目的探讨用猪小肠黏膜下层(SIS)作支架,复合大鼠胰岛体外构建组织工程胰腺的可行性。方法用改良的Ⅴ型胶原酶消化法方法分离纯化大鼠胰岛并培养,再与SIS进行体外复合培养。然后用相差显微镜、透射电镜和扫描电镜检查胰岛的形态,用丫啶橙/碘化丙啶(AO/PI)双染法观察胰岛的生长及细胞活性,用胰岛素释放试验评价胰岛细胞的分泌情况。采用独立t检验,分别比较在高糖、低糖条件下,SIS组与对照组胰岛素释放的差异有无统计学意义。结果胰岛在SIS材料上黏附、生长,细胞功能活跃,AO/PI双染法显示平均存活率92﹪,高糖下胰岛素释放量为(37.79±4.27)mmol/L,明显高于对照组[(28.69±3.58)mmol/L,t=12.678,P=0.000]。胰岛在SIS上成复层生长,复合培养时间明显延长。结论 SIS与胰岛在体外复合培养可以构建出类似生物膜的组织工程胰腺,为进一步研究体内胰岛移植奠定基础。
Objective To investigate the feasibility of artificial pancreas construction with islets in a tissue scaffold of small intestinal submucosa(SIS).Methods Pancreatic islets were isolated and purified using a modified method with collagenase.The islets co-cultivated with SIS were observed by phase contrast microscope,electronic microscope and scanning electronic microscope.Islet growth and secretion function were evaluated by AO/PI staining and high-glucose stimulating test.Results The islets adhered to and grew on SIS,with active cell function.High-glucose stimulating insulin secretion was(37.79 ± 4.27) mmol/L in SIS group,significantly higher than that of control group [(28.69 ± 3.58) mmol/L,t = 12.678,P = 0.000].Islets grew well in layers,with a 92﹪ viability,and the survival time was significantly longer.Conclusion Coculture of SIS and islet in vitro can construct an artificial pancreas similar to the biomembrane,thus providing a base for further study of islet transplantation in vivo by tissue engineering technique.