Exendin-4有益於胰島細胞移植體的作用機轉與以 磁振造影影像偵測及追蹤植入之胰島細胞

Abstract

移植人類胰島細胞對糖尿病是很有療效的方法，而面臨最大的問題是捐贈細胞不足。Exendin-4為糖尿病病人使用的處方藥物，此Glucagon-like peptide-1(GLP-1)異構物的藥理作用在刺激胰島素分泌、抑制升糖素分泌、增加□細胞的再生及減少□細胞的凋亡。本研究探討合併Exendin-4治療，對於不足量胰島細胞移植的效益，結果顯示：移植C57BL/6小鼠150顆胰島細胞至同種STZ藥物誘發糖尿病的小鼠後，合併Exendin-4治療後顯著降低其血糖；在移植後六個星期，合併Exendin-4治療組的β細胞質量約為沒有Exendin-4治療組的2.3倍；更進一步以Ki-67染色發現其胰島細胞移植體在一、四、六週可見細胞的增生標記蛋白，同時以TUNEL染色來分析其胰島細胞移植體在一、四、六週的細胞凋亡體，結果無法判斷細胞的增加，是由增生或凋亡路徑間的動態變化造成。另外，我們發展磁共振成像法觀察胰島細胞移植體的存活，首先，將已標記殼聚醣包覆氧化鐵奈米微粒(chitosan-coated superparamagnetic iron oxide nanoparticles)的胰島細胞移植於C57BL/6小鼠左腎包囊，再以磁共振成像掃描儀進行移植後追蹤；結果顯示移植後至第19週，以MRI掃描仍可偵測到左腎比右腎有30%~50%的訊號降低，進一步取下移植體以免疫組染色可確認鐵與胰島素的位置一致。為了觀察胰島細胞移植體免疫排斥情形，我們將雄性C57BL/6作為胰島細胞捐贈者而以BALB/c小鼠為受贈者，移植後以MRI掃描追蹤左腎發現至45天仍有低訊號，且在第45天時取出移植體以免疫染色確認鐵與胰島素位置一致，證明了移植體在第45天仍未被排斥。

Human islet transplantation is the most effective method for the treatment of diabetes. However, the most encountered problem is the lack of donor cells. Exendin-4, a glucagon-like peptide-1(GLP-1), has been used as a prescribed drug for diabetic patients. The GLP-1 is to stimulate insulin secretion, suppresses glucagons secretion, increases β-cell replication and neogenesis, and reduces β-cell apoptosis. The study aims to investigate if cotreatment of exendin-4 with the islet transplantation of a marginal number of fresh islets is beneficial for the hyperglycemic recipients. The results are as following: Approximately 150 C57BL/6 mouse islets were transplanted into the streptozotocin induced diabetic mice, and then the mice treated with or without exendin-4 for 6 weeks. At the 6th week, the grafts were removed for their β cell mass determination. Blood glucose levels in both groups were progressively decreased after transplantation, and the exendin-4-treated group had had lower blood glucose level than the ones without exendin-4 treatment. When compared to the controls, a 2.3-fold increase of the β-cell mass of the graft was also observed. These results indicated posttransplant exendin-4 treatment in the diabetic recipient with a marginal number of fresh islets not only expands the graft β-cell mass but also improves the transplantation outcome. At the 1st, 4th, and 6th week after transplantation, we used ki-67 staining to investigate cell prolifertation and TUNEL staining to analyze the apoptosis of pancreatic islet cell grafts. Upto now, the data are still unable to clarify the mechanisms of apoptosis or proliferation. In addition, we used a novel MRI contrast agent, chitosan-coated superparamagnetic iron oxide (CSPIO) nanoparticles, to monitor the mouse islet grafts. The male inbred C57BL/6 mice were used as donors and recipients for the islet transplantation. The grafts of CSPIO-labeled islets were found as distinct hypointense spots homogeneously located at the upper pole of the left kidney. The MRI scans revealed that the signal detected from the left kidney was 30% to 50% lower than those of the right kidney. The co-localization of iron and insulin in the removed graft was confirmed by the immunohistochemical stain. In order to investigate if immune rejection occurred in islet cell transplantation, we used C57BL/6 male mice as the islet cell donor and the BALB/c mice as recipient. At the 45th day after transplantation, a low signal detectable from the left kidney MRI scans and the iron and insulin immunohistochemical stain still retained demonstrating the grafts has not yet been excluded.