Construction High-Througout Model To Promote Islet-Progenitor Cells Differentiation Into Insulin-Producing Cells And Compounds Screening

Islet regeneration means new β cells produced from progenitor cell,a promising etiological treatment could potentially cure diabetes through restoring lost beta cell mass and function.Hence,identification of novel and safe differentiation inducer is a prime requisite for islet generation,which could be next generation therapeutics for diabetes.PDX-1 plays a vital role in islet formation,β cells differentiation and maintenance of β cells function.PDX-1 gene was used as a target gene to build a high-throughput screening model in our study,and a compound capable of promoting the expression of PDX-1 was successfully screened.Further studies showed that the compound has the ability to promote the differentiation of islet progenitor cells into insulin-secreting cells.Firstly,PANC-1 cells were induced differentiation into 1slet-like cell clusters,which expressed islet-specific genes and secreted insulin after glucose stimulating.When islet-like cell clusters were transplanted into mice with type 1 diabetes by subrenal capsular transplantation,the blood sugar was reduced to normal level obviously.It implied that we constructed the method of differentiation islet-like progenitor cells into islet cells.Secondly,western blotting analysis and immune-histochemical analysis were used to study PDX-1 expression and distribution in different time points of PANC-1 cell differentiation.We found that PDX-1 expression was increased and it transfered from cytopasma to cell nucleus in the early stage of differentiation.Futhermore,overexpression of PDX-1 could promote PANC-1 cells differentiation and maturation.It implied that PDX-1 plays an important role in PANC-1 cells differentiation.Thirdly,a high-throughput screening model was constructed using PDX-1 as the target gene,and C754,an extract of plant,that promoted the expression of PDX-1 was screened out.Liquid chromatography-mass spectrometry and high performance liquid chromatography were used to authenticate of the main component of C754,results revealed that it was C1037 which also was called andrographolide.We found that C1037 could up-regulated the expression of C1037 in a dose-dependent manner acording to the results of qPCR and WB.Lastly,after C1037 was applied to PANC-1 cells,the expression of islet-specific genes at different time points during differentiation and insulin secretion stimulating by glucose-stimulated insulin of islet-like cell clusters was measured.As a result,C1037 could accelerate PANC-1 cells differentiation into islet-like cell clusters(ILCCs)and promote insulin secretion stimulated by glucose of ILCCs.It showed that C1037 could accelerate the differentiation of PANC-1 cells,and provided a new candidate inducer for clinical treatment of diabetes.