The Role Of HDAC5 In Aerobic Exercise Ameliorating Insulin Resistance

Aim: Insulin resistance(IR) is critical to the pathogenesis of type 2 diabetes mellitus(T2DM). IR is associated with the abnormal lipid metabolism. Aerobic exercise can enhance fatty acid oxidation, and decrease lipid toxicity via increasing metabolic gene expression in skeletal muscle. However, the exact mechanism is not still clear. HDAC5 plays a key role in muscle fiber switch and remodeling. The transcriptional activity of MEF2 was inhibited by HDAC5. A number of genes have been confirmed to possess MEF2-binding sites on the promoters, suggesting that aerobic exercise could regulate the metabolic genes expression by MEF2/HDAC5 interaction. Carnitine palmitoyltransferase 1(CPT1b) plays an important role in skeletal muscle mitochondrial β-oxidation. Whether CPT1 b was regulated by MEF2/HDAC5 in response to exercise was not clear. HDAC5 was regulated by AMPK, the intracellular energy sensor. Exercise training can activate p AMPK Thr172 and promote fatty acid β-oxidation. Together, the purpose of the study is:(1) To determine whether aerobic exercise regulates the activity of HDAC5 in skeletal muscle;(2) To determine whether HDAC5 involves in the regulation of CPT1 b expression in response to aerobic exercise;(3) To determine whether HDAC5 is regulated by exercise training through AMPK. Our project provided the theoretical basis for the mechanism that aerobic exercise regulating the metabolic gene expression and improving IR.Method: 4 week old C57BL/6 mice were divided into normal chow, normal chow exercise, high-fat diet(HFD) control and HFD exercise, the exercise groups were underwent 6-week treadmill training. Oral glucose tolerance test, fasting serum insulin, lipid profiles were detected; Skeletal muscle lipid ectopic deposition was detected by oil red O staining; Citrate synthase activity was determined by enzyme kinetics. The protein levels of HDAC5, p HDAC5 Ser259/498, Ac-H3K9, AMPKα, p AMPKα Thr172, and CPT1 b were determined by western blot; CPT1 b m RNA level was detected by real-time PCR; The interaction of HDAC5 and MEF2 A, AMPKα, p AMPKα Thr172, 14-3-3 was determined by co-immunoprecipitation; The binding ofMEF2A/HDAC5 to the CPTb1 promoter was detected by chromatin immunoprecipitation; MEF2 A, HDAC5 expression vector and luciferase plasmid that contain CPT1 b promoter were co-transfacted into C2C12 cell. CPT1 b m RNA and the activity of luciferase were detected. C2C12 cell were treated by AICAR, or AMPK/DN-AMPK adenovirus. HDAC5, p HDAC5 Ser259/498, AMPKα, p AMPKα Thr172 were determined by western blot; Wild type or AMPKα2 knockout mice were divided into control and exercise group, the exercise group were underwent 1h acute treadmill training. HDAC5, p HDAC5 Ser259/498, AMPKα1, AMPKα2, p AMPKα Thr172, p Ca MKII Thr286, p PKD Ser744/748 in skeletal muscle were detected by western blot.Results:(1) 6-week aerobic exercise enhanced skeletal muscle oxidative capacity; reduced body weight; improved impaired glucose tolerance; and reversed the lipid metabolic disorders;(2) 6-week exercise training increased p HDAC5 Ser259/498; p AMPKα Thr172 phosphorylation levels; promoted the nuclear export of HDAC5; reduced the binding of HDAC5 to MEF2A; increased the binding of HDAC5 to AMPK; and increased the acetylation level of H3K9 in skeletal muscle;(3) 6-week aerobic exercise increased the binding of MEF2 A to the CPT1 b promoter; In C2C12 cell, HDAC5 can inhibit the CPT1 b m RNA and the CPT1 b promoter trancriton activity that were induced by MEF2A; AICAR activated AMPK-HDAC5 signaling pathway, overexpression of AMPKα2/DN-AMPKα2 activited or inhibitd AMPK-HDAC5 signaling pathway;(4) Acute exercise activited AMPK-HDAC5 signaling pathway in wild-type mice skeletal muscle, which was blocked in AMPKα2 KO mice skeletal muscle. Interestingly, both of HDAC5 Ser259/498 and its upstream kinase PKD ser744/748 were increased in AMPKα2 KO mice skeletal muscle.Conclusion: Aerobic exercise improved high-fat diet induced abnormal glucose and lipid metabolism, which was associated with the increased phosphorylation- dependent nuclear HDAC5 export; HDAC5/MEF2 A regulated the transcriptional activity ofCPT1b; Aerobic exercise could enhance the expression of CPT1 b via HDAC5/MEF2 A interaction, so as to improve lipid ectopoc deposition in skeletal muscle; Exercise activited AMPK-HDAC5 signaling pathway. However, the absence of AMPKα2 blocked the increase the activity of HDAC5 induced by exercise training, but increased the compensatory effect of PKD, suggesting that AMPK plays an important role in the regulation of HDAC5 activity in response to exercise training.