同种异体胰岛细胞移植在糖尿病小鼠中的实验研究

Experimental Study of the Islet Cell Allogeneic Graft in the Diabetic Mice

目的通过建立糖尿病小鼠模型,观察同种小鼠胰岛细胞移植前后血糖、胰岛功能的变化,探索胰岛细胞分离、纯化、移植的方法及有效性。方法采用链脲佐菌素(Streptozotocin,STZ)复制糖尿病小鼠模型,经胆囊注入Ⅴ型胶原酶水浴消化胰腺并进行密度梯度离心以分离纯化胰岛细胞,并在消化液和Ficoll分离液中加入了大豆胰酶抑制剂(STI)和牛血清白蛋白组份Ⅴ(BSA),用双硫腙(DTZ)法判定胰岛纯度,锥虫蓝排斥试验评估胰岛存活率;糖尿病小鼠行同种异体胰岛细胞移植后,检测受体血糖、胰岛素及C肽变化,以判定胰岛功能。结果糖尿病小鼠模型胰腺形态有明显病理变化,同种小鼠胰岛纯化后细胞形态完好,使分离纯化后的胰岛产量由原来方法的(40.7±13.2)个提高到了(263.5±32.1)个(P<0.01),活性在95%以上,胰岛移植后,实验组48h后血糖明显下降,C肽水平上升。结论采用加用STI、BSA小鼠胰岛细胞分离纯化方法可增加胰岛细胞产量,同种小鼠胰岛细胞移植能明显降低糖尿病小鼠血糖水平,改善胰岛功能,从而达到治疗目的。

Objective To investigate the change of glucose and C-peptide after the islet transplantation in mice, and explore the methods and efficacy of the islet isolation, purification, and transplantation in mice. Methods The mouse diabetic model was induced by Streptozotocin （STZ）. Type V collagenase was injected through gallbladder to digest the pancreas, and the islets were isolated and purified by density gradient centrifugation method. Soybean trypsin inhibitor （STI） and BSA were added into the digestive and Ficoll separation solutions. The purity of the islets was judged by DTZ method. The viability of islets was determined by trypan blue test. After islet cell allogeneic transplantation in the diabetic mice, the changes in blood glucose, insulin, and C-peptide were tested to assessment the function of islets in recipients. Results There were obviously pathological changes in pancreas of the diabetic mice. The purified islets in the allogeneic mice were intact morphologically. The yield of islets was increased from （40.7±13.2） to （263.5±32.1） （P〈0. 01）, and the viability was beyond 95%. Forty-eight h after islet transplantation, the blood glucose in experimental group was significantly declined, and C-peptide increased. Conclusion The yield of islets was increased after STI and BSA were added into the.digestive and Ficoll separation solutions. Islet allogeneic transplantation can obviously decline the blood glucose and improve the β-cell function in diabetic mice.