2013-08-012024-05-14https://scholars.lib.ntu.edu.tw/handle/123456789/661208摘要：心臟移植為心臟衰竭末期病患之主要治療方式，但移植接受者(recipient)的長期存活率卻受限於心臟植體動脈硬化(transplant arteriosclerosis, TA)。TA 為一種加速性瀰漫性粥狀硬化(accelerated diffuse atherosclerosis)，冠狀動脈因內膜增生而阻塞，導致心肌缺氧。TA 的致病機轉主要為異體免疫反應(allogeneic reaction)，但目前臨床上使用的抗排斥藥物並無法有效防治 TA，是以發展新的免疫調控策略實屬迫切。 調節性 T 細胞 (regulatory T cell, Treg) 被證實可調控異體免疫反應而維持免疫耐受性。目前研究最詳盡的 Treg 包括 CD4+CD25+Foxp3+調節性 T 細胞 (Foxp3+ Treg) 與第一型調節性 T 細胞 (T regulatory type 1 cell, TR1)；前者主要靠 Foxp3 決定其免疫調控能力，後者則仰賴介白素-10 (interleukin-10 , IL-10)為免疫調控的介質。進期的研究發現 TR1 中之一亞群因具有同時分泌 IL-10 與干擾素-γ (interferon-γ, IFN-γ)的特性，而被命名為「類第一型調節性 T 細胞」(TR1-like cell)。本實驗室利用猪隻股動脈同種異體移植模式，已證實並發表骨髓間葉幹細胞 (mesenchymal stem cells, MSC) 可有效抑制 TA，並增加植體血管組織中的 IL-10+ 和 IFN-γ+細胞。我們的先期實驗(NSC101-2314-B-002-175)更進一步顯示，在猪隻周邊血液淋巴細胞混合培養反應(mixed lymphocyte reaction, MLR)中，MSC 可誘發具有 IL-10+IFN-γ+CD4+ 表現型的 T 細胞，此群 T 細胞可抑制異體抗原所引起之細胞增生反應，因而具有 TR1-like cell 的特質。然而，TR1-like cell 不同於 MSC，在於無法分泌前列腺素 E2 (prostaglandin E2 , PGE2)，且其免疫抑制效力僅為 MSC 的 25%。根據上述已發表與先期的實驗結果，我們假設 MSC 在 MLR 系統中誘發的 TR1-like cell 可於活體內維持免疫調控能力以減緩 TA。若補足予 PGE2 則可進一步加強 TR1-like cell 的免疫調控能力，因此，於猪隻股動脈同種異體移植模式中併用 PGE2 與 TR1-like cell，應可促進 TR1-like cell 控制TA 病變的效能。 本研究計畫為期 2 年，其目標為探索 TR1-like cell 治療 TA 的潛能，且發展 PGE2 與TR1-like cell 併用的最佳療程。首先，在 MLR 系統中我們將分析 TR1-like cell 在有無 PGE2存在下的免疫調控能力，同時探討 TR1-like cell 及 PGE2 兩者，於單一或合併存在時對樹突細胞(dendritic cells)與 T 細胞的影響。另一方面，我們將測試 TR1-like cell 在猪隻股動脈同種異體移植模式中對 TA 的抑制效果，並於移植後給予受移植猪隻不同療程的 PGE2，以闡明 PGE2 是否可強化 TR1-like cell 治療 TA 的效益；更進一步分析血管植體中產生的免疫調控相關基因，以釐清併用 PGE2 與 TR1-like cell 影響免疫耐受性的可能機轉。本研究計劃的成果將有助於改善目前植體血管病變之治療困境，展開移植病人長期存活的契機。<br> Abstract: Cardiac transplantation is one of the standard treatment modalities for end-stage heart failure. However, the long-term survival of recipients is limited by the presence of transplant arteriosclerosis (TA). TA is an accelerated, obliterative form of arteriosclerosis characterized by the diffuse, concentric intimal hyperplasia. It has been well accepted that allogeneic immune response plays a critical role in the pathogenesis of TA. However, immunosuppressive regimens that successfully control acute rejection do not prevent TA. This unmet clinic need suggests exploration of new immunomodulatory strategies is warranted. Regulatory T cells (Tregs) are a specialized subset of T cells critically involved in promoting and maintaining immune tolerance. The two most delineated subsets of Tregs are CD4+CD25+Foxp3+ Tregs and T regulatory type 1 (TR1) cells. While Foxp3+ Tregs critically depend on Foxp3 for their regulatory specialization, the signature of TR1 cells is production of IL-10. Furthermore, a subpopulation of TR1 cells that are phenotypically IL-10+IFN-γ+CD4+, are designated as TR1-like cells. We previously demonstrated that bone marrow-derived mesenchymal stem cells (MSCs) effectively control TA by enhancing IL-10+ and IFN-γ+ cells in a porcine femoral arterial transplantation model. In addition, in a mixed lymphocyte reaction (MLR) system composed of porcine peripheral blood mononuclear cells (PBMCs), we found that MSCs induce a subset of IL-10+IFN-γ+CD4+ cells, which confer resistance to allogeneic proliferation, and thus resemble TR1-like cells. However, MSCs constitutively secrete prostaglandin E2 (PGE2) whereas TR1-like cells are deficient in PGE2 and are 4-fold less potent than MSCs in suppressing MLR (supported by NSC101-2314-B- 002-175). Based on our preliminary data, we first propose that TR1-like cells generated by MSCs in vitro have the immunoregulatory potential to attenuate TA in vivo. Second, we hypothesize that PGE2 supplements can enhance the immunosuppressive potency of TR1-like cells, and thus co-administration of PGE2 mimetics may further potentiate the ability of TR1-like cells to control TA. In this 2-year proposal, we aim to investigate the immunomodulatory potential of TR1-like cells on TA, and to develop an optimal protocol by combining PGE2 and TR1-like cells for preventing TA. First, in the MLR, the immunosuppressive potency of TR1-like cells in the presence or absence of PGE2 will be assessed in vitro. In parallel, the phenotypic changes and cytokine production of dendritic cells and responder T cells in response to TR1-like cells and/or PGE2 will be elucidated. In the porcine femoral arterial transplantation model, the efficacy of TR1-like cells on TA will be assessed in vivo. Furthermore, PGE2 will be administered at different time periods after arterial transplantation. If PGE2 dose potentiate TR1-like cells in the treatment of TA, the candidate mediators in the vascular allografts will be analyzed. The output of this proposal will shed light on the underlying mechanisms of immunomodulation mediated by TR1-like cells and/or PGE2, and optimize a new approach for the treatment of TA.植體動脈硬化骨髓間葉幹細胞類第一型調節性 T 細胞前列腺素 E2免疫耐受性transplant arteriosclerosismesenchymal stem cellsTR1-like cellsPGE2immunomodulation