September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Effects of ripasudil (K-115), a Rho kinase inhibitor, on the activation of human conjunctival fibroblasts
Author Affiliations & Notes
  • Akiko Futakuchi
    Ophthalmology, Kumamoto University Hospital, Chuo-ku, Kumamoto city, Kumamoto, Japan
  • Toshihiro Inoue
    Ophthalmology, Kumamoto University Hospital, Chuo-ku, Kumamoto city, Kumamoto, Japan
  • Tomokazu Fujimoto
    Ophthalmology, Kumamoto University Hospital, Chuo-ku, Kumamoto city, Kumamoto, Japan
  • Miyuki Mochita Inoue
    Ophthalmology, Kumamoto University Hospital, Chuo-ku, Kumamoto city, Kumamoto, Japan
  • Hidenobu Tanihara
    Ophthalmology, Kumamoto University Hospital, Chuo-ku, Kumamoto city, Kumamoto, Japan
  • Footnotes
    Commercial Relationships   Akiko Futakuchi, None; Toshihiro Inoue, None; Tomokazu Fujimoto, None; Miyuki Inoue, None; Hidenobu Tanihara, Alcon Japan (S), Kowa (C), MSD (C), Pfizer Japan (S), Santen Pharmaceutical (S), Senju Pharmaceutical (S)
  • Footnotes
    Support  Kowa Company, Ltd., Nagoya, Japan.
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 2929. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to Subscribers Only
      Sign In or Create an Account ×
    • Get Citation

      Akiko Futakuchi, Toshihiro Inoue, Tomokazu Fujimoto, Miyuki Mochita Inoue, Hidenobu Tanihara; Effects of ripasudil (K-115), a Rho kinase inhibitor, on the activation of human conjunctival fibroblasts. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2929.

      Download citation file:


      © 2017 Association for Research in Vision and Ophthalmology.

      ×
  • Supplements
Abstract

Purpose : Ripasudil, a selective Rho kinase inhibitor, is an ophthalmic solution which was approved in Japan for the twice-daily treatment of glaucoma and ocular hypertension in 2014. The purpose of this study is to assess the effects of ripasudil on the activation of human conjunctival fibroblasts.

Methods : Human conjunctival fibroblasts were pretreated with or without different concentrations of ripasudil (25 and 50 μM) for 1 hour and subsequently stimulated with 5 ng/ml TGF-β2 for 48 hours. The effects of ripasudil on α-smooth muscle actin (α-SMA) expression and extracellular matrix (ECM) expression were analyzed by Western blot analysis. Contractile activity was evaluated by collagen gel contraction assay. Cell viability and cytotoxicity were assessed using WST-8 assay and Hoechst 33342/propidium iodide (PI) dual staining, respectively. The human monocytic cell line THP-1 were differentiated into M1- and M2-like macrophages, and fibroblasts were treated with conditioned medium derived from these macrophages in the presence or in the absence of 50 μM ripasudil to quantify the α-SMA expression level.

Results : TGF-β2-treated fibroblasts showed a significant increase (p < 0.0001), 38 ± 4-fold in the expression of α-SMA compared with controls without TGF-β2 stimulation. When fibroblasts were pretreated with ripasudil before TGF-β2 stimulation, α-SMA expression was significantly decreased in a dose-dependent manner (25 and 50 μM, respectively, p < 0.01). Ripasudil pretreatment significantly attenuated TGF-β2-induced fibronectin expression (p < 0.0001). Ripasudil pretreatment significantly attenuated TGF-β2-induced collagen gel contraction (p < 0.0001). TGF-β2 increased the proliferation of fibroblasts (p < 0.0001), and the effect was significantly attenuated by ripasudil (p < 0.0001). Ripasudil did not cause any cellular toxicity at the tested doses. Conditioned medium from M2-like macrophages induced a significant increase (p < 0.001), 3.4 ± 0.7-fold in the expression of α-SMA, and ripasudil suppressed this upregulated expression (p < 0.01).

Conclusions : Ripasudil attenuated the activation of human conjunctival fibroblasts. Our results suggest that ripasudil might have a therapeutic potential in the prevention of excessive scarring after glaucoma filtration surgery.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×