| 摘要: | 移植物對抗宿主反應(GVHD)是一種在異體移植過後常見的排斥反應，特別是在骨髓移植手術後最常發生。在許多研究中指出人類間充質幹細胞(hMSCs)擁有免疫抑制作，所以近期有越來越多研究關於利用hMSCs治療免疫相關疾病，其中一種就是GVHD。在移植手術中利用hMSCs，像是骨隨幹細胞(BMSCs)或是脂肪幹細胞(ADSCs)，有相當顯著的保護能力可以有效保護移植的外來組織以防GVHD的發生。在此研究中，我們主要想要建立一套測試方式來檢測當我們利用不同種類的hMSCs去保護外來組織時是否也能有效的抑制GVHD的發生。我們探討(1)哪一種hMSCs可以更有效的抑制GVHD的發生。在本研究中我們選用人類羊水幹細胞(hAFSCs)、hADSCs和人類誘導多能幹細胞(hiPSCs, HPS0077)分化MSCs；(2)哪一種細胞生長環境(生物材料)能更有利於hMSCs 防止GVHD發生。為了要在與骨髓移植時相近的情況來模擬GVHD，我們先用hMSCs做成骨分化來得到成骨細胞。接下來我們將不同種類的hMSCs分別都與異體單核球共同培養，然後做活死細胞檢測，得知各種hMSCs都具有抗免疫反應。接下來我們更深入探討不同的細胞外基質(ECM)的細胞培養環境 (層粘連蛋白laminin-521, laminin-511, 人類重組-玻璃粘連蛋白rVN, 纖連蛋白fibronectin, 第一型膠原蛋白collagen type I and matrigel) 對於成骨細胞經過異體單核球共同培養後死亡率的減少以及對於hMSCs對於成骨細胞起的保護作用的差異。希望在未來hMSCs用來防止免疫系統作用的這種機制，特別是我們實驗室自己用hiPSCs分化出的MSCs有高度機會能夠應用於免疫相關疾病的治療。;Graft versus host disease (GVHD) is a condition that occurs after allogeneic transplants in patients, especially after bone marrow transplantation. Mesenchymal stem cells (MSC) have been increasingly examined for the treatment of immune-related diseases such as GVHD due to their immunosuppressive properties. The transplantation of human MSCs (hMSCs) such as bone marrow stem cells (BMSCs) and adipose-derived stem cells (ADSCs) has shown great effects to protect transplanted tissues or cells from GVHD. In this study, I aimed to establish the evaluation of GVHD effect on the insertion of various kind of hMSCs. I evaluated (1) which hMSCs were effective to suppress GVHD. The following hMSCs were selected in this study: human amniotic fluid stem cells (hAFSCs), hADSCs and human pluripotent stem cells (hiPSCs, HPS0077)-derived MSCs. I also investigated (2) which cell culture environment (biomaterials) supports GVHD treatment by hMSCs. The osteogenic differentiation from hMSCs was performed to obtain osteoblasts, which mimic the bone cells environment of the most happened GVHD situation. Then, hADSCs, hAFSCs and hiPSCs-derived MSCs were treated with allogenic mononuclear cells and subsequently, the live and dead staining of hMSCs were performed after allogenic mononuclear cell treatment. It is found that hADSCs, hAFSCs, and hiPSCs-derived MSCs had the ability to suppress the inflammation reaction by allogenic mononuclear cells. The effect to suppress the reaction by hMSCs is stronger in the following order: hAFSCs > hADSCs > hiPSCs-derived MSCs. We further evaluated the effect of ECM (extracellular matrix) proteins (laminin-521, laminin-511, recombinant vitronectin, fibronectin, collagen type I and Matrigel) on the decrease of dead cells of osteoblasts on ECM protein-coated dishes by treatment of allogenic mononuclear cells, where several kinds of hMSCs were co-cultured with the osteoblasts. The osteoblasts cultured on Matrigel-coated dishes showed more alive cells than those cultured on other ECM-coated dishes by treatment of allogenic mononuclear cells. For the MSCs which were used to prevent the immune attack, hAFSCs cultured on collagen showed more strongly immune tolerance. In the future, these immunosuppressive effects of hMSC, especially hiPSC-derived MSCs have high possibility to apply in other immune-related diseases. |
