Vascular normalization by ROCK inhibitor: Therapeutic potential of ripasudil (K-115) eye drop in retinal angiogenesis and hypoxia

Muneo Yamaguchi, Shintaro Nakao, Ryoichi Arita, Yoshihiro Kaizu, Mitsuru Arima, Yedi Zhou, Takeshi Kita, Shigeo Yoshida, Kazuhiro Kimura, Tomoyuki Isobe, Yoshio Kaneko, Koh hei Sonoda, Tatsuro Ishibashi

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PURPOSE. In this study, we investigated the therapeutic potential of a Rho-associated coiled-coil–containing protein kinase (ROCK) inhibitor ripasudil (K-115) eye drop on retinal neovascularization and hypoxia. METHODS. In vitro, human retinal microvascular endothelial cells (HRMECs) were pretreated with ripasudil and then stimulated with VEGF. ROCK activity was evaluated by phosphorylation of myosin phosphatase target protein (MYPT)-1. Endothelial migration and cell viability were assessed by cell migration and MTT assay, respectively. The concentration of ripasudil in the retina was measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In vivo, normal saline, 0.4%, or 0.8% ripasudil were administered three times a day to mice with oxygen-induced retinopathy (OIR). The areas of neovascularization and avascular retina were also quantified with retinal flat-mounts at postnatal day (P) 15, P17, or P21. The retinal hypoxic area was evaluated using hypoxia-sensitive drug pimonidazole by immunohistochemistry at P17. The vascular normalization was also evaluated by immunohistochemistry at P17. RESULTS. Ripasudil but not fasudil significantly reduced VEGF-induced MYPT-1 phosphorylation in HRMECs at 30 lmol/L. Ripasudil significantly inhibited VEGF-induced HRMECs migration and proliferation. The concentration of ripasudil in the retina was 3.8 to 10.4 lmol/ L and 6.8 to 14.8 lmol/L after 0.4% and 0.8% ripasudil treatment, respectively. In the 0.4% and 0.8% ripasudil treated OIR mice, the areas of neovascularization as well as avascular area in the retina was significantly reduced compared with those of saline-treated mice at P17 and P21. Pimonidazole staining revealed that treatment with 0.4% and 0.8% ripasudil significantly inhibited the increase in the hypoxic area compared with saline. 0.8% ripasudil could cause intraretinal vascular sprouting and increase retinal vascular perfusion. CONCLUSIONS. Novel ROCK inhibitor ripasudil eye drop has therapeutic potential in the treatment of retinal hypoxic neovascular diseases via antiangiogenic effects as well as vascular normalization.

元の言語英語
ページ(範囲)2264-2276
ページ数13
ジャーナルInvestigative Ophthalmology and Visual Science
57
発行部数4
DOI
出版物ステータス出版済み - 2016

All Science Journal Classification (ASJC) codes

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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