of the grafts in the gld or lpr recipients demonstrated a lesser cellular infiltration and much milder myocyte damage. Proliferative response and cytotoxic T lymphocyte induction against the donor-type alloantigens were not impaired in the gld or lpr recipients. These results indicate a substantial contribution of Fast to cardiac allograft rejection, independent of Fas in the grafts, This raises a possibility that Fast may be more generally involved in tissue damage associated with various diseases than expected from the expression of Fas in the target organs.

Topical immunosuppression in skin grafting with FK 506 ointment
K Yuzawa, H Taniguchi, K Seino, M Otsuka, K Fukao
TRANSPLANTATION PROCEEDINGS, 28, 3, 1387, 1389, APPLETON & LANGE, 1996年06月, ［査読有り］
英語, 研究論文（学術雑誌）

Synergistic effects of mycophenolate mofetil (MMF, RS-61443) and anti-LFA-1/ICAM-1 monoclonal antibodies on the prolongation of heart allograft survival in rats
K Takazawa, Y Hosoda, H Bashuda, K Seino, H Yagita, T Tamatani, M Miyasaka, K Okumura
TRANSPLANTATION PROCEEDINGS, 28, 3, 1980, 1981, APPLETON & LANGE, 1996年06月, ［査読有り］
英語, 研究論文（学術雑誌）

Specific acceptance of cardiac allografts after treatment with antibodies to CD80 and CD86 in mice
H Bashuda, K Seino, M Kano, K Sato, M Azuma, H Yagita, K Okumura
TRANSPLANTATION PROCEEDINGS, 28, 2, 1039, 1041, APPLETON & LANGE, 1996年04月, ［査読有り］
英語, 研究論文（学術雑誌）

CD95 ligand in graft rejection - Reply
RC Duke, A Franzusoff, D Bellgrau
NATURE, 379, 6567, 682, 683, MACMILLAN MAGAZINES LTD, 1996年02月, ［査読有り］
英語

接着分子の制御による末梢性トレランス
場集 田寿, 清野 研一郎, 狩野 基, 佐藤 健志, 八木田 秀雄, 奥村 康
日本臨床免疫学会会誌 = Japanese journal of clinical immunology, 18, 6, 679, 679, 日本臨床免疫学会, 1995年12月31日
日本語

CD86 (B70/B7-2) ON ENDOTHELIAL-CELLS CO-STIMULATES ALLOGENEIC CD4(+) T-CELLS
K SEINO, M AZUMA, H BASHUDA, K FUKAO, T YAGITA, K OKUMURA
INTERNATIONAL IMMUNOLOGY, 7, 8, 1331, 1337, OXFORD UNIV PRESS, 1995年08月, ［査読有り］
英語, 研究論文（学術雑誌）, In vascularized organ transplantation, vascular endothelial cells (EC) confronting recipient T cells are potentially significant APC initiating cellular immune responses that lead to rejection. In the present study, we studied the ability of human EC to stimulate allogeneic T cells and the co-stimulatory molecules involved in this response. On both human umbilical vein endothelial cells (HUVEC) and microvascular endothelial cells (MVEC), MHC class I, intercellular adhesion molecule (ICAM)-1 and CD86 were constitutively expressed as assessed by flow cytometry, After IFN-gamma treatment, MHC class II expression was induced, and MHC class I and ICAM-1 were up-regulated, In contrast, the expression of CD86 was unchanged and CD80 was undetectable even after IFN-gamma treatment. Highly purified CD4(+) T cells proliferated in response to IFN-gamma-treated allogeneic HUVEC and MVEC, and this response was efficiently blocked by mAb to MHC class II, ICAM-1 and CD86. Furthermore, the addition of anti-CD86 mAb to the primary culture with allogeneic EC resulted in the induction of alloantigen-specific anergy. These results suggest that CD86 expressed on EC plays a critical role in initiating cellular immune responses to vascularized allografts and would be an important target for immune intervention.

IL-34, the rationale for its expression in physiological and pathological conditions
Ryo Otsuka, Haruka Wada, Ken-ichiro Seino, SEMINARS IN IMMUNOLOGY, 54, 2021年04月
IL-34 is a cytokine that shares one of its receptors with CSF-1. It has long been thought that CSF-1 receptor (CSF-1R) receives signals only from CSF-1, but the identification of IL-34 reversed this stereotype. Regardless of low structural homology, IL-34 and CSF-1 emanate similar downstream signaling through binding to CSF-1R and provoke similar but different physiological events afterward. In addition to CSF-1R, protein-tyrosine phosphatase (PTP)-. and Syndecan-1 were also identified as IL-34 receptors and shown to be at play. Although IL-34 expression is limited to particular tissues in physiological conditions, previous studies have revealed that it is upregulated in several diseases. In cancer, IL-34 is produced by several types of tumor cells and contributes to therapy resistance and disease progression. A recent study has demonstrated that tumor cell-derived IL-34 abrogates immunotherapy efficacy through myeloid cell remodeling. On the other hand, IL-34 expression is downregulated in some brain and dermal disorders. Despite accumulating insights, our understanding of IL-34 may not be even close to its nature. This review aims to comprehensively describe the physiological and pathological roles of IL-34 based on its similarity and differences to CSF-1 and discuss the rationale for its disease-dependent expression pattern., ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 英語, 書評論文,書評,文献紹介等

IL-34が大腸がんに与える影響の解析
小林拓斗, 韓ナヌミ, 和田はるか, 宮城洋平, 醍醐弥太郎, 醍醐弥太郎, 醍醐弥太郎, 清野研一郎, 日本インターフェロン・サイトカイン学会学術集会抄録集, 85th (CD-ROM), 2021年

ATLLにおけるM-CSFR発現とM-CSFR阻害剤の有効性(Potential anti-lymphoma effect of M-CSFR inhibitor in adult T-cell leukemia/lymphoma)　　　　　　　　　　　　　　　
菰原 義弘, Muhammad Baghdadi, 清野 研一郎, 奥野 豊, 野坂 生郷, 松岡 雅雄, 日本癌学会総会記事, 78回, P, 1014, 2019年09月
日本癌学会, 英語

卵巣癌転移におけるインターロイキン-34の潜在的役割
羽馬直希, バグダーディ ムハンマド, 小林拓斗, 梅山悠伊, 韓ナヌミ, 和田はるか, 清野研一郎, 日本免疫治療学会学術集会プログラム・抄録集, 16th, 2019年

Interleukin-34, がんの治療標的としての可能性
ムハンマド・バグダーディー, 清野研一郎, 臨床免疫・アレルギー科, 70, 4, 422, 428, 2018年10月, ［査読有り］, ［招待有り］
日本語, 記事・総説・解説・論説等（学術雑誌）

再生医療時代におけるあらたな移植免疫制御 (特集 iPS細胞技術を駆使したがん,感染症のあらたな制御)
大塚 亮, 清野 研一郎, 医学のあゆみ, 263, 11, 927, 931, 2017年12月23日
医歯薬出版, 日本語

IL-34 as a prognostic biomarker and a therapeutic target in cancer
Muhammad Baghdadi, Ken-ichiro Seino, CYTOKINE, 100, 58, 58, 2017年12月
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 英語, 研究発表ペーパー・要旨（国際会議）

肺がんにおいてIL-34とM-CSFの共発現が予後に及ぼす影響(Prognostic significance of IL-34 and M-CSF co-expression in lung cancers)　　　　　　　　　　　　　　　
遠藤 拓, Baghdadi Muhammad, 石川 浩三, 和田 はるか, 清野 研一郎, 鈴木 直, 宮城 洋平, 横瀬 智之, 高野 淳, 醍醐 弥太郎, 日本がん免疫学会総会プログラム・抄録集, 21回, 120, 120, 2017年06月
日本がん免疫学会, 英語

放射線刺激により誘発されるインターロイキン34の放射線治療抵抗性における潜在的役割について
遠藤 拓, Muhammad Baghdadi, 石川 浩三, 江澤 永倫子, 梅山 悠伊, 和田 はるか, 鈴木 直, 清野 研一郎, 聖マリアンナ医科大学雑誌, 45, 3, 173, 183, 2017年
<p>放射線療法は手術療法，化学療法と並び，三大がん治療法に挙げられているが，一方で腫瘍再発の起点となる放射線耐性が課題として残されている。がん細胞の薬剤耐性獲得機序には，がん細胞側の内因性機序と腫瘍微小環境（TME）内の骨髄系細胞との外因性機序とが説明されている。近年発見されたInterleukin 34（IL-34）は，マクロファージコロニー刺激因子（M-CSF）と受容体を共有し同様の生理活性を示すが，腫瘍を含む様々な病態への独自の関与が報告されている。最近我々は，肺癌細胞の抗がん剤耐性化にIL-34が上記の内因性，外因性機序の両者に関与することを示した。本研究では，その先行研究を基に，放射線治療適応である前立腺癌および直腸癌細胞株を用いて，放射線の単回および反復照射下にIL-34およびM-CSFの発現変動を調べた。その結果，放射線単回照射では両者の発現誘導が認められたのに対し，反復照射ではIL-34のみが時間経過に伴い顕著な発現誘導を認めた。さらにその機序は抗がん剤刺激と同様にNF-kBを介した経路であることが阻害剤を用いて示された。このことから，長期の放射線ストレスにIL-34が誘導され放射線治療耐性へ関与する可能性が示唆される。今後はさらに放射線暴露によるIL-34のTMEへの作用を精査するとともに，放射線療法との併用としてIL-34標的治療の可能性について探索してゆく。</p>, 学校法人 聖マリアンナ医科大学医学会, 日本語

多能性幹細胞を用いた免疫寛容誘導
大塚 亮, 和田 はるか, バグダーディー ムハンマド, 辻 飛雄馬, 清野 研一郎, 移植, 52, 6, 489, 494, 2017年
<p>Induced pluripotent stem cell (iPSC) -based technologies provide new opportunities in regenerative medicine to generate grafts for transplantation. The banking of iPSCs from donors with homozygous HLA haplotypes is planned in Japan, aiming to reduce immune reaction. Even though HLA-homozygous iPSCs are used, immune suppression would be inevitable because of minor antigen mismatches. A couple of studies concerning such immunological issues have been reported, including gene modification and immune regulatory cell induction from PSCs. Meanwhile, our research group has recently examined the concept that the recipient immune response against PSC-derived graft can be regulated by administration of immunoregulatory cells generated from the same PSC. This therapeutic approach prolonged the survival of PSC-derived allograft by suppressing host T cell proliferation and antibody production by B cells.</p><p>iPSC-based technologies provide us numerous benefits; however, considering their safety from an immunological point of view should be of great importance for the development and clinical translation of this novel techniques.</p>, 一般社団法人 日本移植学会, 日本語

IL-34と腫瘍
Baghdadi Muhammad, 遠藤 拓, 和田 はるか, 清野 研一郎, 臨床免疫・アレルギー科 = Clinical immunology & allergology, 67, 1, 88, 92, 2017年01月
科学評論社, 日本語

オモロイ生き物の分子生物学 長寿・がん化耐性動物ハダカデバネズミiPS細胞の腫瘍化耐性機構の解明　　　　　　　　　　　　　　　
宮脇 慎吾, 河村 佳見, 大岩 祐基, 清野 研一郎, 岡野 栄之, 三浦 恭子, 日本生化学会大会・日本分子生物学会年会合同大会講演要旨集, 88回・38回, [3W8, 1], 2015年12月
(公社)日本生化学会, 日本語

Induction of iNOS Expressed Macrophages from Mouse iPS cells That Contribute to Prolong Same iPS Cells-Derived Graft Survival in Allogeneic Recipients
H. Sasaki, H. Wada, K. Morita, K. -I. Seino, N. Shinohara, AMERICAN JOURNAL OF TRANSPLANTATION, 15, 2015年05月
WILEY-BLACKWELL, 英語, 研究発表ペーパー・要旨（国際会議）

移植における免疫制御の未来動向
清野 研一郎, 移植, 50, 1, 11, 15, 2015年
Transplantation has been developed based on the progress of immunosuppression. During the past half-century, various excellent immunosuppressants have been developed. Furthermore, the induction of immunological tolerance has been investigated to improve the outcome, and it has already been applied clinically. Turning our attention to the future, an age of cellular transplantation using pluripotent stem cells would be realistic. Even in this age, when other-derived pluripotent stem cells as the source of transplantation are used, the idea of immune regulation is important. The investigation of immune regulation using pluripotent stem cells has already been started., 一般社団法人 日本移植学会, 日本語

Induction of Regulatory Macrophage-Like Cells From Mouse iPS Cells That Can Contribute to Suppress Allogeneic Immune Responses.
H. Sasaki, H. Wada, H. Kudo, K. Nonomura, K. Seino, TRANSPLANTATION, 98, 285, 285, 2014年07月
LIPPINCOTT WILLIAMS & WILKINS, 英語, 研究発表ペーパー・要旨（国際会議）

Induction of Regulatory Macrophage-Like Cells From Mouse iPS Cells That Can Contribute to Suppress Allogeneic Immune Responses.
H. Sasaki, H. Wada, H. Kudo, K. Nonomura, K. Seino, AMERICAN JOURNAL OF TRANSPLANTATION, 14, 285, 285, 2014年06月
WILEY-BLACKWELL, 英語, 研究発表ペーパー・要旨（国際会議）

SY-7-5 多能性幹細胞を用いた再生医療時代の免疫制御(SY シンポジウム,第113回日本外科学会定期学術集会)
清野 研一郎, 日本外科学会雑誌, 114, 2, 2013年03月05日
一般社団法人日本外科学会, 日本語

B細胞から誘導したiPS細胞の樹立及び試験管内におけるリンパ球分化
和田 はるか, 香城 諭, 草間 千枝, 岡本 直樹, 佐藤 可野, 石塚 文平, 清野 研一郎, 北海道醫學雜誌 = Acta medica Hokkaidonensia, 87, 2, 2012年04月01日
日本語

免疫療法 (誌上ディベート 癌治療には免疫療法?アンチネオプラストン?)
清野 研一郎, アンチ・エイジング医学, 7, 5, 712, 716, 2011年10月
メディカルレビュー社, 日本語

Induction of Donor Specific hyporesponsiveness by Adoptive Transfer of Ex Vivo Expanded Anergic Cells
I. Koyama, H. Bashuda, K. -I. Seino, J. Yagi, H. Fujii, I. Nakajima, S. Fuchinoue, K. Okumura, S. Teraoka, AMERICAN JOURNAL OF TRANSPLANTATION, 11, 77, 77, 2011年04月
WILEY-BLACKWELL, 英語, 研究発表ペーパー・要旨（国際会議）

新しい再生医療へ向けた新規細胞の作製 : ガンマデルタAPCに関する研究
清野 研一郎, 第一三共生命科学研究振興財団研究報告集, 27, 0, 87, 91, 2011年
第一三共生命科学研究振興財団, 日本語

多能性幹細胞を用いた新しい免疫制御法に関する研究
清野 研一郎, 医科学応用研究財団研究報告, 30, 89, 91, 2011年
鈴木謙三記念医科学応用研究財団, 日本語

NotchシグナルによるNK細胞の分化と機能制御 (特集 NK細胞・NKT細胞の機能制御分子)
和田 はるか, 清野 研一郎, 臨床免疫・アレルギー科, 53, 2, 126, 132, 2010年02月
科学評論社, 日本語

Self-education by NK cells
Haruka Wada, Satoshi Kojo, Ken-Ichiro Seino, Immunotherapy, 1, 5, 738, 2009年09月
英語, 書評論文,書評,文献紹介等

Adjuvant effect of IL-7 in vaccine-mediated cancer immunotherapy
Haruka Wada, Satoshi Kojo, Ken-Ichiro Seino, Immunotherapy, 1, 5, 738, 739, 2009年09月
英語, 書評論文,書評,文献紹介等

Suppressor turns into helper
Haruka Wada, Satoshi Kojo, Ken-ichiro Seino, IMMUNOTHERAPY, 1, 5, 737, 737, 2009年09月
FUTURE MEDICINE LTD, 英語, その他

K3-36 卵巣癌培養細胞中の癌幹細胞の存在と抗癌剤感受性との関連性(高得点演題15 腫瘍,高得点演題プログラム,第61回日本産科婦人科学会学術講演会)
小林 陽一, 清野 研一郎, 細沼 信示, 大原 樹, 鈴木 直, 木口 一成, 石塚 文平, 板持 広明, 紀川 純三, 礒西 成治, 菊池 義公, 日本産科婦人科學會雜誌, 61, 2, 655, 655, 2009年
日本産科婦人科学会, 日本語

19 ナチュラルキラー(NK)T細胞を活性化する新規糖脂質の創製(口頭発表の部)
田代 卓哉, 冨宿 賢一, 清野 研一郎, 渡会 浩志, 谷口 克, 森 謙治, 天然有機化合物討論会講演要旨集, 48, 0, 109, 114, 2006年
NKT cell is a potent producer of immunoregulatory cytokines. T cell receptor of NKT cell recognizes CD1d protein-α-galactosylceramide complex, and activated NKT cell can produce the both Th1 (immunostimulant) and Th2 (immunosuppressant)-types cytokines. KRN7000 (2), developed by Kirin Brewery Co. Ltd., is a derivative of natural α-galactosylceramide, agerasphins (main component is agelasphin-9b, 1). KRN7000 is a strong stimulant to NKT cells. However, it induces both Th1 and Th2-type cytokines production of NKT cell by single stimulation. The analogue OCH (3), possessing shorter alkyl chains as developed by Yamamura et al., induces predominant production of interleukin (IL)-4, a key cytokine of Th2 type, over Th1 type cytokines. The total amount of IL-4, produced by NKT cells by stimulation with OCH, however, was less than that in the case of 2. For improvement of this problem, we developed two kinds of new galactosylceramides. One of them is those possessing a ring structure on the sphingosine chain. This analogue was synthesized from the known epoxide 6. The selective reductive-epoxide opening reaction to 6 afforded alcohol 7 or 12. The four-membered ring part was constructed from alcohol 7, whereas alcohol 12 was converted to the five-membered ring part. The resulting products were converted to the corresponding galactosylceramides respectively. Another kind of the analogues (21) were those possessing sulfonamide chain instead of the acyl chain. Those analogues could be synthesized easily from the commercially available phytosphingosine (17). Both analogues could make complexes with CD1d, and induce NKT cells to produce Th1 type cytokine predominantly., 天然有機化合物討論会実行委員会, 日本語

Preferential enhancement of immunosuppressive function of V alpha 14 NKT cells by liposome-encapsulated alpha-galactosylceramide
Y Ishii, R Nozawa, Y Takamoto, H Nishikawa, KI Seino, M Taniguchi, FASEB JOURNAL, 19, 4, A31, A31, 2005年03月
FEDERATION AMER SOC EXP BIOL, 英語, 研究発表ペーパー・要旨（国際会議）

活性化NKT細胞による寛容誘導性樹状細胞の誘導
香城諭, 清野研一郎, 原田通成, 渡会浩志, 若尾宏, 内田哲郎, 中山俊憲, 谷口克, 日本免疫学会総会・学術集会記録, 34, 336, 336, 2004年11月05日
(NPO)日本免疫学会, 日本語

BCG投与による活性化Vα14 NKT細胞のIgE産生抑制
原田通成, 清野研一郎, 真柄久美子, 石井保之, 香城諭, 若尾宏, 渡会浩志, 中山俊憲, 谷口克, 日本免疫学会総会・学術集会記録, 34, 335, 2004年11月05日
日本語

Prevention of acute and chronic allograft rejection by a novel retinoic acid receptor-alpha-selective agonist (vol 16, pg 665, 2004)
K Seino, T Yamauchi, K Shikata, S Kobayashi, M Nagai, M Taniguchi, K Fukao, INTERNATIONAL IMMUNOLOGY, 16, 7, 1053, 1053, 2004年07月
OXFORD UNIV PRESS, 英語, その他

Bone marrow allograft rejection mediated by the murine NK receptor NKG21/KLRE1 (vol 199, pg 137, 2004)
J Koike, H Wakao, Y Ishizuka, T Sato, M Hamaoki, K Seino, H Koseki, T Nakayama, M Taniguchi, JOURNAL OF EXPERIMENTAL MEDICINE, 199, 3, 435, 435, 2004年02月
ROCKEFELLER UNIV PRESS, 英語, その他

NKT細胞による樹状細胞を介した免疫制御機構の解析
香城諭, 清野研一郎, 原田通成, 渡会浩志, 石井保之, 石井崇太郎, 若尾宏, 中山俊憲, 谷口克, 日本免疫学会総会・学術集会記録, 33, 150, 2003年11月05日
日本語

NKT細胞の活性化によるVα14受容体のdown-regulation　　　　　　　　　　　　　　　
原田 通成, 清野 研一郎, 若尾 宏, 坂田 さくら, 伊藤 俊広, 香城 諭, 中山 俊憲, 谷口 克, 日本免疫学会総会・学術集会記録, 33, 151, 151, 2003年11月
(NPO)日本免疫学会, 日本語

担癌状態におけるNKT細胞低反応性のメカニズム
柳澤 和彦, 黒岩 憲二, 清野 研一郎, 谷口 克, 大河内 信弘, 日本外科学会雑誌, 104, 532, 532, 2003年04月30日
一般社団法人日本外科学会, 日本語

移植免疫寛容誘導におけるNKT細胞の役割　　　　　　　　　　　　　　　
清野 研一郎, 村本 賢三, 谷口 克, 中山 俊憲, 場集田 寿, 竹田 和由, 八木田 秀雄, 奥村 康, 深尾 立, 日本免疫学会総会・学術集会記録, 30, 86, 86, 2000年11月
日本語

PP1454 肺癌術後に血清AFPの高値を示し肝細胞癌との鑑別に難渋した巨大肝転移の一切除例
大城 幸雄, 榎本 剛史, 高田 泰次, 清野 研一郎, 谷口 英樹, 小池 直人, 幸田 圭史, 足立 信也, 大塚 雅昭, 轟 健, 深尾 立, 石川 成美, 飯島 達生, 日本消化器外科学会雑誌, 33, 7, 1258, 1258, 2000年07月01日
一般社団法人日本消化器外科学会, 日本語

An endothelin receptor antagonist ameliorates injuries of sinusoid lining cells in porcine liver transplantation (vol 178, pg 64, 1999)
K Fukanaga, Y Takada, G Mei, H Taniguchi, K Seino, K Yuzawa, M Otsuka, T Todoroki, K Goto, K Fukao, AMERICAN JOURNAL OF SURGERY, 178, 5, 436, 436, 1999年11月
CAHNERS PUBL CO, 英語, その他

移植免疫におけるNKT細胞の役割　　　　　　　　　　　　　　　
清野 研一郎, 村本 賢三, 谷口 克, 中山 俊憲, 八木田 秀雄, 奥村 康, 深尾 立, 日本免疫学会総会・学術集会記録, 29, 55, 55, 1999年10月
日本語

FADD-dominant negative遺伝子導入によるFas Ligand及びTNF誘導性臓器障害の治療　　　　　　　　　　　　　　　
清野 研一郎, 八木田 秀雄, 奥村 康, 深尾 立, 日本外科学会雑誌, 100, 臨増, 78, 78, 1999年02月
日本語

免疫寛容の誘導と維持における最先端 Fasリガンドの発現による免疫寛容誘導に関する研究　　　　　　　　　　　　　　　
清野 研一郎, 八木田 秀雄, 奥村 康, 深尾 立, 移植, 33, 総会臨時, 125, 125, 1998年10月
日本語

皮膚移植拒絶におけるFas/FasLの役割　　　　　　　　　　　　　　　
荻野 隆史, 清野 研一郎, 場集田 寿, 小端 哲二, 八木田 秀雄, 奥村 康, 森下 靖雄, 移植, 33, 総会臨時, 160, 160, 1998年10月
日本語

疾病とFas/Fasリガンド (免疫異常)
清野 研一郎, 八木田 秀雄, 免疫, 1997, 234, 240, 1997年08月
中山書店, 日本語

CD95 ligand in graft rejection
H Yagita, K Seino, N Kayagaki, K Okumura, NATURE, 379, 6567, 682, 682, 1996年02月
MACMILLAN MAGAZINES LTD, 英語, 速報，短報，研究ノート等（学術雑誌）

多発性骨髄腫の骨病変におけるIL-34の作用機序解明と治療応用に向けた研究
科学研究費助成事業
2017年04月01日 - 2020年03月31日
石川 浩三, 清野 研一郎
多発性骨髄腫（Multiple Myeloma : MM）は骨破壊病変を主徴候とする。CSF-1受容体の新規のリガンドであるIL-34と骨破壊病態の関与が近年注目されている。我々は、MMモデルマウスの実験系で、骨髄の炎症性サイトカインや間質細胞がMM細胞のIL-34発現を増強することを示した。またIL-34をknockdownしたところ破骨形成が阻害されることを示した。MM患者の骨髄検体においてCD138+細胞のIL-34発現性は様々であったが、破骨形成誘導能はIL-34中和抗体により抑制された。
以上からMM細胞に起因するIL-34はMM患者の骨破壊治療の有効な標的となる可能性が示唆される。
日本学術振興会, 基盤研究(C), 北海道大学, 17K09913

iPS細胞を用いたアロの壁と時空間を超える免疫制御法に関する研究
科学研究費助成事業
2017年06月30日 - 2019年03月31日
清野 研一郎, バグダーディ ムハンマド, 和田 はるか
ドナー脾臓細胞をレシピエントへ輸注することでドナー特異的に免疫寛容を誘導できることが明らかとなった。さらに脾臓に含まれるB細胞分画のみの移入によってドナー特異的に皮膚移植片の長期生着が達成可能であることがわかった。一方でドナー脾臓中のT細胞の移入は移植片の長期生着に寄与しなかった。また、免疫抑制剤投与下で生着を維持した皮膚移植片からiPS細胞を樹立し、多分化能（テラトーマ形成）、未分化マーカーの発現、増殖能等に異常がないことを確認した。
日本学術振興会, 挑戦的研究(萌芽), 北海道大学, 17K19691

人工多能性幹細胞由来のマクロファージ様免疫抑制性細胞を用いたぶどう膜網膜炎の制御
科学研究費助成事業
2016年04月01日 - 2019年03月31日
南場 研一, 清野 研一郎, 北市 伸義
マウス脾臓B細胞由来の人工多能性幹（iPS）細胞から誘導したマクロファージ様免疫抑制性細胞(iPS-SCs)をヒト視細胞間レチノイド結合蛋白由来合成ペプチド（hIRBPp）1-20抗原から作製したマウス実験的自己免疫性ぶどう膜網膜炎 (EAU)に腹腔内投与することにより、眼底所見による臨床学的重症度とHE染色による組織学的重症度は、ともに対照群に比べ有意に炎症が軽症化した。In vitroの実験でもマウスEAUから採取したhIRBPp1-20抗原特異的なCD4陽性T細胞とhIRBPp1-20抗原と抗原提示細胞にiPS-SCsを加えて共培養すると、CD4陽性T細胞の増殖は有意に抑制された。
日本学術振興会, 基盤研究(C), 北海道大学, 16K11310

ケロイドの免疫細胞治療を目指して！～制御性T細胞は炎症＆線維化を抑制する～
科学研究費助成事業
2014年04月01日 - 2017年03月31日
村尾 尚規, 舟山 恵美, 山本 有平, 小山 明彦, 七戸 龍司, 古川 洋志, 林 利彦, 齋藤 典子, 関堂 充, 清野 研一郎
ケロイドは慢性炎症性線維化疾患であり、CD4陽性T細胞などの炎症性細胞が病態に大きく寄与する。CD4陽性T細胞のサブセットの一つである制御性T細胞は炎症・免疫反応を制御し、抗線維化作用を有する。ケロイド内での炎症制御機構、線維化抑制機構について線維芽細胞ーCD4陽性T細胞共培養モデルを用いて検証した。制御性T細胞などが主に産生する炎症抑制性サイトカインIL-10は、ケロイド線維芽細胞への直接的な作用より、CD4陽性T細胞の各サブセット間の相互作用に寄与し、間接的にケロイド線維芽細胞に対して抗炎症作用、抗線維化作用を示す可能性が示された。
日本学術振興会, 基盤研究(B), 北海道大学, 26293379

悪性黒色腫における腫瘍免疫の破綻を解明する！～マウスリンパ浮腫モデルを用いて～
科学研究費助成事業
2013年04月01日 - 2016年03月31日
山本 有平, 古川 洋志, 小山 明彦, 林 利彦, 舟山 恵美, 村尾 尚規, 清野 研一郎, 七戸 龍司, 大芦 孝平, 齋藤 典子, 関堂 充
本研究は、我々が過去に報告したマウス後肢におけるin transit転移モデルを使用して、予後不良因子であるin transit転移の成立機構を解明することを目的とした。また、in transit転移モデルの基盤となるマウス後肢のリンパ浮腫モデルを改良し、安定的に浮腫が再現することを試みた。更にこのモデルを使用して、後肢浮腫モデルの分子生物学的解析を可能とした。リンパ管機能不全をきたした腫瘍微小環境における腫瘍免疫の破綻が、腫瘍の増殖や転移に影響を与えることが示唆された。
日本学術振興会, 基盤研究(B), 北海道大学, 25293361

ES細胞を用いた新しい免疫制御法に関する研究
科学研究費助成事業
2006年 - 2007年
清野 研一郎
NKT細胞は自然免疫と獲得免疫をつなぐ重要な免疫制御性T細胞の一つである。免疫制御におけるNKT細胞研究の今後の発展を考えた場合、NKT細胞の操作・移入により免疫寛容状態を人為的に誘導できるかどうかが重要なポイントである。またより強固で安全な免疫制御法を確立するためにはその際の分子メカニズムを詳細に知る必要がある。我々は本研究によりNKT細胞の操作・移入による人為的な免疫制御法の確立に向けて、ES細胞など未分化細胞からの樹状細胞やNKT細胞の誘導が可能であるかどうか検討した。またその際の分子メカニズムと制御法について検討を行った。
まず、マウスES細胞を試験管内でOP9細胞とGM-CSFを用いて培養することで樹状細胞様の細胞を分化・誘導し、NKT細胞との共培養に用いた。ES細胞由来樹状細胞はCD11cやCD80/86を発現しており、α-GalCerをパルスしてからNKT細胞と共培養するとサイトカイン産生を誘導することが可能であった。この際、IL-10存在下で培養すると、NKT細胞、樹状細胞ともに炎症性サイトカイン産生の能力が低下し、逆にIL-10産生性が亢進した。NKT細胞の繰り返し刺激による樹状細胞側のIL-10の誘導は、生体内の樹状細胞で見られるよりは軽度であった。
本研究で我々はPLA2のひとつiPLA2の異常により生体内でNKT細胞の比率が高まることを見出した。iPLA2の阻害剤BELを用いて胎仔胸腺培養を行うとNKT細胞の比率が増加し、NKT細胞の分化においてiPLA2の酵素活性が重要な役割を果たしていることが示唆された。骨髄幹細胞や胎児肝臓からの試験管内T細胞誘導においてNKT細胞を分化させることは困難であったが、このiPLA2機能を制御することでNKT細胞の分化効率が変化する可能性が示された。
日本学術振興会, 基盤研究(C), 聖マリアンナ医科大学, 18590478

細胞培養の改善に関する技術開発　　　　　　　　　　　　　　　
競争的資金

細胞分取に関する技術開発　　　　　　　　　　　　　　　
競争的資金

免疫制御に関する創薬　　　　　　　　　　　　　　　
競争的資金

癌の先端的治療　　　　　　　　　　　　　　　
競争的資金

移植に関する諸問題　　　　　　　　　　　　　　　
競争的資金

免疫実験一般　　　　　　　　　　　　　　　
競争的資金

臓器移植、癌、自己免疫疾患などの難病に対する近未来的治療法の一つとして、新しい免疫療法・細胞治療法を開発するための基盤的研究を展開している。現在は、糖脂質を認識し免疫調節作用を担うことが近年明らかとなったNKT 細胞系の生物学の解明に注力しており、その免疫制御のしくみなどについて研究　　　　　　　　　　　　　　　
競争的資金