人α1-抗胰蛋白酶在胰岛β细胞移植中免疫抑制和保护作用的研究

 目的：探讨人α1-抗胰蛋白酶（hAAT）蛋白在胰岛β细胞移植中的免疫抑制和保护作用。方法：构建稳定表达hAAT蛋白的NIT-hAAT细胞系。将NIT-1细胞系和NIT-hAAT细胞系分别2次腹腔注射正常BALB/c小鼠，诱导细胞毒性T淋巴细胞(CTL)产生，将丝裂霉素处理后的2种细胞系与CTL混合培养，流式细胞术检测NIT-hAAT细胞凋亡情况；ELISA检测细胞因子表达；实时荧光定量PCR检测炎症因子mRNA表达。将2种细胞系分别植入糖尿病模型小鼠左肾包膜内，动态观察血糖和体重变化、血清中胰岛素和C肽水平以及移植部位的病理学变化。结果：CTL实验中，NIT-hAAT细胞受体鼠淋巴细胞的细胞毒作用较NIT-1细胞受体鼠明显减轻。hAAT具有减轻细胞凋亡、抑制炎症因子IL-1β、IL-6 mRNA的表达以及调节Th1/Th2细胞因子平衡的作用。NIT-hAAT细胞移植到糖尿病模型小鼠后，血糖明显下降并维持至28 d，血清中胰岛素和C肽含量明显升高，移植部位炎症细胞浸润明显减轻。结论：hAAT蛋白可减轻CTL对β细胞的杀伤作用，抑制炎症因子的表达，短期内可以抑制移植物免疫排斥反应，对胰岛β细胞移植治疗糖尿病具有明显的免疫抑制和保护作用。

Immunosuppressive and protective effects of human α1-antitrypsin on pancreatic β-cell transplantation

AIM：To study the immunosuppressive and protective effects of human α1-antitrypsin (hAAT) on pancreatic β-cell transplantation. METHODS： An NIT-1 cell line (NIT-hAAT) was constructed, which can stably express the protein of hAAT. The BALB/c mice were intraperitoneally injected with NIT-1 and NIT-hAAT cell lines twice to induce cytotoxic T-lymphocytes (CTL). The apoptotic situation, the cytokine expression, and the mRNA expression of inflammatory factors were examined after mixed culture of CTL with NIT-1 or NIT-hAAT cell line pretreated with mitomycin. Both cell lines were transferred into the left renal capsule of the diabetic mice to dynamically observe the changes of blood sugar and body weight, the serum levels of insulin and C-peptide, and the pathological changes of the transplanted sites. RESULTS： The results of extended CTL killing assay showed that the cytotoxic effect on NIT-hAAT cell acceptor mice was significantly reduced compared with NIT-1 cell acceptor mice. hAAT effectively reduced apoptosis, inhibited the mRNA expression of inflammatory factors IL-1β and IL-6, and adjusted the balance of Th1/Th2 cytokine expression. After NIT-hAAT was transplanted into the diabetic mice, blood glucose decreased obviously and maintained for 28 d. The serum levels of insulin and C-peptide increased obviously. The infiltration of the inflammatory cells in the transplanted sites significantly reduced. CONCLUSION：hAAT has the abilities of reducing cytotoxic effect of CTL on the β-cells, inhibiting inflammatory factor expression, and  stopping short-term immunological rejection of the acceptor. hAAT has obvious immunosuppressive and protective effects on pancreatic β-cell transplantation for treatment of diabetes.