MSCs在自组装多肽纳米纤维支架中的三维培养及脂向分化研究

Study on Three-dimensional Culture and Adipogenic Differentiation of MSCs in Self-assembling Peptide Nanofiber Scaffolds

拟采用自组装多肽RADA16水凝胶建立MSCs的三维培养体系,以观察MSCs在RADA16中的黏附、形态及脂向分化情况。将第3代细胞置于RADA16水凝胶中进行三维培养。实验组细胞-材料复合物以脂向诱导液诱导,对照组以DMEM培养液培养,然后进行形态学观察,油红O染色及Westernblot检测。荧光染色显示,绝大多数MSCs在RADA16水凝胶中能存活,且在不同的三维平面上生长。经诱导后,细胞内有大量油红O染色阳性的脂滴聚集,Westernblot分析结果显示有脂肪细胞特异的蛋白PPARγ表达,且表达量随诱导时间的延长而增加。以上结果表明,RADA16水凝胶支架为MSCs的黏附、生长提供良好的三维环境,经定向诱导后支架内的MSCs可向脂肪组织方向分化。

To investigate the adhesion,morphology and differentiation of MSCs in self-assembling peptide RADA16 hydrogel scaffolds,the 3rd passage of MSCs was seeded on RADA16 hydrogel for 3D culture.The morphology and survival of MSCs were observed by inverted phase contrast microscope and fluorescence staining.In the experimental group,adipogenic differentiation was achieved by the induction media.The control MSCs were maintained in DMEM media.Fluorescence staining showed that vast majority of MSCs in the RADA16 hydrogel could adhesive,survive,and distribute uniformly on different planes.After 7 d adipogenic induction intra-cellular lipid droplet appeared,and Oil Red O stained positive cells were identified.Differentiation was further demonstrated by Western blot analysis of adipocytic markers PPARγ expression.The results suggested that self-assembling peptide RADA16 hydrogel scaffolds offered suitable 3D micro-environment for the adhesion,growth,and differentiation of MSCs.