| Muscle fiber is the basic unit of muscle tissue, and its composition affects meat quality. Fibroblast growth factor 21 is a metabolic regulator which has been found in muscle, but the effect on muscle fiber composition is unknown. Therefore, in this experiment, mouse and C2C12 cell line were used as models to explore the effect of FGF21 on muscle fiber types, and further reveal the possible mechanism in vitro.Exp.l Effects of food deprivation on FGF21 expression and muscle fiber types in mouse skeletal muscleThe experiment was conducted to explore the effects of food deprivation on FGF21 expression and muscle fiber types of in mouse skeletal muscle. Sixteen male mice were divided into two groups, namely, control group (Con) and food deprivation group. After food deprivation for 24h. serum, liver, SOL, EDL and GAS were sampled for further measurements. The results showed that:FGF21 mRNA and protein levels in SOL were significantly higher than EDL and GAS (P<0.05). Serum FGF21 levels in fasted mice were significantly higher than that in control, and the expression of FGF21 was much higher in liver after fasting for 24h. Fasting significantly enhanced the mRNA expression of FGF21 in SOL and GAS (P<0.05). However, fasting have no effect on the expression of FGF21 in EDL (P>0.1). Western blot analysis showed that the protein levels of FGF21 in SOL and GAS were significantly increased by fasting (,P<0.05). But the protein level of FGF21 in EDL was not changed (P>0.1). In addition, food deprivation significantly increased the mRNA expression of MyHCI in the SOL and GAS (P<0.05). and induced MyHCIIa and MyHCIIx expression in GAS and EDL (P<0.05). However, the mRNA expression of MyHCI in EDL was not changed (P>0.1). The findings indicated that fasting alters FGF21 expression and the composition of muscle fiber types in SOL and GAS.Exp.2 Regulation of FGF21 on skeletal muscle fiber types in miceThis experiment was conducted to study the effect of FGF21 treatment on skeletal muscle fiber types in mice. Mice were divided into two groups, namely. Con group and FGF21 group. Mouse in Con group were injected with saline, and others were intraperitoneally injected with FGF21 (0.2mg/kg) once daily. After intraperitoneal injection for 7d. serum. SOL. EDL and GAS samples were collected for further determination. The results showed that:serum FGF21 levels were significantly increased in FGF21 injected mice (P<0.05). FGF21 injection significantly decreased FGF21 mRNA expression in SOL and EDL (P<0.05), but the mRNA expression of FGF21 in GAS was not altered (P>0.1). FGF21 injection resulted in increased MyHCI?MyHCIIa?MyHCIIx mRNA expression and decreased MyHCIIb mRNA expression of SOL (P<0.05). The mRNA expression of MyHCIIa in GAS was significantly increased (P<0.05), but MyHCI?MyHCIIx and MyHCIIb mRNA expression was not changed (P>0.1). The mRNA expression of MyHCI and MyHCIIx in EDL was significantly decreased (P<0.05), however, MyHCIIa and MyHCIIb mRNA expression was not changed (P>0.1). Additionally, intraperitoneal injection of FGF21 improved FGFR1 and FGFR4 mRNA expression in SOL (P<0.05), but the mRNA expression of KLB was not significantly changed (P>0.1). Injection of FGF21 for 7d. FGFR1 mRNA expression was significantly higher in GAS (P<0.05). and the expression of FGFR4 was tended to increase (0.05<P<0.1), in contrary, KLB tended to reduce (0.05<P<0.1). FGFR1 mRNA expression was decreased in EDL (P<0.05), and FGFR4 was not changed, however, the expression of KLB was significantly elevated after injection (P<0.05). Besides, we found that FGF21 administration significantly increased PGC1? mRNA expression in SOL (P<0.05) without altering the mRNA expression of PGC1? in GAS (P>0.1). but PGC1? mRNA expression in EDL was significantly decreased (P<0.05). There findings indicated that FGF21 treatment alters muscle fiber types, and AMPK-PGC1? signaling pathway may be involved in this regualtion.Exp.3 The signal transduction mechanism of FGF21 on skeletal muscle fiber types in miceThis experiment was conducted to study the mechanism by which FGF21 regulates mouse skeletal muscle fiber types in vitro. C2C12 myotubes were treated with FGF21 for 3d. The result showed that:FGF21 administration signifiantly decreased FGF21 mRNA expression (P<0.05). Immunostaining of C2C12 myotubes was performed to study the effect of FGF21 on muscle fiber types. The result showed that:the composition of myosin-slow in C2C12 myotubes was significantly increased, and myosin-fast was decreased (P<005). RT-PCR found that FGF21 treatment significantly improved the expression of MyHCI in C2C12 myotubes (P<0.05), and MyHCIIx mRNA expression tended to increase (0.05<P<0.1), the mRNA expression of MyHCIIa was not changed (P>0.1). However, the mRNA expression of MyHCIIb was significantly decreased (P<0.05). Therefore. FGF21 can promote the conversion of type IIb fiber to type I phenotype. PGCla as an important regulater of muscle fiber type conversion. RT-PCR showed that the mRNA expression of PGC1? was significantly inheanced (P<0.05). In order to test that AMPK-PGC1? signaling may be involved in FGF21 regulation of muscle fiber types, FGF21 was added to the culture after Aicar treatment for 30min, then cultured for 3d. The results showed that:FGF21 and AMPK agonist Aicar separately increased the expression of MyHCI (P<0.05). and significantly reduced MyHCIIb mRNA expression (P<0.05). AMPK inhibitor. Compound C (CC) added to the culture for 30min, then FGF21 treated for 2d. Immunofluorescence and qRT-PCR indicated that FGF21 increased the composition of myosin-slow and the mRNA expression of MyHCI, and decreased myosin-fast composition. This effect was inhibited by AMPK inhibitor in C2C12 myotubes (P<0.05). Western blot found that the protein levels of pAMPK and PGC1?, and the ratio of pAMPK/AMPK was significantly increased by FGF21 (P<0.05). CC inhibited the action of FGF21 (P<0.05). It was suggested that AMPK-PGC1? signaling pathway play an important role in FGF21 regulation of muscle fiber types.In summary, food deprivation increased the oxidative fiber composition and FGF21 mRNA expression in SOL and GAS. FGF21 can combine with its receptors on the cell membrane, and then activate the AMPK-PGC1? signaling pathway to regualte muscle fiber types. This study may provide new target and theory of meat quality improvement by nutrition. |
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