Abstract
The cornea is the transparent front part of the eye and comprises three distinct cell layers. One of these cell layers is a self-renewing epithelium long believed to harbor a resident stem cell population. The location and characteristics of corneal epithelial stem cells have now been confirmed by several research groups, and these cells are currently applied therapeutically. The corneal stroma and endothelium are largely quiescent after infancy, and until recently they were not considered to undergo self-renewal or to maintain stem cells. This view was overturned during the last two decades. At present, cell populations with characteristics of adult stem cells are routinely isolated and characterized from the limbal stroma and the corneal endothelium. This chapter describes methods for isolation and culture of limbal stromal cells and corneal endothelial cells.
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References
Secker GA, Daniels JT (2009) Limbal epithelial stem cells of the cornea. StemBook [internet], ed. The Stem Cell Research Community, StemBook, doi:10.3824/stembook.1.48.1
Tseng SC (1989) Concept and application of limbal stem cells. Eye 3:141–157
Kinoshita S, Friend J, Thoft RA (1981) Sex chromatin of donor corneal epithelium in rabbits. Invest Ophthalmol Vis Sci 21:434–441
Figueira EC, Di Girolamo N, Coroneo MT, Wakefield D (2007) The phenotype of limbal epithelial stem cells. Invest Ophthalmol Vis Sci 48:144–156
De Paiva CS, Pflugfelder SC, Li DQ (2006) Cell size correlates with phenotype and proliferative capacity in human corneal epithelial cells. Stem Cells 24:368–375
Akinci MA, Turner H, Taveras M, Barash A, Wang Z, Reinach P et al (2009) Molecular profiling of conjunctival epithelial side-population stem cells: atypical cell surface markers and sources of a slow-cycling phenotype. Invest Ophthalmol Vis Sci 50:4162–4172
Schlotzer-Schrehardt U, Kruse FE (2005) Identification and characterization of limbal stem cells. Exp Eye Res 81:247–264
Itskovitz-Eldor J, Schuldiner M, Karsenti D, Eden A, Yanuka O, Amit M et al (2000) Differentiation of human embryonic stem cells into embryoid bodies compromising the three embryonic germ layers. Mol Med 6:88–95
Schofield R, Lajtha LG (1983) Determination of the probability of self-renewal in haemopoietic stem cells: a puzzle. Blood Cells 9:467–483
Li L, Neaves WB (2006) Normal stem cells and cancer stem cells: the niche matters. Cancer Res 66:4553–4557
Gipson IK (1989) The epithelial basement membrane zone of the limbus. Eye 3: 132–140
Ljubimov AV, Burgeson RE, Butkowski RJ, Michael AF, Sun TT, Kenney MC (1995) Human corneal basement membrane heterogeneity: topographical differences in the expression of type IV collagen and laminin isoforms. Lab Invest 72:461–473
Li DQ, Tseng SC (1995) Three patterns of cytokine expression potentially involved in epithelial-fibroblast interactions of human ocular surface. J Cell Physiol 163:61–79
Dravida S, Pal R, Khanna A, Tipnis SP, Ravindran G, Khan F (2005) The transdifferentiation potential of limbal fibroblast-like cells. Brain Res Dev Brain Res 160:239–251
Du Y, Funderburgh ML, Mann MM, SundarRaj N, Funderburgh JL (2005) Multipotent stem cells in human corneal stroma. Stem Cells 23:1266–1275
Polisetty N, Fatima A, Madhira SL, Sangwan VS, Vemuganti GK (2008) Mesenchymal cells from limbal stroma of human eye. Mol Vis 14:431–442
Criscimanna A, Zito G, Taddeo A, Richiusa P, Pitrone M, Morreale D et al (2011) In vitro generation of pancreatic endocrine cells from human adult fibroblast-like limbal stem cells. Cell Transplant 21:73–90
Nishida T, Nakamura M, Konma T, Ofuji K, Nagano K, Tanaka T et al (1997) Neurotrophic keratopathy–studies on substance P and the clinical significance of corneal sensation. Nippon Ganka Gakkai Zasshi 101:948–974
Bourne WM, Nelson LR, Hodge DO (1997) Central corneal endothelial cell changes over a ten-year period. Invest Ophthalmol Vis Sci 38:779–782
Williams KK, Noe RL, Grossniklaus HE, Drews-Botsch C, Edelhauser HF (1992) Correlation of histologic corneal endothelial cell counts with specular microscopic cell density. Arch Ophthalmol 110:1146–1149
Blake DA, Yu H, Young DL, Caldwell DR (1997) Matrix stimulates the proliferation of human corneal endothelial cells in culture. Invest Ophthalmol Vis Sci 38:1119–1129
Yue BY, Sugar J, Gilboy JE, Elvart JL (1989) Growth of human corneal endothelial cells in culture. Invest Ophthalmol Vis Sci 30:248–253
Miyata K, Drake J, Osakabe Y, Hosokawa Y, Hwang D, Soya K et al (2001) Effect of donor age on morphologic variation of cultured human corneal endothelial cells. Cornea 20:59–63
Amano S (2003) Transplantation of cultured human corneal endothelial cells. Cornea 22:S66–S74
Pistsov MY, Sadovnikova E, Danilov SM (1988) Human corneal endothelial cells: isolation, characterization and long-term cultivation. Exp Eye Res 47:403–414
Zhu C, Joyce NC (2004) Proliferative response of corneal endothelial cells from young and older donors. Invest Ophthalmol Vis Sci 45:1743–1751
Engelmann K, Friedl P (1989) Optimization of culture conditions for human corneal endothelial cells. In Vitro Cell Dev Biol 25:1065–1072
Dawson DG, Kramer TR, Grossniklaus HE, Waring GO, Edelhauser HF (2005) Histologic, ultrastructural, and immunofluorescent evaluation of human laser-assisted in situ keratomileusis corneal wounds. Arch Ophthalmol 123:741–756
Ishino Y, Sano Y, Nakamura T, Connon CJ, Rigby H, Fullwood NJ et al (2004) Amniotic membrane as a carrier for cultivated human corneal endothelial cell transplantation. Invest Ophthalmol Vis Sci 45:800–806
Choi JS, Williams JK, Greven M, Walter KA, Laber PW, Khang G et al (2010) Bioengineering endothelialized neo-corneas using donor-derived corneal endothelial cells and decellularized corneal stroma. Biomaterials 31:6738–6745
Chen KH, Azar D, Joyce NC (2001) Transplant of adult human corneal endothelium ex vivo: a morphologic study. Cornea 20:731–737
Joyce NC, Zhu CC (2004) Human corneal endothelial cell proliferation: potential for use in regenerative medicine. Cornea 23:S8–S19
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Polisetti, N., Joyce, N.C. (2013). The Culture of Limbal Stromal Cells and Corneal Endothelial Cells. In: Wright, B., Connon, C. (eds) Corneal Regenerative Medicine. Methods in Molecular Biology, vol 1014. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-432-6_8
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DOI: https://doi.org/10.1007/978-1-62703-432-6_8
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