Effect Of PPAR? On Muscle Fiber Type During Mouse Myoblast Differentiation

Objective: By adding PPAR delta specific agonist GW0742 and antagonist GSK0660 during the differentiation of mouse myoblasts,the expression level of myosin heavy chain(MHC)subtype during the differentiation of mouse myoblasts was studied,and the influence of PPAR delta on the muscle fiber type during the differentiation of mouse myoblasts was further discussed.Methods: C2C12 myoblasts and the primary myoblasts of kunming mice were cultured.They were divided into three groups: control group(DMSO group),PPAR delta agonist group(GW0742 group),PPAR delta antagonist group(GW0742+GSK0660group).When the confluent degree of cells in each group was up to 90%,C2C12 cell lines were replaced with growth medium(DMEM medium +10%FBS)by differentiation medium(DMEM medium +2%HS),and primary mouse myoblasts were replaced with growth medium(F10 medium +20%FBS+5ng/ml bFGF)by differentiation medium(DMEM medium +2%HS).Cell samples were collected on days 4,8 and 12 of C2C12 myoblasts and days 2,4 and 6 of kunming mice.Identification of myoblasts differentiation by MHC immunofluorescence;The expression of MHC subtypes and PPAR delta was detected by q-rt-pcr.Results:(1)On the fourth day of myoblast differentiation of C2C12,myotube formation was observed in MHC immunofluorescence,and the number of myotubes formed by cell differentiation in the three groups was approximately equal.On day eighth and twelfth of differentiation,the number of myotubes in the PPAR delta group was significantly greater than in the other two groups.(2)On the second day of mouse primary myoblast differentiation,MHC immunofluorescence showed that the number of myotubes formed in the PPAR delta agonist group was significantly greater than that in the control group.On the fourth and sixth day of differentiation,the number of myotubes in the PPAR delta group was significantly greater than in the other two groups.(3)On the fourth day of C2C12 myoblast differentiation,the expression levels of MHC isoforms in PPAR delta agonist group were significantly higher than those in theother two groups.On the eighth day of differentiation,the expression of MHCI and MHCIIx in the PPAR delta agonist group was up-regulated by more than 2-fold and MHCIIa was up-regulated by more than 3-fold compared with the control group.On the twelfth day of differentiation,the expression of MHCI was up-regulated by more than3-fold and MHCIIa was up-regulated by More than 4 times compared with the control group.(4)On the second day of primary myoblast differentiation,the expression of MHCI and MHCIIa in PPAR delta agonist group was up-regulated more than 2-fold compared with the control group.On the fourth day of differentiation,the expression of MHCI and MHCIIx was up-regulated more than 2-fold compared with the control group.The MHCIIa expression level was up-regulated by more than 3 times;on the sixth day of differentiation,the expression of MHCI was up-regulated by more than 4 times compared with the control group,and the expression of MHCIIa was up-regulated by more than 6times compared with the control group.Conclusion:(1)PPAR? can promote mouse myoblast differentiation.(2)PPAR?significantly up-regulated the expression of oxidized fibers during the differentiation of C2C12 myoblasts and mouse primary myoblasts.