The Molecular Mechanism Of The MiR-351-5p Effects 3T3-L1 Preadipocytes Adipogenic Differention

Adipose tissue is an important metabolic organ that is crucial for enery homeostase.The prevalence of obesity and obesity-related diseases has aroused attention to the development of adipose tissue and adipose cells.Differentiation of 3T3-L1 cells into adipocytes involves a orchestrated series of events including clonal expansion,growth arrest,and terminal differentiation.miRNAs are small endogenous RNAs that regulate multiple biological processes including adipogenesis and Lipid metabolism.miR-351-5p may contribute to adipose tissue development according to the other studies.The role of miR-351-5p in the process of 3T3-L1 preadipocyte differentiation and lipid metabolism is not understood.The purpose of this research was to discuss the molecular mechanism of miR-351-5p in 3T3-L1.1? miR-351-5p expression of adipogenic differentiation process in 3T3-L1 cellsqPCR and Oil Red O staining were used to detect the expression of miR-351-5p in the process of the differentiation of 3T3-L1 cells and the expression profile of miR-351-5p in mouse tissues.It was found that the expression of miR-351-5p increased during lipid formation and reached the peak on the 8th day after lipid droplet accumulation.2? Effect of miRNA-351-5p on adipogenic differentiation of 3T3-L1 cellsmiR-351-5p transfected into 3T3-L1 preadipocyte,The qPCR and western blot was used to show the expression of adipocyte-specific molecular marker.It was found that the adipogenic marker gene PPAR? was down-regulated after transfection with miR-351-5p.CCK-8,Ed U,flow cytometry was used to detect the cell activity,proliferation and cell cycle after transfection with miR-351-5p.The results showed that miR-351-5p had no significant effect on cell viability and cell proliferation,but the date of cell cycle showed that cell division was blocked in S phase after transfection with miR-351-5p.Oil red 0 staining and quantitative analysis were used to detect the accumulation of lipid droplets.The results showed that miR-351-5p could inhibit the accumulation of lipid droplets during adipogenesis and differentiation of preadipocytes 3T3-L1.The evdiences showed that the miR-351-5p was a negative regulator of adipogenesis in 3T3-L1 preadipocyte.3? The target gene verification of miR-351-5pIn order to understand the mechanism of miR-351-5p,There was 12 target sites of miR-351-5p predicted by several bioinformatics softwares such as Targeta Scsn,Pic Tar,miRDB.Five genes associated with lipid formation were screened.It has been confirmed that the target gene of miR-351-5p was Necab3 in adipose differentiation by Dual-Luciferase reporter assay and western blot.4? Effect of si-Necab3 on adipogenic differentiation of 3T3-L1 cellsIn order to clarify the effect of Necab3 on adipogenic differentiation of 3T3-L1,3T3-L1 cells were transfected with si RNA,of Necab3.qPCR and western blot was used to show the expression of adipocyte-specific molecular marker,It was found that the adipogenic marker gene PPAR? was down-regulated after transfection with the si RNA of Necab3.Oil red 0 staining and quantitative analysis were used to detect the accumulation of lipid droplets.The results showed that the inhibition of Necab3 could inhibit the accumulation of lipid droplets during adipogenesis and differentiation of preadipocytes 3T3-L1.It was confirmed that miR-351-5p could obstructed the accumulation lipid droplets and the expressions of PPAR? by inhibition of Necab3.The miR-351-5p as a negative regulator effects differentiation of 3T3-L1 cells.