{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T15:30:08Z","timestamp":1787239808586,"version":"build-2736575974"},"reference-count":76,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2023,5,3]],"date-time":"2023-05-03T00:00:00Z","timestamp":1683072000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001349","name":"National Medical Research Council","doi-asserted-by":"publisher","award":["MOH-000197"],"award-info":[{"award-number":["MOH-000197"]}],"id":[{"id":"10.13039\/501100001349","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001349","name":"National Medical Research Council","doi-asserted-by":"publisher","award":["05\/FY2019\/DP(SL)\/51-A101"],"award-info":[{"award-number":["05\/FY2019\/DP(SL)\/51-A101"]}],"id":[{"id":"10.13039\/501100001349","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001349","name":"National Medical Research Council","doi-asserted-by":"publisher","award":["05\/FY2020\/DP\/(SL)\/88-A164"],"award-info":[{"award-number":["05\/FY2020\/DP\/(SL)\/88-A164"]}],"id":[{"id":"10.13039\/501100001349","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001349","name":"National Medical Research Council","doi-asserted-by":"publisher","award":["05\/FY2021\/DP(SL)\/63-A117"],"award-info":[{"award-number":["05\/FY2021\/DP(SL)\/63-A117"]}],"id":[{"id":"10.13039\/501100001349","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Singhealth Duke-NUS Ophthalmology and Visual Sciences Academic Clinical Programme: Academic Medicine Designated Philanthropic Award\u2014Distinguished Professorship in Clinical Innovation in Ophthalmology","award":["MOH-000197"],"award-info":[{"award-number":["MOH-000197"]}]},{"name":"Singhealth Duke-NUS Ophthalmology and Visual Sciences Academic Clinical Programme: Academic Medicine Designated Philanthropic Award\u2014Distinguished Professorship in Clinical Innovation in Ophthalmology","award":["05\/FY2019\/DP(SL)\/51-A101"],"award-info":[{"award-number":["05\/FY2019\/DP(SL)\/51-A101"]}]},{"name":"Singhealth Duke-NUS Ophthalmology and Visual Sciences Academic Clinical Programme: Academic Medicine Designated Philanthropic Award\u2014Distinguished Professorship in Clinical Innovation in Ophthalmology","award":["05\/FY2020\/DP\/(SL)\/88-A164"],"award-info":[{"award-number":["05\/FY2020\/DP\/(SL)\/88-A164"]}]},{"name":"Singhealth Duke-NUS Ophthalmology and Visual Sciences Academic Clinical Programme: Academic Medicine Designated Philanthropic Award\u2014Distinguished Professorship in Clinical Innovation in Ophthalmology","award":["05\/FY2021\/DP(SL)\/63-A117"],"award-info":[{"award-number":["05\/FY2021\/DP(SL)\/63-A117"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>(1) Rho-associated coiled-coil protein kinase (ROCK) signaling cascade impacts a wide array of cellular events. For cellular therapeutics, scalable expansion of primary human corneal endothelial cells (CECs) is crucial, and the inhibition of ROCK signaling using a well characterized ROCK inhibitor (ROCKi) Y-27632 had been shown to enhance overall endothelial cell yield. (2) In this study, we compared several classes of ROCK inhibitors to both ROCK-I and ROCK-II, using in silico binding simulation. We then evaluated nine ROCK inhibitors for their effects on primary CECs, before narrowing it down to the two most efficacious compounds\u2014AR-13324 (Netarsudil) and its active metabolite, AR-13503\u2014and assessed their impact on cellular proliferation in vitro. Finally, we evaluated the use of AR-13324 on the regenerative capacity of donor cornea with an ex vivo corneal wound closure model. Donor-matched control groups supplemented with Y-27632 were used for comparative analyses. (3) Our in silico simulation revealed that most of the compounds had stronger binding strength than Y-27632. Most of the nine ROCK inhibitors assessed worked within the concentrations of between 100 nM to 30 \u00b5M, with comparable adherence to that of Y-27632. Of note, both AR-13324 and AR-13503 showed better cellular adherence when compared to Y-27632. Similarly, the proliferation rates of CECs exposed to AR-13324 were comparable to those of Y-27632. Interestingly, CECs expanded in a medium supplemented with AR-13503 were significantly more proliferative in (i) untreated vs. AR-13503 (1 \u03bcM; * p &lt; 0.05); (ii) untreated vs. AR-13503 (10 \u03bcM; *** p &lt; 0.001); (iii) Y-27632 vs. AR-13503 (10 \u03bcM; ** p &lt; 0.005); (iv) AR-13324 (1 \u03bcM) vs. AR-13503 (10 \u03bcM; ** p &lt; 0.005); and (v) AR-13324 (0.1 \u03bcM) vs. AR-13503 (10 \u03bcM; * p &lt; 0.05). Lastly, an ex vivo corneal wound healing study showed a comparable wound healing rate for the final healed area in corneas exposed to Y-27632 or AR-13324. (4) In conclusion, we were able to demonstrate that various classes of ROCKi compounds other than Y-27632 were able to exert positive effects on primary CECs, and systematic donor-match controlled comparisons revealed that the FDA-approved ROCK inhibitor, AR-13324, is a potential candidate for cellular therapeutics or as an adjunct drug in regenerative treatment for corneal endothelial diseases in humans.<\/jats:p>","DOI":"10.3390\/cells12091307","type":"journal-article","created":{"date-parts":[[2023,5,4]],"date-time":"2023-05-04T01:33:36Z","timestamp":1683164016000},"page":"1307","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":19,"title":["Effects of Rho-Associated Kinase (Rock) Inhibitors (Alternative to Y-27632) on Primary Human Corneal Endothelial Cells"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5387-4209","authenticated-orcid":false,"given":"Gary S. L.","family":"Peh","sequence":"first","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"},{"name":"Singhealth Duke-NUS Ophthalmology & Visual Sciences Academic Clinical Programme , Duke-NUS Graduate Medical School, Singapore 169857, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7298-4315","authenticated-orcid":false,"given":"Francisco","family":"Bandeira","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"},{"name":"Department of Ophthalmology and Visual Sciences, Federal University of S\u00e3o Paulo, S\u00e3o Paulo 04023-062, Brazil"},{"name":"Corneal and External Diseases Department, S\u00e3o Gon\u00e7alo Eye Hospital, Rio de Janeiro 24421-005, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dawn","family":"Neo","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khadijah","family":"Adnan","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7829-9247","authenticated-orcid":false,"given":"Yossa","family":"Hartono","sequence":"additional","affiliation":[{"name":"Bioinformatics Institute, A*STAR, 30 Biopolis Street, Matrix #07-01, Singapore 138671, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5475-8712","authenticated-orcid":false,"given":"Hon Shing","family":"Ong","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"},{"name":"Singhealth Duke-NUS Ophthalmology & Visual Sciences Academic Clinical Programme , Duke-NUS Graduate Medical School, Singapore 169857, Singapore"},{"name":"Singapore National Eye Centre, Singapore 168751, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sacha","family":"Naso","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anandalakshmi","family":"Venkatraman","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jos\u00e9 A. P.","family":"Gomes","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology and Visual Sciences, Federal University of S\u00e3o Paulo, S\u00e3o Paulo 04023-062, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3044-5057","authenticated-orcid":false,"given":"Viridiana","family":"Kocaba","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"},{"name":"Netherlands Institute for Innovative Ocular Surgery, 3071AA Rotterdam, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jodhbir S.","family":"Mehta","sequence":"additional","affiliation":[{"name":"Tissue Engineering and Cell Therapy Group, Singapore Eye Research Institute, Singapore 169856, Singapore"},{"name":"Singhealth Duke-NUS Ophthalmology & Visual Sciences Academic Clinical Programme , Duke-NUS Graduate Medical School, Singapore 169857, Singapore"},{"name":"Singapore National Eye Centre, Singapore 168751, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,5,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"252","DOI":"10.1001\/archopht.121.2.252","article-title":"Gene Expression in Donor Corneal Endothelium","volume":"121","author":"Gottsch","year":"2003","journal-title":"Arch. Ophthalmol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1016\/j.exer.2011.06.004","article-title":"Molecular mechanisms underlying the corneal endothelial pump","volume":"95","author":"Bonanno","year":"2012","journal-title":"Exp. Eye Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"16","DOI":"10.1016\/j.exer.2011.08.014","article-title":"Proliferative capacity of corneal endothelial cells","volume":"95","author":"Joyce","year":"2012","journal-title":"Exp. Eye Res."},{"key":"ref_4","first-page":"3142","article-title":"Role of p27(Kip1) in cAMP- and TGF-beta2-mediated antiproliferation in rabbit corneal endothelial cells","volume":"42","author":"Kim","year":"2001","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_5","first-page":"2152","article-title":"Mechanisms of mitotic inhibition in corneal endothelium: Contact inhibition and TGF-beta2","volume":"43","author":"Joyce","year":"2002","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1097\/00003226-200005000-00002","article-title":"The Resiliency of the Corneal Endothelium to Refractive and Intraocular Surgery","volume":"19","author":"Edelhauser","year":"2000","journal-title":"Cornea"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1749","DOI":"10.1016\/S0140-6736(12)60437-1","article-title":"Corneal transplantation","volume":"379","author":"Tan","year":"2012","journal-title":"Lancet"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1001\/jamaophthalmol.2015.4776","article-title":"Global Survey of Corneal Transplantation and Eye Banking","volume":"134","author":"Gain","year":"2016","journal-title":"JAMA Ophthalmol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1284","DOI":"10.1167\/iovs.15-18586","article-title":"Effect of the Rho-Associated Kinase Inhibitor Eye Drop (Ripasudil) on Corneal Endothelial Wound Healing","volume":"57","author":"Okumura","year":"2016","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"523","DOI":"10.1097\/ICO.0000000000001518","article-title":"Regenerative Therapy for Fuchs Endothelial Corneal Dystrophy","volume":"37","author":"Soh","year":"2018","journal-title":"Cornea"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"382","DOI":"10.1097\/ICO.0000000000001472","article-title":"Selective Endothelial Removal for Peters Anomaly","volume":"37","author":"Soh","year":"2018","journal-title":"Cornea"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1016\/j.survophthal.2017.07.004","article-title":"A review of the evidence for in vivo corneal endothelial regeneration","volume":"63","author":"Bogerd","year":"2018","journal-title":"Surv. Ophthalmol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2646904","DOI":"10.1155\/2017\/2646904","article-title":"Application of Rho Kinase Inhibitors for the Treatment of Corneal Endothelial Diseases","volume":"2017","author":"Okumura","year":"2017","journal-title":"J. Ophthalmol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"529","DOI":"10.1097\/ICO.0000000000001883","article-title":"Use of Topical Rho Kinase Inhibitors in the Treatment of Fuchs Dystrophy after Descemet Stripping Only","volume":"38","author":"Macsai","year":"2019","journal-title":"Cornea"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"6087","DOI":"10.1038\/s41598-019-42493-3","article-title":"Functional Evaluation of Two Corneal Endothelial Cell-Based Therapies: Tissue-Engineered Construct and Cell Injection","volume":"9","author":"Peh","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Ong, H.S., Peh, G., Neo, D.J.H., Ang, H.-P., Adnan, K., Nyein, C.L., Morales-Wong, F., Bhogal, M., Kocaba, V., and Mehta, J.S. (2020). A Novel Approach of Harvesting Viable Single Cells from Donor Corneal Endothelium for Cell-Injection Therapy. Cells, 9.","DOI":"10.3390\/cells9061428"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"995","DOI":"10.1056\/NEJMoa1712770","article-title":"Injection of Cultured Cells with a ROCK Inhibitor for Bullous Keratopathy","volume":"378","author":"Kinoshita","year":"2018","journal-title":"N. Engl. J. Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"589","DOI":"10.1016\/j.str.2005.11.024","article-title":"Molecular Mechanism for the Regulation of Rho-Kinase by Dimerization and Its Inhibition by Fasudil","volume":"14","author":"Yamaguchi","year":"2006","journal-title":"Structure"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1016\/0014-5793(96)00811-3","article-title":"ROCK-I and ROCK-II, two isoforms of Rho-associated coiled-coil forming protein serine\/threonine kinase in mice","volume":"392","author":"Nakagawa","year":"1996","journal-title":"FEBS Lett."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Pipparelli, A., Arsenijevic, Y., Thuret, G., Gain, P., Nicolas, M., and Majo, F. (2013). ROCK Inhibitor Enhances Adhesion and Wound Healing of Human Corneal Endothelial Cells. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0062095"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"C1057","DOI":"10.1152\/ajpcell.00481.2007","article-title":"Novel molecular insights into RhoA GTPase-induced resistance to aqueous humor outflow through the trabecular meshwork","volume":"295","author":"Zhang","year":"2008","journal-title":"Am. J. Physiol. Physiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1495","DOI":"10.1111\/j.1460-9568.1995.tb01144.x","article-title":"Expression of GABA Receptor \u03c11 and \u03c12 Subunits in the Retina and Brain of the Rat","volume":"7","author":"Enz","year":"1995","journal-title":"Eur. J. Neurosci."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e29846","DOI":"10.4161\/sgtp.29846","article-title":"Rho-associated coiled-coil containing kinases (ROCK)","volume":"5","author":"Julian","year":"2014","journal-title":"Small GTPases"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1097\/FJC.0b013e318070d1bd","article-title":"Rho Kinase (ROCK) Inhibitors","volume":"50","author":"Liao","year":"2007","journal-title":"J. Cardiovasc. Pharmacol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1016\/S0968-0004(00)89022-2","article-title":"Rho as a regulator of the cytoskeleton","volume":"20","author":"Takai","year":"1995","journal-title":"Trends Biochem. Sci."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"493","DOI":"10.1038\/sj.cdd.4400987","article-title":"Rho GTPase signalling pathways in the morphological changes associated with apoptosis","volume":"9","author":"Coleman","year":"2002","journal-title":"Cell Death Differ."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"2222","DOI":"10.2174\/0929867325666181031102829","article-title":"Rho-kinase (ROCK) Inhibitors\u2014A Neuroprotective Therapeutic Paradigm with a Focus on Ocular Utility","volume":"27","author":"Abbhi","year":"2020","journal-title":"Curr. Med. Chem."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"704","DOI":"10.2174\/156802609789044452","article-title":"Rho Kinase (ROCK) Inhibitors and Their Application to Inflammatory Disorders","volume":"9","author":"LoGrasso","year":"2009","journal-title":"Curr. Top. Med. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"2383","DOI":"10.1016\/j.bmcl.2016.03.113","article-title":"A review on ROCK-II inhibitors: From molecular modelling to synthesis","volume":"26","author":"Shah","year":"2016","journal-title":"Bioorganic Med. Chem. Lett."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"5025","DOI":"10.1158\/0008-5472.CAN-12-0954","article-title":"RKI-1447 Is a Potent Inhibitor of the Rho-Associated ROCK Kinases with Anti-Invasive and Antitumor Activities in Breast Cancer","volume":"72","author":"Patel","year":"2012","journal-title":"Cancer Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3321","DOI":"10.1158\/0008-5472.CAN-17-1339","article-title":"Rho Kinase Inhibition by AT13148 Blocks Pancreatic Ductal Adenocarcinoma Invasion and Tumor Growth","volume":"78","author":"Rath","year":"2018","journal-title":"Cancer Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1316","DOI":"10.1523\/JNEUROSCI.2336-15.2016","article-title":"Rho Kinase Inhibition as a Therapeutic for Progressive Supranuclear Palsy and Corticobasal Degeneration","volume":"36","author":"Gentry","year":"2016","journal-title":"J. Neurosci."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"281","DOI":"10.1007\/s10792-021-02002-w","article-title":"Status of Rho kinase inhibitors in glaucoma therapeutics\u2014An overview","volume":"42","author":"Saha","year":"2022","journal-title":"Int. Ophthalmol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"9281745","DOI":"10.1155\/2017\/9281745","article-title":"Rho Kinase in Eye Disease","volume":"2017","author":"Okumura","year":"2017","journal-title":"J. Ophthalmol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"2475","DOI":"10.1016\/j.bmcl.2016.03.104","article-title":"Discovery of the ROCK inhibitor netarsudil for the treatment of open-angle glaucoma","volume":"26","author":"Sturdivant","year":"2016","journal-title":"Bioorganic Med. Chem. Lett."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1620","DOI":"10.1007\/s12325-020-01277-2","article-title":"Fixed-Dose Combination of Netarsudil and Latanoprost in Ocular Hypertension and Open-Angle Glaucoma: Pooled Efficacy\/Safety Analysis of Phase 3 MERCURY-1 and -2","volume":"37","author":"Asrani","year":"2020","journal-title":"Adv. Ther."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"338","DOI":"10.1167\/iovs.15-18300","article-title":"Predicative Factors for Corneal Endothelial Cell Migration","volume":"57","author":"Soh","year":"2016","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e22397","DOI":"10.1096\/fj.202101442RR","article-title":"Inhibition of Rho-associated protein kinase activity enhances oxidative phosphorylation to support corneal endothelial cell migration","volume":"36","author":"Ho","year":"2022","journal-title":"FASEB J."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"9167","DOI":"10.1038\/srep09167","article-title":"The effects of Rho-associated kinase inhibitor Y-27632 on primary human corneal endothelial cells propagated using a dual media approach","volume":"5","author":"Peh","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"6067","DOI":"10.1167\/iovs.15-17595","article-title":"Effect of the Rho Kinase Inhibitor Y-27632 on Corneal Endothelial Wound Healing","volume":"56","author":"Okumura","year":"2015","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"273","DOI":"10.1016\/S0076-6879(00)25449-9","article-title":"Use and properties of ROCK-specific inhibitor Y-27632","volume":"325","author":"Narumiya","year":"2000","journal-title":"Methods Enzym."},{"key":"ref_42","first-page":"976","article-title":"Pharmacological properties of Y-27632, a specific inhibitor of rho-associated kinases","volume":"57","author":"Ishizaki","year":"2000","journal-title":"Mol. Pharmacol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"3680","DOI":"10.1167\/iovs.08-2634","article-title":"Enhancement on Primate Corneal Endothelial Cell Survival In Vitro by a ROCK Inhibitor","volume":"50","author":"Okumura","year":"2009","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"318","DOI":"10.1167\/iovs.13-12225","article-title":"Involvement of Cyclin D and p27 in Cell Proliferation Mediated by ROCK Inhibitors Y-27632 and Y-39983 During Corneal Endothelium Wound Healing","volume":"55","author":"Okumura","year":"2014","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"2269","DOI":"10.1021\/acs.jmedchem.5b00683","article-title":"Rho Kinase (ROCK) Inhibitors and Their Therapeutic Potential","volume":"59","author":"Feng","year":"2016","journal-title":"J. Med. Chem."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"980","DOI":"10.1016\/j.ajo.2015.01.026","article-title":"Ocular Hypotensive Safety and Systemic Absorption of AR-13324 Ophthalmic Solution in Normal Volunteers","volume":"159","author":"Levy","year":"2015","journal-title":"Am. J. Ophthalmol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"660","DOI":"10.2174\/1381612822666161205110027","article-title":"The Role of Rho Kinase Inhibitors in Corneal Endothelial Dysfunction","volume":"23","author":"Koizumi","year":"2017","journal-title":"Curr. Pharm. Des."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"287","DOI":"10.3727\/096368913X675719","article-title":"Propagation of Human Corneal Endothelial Cells: A Novel Dual Media Approach","volume":"24","author":"Peh","year":"2015","journal-title":"Cell Transplant."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"834","DOI":"10.1016\/j.ajo.2011.04.012","article-title":"Ocular Hypotensive Effect of the Rho Kinase Inhibitor AR-12286 in Patients with Glaucoma and Ocular Hypertension","volume":"152","author":"Williams","year":"2011","journal-title":"Am. J. Ophthalmol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"6177","DOI":"10.1021\/jm051256o","article-title":"Extra Precision Glide: Docking and Scoring Incorporating a Model of Hydrophobic Enclosure for Protein\u2212Ligand Complexes","volume":"49","author":"Friesner","year":"2006","journal-title":"J. Med. Chem."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Peh, G., Toh, K.-P., Wu, F.-Y., Tan, D.T., and Mehta, J.S. (2011). Cultivation of Human Corneal Endothelial Cells Isolated from Paired Donor Corneas. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0028310"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"14149","DOI":"10.1038\/s41598-017-14723-z","article-title":"Regulatory Compliant Tissue-Engineered Human Corneal Endothelial Grafts Restore Corneal Function of Rabbits with Bullous Keratopathy","volume":"7","author":"Peh","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"26113","DOI":"10.1038\/srep26113","article-title":"Rho kinase inhibitor enables cell-based therapy for corneal endothelial dysfunction","volume":"6","author":"Okumura","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"1006","DOI":"10.1136\/bjo.2010.194571","article-title":"Enhancement of corneal endothelium wound healing by Rho-associated kinase (ROCK) inhibitor eye drops","volume":"95","author":"Okumura","year":"2011","journal-title":"Br. J. Ophthalmol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1009","DOI":"10.3892\/ijmm.2017.3103","article-title":"The ROCK inhibitor, thiazovivin, inhibits human corneal endothelial-to-mesenchymal transition\/epithelial-to-mesenchymal transition and increases ionic transporter expression","volume":"40","author":"Wu","year":"2017","journal-title":"Int. J. Mol. Med."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"5313","DOI":"10.1128\/MCB.16.10.5313","article-title":"The p160 RhoA-Binding Kinase ROK\u03b1 Is a Member of a Kinase Family and Is Involved in the Reorganization of the Cytoskeleton","volume":"16","author":"Leung","year":"1996","journal-title":"Mol. Cell Biol."},{"key":"ref_57","doi-asserted-by":"crossref","unstructured":"Peh, G.S., Toh, K.-P., Ang, H.-P., Seah, X.-Y., George, B.L., and Mehta, J.S. (2013). Optimization of human corneal endothelial cell culture: Density dependency of successful cultures in vitro. BMC Res. Notes, 6.","DOI":"10.1186\/1756-0500-6-176"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1038\/s41598-017-00209-5","article-title":"Human Corneal Endothelial Cell Cultivation From Old Donor Corneas with Forced Attachment","volume":"7","author":"Parekh","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"6843","DOI":"10.1167\/iovs.16-20123","article-title":"Activation of the Rho\/Rho Kinase Signaling Pathway Is Involved in Cell Death of Corneal Endothelium","volume":"57","author":"Okumura","year":"2016","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"2211","DOI":"10.1007\/s40265-014-0333-2","article-title":"Ripasudil: First Global Approval","volume":"74","year":"2014","journal-title":"Drugs"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"642","DOI":"10.1097\/ICO.0000000000001209","article-title":"Descemetorhexis Without Grafting for Fuchs Endothelial Dystrophy\u2014Supplementation with Topical Ripasudil","volume":"36","author":"Moloney","year":"2017","journal-title":"Cornea"},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"e38","DOI":"10.1097\/ICO.0000000000001381","article-title":"ROCK Inhibitor (Ripasudil) as Coadjuvant after Descemetorhexis Without an Endothelial Graft","volume":"36","author":"Galvis","year":"2017","journal-title":"Cornea"},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"7560","DOI":"10.1167\/iovs.15-17887","article-title":"Rho-Associated Kinase Inhibitor Eye Drop (Ripasudil) Transiently Alters the Morphology of Corneal Endothelial Cells","volume":"56","author":"Okumura","year":"2015","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_64","first-page":"289","article-title":"Long-term Side Effects Including Blepharitis Leading to Discontinuation of Ripasudil","volume":"28","author":"Saito","year":"2019","journal-title":"Eur. J. Gastroenterol. Hepatol."},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"1617","DOI":"10.1167\/iovs.17-23637","article-title":"A Cell Culture Approach to Optimized Human Corneal Endothelial Cell Function","volume":"59","author":"Bartakova","year":"2018","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"692","DOI":"10.1038\/s41598-020-80267-4","article-title":"ROCK inhibition reduces morphological and functional damage to rod synapses after retinal injury","volume":"11","author":"Zarbin","year":"2021","journal-title":"Sci. Rep."},{"key":"ref_67","first-page":"CD012097","article-title":"Descemet\u2019s membrane endothelial keratoplasty (DMEK) versus Descemet's stripping automated endothelial keratoplasty (DSAEK) for corneal endothelial failure","volume":"6","author":"Stuart","year":"2018","journal-title":"Cochrane Database Syst. Rev."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"4249","DOI":"10.1167\/iovs.17-22106","article-title":"Allogeneic Descemet\u2019s membrane endothelial keratoplasty (DMEK) versus Descs Membrane Transplantation Enhances Corneal Endothelial Monolayer Formation and Restores Functional Integrity Following Descemet\u2019s Stripping","volume":"58","author":"Bhogal","year":"2017","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"6983","DOI":"10.1038\/s41598-017-07557-2","article-title":"Descemet\u2019s Membrane Supports Corneal Endothelial Cell Regeneration in Rabbits","volume":"7","author":"Chen","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_70","first-page":"54329","article-title":"In Vitro and In Vivo Models to Study Corneal Endothelial-mesenchymal Transition","volume":"114","author":"Ho","year":"2016","journal-title":"J. Vis. Exp."},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1016\/j.exer.2011.08.003","article-title":"Endothelial mesenchymal transformation mediated by IL-1\u03b2-induced FGF-2 in corneal endothelial cells","volume":"95","author":"Lee","year":"2012","journal-title":"Exp. Eye Res."},{"key":"ref_72","doi-asserted-by":"crossref","first-page":"786","DOI":"10.1016\/j.ajpath.2013.05.025","article-title":"Notch Signal Regulates Corneal Endothelial-to-Mesenchymal Transition","volume":"183","author":"Li","year":"2013","journal-title":"Am. J. Pathol."},{"key":"ref_73","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1016\/j.jcjo.2021.06.010","article-title":"Descemet stripping only in Fuchs\u2019 endothelial dystrophy without use of topical Rho-kinase inhibitors: 5-year follow-up","volume":"57","author":"Iovieno","year":"2021","journal-title":"Can. J. Ophthalmol."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"320","DOI":"10.1097\/ICO.0000000000002437","article-title":"Descemet Stripping Only Supplemented With Topical Ripasudil for Fuchs Endothelial Dystrophy 12-Month Outcomes of the Sydney Eye Hospital Study","volume":"40","author":"Moloney","year":"2021","journal-title":"Cornea"},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"1225","DOI":"10.1097\/ICO.0000000000002642","article-title":"A Close Look at the Clinical Efficacy of Rho-Associated Protein Kinase Inhibitor Eye Drops for Fuchs Endothelial Corneal Dystrophy","volume":"40","author":"Kinoshita","year":"2021","journal-title":"Cornea"},{"key":"ref_76","doi-asserted-by":"crossref","first-page":"1188","DOI":"10.1097\/ICO.0000000000002975","article-title":"Surgical Management of Fuchs Endothelial Corneal Dystrophy: A Treatment Algorithm and Individual Patient Meta-Analysis of Descemet Stripping Only","volume":"41","author":"Din","year":"2022","journal-title":"Cornea"}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/9\/1307\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:28:49Z","timestamp":1760124529000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/9\/1307"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,5,3]]},"references-count":76,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2023,5]]}},"alternative-id":["cells12091307"],"URL":"https:\/\/doi.org\/10.3390\/cells12091307","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,5,3]]}}}