Abstract
Objective To evaluate the effect of insulin gene transcription regulators pancreatic and duodenal homeobox-1 (PDX-1),neurogenic differentiation-1 (NeuroD1) and V-maf musculoaponeurotic fibrosarcoma oncogene homologue A (MafA) on the differentiation of induced pluripotent stem cells (iPS)into insulin-producing cells.Methods iPS were infected with adenovirus (Ad-mPDX-1-IRES-GFP、AdmNeuroD1-IRES-GFP and Ad-mMafA-IRES-GFP),and then differentiated into insulin-producing cells in vitro.Reverse transcriptase-polymerase chain reaction (RT-PCR) was applied to detecting target genes and insulin gene expression,immunofluorescence for identifying the presence and location of insulin protein and mouse insulin enzyme-linked immunosorbent assay (ELISA) to evaluating the secretory volume of insulin at different concentration of glucose.Diabetic mice were transplanted with infection iPS under the liver parenchyma.The grafts were analyzed by immunohistochemistry for the presence of insulin-producing cells.Intraperitoneal glucose tolerance test and fasting plasma glucose were used to assess the functions exertion of engrafted cell in vivo and revealed the therapeutic effect.Results RT-PCR results showed that polygenemodified iPS and pancreatic β-cell line MIN6 have similar gene expression.Mouse insulin ELISA showed polygene-modified iPS have a Satisfactory response to different concentrations of glucose.When glucose concentration was 30 mmol/L,insulin release up to (0.309 3 ±0.017 9) rg.Immunohistochemistry was performed to detect the expression of insulin in the liver tissue of diabetic mice and it exhibited positive staining of insulin.The results of intraperitoneal glucose tolerance test and fasting plasma glucose demonstrated the ability of these insulin-producing cells-transplanted mice to dispose of a glucose load.Conclusion The combination of PDX-1,NeuroD1 and MafA markedly induces insulin biosynthesis and secretion in iPS. Key words: Induced pluripotent stem cells; Insulin-producing cells; Pancreatic and duodenal homeobox-1 ; Neurogenic differentiation-1 ; V-maf musculoaponeurotic fibrosarcoma oncogene homologue A
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.