Regulation And Mechanism Of Exercise-incduced Triglyceride Accumulation In Muscle Cells By Lactate/GPR81 Signal Pathway

Objective: The aim of this study is to investigate whether lactate/GPR81 signal pathway is involved in the regulation of triglyceride accumulation and the increase of mitochondrial function protein in skeletal muscle cell,and to investigate whether lactate/GPR81 signal pathway regulates skeletal muscle contraction-induced skeletal muscle triglyceride accumulation and the increase of mitochondrial function protein.Methods: The first part: C2C12,3T3-L1 and 293 cells were treated with lactate,pyruvate,Forskolin and so on.The content of triglyceride,pyruvate and the activity of pyruvate dehydrogenase in the cells were determined.The protein or/and m RNA level of MCAT,CREB,p CREB,lipic acid,PDHE2,p Erk,Erk,GPR81 and so on were determined.In the second part,C2C12 cells were applied by four kinds of Electric pulse stimulation(EPS)protocol,and the cells were treated with lactate and 3-hydroxy-butyrate(3-OBA).The contents of TG,lactate,c AMP and ATP were detected.The protein and m RNA level of MCAT,GPR81,MCT1,MCT4,IL-6 and so on were detected.Results: The first part: 1.Lactate increased the content of TG in C2C12 myotubes.2.Forskolin significantly increased p CREB signaling activity in C2C12 cells.Lactate and 3,5-DHBA significantly reduced p CREB signaling status in C2C12 cells.Lactate could reverse the Forskolin-induced p CREB signal status of C2C12 cells.3.The protein content of MCAT and PDH activity in C2C12 cells were increased by lactate.4.Forskolin reduced MCAT protein content in C2C12 cells.The MCAT inhibition by forskolin was significantly attenuated by pre-treatment of C2C12 cells with lactate.5.Pyruvate did not significantly increase TG content.The protein level of MCAT,lipoylation protein and PDHE2 were also unchanged after pyruvate treatment in C2C12 cells.6.The lactate significantly induced the protein expression of GPR81,MCT1 and MCT4 in C2C12 cells.The second part: 1.The four kinds of EPS protocol(Voltage 30 V,frequency 0.5Hz-10 Hz,a single time 2ms and a total of 3h)significantly increased the lactate concentration in C2C12 cells.Also the level of PGC-1?,Cyt c,IL-6,MCT1,MCT4 and GPR81 m RNA increased.However the content of ATP was reduced after immediate of skeletal muscle construction.Meanwhile,the myosin heavy chain isoforms were altered after EPS.2.EPS protocols,the frequency of which was 0.5Hz,1Hz and 5Hz,all significantly increased the TG content in C2C12 cells.3.c AMP inhibition by lactate(16m M)was significantly attenuated by pre-treatment of C2C12 cells with 3-OBA(6m M and 12 m M).4.EPS(Frequency 1Hz)protocol significantly increased the TG content in C2C12 cells,but this function was weakened by 3-OBA(6m M)pretreatment.5.The EPS protocols(frequency 0.5Hz and 1Hz)significantly increased the protein content of MCAT and GPR81 in contractile cells,but 3-OBA pretreatment can weaken that function.Conclusions: 1.Lactate/GPR81 signaling pathway induces TG accumulation and the increase of the protein level of MCAT and PDH by inhibiting c AMP signaling pathway in C2C12 cells.2.Lactate/GPR81 signaling pathway regulates skeletal muscle cell contraction-induced enhancement of TG accumulation and the increase of mitochondrial function protein.