Contraction of myofibroblasts in granulation tissue is dependent on Rho/Rho kinase/myosin light chain phosphatase activity.
During wound healing and fibrocontractive diseases fibroblasts acquire a smooth muscle cell-like phenotype by differentiating into contractile force generating myofibroblasts. We examined whether regulation of myofibroblast contraction in granulation tissue is dominated by Ca2+-induced phosphorylation of myosin light chain kinase or by Rho/Rho kinase (ROCK)-mediated inhibition of myosin light chain phosphatase, similar to that of cultured myofibroblasts. Strips of granulation tissue obtained from rat granuloma pouches were stimulated with endothelin-1 (ET-1), serotonin, and angiotensin-II and isometric force generation was measured. We here investigated ET-1 in depth, because it was the only agonist that produced a long-lasting and strong response. The ROCK inhibitor Y27632 completely inhibited ET-1-promoted contraction and the phosphatase inhibitor calyculin elicited contraction in the absence of any other agonists, suggesting that activation of the Rho/ROCK/myosn light chain phosphatase pathway is critical in regulating in vivo myofibroblast contraction. Membrane depolarization with K+ also stimulated a long-lasting contraction of granulation tissue; however, the amount of force generated was significantly less compared to ET-1. Moreover, K+-induced contraction was inhibited by Y27632. These results are consistent with inhibition of myosin light chain phosphatase by the Rho/ROCK signaling pathway, which would account for the long-duration contraction of myofibroblasts necessary for wound closure.
Tomasek
James J
JJ
Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA. james-tomasek@ouhsc.edu
Vaughan
Melville B
MB
Kropp
Bradley P
BP
Gabbiani
Giulio
G
Martin
Michael D
MD
Haaksma
Carol J
CJ
Hinz
Boris
B
D000577:Amides / Q000494:pharmacology; D000818:Animals; D019332:Endothelin-1 / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000201:enzymology / Q000502:physiology*; D006097:Granulation Tissue / Q000201:enzymology / Q000473:pathology / Q000503:physiopathology*; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007537:Isometric Contraction / Q000187:drug effects; D008297:Male; D009130:Muscle, Smooth / Q000166:cytology*; D043263:Myosin-Light-Chain Phosphatase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D014945:Wound Healing / Q000502:physiology*; D054460:rho-Associated Kinases
D000577:Amides; D019332:Endothelin-1; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D047428:Protein Kinase Inhibitors; D011725:Pyridines; C108830:Y 27632; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D043263:Myosin-Light-Chain Phosphatase