Differentiation of endometrial stem cells into insulin-producing cells using signaling molecules and zinc oxide nanoparticles, and three-dimensional culture on nanofibrous scaffolds.

Abstract

Diabetes mellitus is the most common metabolic disorder with a high mortality and morbidity rate. A new promising strategy to treat DM is pancreatic tissue engineering. We described a 3D culture system accompanied by signaling factors to differentiate hEnSCs into IPCs in the presence of nZnO. We isolated EnSCs and cultured it in DMEM/F12 medium. Nanofibrous PLA/Cs scaffold was prepared through the electrospinning method. The morphological properties of the scaffolds and cells were evaluated by SEM. MTT assay was used to investigate the metabolic activity of the hEnSCs cultured on the scaffolds and a four-stage protocol was applied to differentiate hEnSCs. The differentiated cells were tested for pancreatic markers by immunocytochemistry, qRT-PCR and DTZ staining. The results of this study revealed that hEnSCs cultured on PLA/Cs scaffold and treated with nZnO can efficiently differentiate into IPCs. The examination of differentiated cell morphology showed their near similarity with pancreatic islet cells, and DTZ staining emphasized the presence of insulin granules inside their cytoplasm. Moreover, qRT-PCR and immunofluorescent staining results showed the efficient expression of specific gene markers of IPCs in resultant differentiated cells. Moreover, PLA/CS and nZnO were able to provide a good nanoenvironment for the differentiation of hEnSCs into IPCS the in presence of other molecules.