海藻酸钠-多聚赖氨酸-海藻酸钠微囊内细胞浓度与胰岛素和C肽的释放

背景：微囊化已经普遍应用于各种实验研究,微囊包裹的干细胞治疗糖尿病问题也成为当今的热点,但是囊内包裹的细胞浓度与胰岛素释放量的关系成为目前需要解决的问题之一。目的：运用海藻酸钠-多聚赖氨酸-海藻酸钠微囊包裹胰岛素产生细胞,观察细胞浓度对胰岛素和C肽释放情况的影响。方法：制备大鼠胰腺损伤提取物,将小鼠骨髓间充质干细胞诱导分化为胰岛素产生细胞。免疫荧光和双硫腙染色鉴定诱导后细胞内胰岛素的表达。然后将胰岛素产生细胞制成浓度分别为1×10⁷ L^-1,5×10⁷ L^-1,1×10⁸ L^-1,5×10⁸ L^-1,1×10⁹ L^-1,5× 10⁹ L^-1的细胞悬液,气体吹喷法制成微囊,用6-羧基乙二酸荧光素检测囊内细胞活力,用葡萄糖刺激微囊内细胞检查其胰岛素和C肽的分泌情况。结果与结论：胰腺损伤提取物诱导后双硫腙染色和细胞免疫荧光鉴定出胰岛素产生细胞内有胰岛素的表达;胰岛素产生细胞制成的微囊直径约为400 µm,大小均一,6-羧基乙二酸荧光素检测到囊内细胞的活力很好。用葡萄糖刺激不同细胞浓度的微囊,发现细胞浓度为1×10⁸ L^-1时,胰岛素和C肽的分泌达到最高。

Release of insulin and C peptide associated with cell concentration in alginate-polylysine- alginate microcapsules

BACKGROUND:Microencapsulation has been widely used in various experimental studies, microencapsulated stem cell therapy for diabetic mellitus has become a hot issue, but the relationship between the wrapped cell concentration and the insulin release becomes one of the problems required to be solved.OBJECTIVE:To investigate the influence to the release of insulin and C peptide by the density of the insulin-producing cells (IPCs) encapsulated by alginate-polylysine-alginate microcapsules. METHODS:The extracts of rat pancreatic injury were prepared, and mouse bone marrow mesenchymal stem cells were induced to differentiate into IPCs. Immunofluorescence and dithizone staining were used to identify the expression of insulin in the induced cells. IPCs were prepared into cell suspension at 1×10⁷/L, 5×10⁴/L, 1×10⁸/L, 5×10⁸/L, 1×10⁹/L, 5×10⁹/L, and then prepared microcapsules by gas blowing spray. Cell viability was detected with 6-carboxyl fluorescein diacetate. Glucose was used to stimulate the cells in microcapsules and to examine the release of insulin and C peptide.RESULTS AND CONCLUSION:After extracts of rat pancreatic injury were induced, the immunofluorescence and dithizone staining have identified the insulin expression in IPCs; IPCs microcapsule was approximately 400 um diameter with uniform size, 6-carboxyl fluorescein diacetate detected that cell vitality was good. Glucose was used to stimulate the microcapsules at different cell densities, results found that the secretion of insulin and C peptide reach a peak at cell density of 1×10⁸/L.