The Survival Influence Experimental Study Of Exendin-4on Mouse Islet Cells Transplantation

Objectives:To clarify the mechanism of Exendin-4(Ex4) injection in mice after islet cells xenotransplantation,investigate the effects of Ex4on islet cells lifetime. Combined the diabetic mice islet celltransplantation under the kidney capsule model, through Ex4injection, look for ways to extend theislet cell lifetime in different species transplantation.Methods:The55SD rats were divided into two groups, determine the time of islet cells isolate bycollagenase digestion, and islet cells separation. During the islet cells isolate by collagenasedigestion. Diphenylthiocarbazne (DTZ) dyeing method was used in each10min from thebeginning, calculate the islet cells number. Choose the most numerous of islet cells time point as anoptimum point. Ficoll-PaqueTMPLUS medium was used in islet cells separate and purify, DTZ andAO-PI florescence analysis were used to identify the cells and evaluate activity.The70ICR mice (blood glucose level steady) were used to establish diabetes model, mice fasting12h before establish model, with Streptozotocin (STZ)200mg/kg by intraperitoneal injections.After treatment mice blood glucose level was determined every day in a week. Until maintained formore than5days of fasting blood glucose>11.1mmol/L or random blood glucose>16.7mmol/Lbut accompany polydipsia, polyphagia, polyuria as a standard for mice diabetes model successbuild.Islet cells were transplanted under the kidney capsule,3groups were random divided (controlgroup, low dose group and high dose group). In low dose group, Ex4were injected subcutaneouslyby0.1μg/(kg*d), In high dose group, Ex4were injected subcutaneously by0.4μg/(kg*d), controlgroup treated with PBS. Observe the change of blood glucose level. Serum were collected todetected insulin, Organ weight/BW coefficients, histopathology and immunohistochemical stainingwere investigated a week after injection. Insulin α chain protein expression was compared usingwestern-blot.Result:①An optimum point of collagenase digestion was30min.②Mice diabetes model success ratiowas72.86%.③After inject Ex4, random blood glucose was <16.7mmol/L in the1rdday in control group，continue to5days; Random blood glucose was <16.7mmol/L in the1rdday in lowdose group and continue to12days; Random blood glucose was <16.7mmol/L in the2thday inhigh dose group and continue to9days. Mice blood glucose was analyzed by statistics, treatgroups blood glucose level was significant lower than control group in day7th,8thand12th(P<0.05).However, there is no difference between high dose and low dose group (P>0.05).④Immunohistochemical staining results showed that the islet cells number in treat groups was morethan control group; Western-blot showed that insulin α chain protein was obviously expressionwithout control group.⑤Both low dose and high dose the insulin level were higher than controlgroup through ELISA assay.Conclusions:Ex4can inhibit the apoptosis of xenogeneic islet cell transplantation, prolong survival time andincrease secretion of insulin. Ficoll-PaqueTM PLUS can be used to extract and purify islet cells;Mice diabetic model success build ratio is high using STZ; Kidney capsule as a position oftransplantation has high feasibility.