The corneal endothelium plays a key role in maintaining corneal transparency. Its dysfunction is currently treated with penetrating or lamellar keratoplasty. Advanced cell therapy methods seek to address the persistent global deficiency of donor corneas by enabling the renewal of the endothelial monolayer with tissue-engineered grafts. This review provides an overview of recently published literature on the preparation of endothelial grafts for transplantation derived from cadaveric corneas that have developed over the last decade (2010-2021). Factors such as the most suitable donor parameters, culture substrates and media, endothelial graft storage conditions, and transplantation methods are discussed. Despite efforts to utilize alternative cellular sources, such as induced pluripotent cells, cadaveric corneas appear to be the best source of cells for graft preparation to date. However, native endothelial cells have a limited natural proliferative capacity, and they often undergo rapid phenotype changes in ex vivo culture. This is the main reason why no culture protocol for a clinical-grade endothelial graft prepared from cadaveric corneas has been standardized so far. Currently, the most established ex vivo culture protocol involves the peel-and-digest method of cell isolation and cell culture by the dual media method, including the repeated alternation of high and low mitogenic conditions. Culture media are enriched by additional substances, such as signaling pathway (Rho-associated protein kinase, TGF-β, etc.) inhibitors, to stimulate proliferation and inhibit unwanted morphological changes, particularly the endothelial-to-mesenchymal transition. To date, this promising approach has led to the development of endothelial grafts for the first in-human clinical trial in Japan. In addition to the lack of a standard culture protocol, endothelial-specific markers are still missing to confirm the endothelial phenotype in a graft ready for clinical use. Because the corneal endothelium appears to comprise phenotypically heterogeneous populations of cells, the genomic and proteomic expression of recently proposed endothelial-specific markers, such as Cadherin-2, CD166, or SLC4A11, must be confirmed by additional studies. The preparation of endothelial grafts is still challenging today, but advances in tissue engineering and surgery over the past decade hold promise for the successful treatment of endothelial dysfunctions in more patients worldwide.

Department of Medical Biochemistry Oslo University Hospital Oslo Norway

Department of Plastic and Reconstructive Surgery Oslo University Hospital Oslo Norway

Laboratory of the Biology and Pathology of the Eye Institute of Biology and Medical Genetics 1st Faculty of Medicine Charles University and General University Hospital Prague Albertov 4 128 00 Prague Czech Republic

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Yee RW, Matsuda M, Schultz RO, et al. Changes in the normal corneal endothelial cellular-pattern as a function of age. Curr Eye Res. 1985;4(6):671–678. doi: 10.3109/02713688509017661. PubMed DOI

Lass JH, Sugar A, Benetz BA, et al. Endothelial cell density to predict endothelial graft failure after penetrating keratoplasty. Arch Ophthalmol. 2010;128(1):63–69. doi: 10.1001/archophthalmol.2010.128.63. PubMed DOI PMC

Edelhauser HF. The balance between corneal transparency and edema: the Proctor Lecture. Investig Ophthalmol Vis Sci. 2006;47(5):1754–1767. doi: 10.1167/iovs.05-0686. PubMed DOI

Krabcova I, Studeny P, Jirsova K. Endothelial cell density before and after the preparation of corneal lamellae for descemet membrane endothelial keratoplasty with a stromal rim. Cornea. 2011;30(12):1436–1441. doi: 10.1097/ICO.0b013e318212497e. PubMed DOI

Joyce NC. Proliferative capacity of corneal endothelial cells. Exp Eye Res. 2012;95(1):16–23. doi: 10.1016/j.exer.2011.08.014. PubMed DOI PMC

McGowan SL, Edelhauser HF, Pfister RR, et al. Stem cell markers in the human posterior limbus and corneal endothelium of unwounded and wounded corneas. Mol Vis. 2007;13(223–27):1984–2000. PubMed

Yam GHF, Seah X, Yusoff NZBM, et al. Characterization of human transition zone reveals a putative progenitor-enriched niche of corneal endothelium. Cells. 2019;8(10). PubMed PMC

He ZG, Campolmi N, Gain P, et al. Revisited microanatomy of the corneal endothelial periphery: new evidence for continuous centripetal migration of endothelial cells in humans. Stem Cells. 2012;30(11):2523–2534. doi: 10.1002/stem.1212. PubMed DOI

Mimura T, Joyce NC. Replication competence and senescence in central and peripheral human corneal endothelium. Investig Ophthalmol Vis Sci. 2006;47(4):1387–1396. doi: 10.1167/iovs.05-1199. PubMed DOI

Senoo T, Joyce NC. Cell cycle kinetics in corneal endothelium from old and young donors. Investig Ophthalmol Vis Sci. 2000;41(3):660–667. PubMed

Parekh M, Peh G, Mehta JS, et al. Effects of corneal preservation conditions on human corneal endothelial cell culture. Exp Eye Res. 2019;179:93–101. doi: 10.1016/j.exer.2018.11.007. PubMed DOI

Gain P, Jullienne R, He Z, et al. Global survey of corneal transplantation and eye banking. JAMA Ophthalmol. 2016;134(2). PubMed

Gage PJ, Rhoades W, Prucka SK, et al. Fate maps of neural crest and mesoderm in the mammalian eye. Investig Ophthalmol Vis Sci. 2005;46(11):4200–4208. doi: 10.1167/iovs.05-0691. PubMed DOI

Jirsova K, Neuwirth A, Kalasova S, et al. Mesothelial proteins are expressed in the human cornea. Exp Eye Res. 2010;91(5):623–629. doi: 10.1016/j.exer.2010.08.002. PubMed DOI

Frausto RF, Le DJ, Aldave AJ. Transcriptomic analysis of cultured corneal endothelial cells as a validation for their use in cell replacement therapy. Cell Transplant. 2016;25(6):1159–1176. doi: 10.3727/096368915X688948. PubMed DOI PMC

Li W, Sabater AL, Chen YT, et al. A novel method of isolation, preservation, and expansion of human corneal endothelial cells. Investig Ophthalmol Vis Sci. 2007;48(2):614–620. doi: 10.1167/iovs.06-1126. PubMed DOI PMC

Choi JS, Kim BY, Kim MJ, et al. Factors affecting successful isolation of human corneal endothelial cells for clinical use. Cell Transplant. 2014;23(7):845–854. doi: 10.3727/096368913X664559. PubMed DOI

Frausto RF, Swamy VS, Peh GSL, et al. Phenotypic and functional characterization of corneal endothelial cells during in vitro expansion. Sci Rep. 2020;10(1):7402. doi: 10.1038/s41598-020-64311-x. PubMed DOI PMC

Peh GSL, Chng ZZ, Ang HP, et al. Propagation of human corneal endothelial cells: a novel dual media approach. Cell Transplant. 2015;24(2):287–304. doi: 10.3727/096368913X675719. PubMed DOI

Ong HS, Peh G, Neo DJH, et al. a novel approach of harvesting viable single cells from donor corneal endothelium for cell-injection therapy. Cells. 2020;9(6). PubMed PMC

Peh GSL, Ong HS, Adnan K, et al. Functional evaluation of two corneal endothelial cell-based therapies: tissue-engineered construct and cell injection. Sci Rep. 2019;9. PubMed PMC

Faye PA, Poumeaud F, Chazelas P, et al. Focus on cell therapy to treat corneal endothelial diseases. Exp Eye Res. 2021;204. PubMed

Gong Y, Duan H, Wang X, et al. Transplantation of human induced pluripotent stem cell-derived neural crest cells for corneal endothelial regeneration. Stem Cell Res Ther. 2021;12(1). PubMed PMC

Kinoshita S, Koizumi N, Ueno M, et al. Injection of cultured cells with a ROCK inhibitor for bullous keratopathy. N Engl J Med. 2018;378(11):995–1003. doi: 10.1056/NEJMoa1712770. PubMed DOI

Numa K, Imai K, Ueno M, et al. Five-year follow-up of first 11 patients undergoing injection of cultured corneal endothelial cells for corneal endothelial failure. Ophthalmology. 2021;128(4):504–514. doi: 10.1016/j.ophtha.2020.09.002. PubMed DOI

Toda M, Ueno M, Hiraga A, et al. Production of homogeneous cultured human corneal endothelial cells indispensable for innovative cell therapy. Investig Ophthalmol Vis Sci. 2017;58(4):2011–2020. doi: 10.1167/iovs.16-20703. PubMed DOI

Ueno M, Asada K, Toda M, et al. MicroRNA profiles qualify phenotypic features of cultured human corneal endothelial cells. Investig Ophthalmol Vis Sci. 2016;57(13):5509–5517. doi: 10.1167/iovs.16-19804. PubMed DOI

Frausto RF, Wang C, Aldave AJ. Transcriptome analysis of the human corneal endothelium. Investig Ophthalmol Vis Sci. 2014;55(12):7821–7830. doi: 10.1167/iovs.14-15021. PubMed DOI PMC

Jirsova K, Dahl P, Armitage WJ. Corneal storage, hypothermia, and organ culture. In Light and specular microscopy of the cornea. 2017. p. 41–57.

Peh GSL, Ang H-P, Lwin CN, et al. Regulatory compliant tissue-engineered human corneal endothelial grafts restore corneal function of rabbits with bullous keratopathy. Sci Rep. 2017;7(1). PubMed PMC

Xia X, Atkins M, Dalal R, et al. Magnetic human corneal endothelial cell transplant: delivery, retention, and short-term efficacy. Investig Ophthalmol Vis Sci. 2019;60(7). PubMed PMC

Zavala J, López Jaime GR, Rodríguez Barrientos CA, et al. Corneal endothelium: developmental strategies for regeneration. Eye. 2013;27(5):579–588. doi: 10.1038/eye.2013.15. PubMed DOI PMC

He Z, Forest F, Gain P, et al. 3D map of the human corneal endothelial cell. Sci Rep. 2016;6:29047. doi: 10.1038/srep29047. PubMed DOI PMC

Merjava S, Neuwirth A, Mandys V, et al. Cytokeratins 8 and 18 in adult human corneal endothelium. Exp Eye Res. 2009;89(3):426–431. doi: 10.1016/j.exer.2009.04.009. PubMed DOI

Foets BJJ, Vandenoord JJ, Desmet VJ, et al. Cytoskeletal filament typing of human corneal endothelial-cells. Cornea. 1990;9(4):312–317. doi: 10.1097/00003226-199010000-00008. PubMed DOI

Bonanno JA. Molecular mechanisms underlying the corneal endothelial pump. Exp Eye Res. 2012;95(1):2–7. doi: 10.1016/j.exer.2011.06.004. PubMed DOI PMC

Zhang W, Li H, Ogando DG, et al. Glutaminolysis is essential for energy production and ion transport in human corneal endothelium. EBioMedicine. 2017;16:292–301. doi: 10.1016/j.ebiom.2017.01.004. PubMed DOI PMC

Dawson DG, Edelhauser HF. Corneal edema. In Ocular Disease. 2010. p. 64–73.

Parekh M, Romano V, Ruzza A, et al. Increasing donor endothelial cell pool by culturing cells from discarded pieces of human donor corneas for regenerative treatments. J Ophthalmol. 2019;2019. PubMed PMC

Bartakova A, Kuzmenko O, Alvarez-Delfin K, et al. A cell culture approach to optimized human corneal endothelial cell function. Investig Ophthalmol Vis Sci. 2018;59(3):1617–1629. doi: 10.1167/iovs.17-23637. PubMed DOI PMC

Zhang W, Chen J, Fu Y, et al. The signaling pathway involved in the proliferation of corneal endothelial cells. J Recept Signal Transduct Res. 2015;35(6):585–591. PubMed

Hirata-Tominaga K, Nakamura T, Okumura N, et al. Corneal endothelial cell fate is maintained by LGR5 through the regulation of Hedgehog and Wnt pathway. Stem Cells. 2013;31(7):1396–1407. doi: 10.1002/stem.1390. PubMed DOI

Amann J, Holley GP, Lee SB, et al. Increased endothelial cell density in the paracentral and peripheral regions of the human cornea—Author reply. Am J Ophthalmol. 2003;136(4):774–775. doi: 10.1016/S0002-9394(03)00808-0. PubMed DOI

Meir Y-JJ, Chen H-C, Chen C-C, et al. Revisiting existing evidence of corneal endothelial progenitors and their potential therapeutic applications in corneal endothelial dysfunction. Adv Ther. 2020;37(3):1034–1048. PubMed

Parekh M, Ahmad S, Ruzza A, et al. Human corneal endothelial cell cultivation from old donor corneas with forced attachment. Sci Rep. 2017;7. PubMed PMC

Peh GSL, Toh KP, Wu FY, et al. Cultivation of human corneal endothelial cells isolated from paired donor corneas. PLoS ONE. 2011;6(12). PubMed PMC

Zhu C, Rawe I, Joyce NC. Differential protein expression in human corneal endothelial cells cultured from young and older donors. Mol Vis. 2008;14:1805–1814. PubMed PMC

Bennett A, Mahmoud S, Drury D, et al. Impact of donor age on corneal endothelium-descemet membrane layer scroll formation. Eye Contact Lens. 2015;41(4):236–239. doi: 10.1097/ICL.0000000000000108. PubMed DOI PMC

Okumura N, Inoue R, Kakutani K, et al. Corneal endothelial cells have an absolute requirement for cysteine for survival. Cornea. 2017;36(8):988–994. doi: 10.1097/ICO.0000000000001242. PubMed DOI

Doutch JJ, Quantock AJ, Joyce NC, et al. Ultraviolet light transmission through the human corneal stroma is reduced in the periphery. Biophys J. 2012;102(6):1258–1264. doi: 10.1016/j.bpj.2012.02.023. PubMed DOI PMC

Patel P and Chatterjee S. Peroxiredoxin 6 in endothelial signaling. Antioxidants. 2019;8(3). PubMed PMC

Corwin WL, Baust JM, Van Buskirk RG, et al. 47. In vitro assessment of apoptosis and necrosis following cold storage in human corneal endothelial cells. Cryobiology. 2013;66(3).

He Z, Okumura N, Sato M, et al. Corneal endothelial cell therapy: feasibility of cell culture from corneas stored in organ culture. Cell Tissue Bank. 2021. PubMed

Nejepinska J, Juklova K, Jirsova K. Organ culture, but not hypothermic storage, facilitates the repair of the corneal endothelium following mechanical damage. Acta Ophthalmol. 2010;88(4):413–419. doi: 10.1111/j.1755-3768.2008.01490.x. PubMed DOI

Corwin WL, Baust JM, Baust JG, et al. The unfolded protein response in human corneal endothelial cells following hypothermic storage: implications of a novel stress pathway. Cryobiology. 2011;63(1):46–55. doi: 10.1016/j.cryobiol.2011.04.008. PubMed DOI PMC

Fuchsluger TA, Jurkunas U, Kazlauskas A, et al. Anti-apoptotic gene therapy prolongs survival of corneal endothelial cells during storage. Gene Ther. 2011;18(8):778–787. doi: 10.1038/gt.2011.20. PubMed DOI PMC

Schmid R, Tarau I-S, Rossi A, et al. In vivo-like culture conditions in a bioreactor facilitate improved tissue quality in corneal storage. Biotechnol J. 2018;13(1). PubMed

Zhu YT, Chen HC, Chen SY, et al. Nuclear p120 catenin unlocks mitotic block of contact-inhibited human corneal endothelial monolayers without disrupting adherent junctions. J Cell Sci. 2012;125(15):3636–3648. PubMed PMC

Spinozzi D, Miron A, Bruinsma M, et al. Improving the success rate of human corneal endothelial cell cultures from single donor corneas with stabilization medium. Cell Tissue Bank. 2018;19(1):9–17. doi: 10.1007/s10561-017-9665-y. PubMed DOI PMC

Peh GS, Toh KP, Ang HP, et al. Optimization of human corneal endothelial cell culture: density dependency of successful cultures in vitro. BMC Res Notes. 2013;6:176. doi: 10.1186/1756-0500-6-176. PubMed DOI PMC

Formisano N, Sahin G, Català P, et al. Nanoscale topographies for corneal endothelial regeneration. Appl Sci. 2021;11(2).

Zhu Q, Sun H, Yang DM, et al. Cellular substrates for cell-based tissue engineering of human corneal endothelial cells. Int J Med Sci. 2019;16(8):1072–1077. doi: 10.7150/ijms.34440. PubMed DOI PMC

Parekh M, Romano V, Hassanin K, et al. Biomaterials for corneal endothelial cell culture and tissue engineering. J Tissue Eng. 2021;12. PubMed PMC

Navaratnam J, Utheim TP, Rajasekhar VK, et al. Substrates for expansion of corneal endothelial cells towards bioengineering of human corneal endothelium. J Funct Biomater. 2015;6(3):917–945. doi: 10.3390/jfb6030917. PubMed DOI PMC

Kabosova A, Azar DT, Bannikov GA, et al. Compositional differences between infant and adult human corneal basement membranes. Investig Ophthalmol Vis Sci. 2007;48(11):4989–4999. doi: 10.1167/iovs.07-0654. PubMed DOI PMC

Bhogal M, Lwin CN, Seah XY, et al. Allogeneic descemet's membrane transplantation enhances corneal endothelial monolayer formation and restores functional integrity following descemet's stripping. Investig Ophthalmol Vis Sci. 2017;58(10):4249–4260. doi: 10.1167/iovs.17-22106. PubMed DOI

Soh YQ, Mehta JS. Regenerative therapy for Fuchs endothelial corneal dystrophy. Cornea. 2018;37(4):523–527. doi: 10.1097/ICO.0000000000001518. PubMed DOI

Wahlig S, Peh GSL, Adnan K, et al. Optimisation of storage and transportation conditions of cultured corneal endothelial cells for cell replacement therapy. Sci Rep. 2020;10(1):1681. doi: 10.1038/s41598-020-58700-5. PubMed DOI PMC

Mimura T, Yamagami S, Yokoo S, et al. Selective isolation of young cells from human corneal endothelium by the sphere-forming assay. Tissue Eng Part C Methods. 2010;16(4):803–812. doi: 10.1089/ten.tec.2009.0608. PubMed DOI

Katikireddy KR, Schmedt T, Price MO, et al. Existence of neural crest-derived progenitor cells in normal and fuchs endothelial dystrophy corneal endothelium. Am J Pathol. 2016;186(10):2736–2750. doi: 10.1016/j.ajpath.2016.06.011. PubMed DOI PMC

Sie NM, Yam GH-F, Soh YQ, et al. Regenerative capacity of the corneal transition zone for endothelial cell therapy. Stem Cell Res Ther. 2020;11(1). PubMed PMC

Mimura T, Yamagami S, Usui T, et al. Necessary prone position time for human corneal endothelial precursor transplantation in a rabbit endothelial deficiency model. Curr Eye Res. 2007;32(7–8):617–623. doi: 10.1080/02713680701530589. PubMed DOI

Ishino Y, Sano Y, Nakamura T, et al. Amniotic membrane as a carrier for cultivated human corneal endothelial cell transplantation. Investig Ophthalmol Vis Sci. 2004;45(3):800–806. doi: 10.1167/iovs.03-0016. PubMed DOI

Jackel T, Knels L, Valtink M, et al. Serum-free corneal organ culture medium (SFM) but not conventional minimal essential organ culture medium (MEM) protects human corneal endothelial cells from apoptotic and necrotic cell death. Br J Ophthalmol. 2011;95(1):123–130. doi: 10.1136/bjo.2010.183418. PubMed DOI

Vianna LMM, Kallay L, Toyono T, et al. Use of human serum for human corneal endothelial cell culture. Br J Ophthalmol. 2015;99(2):267–271. doi: 10.1136/bjophthalmol-2014-306034. PubMed DOI

Thieme D, Reuland L, Lindl T, et al. Optimized human platelet lysate as novel basis for a serum-, xeno-, and additive-free corneal endothelial cell and tissue culture. J Tissue Eng Regen Med. 2018;12(2):557–564. doi: 10.1002/term.2574. PubMed DOI

Khalili M, Asadi M, Kahroba H, et al. Corneal endothelium tissue engineering: An evolution of signaling molecules, cells, and scaffolds toward 3D bioprinting and cell sheets. J Cell Physiol. 2020;236(5):3275–3303. doi: 10.1002/jcp.30085. PubMed DOI

Engelmann K, Friedl P. Growth of human corneal endothelial cells in a serum-reduced medium. Cornea. 1995;14(1):62–70. doi: 10.1097/00003226-199501000-00011. PubMed DOI

Zhu C, Joyce NC. Proliferative response of corneal endothelial cells from young and older donors. Investig Ophthalmol Vis Sci. 2004;45(6):1743–1751. doi: 10.1167/iovs.03-0814. PubMed DOI

Hopenreijs VPT, Pels E, Vrensen GFJM, et al. Effects of human epidermal growth-factor on endothelial wound-healing of human corneas. Investig Ophthalmol Vis Sci. 1992;33(6):1946–1957. PubMed

Roy O, Leclerc VB, Bourget JM, et al. Understanding the process of corneal endothelial morphological change in vitro. Investig Ophthalmol Vis Sci. 2015;56(2):1228–1237. doi: 10.1167/iovs.14-16166. PubMed DOI

Lee JG, Jung E, Heur M. Fibroblast growth factor 2 induces proliferation and fibrosis via SNAI1-mediated activation of CDK2 and ZEB1 in corneal endothelium. J Biol Chem. 2018;293(10):3758–3769. doi: 10.1074/jbc.RA117.000295. PubMed DOI PMC

Hongo A, Okumura N, Nakahara M, et al. The effect of a p38 mitogen-activated protein kinase inhibitor on cellular senescence of cultivated human corneal endothelial cells. Investig Ophthalmol Vis Sci. 2017;58(9):3325–3334. doi: 10.1167/iovs.16-21170. PubMed DOI

Nakahara M, Okumura N, Nakano S, et al. Effect of a p38 mitogen-activated protein kinase inhibitor on corneal endothelial cell proliferation. Investig Ophthalmol Vis Sci. 2018;59(10):4218–4227. doi: 10.1167/iovs.18-24394. PubMed DOI

Okumura N, Kay EP, Nakahara M, et al. Inhibition of TGF-beta signaling enables human corneal endothelial cell expansion in vitro for use in regenerative medicine. PLoS ONE. 2013;8(2):e58000. PubMed PMC

Sabater AL, Andreu EJ, García-Guzmán M, et al. Combined PI3K/Akt and Smad2 activation promotes corneal endothelial cell proliferation. Investig Ophthalmol Vis Sci. 2017;58(2). PubMed

Okumura N, Nakano S, Kay EP, et al. Involvement of Cyclin D and p27 in cell proliferation mediated by ROCK inhibitors Y-27632 and Y-39983 during corneal endothelium wound healing. Investig Ophthalmol Vis Sci. 2014;55(1). PubMed

Pipparelli A, Arsenijevic Y, Thuret G, et al. ROCK inhibitor enhances adhesion and wound healing of human corneal endothelial cells. PLoS ONE. 2013;8(4). PubMed PMC

Sturdivant JM, Royalty SM, Lin CW, et al. Discovery of the ROCK inhibitor netarsudil for the treatment of open-angle glaucoma. Bioorg Med Chem Lett. 2016;26(10):2475–2480. doi: 10.1016/j.bmcl.2016.03.104. PubMed DOI

Schlötzer-Schrehardt U, Zenkel M, Strunz M, et al. Potential functional restoration of corneal endothelial cells in Fuchs endothelial corneal dystrophy by ROCK inhibitor (Ripasudil) Am J Ophthalmol. 2021;224:185–199. doi: 10.1016/j.ajo.2020.12.006. PubMed DOI

Sun P, Shen L, Zhang CW, et al. Promoting the expansion and function of human corneal endothelial cells with an orbital adipose-derived stem cell-conditioned medium. Stem Cell Res Ther. 2017;8. PubMed PMC

Liu J, Wen Y, Luo W, et al. Human amniotic epithelial cells promote the proliferation of human corneal endothelial cells by regulating telomerase activity via the Wnt/beta-catenin pathway. Curr Eye Res. 2020:1–9. PubMed

Nakahara M, Okumura N, Kay EP, et al. corneal endothelial expansion promoted by human bone marrow mesenchymal stem cell-derived conditioned medium. PLoS ONE. 2013;8(7). PubMed PMC

Van den Bogerd B, Zakaria N, Matthyssen S, et al. Exploring the mesenchymal stem cell secretome for corneal endothelial proliferation. Stem Cells Int. 2020;2020. PubMed PMC

Kremer I, Rapuano CJ, Cohen EJ, et al. Retrocorneal fibrous membranes in failed corneal grafts. Am J Ophthalmol. 1993;115(4):478–483. doi: 10.1016/S0002-9394(14)74450-2. PubMed DOI

Katikireddy KR, White TL, Miyajima T, et al. NQO1 downregulation potentiates menadione-induced endothelial-mesenchymal transition during rosette formation in Fuchs endothelial corneal dystrophy. Free Radic Biol Med. 2018;116:19–30. doi: 10.1016/j.freeradbiomed.2017.12.036. PubMed DOI PMC

Kim Y, You HJ, Park SH, et al. A mutation in ZNF143 as a novel candidate gene for endothelial corneal dystrophy. J Clin Med. 2019;8(8). PubMed PMC

Wheelock MJ, Shintani Y, Maeda M, et al. Cadherin switching. J Cell Sci. 2008;121(6):727–735. doi: 10.1242/jcs.000455. PubMed DOI

Maurizi E, Schiroli D, Zini R, et al. A fine-tuned β-catenin regulation during proliferation of corneal endothelial cells revealed using proteomics analysis. Sci Rep. 2020;10(1). PubMed PMC

Li C, Dong F, Jia YN, et al. Notch signal regulates corneal endothelial-to-mesenchymal transition. Am J Pathol. 2013;183(3):786–795. doi: 10.1016/j.ajpath.2013.05.025. PubMed DOI

Zhang ZH, Miao YY, Ke BL, et al. LY2109761, transforming growth factor beta receptor type I and type II dual inhibitor, is a novel approach to suppress endothelial mesenchymal transformation in human corneal endothelial cells. Cell Physiol Biochem. 2018;50(3):963–972. doi: 10.1159/000494480. PubMed DOI

Li ZY, Duan HY, Li WJ, et al. Nicotinamide inhibits corneal endothelial mesenchymal transition and accelerates wound healing. Exp Eye Res. 2019;184:227–233. doi: 10.1016/j.exer.2019.04.012. PubMed DOI

Ho WT, Chang JS, Su CC, et al. Inhibition of matrix metalloproteinase activity reverses corneal endothelial-mesenchymal transition. Am J Pathol. 2015;185(8):2158–2167. doi: 10.1016/j.ajpath.2015.04.005. PubMed DOI

Su CC, Ho WT, Peng FT, et al. Exploring a peptidomimetic approach of N-cadherin in modulating fibroblast growth factor receptor signaling for corneal endothelial regeneration. FASEB J. 2020. PubMed

Frausto RF, Chung DD, Boere PM, et al. ZEB1 insufficiency causes corneal endothelial cell state transition and altered cellular processing. PLoS ONE. 2019;14(6):e0218279. PubMed PMC

Jirsova K, Merjava S, Martincova R, et al. Immunohistochemical characterization of cytokeratins in the abnormal corneal endothelium of posterior polymorphous corneal dystrophy patients. Exp Eye Res. 2007;84(4):680–686. doi: 10.1016/j.exer.2006.12.006. PubMed DOI

Zhang Y, Liu X, Liang W, et al. Expression and function of ZEB1 in the cornea. Cells. 2021;10(4). PubMed PMC

Bartakova A, Alvarez-Delfin K, Weisman AD, et al. Novel identity and functional markers for human corneal endothelial cells. Investig Ophthalmol Vis Sci. 2016;57(6):2749–2762. doi: 10.1167/iovs.15-18826. PubMed DOI PMC

Chng ZZ, Peh GSL, Herath WB, et al. High throughput gene expression analysis identifies reliable expression markers of human corneal endothelial cells. PLoS One. 2013;8(7). PubMed PMC

Canals M, Costa-Vila J, Potau JM, et al. Morphological study of cryopreserved human corneal endothelium. Cells Tissues Organs. 1999;164(1):37–45. doi: 10.1159/000016641. PubMed DOI

Eskandari N, Marquez-Curtis LA, McGann LE, et al. Cryopreservation of human umbilical vein and porcine corneal endothelial cell monolayers. Cryobiology. 2018;85:63–72. doi: 10.1016/j.cryobiol.2018.10.001. PubMed DOI

Okumura N, Kagami T, Watanabe K, et al. Feasibility of a cryopreservation of cultured human corneal endothelial cells. PLoS ONE. 2019;14(6). PubMed PMC

Xia X, Atkins M, Dalal R, et al. Magnetic human corneal endothelial cell transplant: delivery, retention, and short-term efficacy. Investig Ophthalmol Vis Sci. 2019;60(7):2438–2448. doi: 10.1167/iovs.18-26001. PubMed DOI PMC

Patel SV, Bachman LA, Hann CR, et al. human corneal endothelial cell transplantation in a human ex vivo model. Investig Ophthalmol Vis Sci. 2009;50(5):2123–2131. doi: 10.1167/iovs.08-2653. PubMed DOI PMC

Okumura N, Sakamoto Y, Fujii K, et al. Rho kinase inhibitor enables cell-based therapy for corneal endothelial dysfunction. Sci Rep. 2016;6. PubMed PMC

Mimura T, Shimomura N, Usui T, et al. Magnetic attraction of iron-endocytosed corneal endothelial cells to Descemet's membrane. Exp Eye Res. 2003;76(6):745–751. doi: 10.1016/S0014-4835(03)00057-5. PubMed DOI

Parikumar P, Haraguchi K, Senthilkumar R, et al. Human corneal endothelial cell transplantation using nanocomposite gel sheet in bullous keratopathy. Am J Stem Cells. 2018;7(1):18–24. PubMed PMC

Choi JS, Williams JK, Greven M, et al. Bioengineering endothelialized neo-corneas using donor-derived corneal endothelial cells and decellularized corneal stroma. Biomaterials. 2010;31(26):6738–6745. doi: 10.1016/j.biomaterials.2010.05.020. PubMed DOI

Spinozzi D, Miron A, Lie JT, et al. In vitro evaluation and transplantation of human corneal endothelial cells cultured on biocompatible carriers. Cell Transplant. 2020;29:963689720923577. doi: 10.1177/0963689720923577. PubMed DOI PMC

Van den Bogerd B, Ni Dhubhghaill S, Zakaria N. Characterizing human decellularized crystalline lens capsules as a scaffold for corneal endothelial tissue engineering. J Tissue Eng Regen Med. 2018;12(4):E2020–E2028. doi: 10.1002/term.2633. PubMed DOI PMC

Yamaguchi M, Ebihara N, Shima N, et al. Adhesion, migration, and proliferation of cultured human corneal endothelial cells by Laminin-5. Invest Ophthalmol Vis Sci. 2011;52(2):679–684. doi: 10.1167/iovs.10-5555. PubMed DOI

Okumura N, Kakutani K, Numata R, et al. Laminin-511 and-521 enable efficient in vitro expansion of human corneal endothelial cells. Investig Ophthalmol Vis Sci. 2015;56(5):2933–2942. doi: 10.1167/iovs.14-15163. PubMed DOI

Palchesko RN, Funderburgh JL, Feinberg AW. Engineered basement membranes for regenerating the corneal endothelium. Adv Healthc Mater. 2016;5(22):2942–2950. PubMed PMC

Kruse M, Walter P, Bauer B, et al. Electro-spun membranes as scaffolds for human corneal endothelial cells. Curr Eye Res. 2018;43(1):1–11. doi: 10.1080/02713683.2017.1377258. PubMed DOI

Kennedy S, Lace R, Carserides C, et al. Poly-epsilon-lysine based hydrogels as synthetic substrates for the expansion of corneal endothelial cells for transplantation. J Mater Sci Mater Med. 2019;30(9):102. doi: 10.1007/s10856-019-6303-1. PubMed DOI PMC

Van Hoorick J, Delaey J, Vercammen H, et al. Designer descemet membranes containing PDLLA and functionalized gelatins as corneal endothelial scaffold. Adv Healthc Mater. 2020. PubMed

Salehi S, Czugala M, Stafiej P, et al. Poly (glycerol sebacate)-poly (ε-caprolactone) blend nanofibrous scaffold as intrinsic bio- and immunocompatible system for corneal repair. Acta Biomater. 2017;50:370–380. doi: 10.1016/j.actbio.2017.01.013. PubMed DOI

Peh GSL, Adnan K, George BL, et al. The effects of Rho-associated kinase inhibitor Y-27632 on primary human corneal endothelial cells propagated using a dual media approach. Sci Rep. 2015;5. PubMed PMC

Okumura N, Hirano H, Numata R, et al. Cell surface markers of functional phenotypic corneal endothelial cells. Investig Ophthalmol Vis Sci. 2014;55(11):7610–7618. doi: 10.1167/iovs.14-14980. PubMed DOI

Ding V, Chin A, Peh G, et al. Generation of novel monoclonal antibodies for the enrichment and characterization of human corneal endothelial cells (hCENC) necessary for the treatment of corneal endothelial blindness. MAbs. 2014;6(6):1439–1452. doi: 10.4161/mabs.36249. PubMed DOI PMC

Hamuro J, Ueno M, Toda M, et al. Cultured human corneal endothelial cell aneuploidy dependence on the presence of heterogeneous subpopulations with distinct differentiation phenotypes. Investig Ophthalmol Vis Sci. 2016;57(10):4385–4392. doi: 10.1167/iovs.16-19771. PubMed DOI