ABSTRACT
A possible cure for diabetes is explored by using non-pancreatic cells such as fetal hepatocytes. The expression of insulin and transcription factors for insulin is investigated in mouse fetal liver. We detected mRNAs for insulin I (Ins1) and insulin II (Ins2) and proinsulin- and mature insulin-positive cells in mouse fetal liver by reverse transcription plus the polymerase chain reaction and immunohistochemistry. Glucagon, somatostatin and pancreatic polypeptide were not expressed throughout development. Mouse Ins2 and Ins1 promoters were transiently activated in mouse fetal hepatocytes of embryonic days 13.5 and 16.5, respectively. Pancreatic and duodenal homeobox 1 (Pdx1) mRNA was not expressed during development of the liver. In contrast, mRNAs and proteins of neurogenic differentiation (NeuroD)/ß cell E-box transactivator 2 (Beta2) and v-maf musculoaponeurotic fibrosarcoma oncogene homolog (MafA) were almost simultaneously expressed with insulin genes in the liver. Ins2 and Ins1 promoters were activated in hepatoma cells by the transfection of the expression vector for NeuroD/Beta2 alone and by the combination of NeuroD/Beta2 and MafA, respectively. These results indicate that the expression of NeuroD/Beta2 and MafA is linked temporally with the transcription of Ins2 and Ins1 genes in mouse fetal liver and suggest the potential usage of fetal hepatocytes to make insulin-producing ß cells by introducing transcription factors.
Subject(s)
Fetus/metabolism , Gene Expression Regulation, Developmental , Insulin/genetics , Liver/embryology , Liver/metabolism , Animals , Basic Helix-Loop-Helix Transcription Factors/genetics , Basic Helix-Loop-Helix Transcription Factors/metabolism , Electrophoretic Mobility Shift Assay , Female , Glucagon/metabolism , Hepatocytes/metabolism , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Immunohistochemistry , Insulin/metabolism , Maf Transcription Factors, Large/genetics , Maf Transcription Factors, Large/metabolism , Mice , Mice, Inbred ICR , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Pregnancy , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Reverse Transcriptase Polymerase Chain Reaction , Somatostatin/metabolism , Trans-Activators/genetics , Trans-Activators/metabolism , Transcriptional Activation/geneticsABSTRACT
Regenerating gene product (Reg) is induced in pancreatic beta-cells and acts as an autocrine/paracrine growth factor for regeneration via a cell surface Reg receptor. However, the manner by which Reg induces beta-cell regeneration was unknown. In the present study, we found that Reg increased phospho-ATF-2, which binds to -57 to -52 of the cyclin D1 gene to activate the promoter. The Reg/ATF-2-induced cyclin D1 promoter activation was attenuated by PI(3)K inhibitors such as LY294002 and wortmannin. In Reg knockout mouse islets, the levels of phospho-ATF-2, cyclin D1, and phospho-Rb were greatly decreased. These results indicate that the Reg-Reg receptor system stimulates the PI(3)K/ATF-2/cyclin D1 signaling pathway to induce beta-cell regeneration.
