title: Angiotensin II regulates the LARG/RhoA/MYPT1 axis in rat vascular smooth muscle in vitro.
abstract: OBJECTIVE
  To identify a key protein that binds monomeric G protein RhoA and activates the RhoA/Rho kinase/MYPT1 axis in vascular smooth muscle cells (VSMCs) upon angiotensin II (Ang II) stimulation.
  
  
  METHODS
  Primary cultured VSMCs from Sprague-Dawley rats were transfected with siRNAs against leukemia-associated RhoGEF (LARG), and then treated with Ang II, losartan, PD123319, or Val(5)-Ang II. The target mRNA and protein levels were determined using qPCR and Western blot analysis, respectively. Rat aortic rings were isolated, and the isometric contraction was measured with a force transducer and recorder.
  
  
  RESULTS
  Stimulation with Ang II (0.1 μmol/L) for 0.5 h significantly increased the level of LARG mRNA in VSMCs. At 3, 6, and 9 h after the treatment with Ang II (0.1 μmol/L) plus AT(2) antagonist PD123319 (1 μmol/L) or with AT(1) agonist Val(5)-Ang II (1 μmol/L), the LARG protein, RhoA activity, and phosphorylation level of myosin phosphatase target subunit 1 (MYPT1) in VSMCs were significantly increased. Knockdown of LARG with siRNA reduced these effects caused by AT(1) receptor activation. In rat aortic rings pretreated with LARG siRNA, Ang II-induced contraction was diminished.
  
  
  CONCLUSIONS
  Ang II upregulates LARG gene expression and activates the LARG/RhoA/MYPT1 axis via AT(1), thereby maintaining vascular tone.
authors.affiliation: Graduate Institute of Physiology, National Taiwan University College of Medicine, Taipei, Taiwan, China.
authors.forename: Wei-chiao
authors.initials: WC
authors.lastname: Chiu
authors.forename: Jyh-ming
authors.initials: JM
authors.lastname: Juang
authors.forename: Shen-nan
authors.initials: SN
authors.lastname: Chang
authors.forename: Cho-kai
authors.initials: CK
authors.lastname: Wu
authors.forename: Chia-ti
authors.initials: CT
authors.lastname: Tsai
authors.forename: Yung-zu
authors.initials: YZ
authors.lastname: Tseng
authors.forename: Fu-tien
authors.initials: FT
authors.lastname: Chiang
mesh_terms: D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000503:physiopathology; D015153:Blotting, Western; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics*; D007537:Isometric Contraction / Q000187:drug effects; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D061251:Primary Cell Culture; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D060888:Real-Time Polymerase Chain Reaction; D064067:Rho Guanine Nucleotide Exchange Factors; D015854:Up-Regulation; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: C577224:Arhgef12 protein, rat; D020662:Guanine Nucleotide Exchange Factors; D034741:RNA, Small Interfering; D064067:Rho Guanine Nucleotide Exchange Factors; D000804:Angiotensin II; C499850:Ppp1r12a protein, rat; D054645:Protein Phosphatase 1; D020742:rhoA GTP-Binding Protein