小鼠胰岛α细胞的缺失对移植胰岛功能的影响

目的　探讨胰岛α细胞的丢失对胰岛素分泌功能的影响。方法　在人胰岛分离过程中 ,经胶原酶的过度消化造成胰岛周边α细胞缺失 ,用免疫组化及胰岛素 /胰高糖素含量分析予以证实。在体外 ,检测葡萄糖刺激引起的实验性糖尿病小鼠胰岛素的分泌 ;在体内 ,评价胰岛移植后对糖尿病小鼠血糖的调节 ,并研究胰高糖素对α细胞缺失胰岛功能的影响。结果　胶原酶的过度消化引起胰岛周边的α细胞丢失 ,使分离胰岛内胰岛素 /胰高糖素比值明显升高。与正常胰岛相比 ,α细胞缺失胰岛对葡萄糖刺激产生的胰岛素明显降低 ,胰岛素释放在正常胰岛和α细胞缺失胰岛分别为2 2 2 7uU/ml± 32 1uU/ml和 12 4 6uU/ml± 12 6uU/ml(P <0 0 1)。在体内 ,移植α细胞缺失的胰岛到糖尿病的C5 7/BL小鼠后不能有效地纠正动物的高血糖 ,胰岛移植后的平均血糖浓度在正常胰岛组和α细胞缺失胰岛组分别为 :8 9mmol/L± 1 98mmol/L和 2 1 3mmol/L± 2 2mmol/L(P <0 0 1) ,而给予外源性的胰高糖素可以在体外及体内明显改善α细胞缺失胰岛的胰岛素分泌功能。结论　胰岛α细胞的缺失明显降低胰岛的胰岛素分泌功能 ,用外源性的胰高糖素可以改善α细胞缺失胰岛的胰岛素分泌功能。

Impact of islet alpha cell loss on insulin secretion

OBJECTIVE: To investigate the impact of islet alpha cell loss on insulin secretion. METHODS: Human pancreatic tissues underwent over-digestion of collagenase. Digestion was terminated in half of the tissue suspension, and half of the suspension was put into solution of collagenase to undergo over-digestion so as to cause loss of islet alpha cells. The tissues of digested human pancreas and over-digested human pancreas were fixed and immunohistochemical staining was made to observe the loss of islet alpha cells. The contents of insulin and glucagon in solutions of normal human pancreas, human islet of pancreas with normal enzyme secretion, and over-digested human islet were measured by radioimmunoassay. The ratios of insulin/glucagon in these three solutions were calculated so as to analyze the loss of alpha cells. Different human pancreatic tissues were inoculated in 24-well plate. Glucose solutions of different concentrations were added into the suspensions. The concentration of insulin in the supernatant was measured after incubation. C57/BL Normal human pancreas and human pancreas with islet alpha cell loss were transplanted into the renal capsules of two groups of 5 C57/BL mice with experimental diabetes mellitus respectively. The blood insulin was measured for two days. On the 3(rd) day, the left kidneys of the mice were taken to undergo insulin/glucagon immunohistochemical staining to observe the loss of alpha cells. Human islets with alpha cell loss were transplanted into the renal capsules of another 10 mice. Then 5 of them received injection of normal saline and the other 5 received glucagon for 7 days, then glucagon was injected into the mice of normal saline group, and vise versa. The blood sugar was measured every day. RESULTS: Glucagon immunohistochemistry showed alpha cell loss from islet periphery in over-digested human pancreas. The insulin/glucagons ratios were 100:2.34 and 100:2.05 in the normal pancreas and isolated normal islets respectively, and was 100:0.314 in the over-digested islets, showing a loss of > 80% of alpha cells. As compared with normal islets, glucose-induced insulin secretion was significantly decreased. The insulin concentration was 222.7 micro U/ml +/- 32.1 micro U/ml in the suspension of cultured normal pancreatic cells and was 124.6 micro U/ml +/- 12.6 micro U/ml in the supernatant of suspension of pancreatic islets with alpha cell loss (P < 0.01). The average blood sugar was 8.9 micro U/ml +/- 1.98 micro U/ml in mice grafted with normal islets and was 21.3 micro U/ml +/- 2.2 micro U/ml in mice grafted with alpha cell deficient islets (P < 0.01). However, insulin secretion in islets with alpha cell loss was significantly improved by exogenous glucagon in vitro and in vivo. CONCLUSION: The function of insulin secretion in islets with alpha cell loss is significantly decreased. Insulin secretion of islets with alpha cell loss can be improved by exogenous glucagon.