Study On The Relationship Between Wnt10b And PTEN In Inhibiting BMP9-induced Osteogenic Differentiation Of Mesenchymal Stem Cells

Bone morphogenetic protein 9(BMP9)is one of the most efficacious cytokines to induce osteogenic differentiation in mesenchymal stem cells.Both PTEN and Wnt10 b were implicated in regulating the BMP9’s osteogenic potential,but the relationship between them keeps unknown.Based on our previous work,this study analyzed the relationship between Wnt10 b and PTEN when regulating BMP9-induced osteogensis in C3H10T1/2 cells,and the possible molecular mechanism for this biological process.In this experiment,we used RT-PCR,Alizarin Red S staining,IP,CHIP,histochemical staining,ectopic osteogenesis and so on to analyze the effects of BMP9 on the expression of PTEN and Wnt10 b in C3H10T1/2 cells.Then,we analyzed the effects of PTEN and BMP9 on the expression of Wnt10 b in C3H10T1/2 cells.Finally,we investigated the relationship between Wnt10 b and PTEN about inhibiting BMP9-induced osteogenic differentiation,as well as the possible molecular mechanism in C3H10T1/2 cells.Results showed that PTEN was down-regulated by BMP9 in C3H10T1/2 cells,but Wnt10 b was up-regulated.Exogenous PTEN inhibited the ALP activities,expression of early and late osteogenic markers and mineralization induced by BMP9,while silencing PTEN exhibited reversal effects.Exogenous of Wnt10 b reversed PTEN’s inhibitory effect on ALP activity and mineralization in BMP9-induced C3H10T1/2 cells.Conversely,the BMP9’s potential to induce osteogenic markers was promoted by silencing PTEN,which was almost deminished by silencing Wnt10 b.Ectopic bone formation of BMP9-induced in C3H10T1/2 cells was enhanced by silencing PTEN,while silencing Wnt10 b reduced this effect of silencing PTEN.BMP9 increased the level of Wnt10 b and p-CREB,which were decreased by PTEN,rapamycin or Ly294002.However,it was enhanced by silencing PTEN.The level of p-Smad1/5/9 induced by BMP9 was reduced by PTEN,but increased by silencing PTEN.There exists interaction between p-Smad1/5/9 and p-CREB in MSCs.Meanwhile,p-CREB and p-Smad1/5/9 were both enriched at the promoter region of Wnt10 b.Our data indicated that effect of PTEN on BMP9’s osteogenic capacity may be mediated through regulating Wnt10 b via the interaction between BMP / Smad and PI3K/MTOR/CREB.