Scientific Online Resource System

Bulgarian Review of Ophthalmology

Limbal stem cell deficiency - concept and treatment

R. Hristova, I. Tanev


Limbal stem cell deficiency is implicated in many diseases and can be a reason for failure of different ocular surface reconstruction interventions, including penetrating keratoplasty. Classic clinical presentation includes the triad conjunctivalization, neovascularization, and chronic inflammation demonstrated by clinical and instrumental methods like impression cytology, confocal microscopy, and other imaging techniques. Management of limbal epithelial stem cell deficiency requires a complex approach and providing optimal conditions for limbal stem cell survival by correcting all coexistent pathophysiologic aberrations. A new direction in operative treatment is ex vivo expansion of autologous limbal epithelial stem cells obtained by minimally invasive limbal biopsy. With growing practical significance of the disease new innovative methods for ocular surface reconstruction continue to arise, aiming at the best visual outcome.


ocular surface; limbus; stem cells; transplantation

Full Text


Alio JL, Rodriguez AE, WróbelDudzinska D. Eye platelet-rich plasma in the treatment of ocular surface disorders. Curr Opin Ophthalmol. 2015, 26, 325-332.

Azar DT. Corneal angiogenic privilege: angiogenic and antiangiogenic factors in corneal avascularity, vasculogenesis, and wound healing (an American Ophthalmological Society thesis). Trans Am Ophthalmol Soc. 2006, 104, 264-302.

Baluk P, McDonald DM. Markers for microscopic imaging of lymphangiogenesis and angiogenesis. Ann N Y Acad Sci. 2008, 1131, 1-12.

Baudouin C, Brignole F, Becquet F, et al. Flow cytometry in impression cytology specimens. A new method for evaluation of conjunctival inflammation. Investig Ophthalmol Vis Sci. 1997, 38, 1458-1464.

Beyazyildiz E, Pinarli FA, Beyazyildiz Ö, et al. Efficacy of Topical Mesenchymal Stem Cell Therapy in the Treatment of Experimental Dry Eye Syndrome Model. Stem Cells International, 2014, 230-250.

Calonge M, Diebold Y, Saez V, et al. Impression cytology of the ocular surface: a review. Exp Eye Res. 2004, 78, 457-472.

Chen S-Y, Hayashida Y, Chen M-Y, et al. A New Isolation Method of Human Limbal Progenitor Cells by Maintaining Close Association with Their Niche Cells. Tissue Engineering Part C, Methods. 2011, 17, 537-548.

Cheung AT, Harmatz P, Wun T, et al. Correlation of abnormal intracranial vessel velocity, measured by transcranial Doppler ultrasonography, with abnormal conjunctival vessel velocity, measured by computer-assisted intravital microscopy, in sickle cell disease. Blood. 2001 97, 3401-3404.

Coster DL, Aggarwal RK, Williams KA. Surgical managementof ocular surface disorders using conjunctival and stem cells allografts. Br J Ophthalmol. 1995, 79, 977-982.

Cursiefen C, Maruyama K, Jackson DG, et al. Time course of angiogenesis and lymphangiogenesis after brief corneal inflammation. Cornea. 2006, 25, 443-447.

Cursiefen C, Reinhard T, Bock F,Prokosch HU. Subconjunctival Cyclosporine A implants do not affect corneal neovascularisation after transplantation: results of a randomized clinical trial. Invest. Ophthalmol. Vis. Sci. 2013, 54, 2090.

Dart J. Impression cytology of the ocular surface- research tool or routine clinical investigation? Br J Ophthalmol. 1997, 81, 930.

De Luca M, Pellegrini G, Green H. Regeneration of squamous epithelia from stem cells of cultured grafts. Regen Med 2006, 1, 45-57.

Del Buey MA, Casas, P, Caramello CM, et al. Analysis of the efficacy of the tissue regenerating agent (RGTA) 0.01% poly-carboxymethylglucose sulfate in the treatment of neurotrophic corneal ulcers and persistent epithelial defects. Acta Ophthalmologica, 2015, 93, S255.

Di Iorio E, Barbaro V, Ruzza A, et al. Isoforms of DeltaNp63 and the migration of ocular limbal cells in human corneal regeneration. Proc Natl Acad Sci U S A. 2005, 102, 9523-9528.

Di Iorio E, Ferrari S, Fasolo A, et al. Techniques for culture and assessment of limbal stem cell grafts. Ocul Surf. 2010, 8, 146-153.

Djalilian 􀈺R, Mahesh SP, Koch CA, et al. Survival of Donor Epithelial Cells after Limbal Stem Cell Transplantation. Invest Ophthalmol Vis Sci. 2005, 46, 803-807.

Dua HS, Saini JS, Azuara-Blanco A, Gupta P. Limbal stem cell deficiency: concept, aetiology, clinical presentation, diagnosis and management. Indian J. Ophthalmol. 2000, 48, 83-92.

Dua HS, Gomes JAP, Jindal V, et al. Mucosa specific lymphocytes in the human conjunctiva, corneoscleral limbus and lacrimal gland. Curr Eye Res,1994, 13, 87-93.

Dua HS, Gomes JAP, Singh A. Corneal epithelial wound healing. Br J Ophthalmol 1994, 78, 401-408.

Dua HS, Joseph A,Shanmuganathan VA, Jones RE. Stem cell differentiation and effects of deficiency. Eye. 2003, 17, 877-885.

Dua HS, Shanmuganathan VA, Powell- Richards AO, et al. Limbal epithelial crypts - A novel anatomical structure and a putative limbal stem cell niche. Br J Ophthalmol. 2005, 89, 529- 532.

Dua HS. Sequential sectoral conjunctival epitheliectomy. In: Ocular surface disease medical and surgical management, eds. Holland EJ, Mannis MJ, Berlin, Heidelberg, New York, Tokyo, Springer, 2002, 168-174.

Dua HS. The conjunctiva in corneal epithelial wound healing. Br J Ophthalmol 1998, 82, 1407-1411.

Ebrahem Q, Minamoto A, Hoppe G, et al. Triamcinolone acetonide inhibits IL-6- and VEGF induced angiogenesis downstream of the IL-6 and VEGF receptors. Invest Ophthalmol Vis Sci. 2006, 47, 4935 4941.

Ellenberg D, Azar DT, Hallak JA, et al. Novel aspects of corneal angiogenic and lymphangiogenic privilege. Prog Retin Eye Res. 2010, 29, 208-248.

Espana EM, Di PM, Grueterich M et al. Keratolimbal allograft in corneal reconstruction. Eye. 2004, 18, 406-417.

Freire V, Andollo N, Etxebarria J, et al. Corneal wound healing promoted by 3 blood derivatives: an in vitro and in vivo comparative study. Cornea. 2014, 33, 614-620.

Gao J, Morgan G, Tieu D, et al. ICAM-1 expression predisposes ocular tissues to immunebased inflammation in dry eye patients and Sjogrens syndrome-like MRL/lpr mice. Exp Eye Res. 2004, 78, 823-835.

García I, Etxebarria J, Merayo-Lloves J, et al. Novel Molecular Diagnostic System of Limbal Stem Cell Deficiency Based on MUC5AC Transcript Detection in Corneal Epithelium by PCR-Reverse Dot Blot. Invest Ophthalmol Vis Sci. 2013, 54, 5643-5652.

Ghoubay-Benallaoua D, Sandali O, Goldschmidt P, Borderie V. Kinetics of expansion of human limbal epithelial progenitor cells in primary culture of explants without feeders. PloS One. 2013, 8, e81965.

Giebel SJ, Menicucci G, McGuire PG, Das A. Matrix metalloproteinases in early diabetic retinopathy and their role in alteration of the blood-retinal barrier. Lab Invest. 2005, 85, 597-607.

Gong Y, Koh DR. Neutrophils promote inflammatory angiogenesis via release of preformed VEGF in an in vivo corneal model. Cell Tissue Res. 2010, 339, 437-448.

Grus FH, Podust VN, Bruns K, et al. SELDI-TOF-MS Protein- Chip array profiling of tears from patients with dry eye. Investig Ophthalmol Vis Sci. 2005, 46, 863-876.

Henderson HW, Collin JR. Mucous membrane grafting. Dev Ophthalmol. 2008, 41, 230- 242.

Hirayama M, Tsubota K, Tsuji T. Bioengineered Lacrimal Gland Organ Regeneration in Vivo. J Funct Biomater. 2015, 6, 634-649.

Huang AJ, Tseng SC, Kenyon KR. Alteration of epithelial paracellular permeability during corneal epithelial wound healing. Invest Ophthalmol Vis Sci. 1990, 31, 429-435.

Huang AJW, Tseng SCG. Corneal epithelial wound healing in the absence of limbal epithelium. ARVO Abstracts. Invest Ophthalmol Vis Sci. 1988, 29, 190.

Ilari L, Daya SM. Long-term outcomes of keratolimbal allograft for the treatment of severe ocular surface disorders. Ophthalmology. 2002, 109, 1278-1284.

ackson DG, Prevo R, Clasper S, et al. LYVE-1, the lymphatic system and tumor lymphangiogenesis. Trends Immunol. 2001, 22, 317-321.

Jackson DG. Immunological functions of hyaluronan and its receptors in the lymphatics. Immunol Rev. 2009, 230, 216-231.

Jenkins C, Tuft S, Liu C, et al. Limbal transplantation in the management of chronic contact-lens-associated epitheliopathy. Eye. 1993, 7, 629-633.

Jiang H, Ye Y, DeBuc DC, et al. Human conjunctival microvasculature assessed with a retinal function imager (RFI). Microvasc Res. 2013, 85, 134-137.

Jirsova K, Dudakova L, Kalasova S, et al. The OV-TL 12/30 clone of anti-cytokeratin 7 antibody as a new marker of corneal conjunctivalization in patients with limbal stem cell deficiency. Invest Ophthalmol Vis Sci. 2011, 52, 5892-5898.

Joseph A, Hossain P, Jham S, et al. Expression of CD34 and L-selectin on human corneal keratocytes. Invest Ophthalmol Vis Sci. 2003, 44, 4689-4692.

Kenyon KR, Tseng SC. Limbal autograft transplantation for ocular surface disorders. Ophthalmology. 1989, 96, 709-722.

Kwitko S, Marinho D, Barcaro S, et al. Allograft conjunctival transplantation for bilateral ocular surface disorders. Ophthalmology. 1995, 102, 1020-1025.

Lambiase S, Manni L, Bonini S, et al. Nerve growth factor promotes corneal healing: structural, biochemical, and molecular analyses of rat and human corneas. Invest Ophthalmol Vis Sci. 2000, 41, 1063-1069.

Mackie IA. Neuroparalytic keratitis. In: Current ocular therapy 4, eds. Fraunfelder FT, Roy FH, Philadelphia, W.B. Saunders, 1995, 506-508.

Massingale ML, Li X, Vallabhajosyula M, et al. Analysis of inflammatory cytokines in the tears of dry eye patients. Cornea. 2009, 28, 1023-1027.

Meyer PA, Watson PG. Low dose fluorescein angiography of the conjunctiva and episclera. Br J Ophthalmol. 1987, 71, 2-10.

Nakamura T, Inatomi T, Sotozono C et al. Transplantation of autologous serum-derived cultivated corneal epithelial equivalents for the treatment of severe ocular surface disease. Ophthalmology. 2006, 113, 1765-1772.

Nubile M, Lanzini M, Miri A, Pocobelli A, Calienno R, Curcio C, Mastropasqua R, Dua HS, Mastropasqua L. In vivo confocal microscopy in diagnosis of limbal stem cell deficiency. Am J Ophthalmol. 2013, 155, 220-232.

Paolin A, Cogliati E, Trojan D, et al. Amniotic membranes in ophthalmology: long term data on transplantation outcomes. Cell Tissue Bank. 2015, 11, 1-8.

Pellegrini G, Traverso CE, Franzi AT, et al. Long-term restoration of damaged corneal surfaces with autologous cultivated corneal epithelium. Lancet. 1997, 349, 990-993.

Poli M, Burillon C, Auxenfans C, et al. Immunocytochemical Diagnosis of Limbal Stem Cell Deficiency: Comparative Analysis of Current Corneal and Conjunctival Biomarkers. Cornea. 2015, 34, 817-823.

Puangsricharern V, Tseng SC. Cytologic evidence of corneal diseases with limbal stem cell deficiency. Ophthalmology. 1995, 102, 1476-1485.

Rama P, Bonini S, Lambiase A et al. Autologous fibrin cultured limbal stem cells permanently restore the corneal surface of patients with total limbal stem cell deficiency. Transplantation. 2001, 72, 1478-1485.

Ramos T, Scott D, Ahmad S. An Update on Ocular Surface Epithelial Stem Cells: Cornea and Conjunctiva. Stem Cells International. 2015.

Rao SK, Rajagopal R, Sitalakshmi G, et al. Limbal allografting from related live donors for corneal surface reconstruction. Ophthalmology. 1999, 106, 822-828.

Romano MR, Biagioni F, Carrizzo A, et al. Effects of vitamin B12 on the corneal nerve regeneration in rats. Exp Eye Res. 2014, 120, 109-117.

Rosenthal P, Cotter J. The Boston Scleral Lens in the management of severe ocular surface disease. Ophthalmol Clin North Am. 2003, 16, 89-93.

Sambursky R, Davitt WF, Latkany R, et al. Sensitivity and specificity of a point-of-care matrix metalloproteinase 9 immunoassay for diagnosing inflammation related to dry eye. JAMA Ophthalmol. 2013, 131, 24-28.

Sangwan VS, Vemuganti GK, Iftekhar G, et al. Use of autologous cultured limbal and conjunctival epithelium in a patient with severe bilateral ocular surface disease induced by acid injury: a case report of unique application. Cornea. 2003, 22, 478-481.

Santos MS, Gomes JA, Hofling-Lima AL, et al. Survival analysis of conjunctival limbal graft s and amniotic membrane transplantation in eyes with total limbal stem cell deficiency. Am J Ophthalmol. 2005, 140, 223-230.

Schoenbeck U, Mach F, Libby P. Generation of biologically active IL-1 beta by matrix metalloproteinases: a novel caspase-1-independent pathway of IL-1 beta processing. J Immunol. 1998, 161, 3340-3346.

Tatlipinar S, Akpek EK. Topical ciclosporin in the treatment of ocular surface disorders. Br J Ophthalmol. 2005, 89, 1363-1367.

Tseng SCG, Chen SY, Shen YC, et al. Critical appraisal of ex vivo expansion of human limbal epithelial stem cells. Current Molecular Medicine. 2010, 10, 841-850.

Tseng SC, Prabhasawat P, Barton K, et al. Amniotic membrane transplantation with or without limbal allograft s for corneal surface reconstruction in patients with limbal stem cell deficiency. Arch Ophthalmol. 1998, 116, 431-441.

Tseng SCG, Tsubota K. Important concepts for treating ocular surface and tear disorders. Am J Opthalmol 1997, 124, 825-835.

Tsubota K, Fujihara T, Saito K, Takeuchi T. Conjunctival epithelium expression of HLADR in dry eye patients. Ophthalmologica. 1999, 213, 16-19.

Tsubota K, Satake Y, Ohyama M et al. Surgical reconstruction of the ocular surface in advanced ocular cicatricial pemphigoid and Stevens-Johnson syndrome. Am J Ophthalmol. 1996, 122, 38-52.

Tsubota K, Toda I, Saito H, et al. Reconstruction of the corneal epithelium by limbal allograft transplantation for severe ocular surface disorders. Ophthalmology. 1996, 102, 1486- 1496.

Utheim O, Islam R, Lyberg T et al. Serumfree and xenobiotic-free preservation of cultured human limbal epithelial cells. PLoS One. 2015, 10, e0118517.

Watanabe K, Nishida K, Yamato M, et al. Human limbal epithelium contains side population cells expressing the ATP-binding cassette transporter ABCG2. FEBS Lett. 2004, 565, 6-10.

Williams KA, Brereton HM, Aggarwal R, et al. Use of DNA polymorphisms and the polymerase chain reaction to examine the survival of a human limbal stem cell allograft. Am J Ophthalmol 1995, 120, 342 350.

Yao L, Bai H. Review: mesenchymal stem cells and corneal reconstruction. Molecular Vision. 2013, 19, 2237-2243.

Yoon KC, Jeong IY, Park YG, Yang SY. Interleukin-6 and tumor necrosis factor-alpha levels in tears of patients with dry eye syndrome. Cornea. 2007, 26, 431-437.

Zhang J, Cao R, Zhang Y, et al. Differential roles of PDGFR-alpha and PDGFR-beta in angiogenesis and vessel stability. FASEB J. 2009, 23, 153-163.

Zhao X, Das AV, Thoreson WB, et al. Adult corneal limbal epithelium: a model for studying neural potential of non-neural stem cells/progenitors. Dev Biol. 2002, 250, 317-331.



Font Size

Article Tools
Email this article (Login required)
About The Authors

R. Hristova
Medical University - Sofia, University Hospital Alexandrovska, Department of Ophthalmology

I. Tanev
Medical University - Sofia, University Hospital Alexandrovska, Department of Ophthalmology