title: A selective inhibitor of the Rho kinase pathway, Y-27632, and its influence on wound healing in the corneal stroma.
abstract: OBJECTIVE
  Our study examined the effect of a selective Rho kinase inhibitor, Y-27632, on corneal wound healing and potential stromal scarring after superficial keratectomy.
  
  
  METHODS
  Rabbit keratocytes were induced into myofibroblasts by transforming growth factor β1 (TGFβ1) either with or without Y-27632. Then α-smooth muscle actin (α-SMA) was examined by immunohistochemistry and western blotting, and the contractility of the seeded collagen gels was measured. Y-27632 eye drops (or vehicle only) were administered to eyes after a superficial keratectomy, and the tissue was examined by immunohistochemistry for α-SMA, collagen types I, II, and III, and keratan sulfate. Electron microscopy was conducted with and without histochemical contrasting of sulfated proteoglycans.
  
  
  RESULTS
  Spindle-like cells in culture constituted 99.5±1.1% with TGFβ1 stimulation, but 3.5±1.0% after TGFβ1 and Y-27632 treatment (p<0.01, n=6). α-SMA was seen in 4% of TGFβ1-treated cells, but in only 0.3% of cells with Y-27632 added (p<0.01, n=6), which was confirmed by western blotting. Y-27632 also inhibited the TGFβ1-induced contraction of seeded collagen gels. After superficial keratectomies, collagen type I and keratan sulfate were unchanged by Y-27632 application. Collagen type II was not detected in Y-27632 or vehicle-only corneas. With Y-27632 treatment, α-SMA expression increased and the collagen type III signal became in the weaker subepithelial area. Interestingly, bundles of aligned and uniformly spaced collagen fibrils were more prevalent in keratocytes in Y-27632-treated corneas, which is reminiscent of fibripositor-like structures that have been proposed as a mechanism of matrix deposition in embryonic connective tissues.
  
  
  CONCLUSIONS
  Y-27632 inhibits keratocyte-to-myofibroblast transition, and its topical application after a superficial lamellar keratectomy elicits an altered wound healing response, with evidence of an embryonic-type deposition of collagen fibrils.
authors: Faculty of Life and Medical Sciences, Doshisha University, Kyotanabe, Kyoto, Japan.
authors: Mayumi
authors: M
authors: Yamamoto
authors: Andrew J
authors: AJ
authors: Quantock
authors: Robert D
authors: RD
authors: Young
authors: Naoki
authors: N
authors: Okumura
authors: Morio
authors: M
authors: Ueno
authors: Yuji
authors: Y
authors: Sakamoto
authors: Shigeru
authors: S
authors: Kinoshita
authors: Noriko
authors: N
authors: Koizumi
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D000577:Amides / Q000494:pharmacology* / Q000627:therapeutic use; D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D002921:Cicatrix / Q000188:drug therapy* / Q000378:metabolism / Q000517:prevention & control; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D060527:Corneal Keratocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D003319:Corneal Stroma / Q000187:drug effects* / Q000378:metabolism / Q000601:surgery; D004791:Enzyme Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D005782:Gels; D015870:Gene Expression; D007632:Keratan Sulfate / Q000235:genetics / Q000378:metabolism; D008297:Male; D058628:Myofibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009883:Ophthalmic Solutions / Q000494:pharmacology* / Q000627:therapeutic use; D013508:Ophthalmologic Surgical Procedures; D011725:Pyridines / Q000494:pharmacology* / Q000627:therapeutic use; D011817:Rabbits; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology; D014945:Wound Healing / Q000187:drug effects*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics
chemical_list: D000199:Actins; D000577:Amides; D004791:Enzyme Inhibitors; D005782:Gels; D009883:Ophthalmic Solutions; D011725:Pyridines; D053773:Transforming Growth Factor beta1; C108830:Y 27632; D003094:Collagen; D007632:Keratan Sulfate; D054460:rho-Associated Kinases