The regulatory mechanism of insulin-producing cells (IPCs) differentiation from induced pluripotent stem cells (iPSCs) in vitro is very important in the phylogenetics of pancreatic islets, the molecular pathogenesis of diabetes, and the acquisition of high-quality pancreatic β-cells derived from stem cells for cell therapy. miPSCs were induced for IPCs differentiation. miRNA microarray assays were performed by using total RNA from our iPCs-derived IPCs containing undifferentiated iPSCs and iPSCs-derived IPCSs at day 4, day 14, and day 21 during step 3 to screen the differentially expressed miRNAs (DEmiRNAs) related to IPCs differentiation, and putative target genes of DEmiRNAs were predicted by bioinformatics analysis. miR-690 was selected for further research, and MPCs were transfected by miR-690-agomir to confirm whether it was involved in the regulation of IPCs differentiation in iPSCs. Quantitative Real-Time PCR (qRT-PCR), Western blotting, and immunostaining assays were performed to examine the pancreatic function of IPCs at mRNA and protein level respectively. Flow cytometry and ELISA were performed to detect differentiation efficiency and insulin content and secretion from iPSCs-derived IPCs in response to stimulation at different concentration of glucose. The targeting of the 3′-untranslated region of Sox9 by miR-690 was examined by luciferase assay. We found that miR-690 was expressed dynamically during IPCs differentiation according to the miRNA array results and that overexpression of miR-690 significantly impaired the maturation and insulinogenesis of IPCs derived from iPSCs both in vitro and in vivo. Bioinformatic prediction and mechanistic analysis revealed that miR-690 plays a pivotal role during the differentiation of IPCs by directly targeting the transcription factor sex-determining region Y (SRY)-box9. Furthermore, downstream experiments indicated that miR-690 is likely to act as an inactivated regulator of the Wnt signaling pathway in this process. We discovered a previously unknown interaction between miR-690 and sox9 but also revealed a new regulatory signaling pathway of the miR-690/Sox9 axis during iPSCs-induced IPCs differentiation.

Islet transplantation is considered as one of the promising treatment options for curing diabetes. However, the extracellular matrix (ECM) is destroyed during the process of islet isolation and extraction, which leads to decreased islet activity in vitro. ECM-based biomaterials which used to reconstruct the microenvironment of cells have been applied in various fields. In this study, an electrospinning hybrid scaffolds with silk fibroin (SF) and pig pancreatic decellularized extracellular matrix (P-dECM) have been prepared to mimic the islet ECM in vivo. Furthermore, the activity and function of islet were evaluated in vitro. The microstructures, hydrophilia and the main components of scaffolds were characterized by SEM, contact angle analysis and immunohistochemical experiment. The toxicity of stents was assessed by MTT assay. Cell activity and function were estimated by the live-dead cell staining, immunofluorescence, glucose-stimulated insulin secretion assay and q-PCR. A nanofiber scaffold with good hydrophilicity, non-toxic and retention of key ECM components has been obtained, which can improve the survival and promote and function of islets. This scaffold can be a promising candidate for pancreatic tissue engineering and provides a new strategy for islet transplantation.

Pancreatic ductal adenocarcinoma (PDAC) has a signature of extremely high matrix stiffness caused by a special desmoplastic reaction, which dynamically stiffens along with the pathological process. The poor prognosis and low five-year survival rate of PDAC are partly owing to chemoresistance triggered by substrate stiffness. Understanding the potential mechanisms of matrix stiffness causing PDAC chemoresistance is of great significance. In this study, methacrylated gelatin hydrogel was used as platform for PANC-1 and MIA-PaCa2 cell culture. The results indicated that compared to soft substrate, stiff substrate distinctively reduced the gemcitabine sensitivity of pancreatic cancer. Intriguingly, transmission electron microscopy, immunofluorescence staining, western blot and qRT-PCR assay showcased that the number of autophagosomes and the expression of LC3 were elevated. The observations indicate that matrix stiffness may regulate the autophagy level, which plays a vital role during chemoresistance. In brief, soft substrate exhibited low autophagy level, while the counterpart displayed elevated autophagy level. In order to elucidate the underlying interaction between matrix stiffness-mediated cell autophagy and chemoresistance, rescue experiments with rapamycin and chloroquine were conducted. We found that inhibiting cell autophagy dramatically increased the sensitivity of pancreatic cancer cells to gemcitabine in the stiff group, while promoting autophagy-driven chemoresistance in the soft group, demonstrating that matrix stiffness modulated chemoresistance via autophagy. Furthermore, RNA-seq results showed that miR-1972 may regulate autophagy level in response to matrix stiffness. Overall, our research shed light on the synergistic therapy of PDAC combined with gemcitabine and chloroquine, which is conducive to promoting a therapeutic effect.