Limbal stem cells (LSC), which reside in the basal layer of the limbus, are thought to be responsible for corneal epithelial healing after injury. When the cornea is damaged, LSC start to proliferate, differentiate, and migrate to the site of injury. To characterize the signaling molecules ensuring communication between the cornea and LSC, we established a mouse model of mechanical corneal damage. The central cornea or limbal tissue was excised at different time intervals after injury, and the expression of genes in the explants was determined. It was observed that a number of genes for growth and differentiation factors were significantly upregulated in the cornea rapidly after injury. The ability of these factors to regulate the differentiation and proliferation of limbal cells was tested. It was found that the insulin-like growth factor-I (IGF-I), which is rapidly overexpressed after injury, enhances the expression of IGF receptor in limbal cells and induces the differentiation of LSC into cells expressing the corneal cell marker, cytokeratin K12, without any effect on limbal cell proliferation. In contrast, the epidermal growth factor (EGF) and fibroblast growth factor-β (FGF-β), which are also produced by the damaged corneal epithelium, supported limbal cell proliferation without any effect on their differentiation. Other factors did not affect limbal cell differentiation or proliferation. Thus, IGF-I was identified as the main factor stimulating the expression of IGF receptors in limbal cells and inducing the differentiation of LSC into cells expressing corneal epithelial cell markers. The proliferation of these cells was supported by EGF and FGF.

Institute of Molecular Genetics Academy of Sciences of the Czech Republic Prague Czech Republic

Zobrazit více v PubMed

Majo F. Rochat A. Nicolas M. Jaoude GA. Barrandon Y. Oligopotent stem cells are distributed throughout the mammalian ocular surface. Nature. 2008;456:250–254. PubMed

Chang C-Y. Green CR. McGhee CNJ. Sherwin T. Acute wound healing in the human central corneal epithelium appears to be independent of limbal stem cell influence. Invest Ophthalmol Vis Sci. 2008;2008;49:5279–5286. PubMed

Dua HS. Miri A. Alomar T. Yeung AM. Said DG. The role of limbal stem cells in corneal epithelial maintenance: testing the dogma. Ophthalmology. 2009;116:856–863. PubMed

Tseng SC. Regulation and clinical implication of corneal epithelial stem cells. Mol Biol Rep. 1996;23:47–58. PubMed

Gomes JA. dos Santos MS. Cunha MC. Mascaro VL. Barros JN. de Sousa LB. Amniotic membrane transplantation for partial and total limbal stem cell deficiency secondary to chemical burn. Ophthalmology. 2003;110:466–473. PubMed

Du Y. Chen J. Funderburgh JL. Zhu X. Li L. Functional reconstruction of rabbit corneal epithelium by human limbal cells cultured on amniotic membrane. Mol Vis. 2003;9:635–643. PubMed PMC

Rama P. Matuska S. Paganoni G. Spinelli A. De Luca M. Pellegrini G. Limbal stem-cell therapy and long-term corneal regeneration. N Engl J Med. 2010;363:147–155. PubMed

Lehrer MS. Sun TT. Lavker RM. Strategies of epithelial repair: modulation of stem cell and transit amplifying cell proliferation. J Cell Sci. 1998;111:2867–2875. PubMed

Jia C. Zhu W. Ren S. Xi H. Li S. Wang Y. Comparison of genome-wide gene expression in suture- and alkali burn-induced murine corneal neovascularization. Mol Vis. 2011;17:2386–2399. PubMed PMC

Cheng CC. Wang DY. Kao MH. Chen JK. The growth-promoting effect of KGF on limbal epithelial cells is mediated by upregulation of DeltaNp63alpha through the p38 pathway. J Cell Sci. 2009;122:4473–4480. PubMed

Li DQ. Tseng SC. Three patterns of cytokine expression potentially involved in epithelial-fibroblast interactions of human ocular surface. J Cell Physiol. 1995;163:64–79. PubMed

Wilson SE. He YG. Weng J. Zieske JD. Jester JV. Schultz GS. Effect of epidermal growth factor, hepatocyte growth factor, and keratinocyte growth factor, on proliferation, motility and differentiation of human corneal epithelial cells. Exp Eye Res. 1994;59:665–678. PubMed

Jester JV. Ho-Chang J. Modulation of cultured corneal keratocyte phenotype by growth factors/cytokines control in vitro contractility and extracellular matrix contraction. Exp Eye Res. 2003;77:581–592. PubMed

Ko JA. Yanai R. Nishida T. IGF-1 released by corneal epithelial cells induces up-regulation of N-cadherin in corneal fibroblasts. J Cell Physiol. 2009;221:254–261. PubMed

Hassell JR. Birk DE. The molecular basis of corneal transparency. Exp Eye Res. 2010;91:326–335. PubMed PMC

Etheredge L. Kane BP. Hassell JR. The effect of growth factor signaling on keratocytes in vitro and its relationship to the phases of stromal wound repair. Invest Ophthalmol Vis Sci. 2009;50:3128–3136. PubMed

Pollak M. Insulin and insulin-like growth factor signalling in neoplasia. Nat Rev Cancer. 2009;8:915–928. PubMed

Liu CY. Zhu G. Converse R. Kao CW. Nakanuta H. Tseng SC. Mui MM. Seyer J. Justice MJ. Stech ME, et al. Characterization and chromosomal localization of the cornea-specific murine keratin gene Krt1.12. J Biol Chem. 1994;269:24627–24636. PubMed

Matic M. Petrov IN. Chen S. Wang C. Dimitrijevich SD. Wolosin JM. Stem cells of the corneal epithelium lack connexins and metabolite transfer capacity. Differentiation. 1997;61:251–260. PubMed

Blazejewska EA. Schlötzer-Schrehardt U. Zenkel M. Bachmann B. Chankiewitz E. Jacobi C. Kruse FE. Corneal limbal microenvironment can induce transdifferentiation of hair follicle stem cells into corneal epithelial-like cells. Stem Cells. 2009;27:642–652. PubMed PMC

Gu S. Xing C. Han J. Tso MO. Hong J. Differentiation of rabbit bone marrow mesenchymal stem cells into corneal epithelial cells in vivo and ex vivo. Mol Vis. 2009;15:99–107. PubMed PMC

Homma R. Yoshikawa H. Takeno M. Kurokawa MS. Masuda C. Takada E. Tsubota K. Ueno S. Suzuki N. Induction of epithelial progenitors in vitro from mouse embryonic stem cells and application for reconstruction of damaged cornea in mice. Invest Ophthalmol Vis Sci. 2004;45:4320–4326. PubMed

Jiang TS. Cai L. Ji WY. Hui YN. Wang YS. Hu D. Zhu J. Reconstruction of the corneal epithelium with induced marrow mesenchymal stem cells in rats. Mol Vis. 2010;16:1304–1316. PubMed PMC

Krulova M. Pokorna K. Lencova A. Zajicova A. Fric J. Filipec M. Forrester JV. Holan V. A rapid separation of two distinct populations of corneal epithelial cells with limbal stem cell characteristics in the mouse. Invest Ophthalmol Vis Sci. 2008;49:3903–3908. PubMed

Holan V. Pokorna K. Prochazkova J. Krulova M. Zajicova A. Immunoregulatory properties of mouse limbal stem cells. J Immunol. 2010;184:2124–2129. PubMed

Svobodova E. Krulova M. Zajicova A. Prochazkova J. Trosan P. Holan V. The role of mouse mesenchymal stem cells in differentiation of naive T cells into anti-inflammatory regulatory T cell and proinflammatory helper T-cell 17 population. Stem Cells Dev. 2012;21:901–910. PubMed PMC

Schermer A. Galvin S. Sun TT. Differentiation-related expression of a major 64K corneal keratin in vivo and in culture suggests limbal location of corneal epithelial stem cells. J Cell Biol. 1986;103:49–62. PubMed PMC

Li W. Hayashida Y. Chen Y-T. Tseng SCG. Niche regulation of corneal epithelial stem cells at the limbus. Cell Res. 2007;17:26–36. PubMed PMC

Ahmad S. Stewart R. Yung S. Kolli S. Armstrong L. Stojkovic M. Figueiredo F. Lako M. Differentiation of human embryonic stem cells into corneal epithelial-like cells by in vitro replication of the corneal epithelial stem cell niche. Stem Cells. 2007;5:1145–1155. PubMed

Izumi K. Kurosaka D. Iwata T. Oguchi Y. Tanaka Y. Mashima Y. Tsubota K. Involvement of insulin-like growth factor-I and insulin-like growth factor binding protein-3 in corneal fibroblasts during corneal wound healing. Invest Ophthalmol Vis Sci. 2006;47:591–598. PubMed

Maiter D. Underwood LE. Maes M. Ketelslegers JM. Acute downregulation of the somatogenic receptors in rat liver by a single injection of growth hormone. Endocrinology. 1988;122:1291–1296. PubMed

Smith TJ. Insuline-like growth factor-I regulation of immune function: a potential therapeutic target in autoimmune diseases? Pharmacol Rev. 2010;62:199–236. PubMed PMC

Nakamura M. Chikama TI. Nishida T. Characterization of insulin-like growth factor-1 receptors in rabbit corneal epithelial cells. Exp Eye Res. 2000;70:199–204. PubMed

Yamada N. Yanai R. Nakamura M. Inui M. Nishida T. Role of the C domain of IGFs in synergistic promotion, with a substance P-derived peptide, of rabbit corneal epithelial wound healing. Invest Ophthalmol Vis Sci. 2004;45:1125–1131. PubMed

Ayatollahi M. Soleimani M. Geramizadeh B. Imanieh MH. Insulin-like growth factor 1 (IGF-I) improves hepatic differentiation of human bone marrow-derived mesenchymal stem cells. Cell Biol Int. 2011;35:1169–1176. PubMed

Matheny RW., Jr. Nindl BC. Loss of IGF-IEa or IGF-IEb impairs myogenic differentiation. Endocrinology. 2011;152:1923–1934. PubMed

Shima A. Pham J. Blanco E. Barton ER. Sweeney HL. Matsuda R. IGF-I and vitamin C promote myogenic differentiation of mouse and human skeletal muscle cells at low temperatures. Exp Cell Res. 2011;317:356–366. PubMed

Kang HM. Park S. Kim H. Insulin-like growth factor 2 enhances insulinogenic differentiation of human eyelid adipose stem cells via the insulin receptor. Cell Prolif. 2011;44:254–263. PubMed PMC

D'Amario D. Cabral-Da-Silva MC. Zheng H. Fiorini C. Goichberg P. Steadman E. Ferreira-Martins J. Sanada F. Piccoli M, et al. Insulin-like growth factor-1 receptor identifies a pool of human cardiac stem cells with superior therapeutic potential for myocardial regeneration. Circ Res. 2011;108:1467–1481. PubMed PMC

Ellison GM. Torella D. Dellegrottaglie S. Perez-Martinez C. Perez de Prado A. Vicinanza C. Purushothaman S. Galuppo V. Iaconetti C, et al. Endogenous cardiac stem cell activation by insulin-like growth factor-1/hepatocyte growth factor intracoronary injection fosters survival and regeneration of the infarcted pig heart. J Am Coll Cardiol. 2011;58:977–986. PubMed

Zajicova A. Pokorná K. Lencova A. Krulova M. Svobodova E. Kubinova S. Sykova E. Pradny M. Michalek J, et al. Treatment of ocular surface injuries by limbal and mesenchymal stem cells growing on nanofiber scaffolds. Cell Transplant. 2010;19:1281–1290. PubMed

Interleukin-13 increases the stemness of limbal epithelial stem cells cultures

Mesenchymal Stem Cell-Based Therapy for Retinal Degenerative Diseases: Experimental Models and Clinical Trials

The enzymatic de-epithelialization technique determines denuded amniotic membrane integrity and viability of harvested epithelial cells

Molecular Hydrogen Effectively Heals Alkali-Injured Cornea via Suppression of Oxidative Stress

Application of Concanavalin A during immune responsiveness skin-swelling tests facilitates measurement interpretation in mammalian ecology

The Favorable Effect of Mesenchymal Stem Cell Treatment on the Antioxidant Protective Mechanism in the Corneal Epithelium and Renewal of Corneal Optical Properties Changed after Alkali Burns

A Comparative Study of the Therapeutic Potential of Mesenchymal Stem Cells and Limbal Epithelial Stem Cells for Ocular Surface Reconstruction

Modulation of the early inflammatory microenvironment in the alkali-burned eye by systemically administered interferon-γ-treated mesenchymal stromal cells

Distinct cytokines balance the development of regulatory T cells and interleukin-10-producing regulatory B cells

Mesenchymal stem cells, nanofiber scaffolds and ocular surface reconstruction