The Mechanism Of Mirna Regulation In IR And Lipids Ectopic Deposition Induced By Resistin

As the most organ of energy storage, adipose tissue also can secrete a lot of cytokines which named adipocytokines. Resistin is a newly discovered important adipocytokine which can regulate glucose and lipid metabolism and insulin sensitivity. Previous studies have revealed that resistin plays important role in insulin resistance and lipids ectopic deposition. Micro-RNAs (miRNAs) are short, endogenously produced non-coding RNA molecules that regulate the expression of target genes by pairing with sites in the 3’untranslated region (3’-UTR). Increasingly evidence has suggested that miRNAs have an important function in many physiological and pathological processes. Consequently, dysfunction of miRNAs and their target genes can lead to a variety of disorders. In this study, human and mouse recombinant resistin were selected as the object and researched the miRNA involved mechanism of insulin resistance and fat ectopic deposition induced by resistin using the technology of quantitative real time polymerase chain reaction, western blot, cells transient transfection, dual-luciferase reporter system and chromatin immunoprecipitation and so on. The results as follows:1. Analyzed the differential expression of miRNA in mouse liver treated with or without resistin by miRNA microarray, and the results showed that 25 miRNA were up-regulated and 51 miRNA were down-regulated which indicated that miRNA expression were changed by resistin. Confirmation results using quantitative real time PCR were consistent with the result from microarray.2. Bioinformatics technology were used to predict the target genes and signaling pathway of differential expressed miRNA, and results displayed that differential expressed miRNA induced by resistin involved in a lot of glucose, lipids, mitochondrial and inflammation related genes(such as C/EBP &, GSK3 β, PPAR a, ACSL1, SREBP1, NRF1, IRS-1, IRS-2, Sirtl, TNF etc) and the transduction of many signaling pathway(such as AMPK, insulin, adipocytokine, PPAR a, fatty acid metabolism and oxidative phosphorylation signaling pathway etc).3. Analyzed the mechanism of insulin resistance induced by resistin which has miR-145 involved in using the HepG2 cell line, and results indicated that human recombinant resistin up-regulated miR-145 through p65 in a dose-dependent manner. miR-145 targets IRS-1 and resulted in the development of insulin resistance by impaired the transduction of insulin signaling pathway, inhibited the glucose uptake.4. The results of roles of miR-34a in the ectopic deposition of lipids induced by resistin showed that miR-34a was up-regulated in the fatty liver model mouse. Human resistin also stimulates the expression of miR-34a in HepG2 cells. The overexpression of miR-34a increased the TG contents, however decreased the mitochondrial contents as well as the mitochondrial membrane potential. Dual-luciferase reporter system and western blot confirmed that adipoR2 act as the target of miR-34a.5. MiR-34a inhibits the PPAR a signaling pathway through adipoR2 and stimulated the SREBP-lc, thus increased the TG contents. The overexpression of miR-34a also impaired the transduction of AMPK signaling pathway and inhibited the phosphorylated-ACC1 a and CPT1 a, thereby inhibited fatty acid oxidation reaction which further increased the TG deposition.6. Western blot showed that the acetylation of PGC1 a was strengthened through inhibition of sirtl by miR-34a, and inhibited the transcript activation of PGC1 α to NRF1, and further decreased the mitochondrial contents.7. The result from ChIP showed that C/EBPβ up-regulated miR-34a through binding on the promoter of miR-34a directly, however siC/EBP β attenuated the increase of miR-34a stimulated by resistin, which indicated that resistin up-regulated miR-34a through the dependence on C/EBP β.