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Leslie Ramos, Dhanesh Amarnani, Nakul Singh, Leo A Kim; Effect of Rho-kinase Inhibition on a Patient-Derived Model of Proliferative Vitreoretinopathy. Invest. Ophthalmol. Vis. Sci. 2021;62(8):3641.
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© ARVO (1962-2015); The Authors (2016-present)
Proliferative vitreoretinopathy (PVR) is a common complication of open globe trauma and recurrent retinal detachment currently lacking medical treatment. Migration and proliferation of retinal cells, the formation of contractile membranes, and subsequent retinal detachment are hallmarks of PVR. This study aims to investigate the effect of Rho-kinase inhibition using an in vitro patient-derived cell and ex vivo explant model of PVR.
PVR membranes from human donors were cultured into a single cell suspension after digestion with enzymes creating primary PVR cell cultures. Explants were made from fragments of PVR membranes embedded into Matrigel. The effect of Rho-kinase inhibition using ROCK1 and ROCK2 inhibitors: ripasudil, netarsudil, fasudil and Y-2762 were tested. We examined the response of these inhibitors on cell proliferation of PVR cell cultures at two concentrations, selected from the IC50 . The effects of ripasudil (150 nM and 300 nM), netarsudil (30 nM and 60 nM), fasudil (500 nM and 1μM), and Y-2762 (500 nM and 1μM) were evaluated at 24 hours. The effect of Rho-kinase inhibitors at 1 μM was tested on our PVR explant. Phase contrast images were taken at 7 and 14 days, and outgrowths were measured.
At 24 hours, ripasudil (300 nM) and netarsudil (60 nM) revealed a 11% and 13% reduction in proliferation in PVR cells, whereas fasudil showed a 39% reduction in proliferation at 500 nM and a 87% reduction at 1 μM concentration. Similarly, Y-2762 showed a significant 80% reduction at 500 nM and an almost complete (94%) reduction in the proliferation of PVR cells. Robust outgrowths were observed growing from the freshly isolated PVR explant samples at 7 and 14 days (28.58 mm and 207 mm respectively) post embedding in Matrigel in culture. Ripasudil (0.8 mm and 15 mm), and netarsudil (4.2 mm and 37 mm) successfully inhibited and reduced explant growth at 7 and 14 days. The explants treated with fasudil (1 μM) and Y-2762 (1 μM) showed no outgrowths and almost complete inhibition of migration at all time points.
Currently, there are no specific therapeutic agents to prevent PVR. Common in vitro models of PVR rely primarily on RPE cell cultures to mimic early PVR, whereas our model relies on patient-derived PVR membranes. Our assays, when used in drug screens, may be beneficial at identifying potential therapies likely to work in preclinical and clinical trials for PVR.
This is a 2021 ARVO Annual Meeting abstract.
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