MiR-455b-3p Regulates APN Expression In Adipocytes And Its Mechanism

Objective: To investigate CHD(Coronary Artery Disease,CAD)patients epicardial adipose tissue(Epicardial Adipose Tissue,EAT),and serum adiponectin(adiponectin,APN,ADIPOQ)Differential expression of mi R-455b-3p adipocytokines Effects of APN,Kruppel-like factor 4(KLF4),interleukin-6(IL-6)and monocyte chemokine 1(MCP-1)secretion and adipocyte differentiation,elucidating mi R-455b-3p in CAD Possible mechanisms of adipocyte differentiation and inflammatory responses during development.Methods: 1.Selecting the AHP-related significant up-regulation of miR-455b-3p in the EAT tissues of CAD patients in the early stage of the study group,and collecting EAT and serum from patients with coronary heart disease(CAD)and non-coronary heart disease(Non-CAD).Specimens were used to verify the expression of mi R-455b-3p in EAT and serum samples from CAD and Non-CAD patients by real-time quantitative PCR(q RT-PCR).2,3T3-L1 preadipocytes were cultured in vitro,the experimental components were mi R-455b-3p overexpression group(transfected mi R-455b-3p mimic)and inhibitor group(transfected mi R-455b-3p inhibitor)And a blank control group.mi R-455b-3p mimics,inhibitor and NC were transfected into induced mature 3T3-L1 adipocytes in vitro,and the expression levels of inflammatory factors APN,KLF4,IL-6 and MCP-1 in adipocytes were detected by q RT-PCR.3,mi R-455b-3p agomir,antagomir and NC were transfected into 3T3-L1 preadipocytes cultured in vitro,and lipid droplet accumulation was observed after oil red O staining for 8 days.q RT-PCR was used to detect adipocyte differentiation and maturation in adipocytes.Expression levels of APN,FABP4,PPARγ and C/EBPα m RNA and APN and FABP4 proteins.Results: 1.Human tissue experiment(1)Comparison of general data,blood lipids and plasma inflammatory factors levels of the tested individuals The LDL(low density lipoprotein,LDL-c)and C-reactive protein of the patients with CAD is higher than the non-CAD group,HDL(high-density lipoprotein,HDL-c)is lower than the non-CAD ADIPOQ group.The differences were statistically significant(P<0.05).(2)Expression levels of mi R-455b-3p in EAT tissues and serum of test subjects Compared with non-CAD,mi R-455b-3p was elevated in EAT and serum of CAD patients(P<0.01).2.Experiments in vitro(1)Culture and differentiation of 3T3-L1 adipocytes The 3T3-L1 cells that were not induced to differentiate were fusiform,and there was no lipid droplet in the cytoplasm.When induced to differentiate to the 8th day,the cells were oval,and more than 90% of the cells were differentiated and mature,and more lipid droplets were aggregated.Oil red O staining showed that the lipid droplets in the adipocytes were stained red,and the 3T3-L1 cells were successfully induced todifferentiate into adipocytes.(2)Detection of mi R-455b-3p regulation of adipocyte inflammatory factors Compared with the control group,the expression of APN and KLF4 m RNA and the expression of IL-6 and MCP-1 were decreased in mature 3T3-L1 adipocytes in the mi R-455b-3p overexpression group(P<0.05);mi R-455b-3p inhibition was observed.In the agent group,the expression of APN and KLF4 increased,and the expression of IL-6 and MCP-1 decreased(P<0.05).(3)Detection of the effect of mi R-455b-3p on adipocyte differentiation During the differentiation of 3T3-L1 preadipocytes,the expression level of mi R-455b-3p increased first and then decreased,and finally increased rapidly.Overexpression of mi R-455b-3p in uninduced 3T3-L1 preadipocytes and induced maturation,increased lipid droplet accumulation compared with the control group,and expression of FABP4,PPARγ,C/EBPα m RNA in adipocyte differentiation markers The expression of FABP4 protein was increased and the expression of APN protein was decreased(P<0.05).Conclusions:(1)Differentially expressed miRNAs in EAT of CAD patients,in which mi R-455b-3p is highly expressed;(2)Highly expressed mi R-455b-3p negatively regulates APN expression and affects adipocytokines KLF4,IL-6 and the expression of MCP-1,so that the inflammation is aggravated;(3)in the adipocytes,mi R-455b-3p promotes the differentiation and maturation of adipocytes.