LncRNA-MEG3 Regulates Osteogenesis And Adipogenic Differentiation Of Stressed BMSCs Through MiRNA-140-5P

Objective one:To study the effect of stress on the differentiation of rat bone marrow stem cells and the change of LncRNA-MEG3 expression during this period;Objective two:To study the effect of LncRNA-MEG3 on the osteogenesis and adipogenic differentiation of rat bone marrow stem cells under stress;three:Study the effect of cell stress on miRNA-140-5P expression and explore the relationship between LncRNA-MEG3 and miRNA-140-5P.Methods:Step one:Isolate rat bone marrow stem cells in vitro,and culture them in osteogenic and adipogenic induction.The experimental group is loaded with appropriate cell stress.After induction for a certain period of time,the expression level of osteogenic differentiation component bone markers and the expression of adipogenic differentiation lipid markers were detected,and the expression level of"LncRNA-MEG3" in each group at different periods was also detected.Step two:Construct LncRNA-MEG3 over-expressing and low-expressing BMSCs models,and load them seperately into groups for osteogenesis and adipogenic differentiation.After a period of induction,the expression levels of osteogenic differentiation markers and adipogenic differentiation markers in the LncRNA-MEG3 overexpression and low expression models were detected by PCR and WB,respectively.Step three:Step 3:Isolate rat bone marrow stem cells in vitro and induce osteogenic and adipogenic differentiation while loading stress,and detect the expression level of miRNA-140-5P respectively.The BMSCs model of LncRNA-MEG3 over-expression and low-expression was constructed,and the cells were grouped under stress to induce osteogenic and adipogenic differentiation culture,and the expression levels of miRNA-140-5P were detected respectively.The dual luciferase reporter gene method was used to verify the targeting relationship between miRNA-40-5p and lncRNA-MEG3.Results:In step one:When BMSCs were induced into osteogenic differentiation,the expression of osteogenic differentiation indicators increased after loading with tensile stress,and the index of osteogenic differentiation decreased;In the case of adipogenic differentiation,the expression of osteogenic differentiation indexes decreased after loading with the tensile stress,and the index of adipogenic differentiation increased;The expression of LncRNA-MEG3 in BMSCs was up-regulated after loading with tensile stress.In step two:When BMSCs under stretch stress,osteogenic differentiation markers were promoted and adipogenic differentiation marker expression was inhibited when LncRNA-MEG3 overexpression was overexpression;when LncRNA-MEG was silenced,osteogenic differentiation markers expression was inhibited and adipogenic differentiation markers expression were promoted.In third step:The expression of miRNA-140-5P in BMSCs was down-regulated after BMSCs were loaded with a stretch stress;the expression of miRNA-140-5P was down-regulated when LncRNA-MEG3 was overexpressed;the expression of miRNA-140-5P was increased when LncRNA-MEG was silenced;There is a targeting relationship between miRNA-40-5p and lncRNA-MEG3.Conclusion:Stress can promote osteogenic differentiation of bone marrow stem cells while inhibiting adipogenic differentiation;LncRNA-MEG3 expression is promoted during osteogenic differentiation,and LncRNA-MEG3 expression is inhibited during adipogenic differentiation.Stress can promote the expression of LncRNA-MEG3.When LncRNA-MEG3 is over-expressed,osteogenic differentiation of BMSCs is promoted and adipogenic differentiation is suppressed;when LncRNA-MEG3 is silenced,osteogenic differentiation of BMSCs is inhibited and adipogenic differentiation is promoted.LncRNA-MEG3 has a negative regulatory relationship with miRNA-140-5P,and there is a targeting relationship between miRNA-40-5p and lncRNA-MEG3.