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目的探讨细胞毒性T淋巴细胞相关抗原4免疫球蛋白(CTLA4Ig)和T细胞分化群40补体免疫球蛋白(CD40LIg)基因修饰骨髓间充质干细胞(MSCs)在异种胰岛移植排斥反应中的作用及其可能的机制。方法将SD大鼠胰岛细胞移植到Wistar糖尿病大鼠肾包膜下,建立SD-Wistar大鼠的异种胰岛移植模型,用携带CTLA4Ig和CD40LIg基因重组腺病毒感染的MSCs进行干预治疗。取造模成功的糖尿病大鼠45只,按随机数字表法分为3组(每组15只):对照组只植入胰岛细胞;单纯MSCs组植入胰岛细胞和MSCs;基因转染MSCs组植入胰岛细胞及CTLA4Ig和CD40LIg基因转染的MSCs。观察胰岛移植后糖尿病大鼠的生存情况、血糖变化和移植物病理形态学的改变,检测移植物CTLA4Ig、CD40LIg和胰岛素的表达以及胰岛移植大鼠细胞因子水平的变化。结果①糖尿病大鼠血糖在移植后2 d降至正常,对照组血糖平均在移植后8 d升高,单纯MSCs组和基因转染MSCs组血糖分别在21 d和49 d升高。②对照组、单纯MSCs组和基因转染MSCs组,移植物存活时间分别为(10.2±2.1)、(23.5±6.8)和(52.1±7.3)d,各组间比较差异有统计学意义(P<0.05);移植大鼠的生存时间分别为(21.3±5.3)、(49.7±8.5)、(95.8±13.8)d,各组间比较和移植物存活时间相似(P<0.05)。③对照组TNF-α和IL-2的水平在移植后1周内均急剧上升,而IL-4和IL-10的水平较移植前显著下降(P<0.01);2个治疗组细胞因子水平的变化和对照组正好相反。④2个治疗组移植物内可见成片的胰岛细胞团,未见明显淋巴细胞浸润,基因转染MSCs组移植物内可见CTLA4Ig、CD40LIg和胰岛素的表达。结论转移CTLA4Ig和CD40LIg基因修饰MSCs可抑制异种胰岛移植排斥反应,其效果优于MSCs单独应用。
Objective To investigate the role of cytotoxic T lymphocyte associated antigen 4 immunoglobulin (CTLA4Ig) and T cell differentiation group 40 complement immunoglobulin (CD40LIg) gene modified bone marrow mesenchymal stem cells (MSCs) in xenotransplantation rejection Possible mechanism. Methods SD rat islet cells were transplanted into the renal capsule of Wistar diabetic rats. The islet xenograft model of SD-Wistar rats was established. The MSCs infected with CTLA4Ig and CD40LIg recombinant adenovirus were used for intervention. Forty-five diabetic rats were randomly divided into three groups (n = 15): control group, only islet cells were implanted; MSCs group was implanted with islet cells and MSCs; MSCs group Transplantation of islet cells and MSCs transfected with CTLA4Ig and CD40LIg genes. The survival, blood glucose and histopathological changes of diabetic rats after islet transplantation were observed. The expression of CTLA4Ig, CD40LIg and insulin in the graft and the changes of cytokines in islet transplantation rats were observed. Results ① The blood glucose of diabetic rats decreased to normal 2 days after transplantation, and the average blood glucose of control group increased 8 days after transplantation. The blood glucose of MSCs group and MSCs group transfected with MSCs increased at 21 d and 49 d respectively. ② The survival time of MSCs group, MSCs group and gene transfected MSCs group were (10.2 ± 2.1), (23.5 ± 6.8) and (52.1 ± 7.3) days, respectively. The difference between the two groups was statistically significant (P <0.05). The survival time of the transplanted rats were (21.3 ± 5.3), (49.7 ± 8.5) and (95.8 ± 13.8) d, respectively. The survival time of the grafted rats was similar to that of the grafts (P <0.05). ③ The levels of TNF-α and IL-2 in control group increased sharply within 1 week after transplantation, while the levels of IL-4 and IL-10 decreased significantly (P <0.01); the levels of cytokines The change is opposite to the control group. ④ There were no obvious lymphocyte infiltration in the two treatment groups, and the expression of CTLA4Ig, CD40LIg and insulin were found in the MSCs group. Conclusion CTLA4Ig and CD40LIg gene-modified MSCs can inhibit the rejection of xenotransplanted islet allografts, and its effect is better than MSCs alone.