The Regulation Of Adiponectin Expression And Secretion

Adipose tissue as an energy storage organ has an irreplaceable role in triglyceride storage, mobilization and its regulation, recent studies show that the adipose tissue also plays an important role in the endocrine and paracrine, can secrete a large number of hormones and cytokines to regulate the metabolism of the tissue and cell functions.Adiponectin is a recently discovered key adipokine, it play an important role in the regulation of lipid and glucose metabolism. Adiponectin stimulates fatty acid oxidation, suppresses gluconeogenesis in liver, increases insulin sensitivity, exerts a protective role against chronic inflammation, possess anti-atherosclerosis and regulates feeding and body weight through the nervous system.Adiponectin is secreted from adipocytes into the circulation as three oligomeric isoforms,. including trimeric, hexameric and the high-molecular-mass oligomeric complex. Each oligomeric isoform of adiponectin exerts distinct biological properties in its various target tissues. The high-molecular-mass oligomer is the major active form mediating the insulin-sensitizing effects of adiponectin, whereas the central actions of this adipokine are attributed primarily to the hexameric and trimeric oligomers. The biosynthesis of the adiponectin oligomers is a complex process involving extensive post-translational modifications. Hydroxylation and glycosylation of several conserved lysine residues in the collagenous domain of adiponectin are necessary for the intracellular assembly and stabilization of its high-order oligomeric structures.Previous studies reported that adiponectin is transcriptionally regulated by various transcription factors through cis-regulatory elements in its promoter region or first intron. Secretion of the adiponectin oligomers is tightly controlled by a pair of molecular chaperones in the endoplasmic reticulum (ER), including ERp44 (ER protein of 44 kDa) and EROl-Lα(ER oxidoreductase 1-Lα). But the systematic study on the expressional and secretional regulation of adiponectin is yet to be elaborated.In this study, by choosing adiponectin stable transfected HEK293 cells and isolated pig bone marrow recharge stem cells (MSCs), using real-time quantitative PCR, Western blot, chromatin immunoprecipitation, cell transfection and promoter fluorescence luciferase reporter detection techniques, the expression, secretion and regulation mechanism of adiponectin was analysis, the following results: 1. Using homologous comparative analysis, we cloned the pig ERp44 and EROl-Lαgene. Real-time quantitative PCR detected the tissue expression patten of ERp44 and EROl-Lαgene in 12 tissue samples, ERp44 and EROl-Lαgene expressed mainly in adipose tissue. By in silico mapping analysis, pig ERp44 and EROl-La was located in chromosome 1q29 and 1q21, respectively.2. Bioinformatic analysis predicted the existence of PPARy binding sites in the ERp44 gene promoter; co-transfection found that ERp44 gene expression regulation by PPARy. Using chromatin immunoprecipitation and luciferase reporter system, we confirmed that PPARy can bind to the PPRE site in 981 to-1004 region of ERp44 gene promoter, and then repress the expression of ERp44 gene.3. Rosiglitazone (PPARy agonist) treatment or over-expression of PPARy could reduce the ERp44 gene transcription, thereby causing the secretion of adiponectin were significantly increased.4. Bone marrow MSCs was isolated from a month-old Meishan pig, by cell morphology, growth characteristics and marker genes (PouV, Sox2, and Nanog, etc.) analysis showed that the MSCs have mesenchymal stem cell morphology and cell pluripotency (for example, to differentiate into mature adipocyte).5. Bioinformatic analysis predicted the KLF binding sites in adiponectin gene promoter, and the adiponectin gene expression was regulated by KLF 15 during the porcine bone marrow MSCs differentiated into adipocyte,. Using chromatin immunoprecipitation and luciferase reporter gene system to analysis, the results shows that KLF 15 can specifically bind to the adiponectin gene promoter-93 to-77 region, and then activate the expression of adiponectin gene.