脂肪干细胞在同种异体复合组织移植中的免疫调节作用及相关机制研究

Vascularized composite tissue allotransplantation (VCA) gained rapid development in transplantation field recently. More than hundreds cases had been performed worldwide, the key problem is rejection post-transplantation. Cytotherapy became a new immunosuppression method which also applied in VCA. Because of low expression of costimulatory molecules and immunomodulatory properties, BMSCs has often been used in transplantation. However, it is difficult for BMSCs to be largely harvested in clinical and need long proliferation period in vitro limit its use in clinical. ADSCs have a multilineage mesenchymal potential similar to BMSCs, it is abundant in fatty tissue which enable them to be largely harvested and less injury to donor. ADSCs, also, was proved possessing immunomodulatory ability, however, the mechanism has not been clearly answered, its application in VCA is rarely reported. In this study, we try to investigate the function of NF-κB pathway in ADSCs’ immunomodulatory process. Then we regulate the expression of key molecular targets IDO and PGE-2 in this pathway using transgenic techniques and investigate its function in VCA. Lastly, ADSCs and Triptolide was combined used for VCA immunosuppression.

同种异体复合组织移植（VCA）能修复重建复杂的组织缺损，其主要问题是术后的排斥反应。细胞治疗是移植领域新的治疗手段，也是目前唯一成功诱导免疫耐受的治疗方法。脂肪间充质干细胞（ADSCs）免疫原性较低且被证实具有免疫调节功能使其能够用于移植后的细胞治疗，但其免疫调节机制却一直未能清晰的阐述，而ADSCs在VCA中的免疫调节作用更是鲜有报道。NF-κB通路参与免疫反应早期和各种炎症反应，ADSCs表面的TLR受体也参与该通路，TLR下游IKK复合物是该通路的重要组成部分，而NF-κB激活后的靶基因IDO与PGE2也是免疫调节过程中的重要因子。本课题拟研究NF-κB信号通路在ADSCs免疫调节中的作用，观察ADSCs对T细胞的影响，并利用IDO和PGE-2构建转基因ADSCs进行免疫调节。之后进行体内试验观察ADSCs在VCA中的作用及其对嵌合体等指标的影响

研究内容：本实验通过脂肪干细胞与淋巴细胞系共培养体系，研究脂肪干细胞对淋巴细胞增殖的影响，并明确其中炎症相关蛋白、基因以及通路的作用机制。为进一步了解机制，通过脂肪干细胞条件培养液培养淋巴细胞，检测与淋巴细胞增殖相关的蛋白、基因的表达。明确NF-kB通路在免疫抑制中的作用后，利用慢病毒载体，构建过表达IDO的脂肪干细胞，研究其对淋巴细胞增殖和炎症反应的影响。最后通过大鼠后肢移植模型，研究脂肪干细胞在同种异体复合组织移植中的免疫抑制作用。.重要成果及科学意义：本实验研究结果发现，无论是脂肪干细胞还是脂肪干细胞条件培养液均能够明显抑制淋巴细胞的增殖，同时脂肪干细胞还能够抑制淋巴细胞 NF-kB通路相关分子的表达。我们成功构建了过表达IDO的脂肪干细胞，证实这一细胞能进一步抑制淋巴细胞增殖。通过动物体内实验，进一步证实了脂肪干细胞在同种异体复合组织移植中具有有效的免疫抑制作用。我们的实验不仅证明了脂肪干细胞对淋巴细胞的抑制作用，还发现了其作用相关的机制通路，这为脂肪干细胞在移植免疫、疾病免疫等方面提供了一定的科学基础，同时也为免疫药物的研发提供了一定的科学依据。