Study Of Biological Scaffold Application In Stem Cell Differentiation And Islet Culture

Background and Objective It is one of the hot spots to study adult stem cells (ASCs) in the field of type1diabetes research. And there is no further study on the induction of ASCs to islet-like tissue with3D culture. This study is attempted to explore the feasibility of a new culture method for the differentiation of ASCs to islet-like tissue, i.e. PLGA scaffold combined with ECM, for a better technical resource of tissue engineering. Method Bone marrow derived stem cells (BMSC) were isolated from mouse, and divided into3groups:2D culture、ECM culture and PLGA scaffold combined with ECM culture. The induction period lasted10days, and the culture cells were collected for further identification, via cell morphology and immunology and function tests. Results Those were demonstrated by inverted microscope, scanning and transmission electron microscopy, immunohistochemistry, and radio immunoassay of insulin stimulated by glucose as follows. Many cells developed DTZ-positive vesicles in the cytoplasm, and held an ultrastructure in accordance with β cells. The results from scanning electron microscopy showed the compatibility of the cells and the scaffolds. With immunohistochemistry staining, some of the cells showed positive for insulin, C-peptide, glucagon, and somatostatin. The glucose stimulating tests suggested there was a statistically significant elevation of insulin secretion of BMSCs by2D culture (p<0.05), ECM culture and scaffold-ECM culture (p<0.01). Under high glucose, insulin secretion of scaffold-ECM cultured cells had a statistically significant elevation compared with those of2D cultured cells and ECM cultured cells. Conclusion It is feasible to induce mouse BMSCs in vitro to differentiate to cell clusters that secret several islet-specific hormones, which has preliminarily verified by morphology, functional tests and immunology methods. The results also demonstrated the advantage of3D culture using PLGA scaffold, and the good compatibility of BMSCs and PLGA scaffolds. Background and Objective There is a hot field in researches of islet-transplantation treating type1diabetes, which concerns to expand the life span and to preserve the activity of islets in vitro by co-culture system with stem cells. In recent years, adipose tissue derived stem cells (ATDSCs) have become more popular for its sufficient and easy aquirement. There are still no reports on extending islets’ life span by co-culturing with ATDSCs. Poly-lactic-co-glycolic acid (PLGA), as a medical biologically-adaptive polymer approved by FDA, is one of the most studied bio-degradation materials. We cultured mouse islets on PLGA-ECM scaffolds, and co-cultured with ATDSCs, in order to find a better islet culture system in vitro and to use it in consequent researches on transplantation. Method In vitro, mouse islets were cultured2D in group1, on the modified PLGA-ECM scaffold in group2, and they were co-cultured with ATDSCs on PLGA-ECM scaffolds in group3. And we compared3groups to evaluate the effects of the scaffold and ATDSCs. Results Compared to islets from2D culture by inverted microscopy, those cultured in PLGA-ECM scaffolds of both two groups had good shapes and growths, which adhered tightly to scaffolds and had a smaller number of dying cells. Dithizon (DTZ) staining results indicated that the purity of islets on PLGA-ECM scaffolds was quite high. Acridine Orange (AO) and Propidium Iodide (PI) fluorescence double-staining results suggested that islets co-cultured with ATDSCs on PLGA-ECM scaffolds had a higher survival rate. MTT results demonstrated a better compatibility between islets and PLGA-ECM scaffolds when co-cultured with ATDSCs. Under scanning electron microscopy, islets showed good adhesive growth to PLGA-ECM scaffolds in morphology and big gaps between islets. Under laser confocal microscopy, green β cells and red a cells of islets were clearly seen on PLGA scaffolds of last2groups. Conclusion Results from the experiment demonstrated that good growth environment for islet cells in vitro can be provided by ATDSCs as co-cultured seeds and PLGA-ECM scaffolds as extracellular matrix.