title: Suppression of Rho-kinase 1 is responsible for insulin regulation of the AMPK/SREBP-1c pathway in skeletal muscle cells exposed to palmitate.
abstract: OBJECTIVE
  Clinical and experimental data suggest that early insulin therapy could reduce lipotoxicity in subjects and animal models with type 2 diabetes mellitus. However, the underlying mechanisms need to be clarified. Sterol regulatory element-binding protein 1c (SREBP-1c), which is negatively regulated by AMP-activated protein kinase (AMPK), plays a critical role in lipotoxicity and insulin resistance in skeletal muscle cells. Here, we investigated the effect and molecular mechanism of insulin intervention on the AMPK/SREBP-1c pathway in skeletal muscle cells with chronic exposure to palmitic acid (PA).
  
  
  METHODS
  Male C57BL/6 mice were fed with a high-fat diet for 12 weeks and were then treated with insulin, AMPK inhibitor, or metformin. L6 myotubes incubated with palmitic acid (PA) were treated with insulin or metformin. Dominant-negative AMPKα2 (DN-AMPKα2) lentivirus, AMPKα2 siRNA, or Rho-kinase 1 (ROCK1) siRNA were transfected into PA-treated L6 myotubes.
  
  
  RESULTS
  We found that the ability of PA to stimulate SREBP-1c and inhibit AMPK was reversed by insulin in L6 cells. Moreover, DN-AMPKα2 lentivirus and AMPKα2 siRNA were transfected into PA-treated L6 myotubes, and the decrease in SREBP-1c expression caused by insulin was blocked by AMPK inhibition independent of the phosphatidylinositol-4,5-biphosphate-3-kinase (PI3K)/AKT pathway. The serine/threonine kinase Rho-kinase (ROCK) 1, a downstream effector of the small G protein RhoA, was activated by PA. Interestingly, knockdown of ROCK1 by siRNA blocked the downregulation of AMPK phosphorylation under PA-treated L6 myotubes, which indicated that ROCK1 mediated the effect of insulin action on AMPK.
  
  
  CONCLUSIONS
  Our study indicated that insulin reduced lipotoxicity via ROCK1 and then improved AMPK/SREBP-1c signaling in skeletal muscle under PA-induced insulin resistance.
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China.
authors.forename: Sunyinyan
authors.initials: S
authors.lastname: Tang
authors.affiliation: Department of Endocrinology, Wuxi People's Hospital of Nanjing Medical University, Wuxi, Jiangsu, People's Republic of China.
authors.forename: Wenjun
authors.initials: W
authors.lastname: Wu
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China.
authors.forename: Wenjuan
authors.initials: W
authors.lastname: Tang
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China.
authors.forename: Zhijuan
authors.initials: Z
authors.lastname: Ge
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China.
authors.forename: Hongdong
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China.
authors.forename: Ting
authors.initials: T
authors.lastname: Hong
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China. zhudalong@nju.edu.cn.
authors.forename: Dalong
authors.initials: D
authors.lastname: Zhu
authors.affiliation: Department of Endocrinology, Drum Tower Hospital Affiliated to the Nanjing University Medical School, Nanjing, Jiangsu, People's Republic of China. biyan@nju.edu.cn.
authors.forename: Yan
authors.initials: Y
authors.lastname: Bi
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology*; D008297:Male; D008687:Metformin / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D018485:Muscle Fibers, Skeletal / Q000187:drug effects* / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D019308:Palmitic Acid / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D051780:Sterol Regulatory Element Binding Protein 1 / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D007328:Insulin; D051780:Sterol Regulatory Element Binding Protein 1; D019308:Palmitic Acid; D008687:Metformin; D054460:rho-Associated Kinases; D055372:AMP-Activated Protein Kinases
keywords: AMPK; Insulin; Rho-kinase 1; SREBP-1c; Skeletal muscle