Overexpression Of Atg5 Inhibits Osteogenic Differentiation Of VICs By Upregulating Autophagy Levels

Our study was to investigate the effect of autophagy related protein Atg5 on autophagy and osteogenic differentiation of aortic valve interstitial cells,which was order to provide new methods and ideas for the study of the calcified aortic valve disease.Primary valve interstitial cells were isolated from the porcine aortic valve.Cells phenotype identification was used by immunofluorescent staining.The expression of Runx2,OPN,p62 and LC3 B I/II was detected by Western blot to identified the relationship between autophagy and osteogenesis in valvular interstitial cells.The plasmid p Ad Track-ATG5 overexpressing Atg5 was constructed and transfected into VICs.After transfecting with control plasmid or p Ad Track-ATG5,the VICs were cultured with osteogenic medium.The osteogenic differentiation ability of valve interstitial cells was detected by Western blot,alkaline phosphatase staining and alizarin red S staining.For immunofluorescent staining,Alpha-SMA and Vimentin were positive and v WF was negative in VICs.Compared with the CM group,the expression of Runx2,OPN and p62 protein in the OM group was increased but LC3 B ?/? decreased.The expression of Atg5 and LC3 B ?/? in cells transfeced with p Ad Track-ATG5 was raised but p62 was decreased compared with the negative control.The expression level of Runx2 and OPN in the OM+Atg5 group was lower than that in the OM+NC group.Alkaline phosphatase staining and alizarin red S staining were also showed that the osteogenic differentiation ability in the OM+Atg5 group was weaker than the OM+NC group.Take together,overexpression of Atg5 inhibits osteogenic differentiation of valvular interstitial cells by upregulating autophagic activity.