[MicroRNA-22-3p Regulates the Expression of Kruppel-like Factor 6 to Affect the Cardiomyocyte-like Differentiation of Bone Marrow Mesenchymal Stem Cell].

Objective To explore the effect of microRNA-22-3p (miR-22-3p) regulating the expression of Kruppel-like factor 6 (KLF6) on the cardiomyocyte-like differentiation of bone marrow mesenchymal stem cell (BMSC). Methods Rat BMSC was isolated and cultured,and the third-generation BMSC was divided into a control group,a 5-azacytidine(5-AZA)group,a mimics-NC group,a miR-22-3p mimics group,a miR-22-3p mimics+pcDNA group,and a miR-22-3p mimics+pcDNA-KLF6 group.Real-time fluorescent quantitative PCR (qRT-PCR) was carried out to determine the expression of miR-22-3p and KLF6 in cells.Immunofluorescence staining was employed to detect the expression of Desmin,cardiac troponin T (cTnT),and connexin 43 (Cx43).Western blotting was employed to determine the protein levels of cTnT,Cx43,Desmin,and KLF6,and flow cytometry to detect the apoptosis of BMSC.The targeting relationship between miR-22-3p and KLF6 was analyzed by dual luciferase reporter gene assay. Results Compared with the control group,5-AZA up-regulated the expression of miR-22-3p (=7.971,<0.001),Desmin (=7.876,<0.001),cTnT (=10.272,<0.001),and Cx43 (=6.256,<0.001),increased the apoptosis rate of BMSC (=12.708,<0.001),and down-regulated the mRNA (=20.850,<0.001) and protein (=11.080,<0.001) levels of KLF6.Compared with the 5-AZA group and the mimics-NC group,miR-22-3p mimics up-regulated the expression of miR-22-3p (=3.591,<0.001;=11.650,<0.001),Desmin (=5.975,<0.001;=13.579,<0.001),cTnT (=7.133,<0.001;=17.548,<0.001),and Cx43 (=4.571,=0.037;=11.068,<0.001),and down-regulated the mRNA (=7.384,<0.001;=28.234,<0.001) and protein (=4.594,=0.036;=15.945,<0.001) levels of KLF6.The apoptosis rate of miR-22-3p mimics group was lower than that of 5-AZA group (=8.216,<0.001).Compared with the miR-22-3p mimics+pcDNA group,miR-22-3p mimics+pcDNA-KLF6 up-regulated the mRNA(=23.891,<0.001) and protein(=13.378,<0.001)levels of KLF6,down-regulated the expression of Desmin (=9.505,<0.001),cTnT (=10.985,<0.001),and Cx43 (=8.301,<0.001),and increased the apoptosis rate (=4.713,=0.029).The dual luciferase reporter gene experiment demonstrated that KLF6 was a potential target gene of miR-22-3p. Conclusion MiR-22-3p promotes cardiomyocyte-like differentiation of BMSC by inhibiting the expression of KLF6.