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Year : 2015  |  Volume : 3  |  Issue : 2  |  Page : 71-75

Limbal stem cell deficiency: A review

Consultant Eye Surgeon, Cornea and Oculoplasty Services, Riti Eye Care Hospital, Rewari, Haryana, India

Date of Submission11-Jan-2014
Date of Acceptance04-Nov-2014
Date of Web Publication7-May-2015

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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/2320-3897.156582

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Limbal stem cell deficiency is commonly encountered entity in routine practice. Most of the cases go unidentified at the first visit and progress to the severe disease when the problem is actually realized. In limbal stem cell deficiency, corneal limbal stem cells fail to maintain and renew corneal epithelial surface and the corneal surface is encroached by conjunctival epithelium, compromising the transparency and visual acuity. First step in the management of limbal stem cell deficiency is early control of the ocular surface inflammation and elimination of the causative factor. Surgical intervention in terms of limbal stem cell transplantation is the definitive treatment for most of the cases. Successful limbal stem cell transplantation can achieve rapid surface healing, stable ocular surface without recurrent erosions or persistent epithelial defects, regression of corneal vascularization, and restoration of a smooth and optically improved ocular surface, resulting in improved visual acuity and probably increased success for subsequent keratoplasty. Here is a brief review covering etiology, clinical presentation, and management options for the cases of limbal stem cell deficiency.

Keywords: Limbal stem cell deficiency, ocular surface reconstruction, stem cell transplantation

How to cite this article:
Jhagta HS, Jain P. Limbal stem cell deficiency: A review. J Clin Ophthalmol Res 2015;3:71-5

How to cite this URL:
Jhagta HS, Jain P. Limbal stem cell deficiency: A review. J Clin Ophthalmol Res [serial online] 2015 [cited 2023 Mar 23];3:71-5. Available from: https://www.jcor.in/text.asp?2015/3/2/71/156582

Limbal stem cell deficiency (LSCD) is commonly encountered entity in routine practice. Most of the cases go unidentified at the first visit and progress to the severe disease when the problem is actually realized. In LSCD, corneal limbal stem cells fail to maintain and renew corneal epithelial surface and the corneal surface is encroached by conjunctival epithelium, compromising the transparency and visual acuity. First step in the management of LSCD is early control of the ocular surface inflammation and elimination of the causative factor. Surgical intervention in terms of limbal stem cell transplantation is the definitive treatment for most of the cases. Successful limbal stem cell transplantation can achieve rapid surface healing, stable ocular surface without recurrent erosions or persistent epithelial defects, regression of corneal vascularization, and restoration of a smooth and optically improved ocular surface, resulting in improved visual acuity and probably increased success for subsequent keratoplasty. Here is a brief review covering etiology, clinical presentation, and management options for the cases of LSCD.

Corneal limbus is a specialized structure that harbors corneal epithelial stem cells in its vicinity. Anatomically, it is a transition zone between cornea, conjunctiva, and sclera. Limbal epithelium dips down into the underlying loose connective tissue to form palisades of Vogt where the basal stem cells are supported by a unique microenvironment. Along with blood vessels, lymphatics, mast cells, lymphocytes, and macrophages, melanocytes are also present to protect underlying stem cells from ultraviolet (UV) radiations. It also acts as a barrier for the growth of conjunctival epithelium over cornea. [1]

LSCD occurs when the limbal epithelial stem cells are destroyed or their supporting microenvironment becomes dysfunctional. LSCD is characterized by intrusion of corneal surface by conjunctival epithelium containing goblet cells, neovascularization, chronic inflammation, and recurrent or persistent corneal epithelial defects [Figure 1], [Figure 2], [Figure 3], [Figure 4]. [2] Pathologically, such corneas exhibit destruction of basement membrane, superficial neovascularization, infilteration, scarring and poor epithelial integrity. Such patients are poor candidates for keratoplasty surgery due to increased chances of graft failure.

  Etiology of LSCD Top

LSCD can be primary, related to an insufficient stromal microenvironment to support stem cell function, such as aniridia, congenital erythrokerato-dermia, keratitis associated with multiple endocrine deficiencies, neurotrophic (neural and ischemic) keratopathy, and chronic limbitis; or secondary (more common) related to external factors that destroy limbal stem cells such as chemical (most common) or thermal injuries, ocular cicatricial pemphigoid (OCP), Stevens Johnson syndrome (SJS), multiple surgeries or cryotherapies, contact lens wear, or extensive microbial infection. [3],[4] LSCD can be diffuse (total) or sectoral (partial). In the latter case, conjunctivalization of the corneal epithelium affects only part of the corneal surface. In some patients, stem cell deficiency may be subclinical at the time of the insult, and may eventually progress to an advanced stage as the stem cell population depletes further, over time.

Aniridia is rare, bilateral, pan-ocular disorder with an underlying mutation in PAX-6 gene. It involves cornea, iridocorneal angle, iris, lens, fovea, and optic nerve. Aniridia associated keratopathy begins in first decade of life and characterized by progressive thickening and vascularization of cornea, recurrent erosions, ulcerations, sustained inflammation, and subepithelial fibrosis. [5] Studies in animal models revealed altered expression of CK 12 keratin, presence of goblet cells on corneal surface, depleted limbal stem cells with reduced production of corneal epithelial progeny. [6]

Chemical and thermal injuries by causing damage to limbal vasculature and intense inflammation causes irreversible damage to limbal stem cell population. [7] Long-term contact lens wear is also implicated in causing limbal ischemia and depletion of stem cell population. Repeated cautery on limbus, cryotherapy, and repeated ocular surgeries cause direct damage to limbal stem cells.

OCP is an autoimmune disease in which autoantibodies are directed against structural proteins of epithelial-subepithelial junction of mucosae and skin. It leads to chronic inflammation of conjunctiva, severe dry eye, symblephera, epithelial keratinization, and subsequent LSCD. [8] All other conditions causing prolonged ocular surface inflammation destroy stromal microenvironment and eventually deplete limbal stem cell population.

  Clinical Features and Diagnosis of LSCD Top

LSCD is characterized by decreased vision, photophobia, redness, watering, and recurrent attacks of pain. [1] The hallmark of LSCD is a triad of conjunctivalization, neovascularization, and chronic inflammation [Figure 1], [Figure 2], [Figure 3], [Figure 4]. [3] Cornea loses its normal transparency and frequently stained abnormally by fluorescein giving stippled appearance. In some cases of partial LSCD, a clear line of demarcation is visible between corneal and conjunctival cells. Diagnosis of LSCD can be confirmed clinically by persistant corneal epithelial defects and loss of limbal palisades of Vogt. [9] Impression cytology is a non invasive investigative tool where superficial layers of ocular surface epithelium can be removed by application of nitrocellulose acetate paper to the ocular surface. The cells thus removed can be subjected to histological, immunohistological, or molecular analysis. [10],[11] Immunohistochemically, the absence of a cornea-type differentiation (such as absence of CK3), and the presence of mucin in goblet cells, has been shown by monoclonal antibodies. [12] Histopathology of resected pannus from corneal surface is confirmatory in most of the cases. Histopathology shows presence of conjunctival tissue along with fibrovascular components.
Figure 1: Showing recurrent scarring and vascularization leading to graft failure

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Figure 2: Limbal stem cell deficiency in a case of vernal conjunctivitis

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Figure 3: Limbal stem cell deficiency followed by chemical injury with lime

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Figure 4: Extensive corneal vascularization and conjunctivalization in a case of recurrent viral keratitis

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  Management of LSCD Top

Currently no medical treatment is available to restore limbal stem cells. After the acute inflammation and inciting event is controlled, stem cell transplantation is the definitive treatment to correct LSCD.

Asymptomatic patients with partial and peripheral conjunctivalization of the corneal surface can be kept under observation and may not require intervention. Corneal and conjunctival epithelial cell phenotypes have been known to co-exist on the corneal surface for prolonged periods without significant extension of the conjunctivalized area. [13] Any tendency of conjunctiva-like epithelium to encroach on to corneal surface can be prevented by sequential sectoral conjunctival epitheliectomy (SSCE) and scraping. [13],[14],[15] Conjunctival-limbal and keratolimbal grafts are other modalities of treatment. The transplants that contain mostly conjunctival epithelium and limbal tissue are called conjunctival-limbal grafts, for kerato-limbal grafts the biggest part is corneal and limbal tissue. The conjunctival-limbal grafts are mostly used when there is a living donor, i.e., autologous transplants in unilateral affection and living related donors (a relative of the patient) in bilateral cases. The kerato-limbal transplant probably contains more limbal stem cell; however, it is more damaging to the donor eye and is therefore more often used when there is cadaveric tissue often in cases with bilateral disease.

Sectoral sequential conjunctival epitheliectomy (SSCE)

It is well-known that any corneal epithelial defect involving the limbus heals by centripetal as well as circumferential migration of dividing epithelial cells. However, in cases of sectoral LSCD, often, conjunctival cells intrude the adjacent area of cornea thereby preventing expansion of the remaining nearby stem cells in the affected area. In these circumstances, SSCE is performed where repeated removal of sectoral conjunctiva covering the cornea or bulbar conjunctiva adjacent to the affected area prevents conjunctiva from reaching the limbus and allow healing of defect with epithelial cells of corneal phenotype. [14] Through this procedure conjunctiva is effectively held back until repopulation of the limbus from surviving limbal cells or donor limbus is achieved.

Dua et al. reported a series of 10 patients with conjunctivalization of cornea which were followed for an average of 7.5 months. Five patients in this group had their conjunctival epithelium removed from the corneal surface and allowed to heal from the remaining intact corneal epithelium. In another 4 paients with corneal epithelial defect, the conjunctival epithelium was actively prevented from crossing the limbus by mechanically scraping it off. Removal of conjunctival epithelium from the cornea allowed cells of corneal epithelial phenotype to cover the denuded area with improvement of symptoms and vision. [15]

Conjuntival-limbal autograft (CLAU)

In this procedure, 3-4 mm of conjunctiva along with limbal epithelium is harvested from fellow eye of the patient. It is imperative to exclude LSCD in the donor eye especially in bilateral chemical injuries where other eye may be subclinically involved. Basti et al. noted progressive conjunctivalization in three cases following conjunctival limbal autograft, probably due to attenuation of limbal stem cell function with time. [16] Careful case selection is very important aspect of this procedure. [17] In their series, pseudopterygium formation was seen in one donor eye of a patient with history of bilateral chemical injury but without clinical evidence of LSCD. The amount of harvested tissue should not exceed 6 clock hours for fear of precipitating LSCD in donor eye. Care is taken to avoid damage to limbus while dissection. Buttonholing and dessication of donor tissue should be avoided.

Kenyon and Tseng first described the technique of limbal autograft transplantation in a series of 26 patients. The indications were chemical and thermal injuries in 22 (84.6%), contact lens-induced keratopathy in 3 (11.5%), and stem cell deficiency following multiple surgeries in 1 (3.8%). They fashioned two free limbal grafts that encompassed 8 clock hours of the recipient limbus from the apparently healthy contralateral eye. The mean follow-up was 18 months (2-45 months). In 10 (38.5%) patients with persistent epithelial defects for 3 weeks to 4 years, there was rapid re-epithelialization (1-4 weeks). Failure of epithelialization was seen in 3 (11.5%) eyes. Substantial improvement was noted in vision and reduction in stromal vascularization. [18]

Keratolimbal allograft (KLAL)

In this technique, the cadaveric donor tissue incorporated part of cornea along with limbal tissue. Tissue from the youngest possible donor with an upper limit of 50 years is recommended. Surgery should be performed within 72 hours as the cells are expected to be more active and vital. [19] While harvesting the donor tissue damage to epithelium should be avoided and a peripheral skirt of adjoining conjunctiva and partial thickness sclera of about 3-4 mm should be included. If the whole globe is used, a vaccum trephine may be used centrally, set to a depth of 150 microns and the dissection of peripheral cornea and limbus may be carried out with a diamond knife. [20] Amniotic membrane has been used as an adjunct to limbal transplantation as it helps to reconstruct perilimbal stroma and reduces inflammation and vascularization. [18]

Two recent reports [21],[22] have described the long-term outcome of keratolimbal allograft with an average follow-up of 3 and 5 years each. Both groups noted a progressive decline in the visual acuity and graft survival with time. The overall change in ambulatory vision (best-corrected visual acuity, 6/60) was 76.6% ± 6.8% after 1 year. This decreased to 53.6% ± 9.9% and 44.6% ± 11.6% at 3 and 5 years, respectively. [22] SJS patients fared worse than the others. The survival of the allograft also decreased with time from 76.9% ± 6.7% at one year to 47.4% ± 11.7% at 3 years and 23.7% ± 17.5% at 5 years. Simultaneous penetrating keratoplasty and limbal transplant resulted in a less favorable outcome after one year though no difference was noted at 3 years. The survival of a second limbal transplant was better than the first. [21]

Amniotic membrane transplantation (AMT)

Role of amniotic membrane (AM) in cases of recent chemical and thermal injuries is well-established. It not only controls active inflammation that is harmful for surviving limbal stem cells but also provides stable and quite ocular surface for stem cell transplantation. Tseng et al. have successfully used AMT and mechanical debridement and advocate its use to treat patients with partial stem cell deficiency. [23] Their study showed that with partial or focal limbal stem cell deficiency, AMT improves both the corneal surface and the vision. Both fresh and preserved AM have been found to function equally well when transplanted onto the ocular surface. [24] Advantage with preserved AM is that it is well-processed and thoroughly screened against transmissible diseases thereby eliminating the slightest risk of disease transmission that may be associated with fresh AM. In addition to this 30 preserved grafts can be prepared from one placenta unlike tissue wastage while using fresh AM. Dry preserved AM is easily available in market and can be stored at room temperature for 2 to 5 years unlike cryopreserved AM which requires stringent storage conditions.

Tseng et al. used AMT associated with limbal transplantation in cases with total stem cell deficiency. [23] AM as a carrier in this instance has several distinct advantages over the other substrates that have been used. The basement membrane of AM contains Type IV collagen and laminin which plays an important role in cell adhesion. [25] Further, it acts as a natural substrate for the cell growth and when transplanted gets integrated onto the corneal surface. It also enables easier handling during transplantation. Tseng et al. proposed that culturing the explants on an intact AM with devitalized epithelium favors expansion of an epithelial phenotype that closely resembles limbal stem cells. [23]

Cultivated limbal epithelial transplantation (CLET)

The technique was first described by Pelligrini et al.[26] after which various modifications were suggested by several groups. In this procedure autologous limbal epithelial stem cells are expanded over a carrier tissue (AM mostly) in a special explants cell culture system for 2 weeks and then transplanted onto the diseased ocular surface. Various animal derived products are used in xenogenic explants culture system, eg., murine feeder cells, fetal bovine serum, which theoretically carries a risk of rejection or transmission of prion diseases. To avoid this autologus human serum has been proposed as an alternative constituent in explant cultures.

Simple limbal epithelial transplantation (SLET)

limbal tissue is divided into small pieces and transplanted over amniotic membrane sutured over recipient cornea followed by bandage contact lens application. SLET enjoys advantage over CLET in being a single staged procedure without any extra requirements of stringent laboratory conditions to propagate and maintain cellular growth. Sangwan et al.[27] has reported series of six patients having total LSCD secondary to ocular burns who underwent SLET. All their cases maintained stable corneal epithelial surface by 6 weeks post-operatively and maintained so during follow-up visits. Visual acuity improved from worse than 20/200 in all cases to 20/60 or better in 66.6% cases without any complications. They concluded that SLET is an effective modality for treating total LSCD secondary to ocular burns.

Cultivated oral mucosal epithelial transplantation (COMET)

COMET was first described by Nakamura et al.[28],[29] in 2004. Various cases with bilateral LSCD demonstrated stable corneal surface and reduced inflammation, making them suitable candidate for future corneal transplantation.


Patients having SJS, OCP, or other autoimmune diseases are poor candidates for stem cell therapy and corneal transplantation as the severe and long standing ocular surface inflammation hinders the survival of stem cells. Such candidates can be given visual outcome with the help of keratoprosthesis. Boston type I keratoprosthesis and Osteo-odonto Keratoprosthesis have shown satisfactory visual outcomes in such cases. [30] However, various complications related to this are also a point of concern. [31]

  References Top

Dua HS, Azuara-Blanco A. Limbal stem cells of the corneal epithelium. Surv Ophthalmol 2000;44:415-25.  Back to cited text no. 1
Tseng SC. Regulation and clinical implications of corneal epithelial stem cells. Mol Biol Rep 1996;23:47-58.  Back to cited text no. 2
Pfister RR. Corneal stem cell disease: Concepts, categorization and treatment by auto and homotransplantation of limbal stem cells. CLAO J 1994;20:64-72.  Back to cited text no. 3
Dua HS. Stem cells of the ocular surface: Scientific principles and clinical applications. Br J Ophthalmol 1995;79:968-9.  Back to cited text no. 4
Lee H, Khan R, Okeefe M. Aniridia: Current pathology and management. Acta Ophthalmol 2008;86:708-15.  Back to cited text no. 5
Ramaesh T, Collinson JM, Ramaesh K, Kaufman MH, West JD, Dhillon B. Corneal abnormalities in PAX6+/- small eye mice mimic human aniridia- related keratopathy. Invest Ophthalmol Vis Sci 2003;44:1871-8.  Back to cited text no. 6
Wagoner MD. Chemical injuries of the eye: Current concepts in pathophysiology and therapy. Surv Ophthalmol 1997; 41:275-313.  Back to cited text no. 7
Foster CS, Sainz M. Ocular cicatricial pemphigoid review. Curr Opin Allergy Clin Immunol 2004;4:435-9.  Back to cited text no. 8
Chen JJ, Tseng SC. Abnormal corneal epithelial wound healing in partial-thickness removal of limbal epithelium. Invest Ophthalmol Vis Sci 1991;32:2219-33.  Back to cited text no. 9
Kinoshita S, Kiritoshi A, Ohji M, Ohashi Y, Manabe R. Disappearance of palisades of Vogt in ocular surface disease. Jpn J Clin Ophthalmol 1986;40:363-6.  Back to cited text no. 10
Puangsricharern V, Tseng SC. Cytologic evidence of corneal diseases with limbal stem cell deficiency. Ophthalmology 1995;102:1476-85.  Back to cited text no. 11
Kenyon KR, Bulusoglu G, Ziske JD. Clinical pathologic correlations of limbal autograft transplantation. Am J Ophthalmol 1990;31:l-12.  Back to cited text no. 12
Coster DL, Aggarwal RK, Williams KA. Surgical management of ocular surface disorders using conjunctival and stem cells allografts. Br J Ophthalmol 1995;79:977-82.  Back to cited text no. 13
Dua HS, Forrester JV. The corneoscleral limbus in human corneal epithelial wound healing. Am J Ophthalmol 1990;110:646-56.  Back to cited text no. 14
Dua HS. The conjunctiva in corneal epithelial wound healing. Br J Ophthalmol 1998;82:1407-11.  Back to cited text no. 15
Basti S, Mathur U. Unusual intermediate-term outcome in 3 cases of limbal autograft transplantation. Ophthalmology 1999;106:958-63.  Back to cited text no. 16
Basti S, Rao SK. Current status of limbal conjunctival autograft. Curr Opin Ophthalmology 2000;11:224-32.  Back to cited text no. 17
Kenyon KR, Tseng SC. Limbal autograft transplantation for ocular surface disorders. Ophthalmology 1989;96:709-22.  Back to cited text no. 18
Croasdale CK, Schwartz GS, Malling JV, Holland EJ. Keratolimbal allograft: Recommendations for tissue procurement and preparation by eye banks and standard surgical technique. Cornea 1999;18:52-8.  Back to cited text no. 19
Dua HS, Azuara-Blanco A. Allo-limbal transplantation in patients with limbal stem cell deficiency. Br J Ophthalmol 1999;83:414-9.  Back to cited text no. 20
Solomon A, Ellies P, Anderson DF, Touhami A, Grueterich M, Espana EM, et al. Long-term outcome of keratolimbal allograft with and without penetrating keratoplasty for total limbal stem cell deficiency. Ophthalmology 2002;109:1159-66.  Back to cited text no. 21
Ilari L, Daya SM. Long-term outcomes of keratolimbal allografts for the treatment of severe ocular surface disorders. Ophthalmology 2002;109:1278-84.  Back to cited text no. 22
Tseng SC, Prabhasawat P, Barton K, Gray T, Meller D. Amniotic membrane transplantation with or without limbal allografts for corneal surface reconstruction in patients with limbal stem cell deficiency. Arch Ophthalmol 1998;116:431-41.  Back to cited text no. 23
Adds PJ, Hunt CJ, Dart JK. Amniotic membrane grafts, "fresh" or frozen? A clinical and in vitro comparison. Br J Ophthalmol 2001;85:905-7.  Back to cited text no. 24
Grueterich M, Espana EM, Tseng SC. Ex vivo expansion of limbal epithelial stem cells: Amniotic membrane serving as a stem cell niche. Surv Ophthalmol 2003;48:631-46.  Back to cited text no. 25
Pelligrini G, Traverso CE, Franzi AT, Zingirian M, Cancedda R, De Luca M. Long term restoration of damaged corneal surfaces with autologus cultivated corneal epithelium. Lancet 1997;349:990-3.  Back to cited text no. 26
Sangwan VS, Basu S, MacNeil S, Balasubramanian D. Simple limbal epithelial transplantation: A novel surgical technique for the treatment of unilateral limbal stem cell deficiency. Br J Opthalmol 2012;96:931-4.  Back to cited text no. 27
Nakamura T, Endo K, Cooper LJ, Fullwood NJ, Tanifuji N, Tsuzuki M, et al. The successful culture and autologus transplantation of rabbit oral mucosal epithelial cells on amniotic membrane. Invest Ophthalmol Vis Sci 2003;44:106-16.  Back to cited text no. 28
Nakamura T, Inatomi T, Cooper LJ, Rigby H, Fullwood NJ, Kinoshita S. Phenotypic investigation of human eyes with transplanted autologous cultivated oral mucosal epithelial sheets for severe ocular surface diseases. Ophthalmology 2007;114:1080-8.  Back to cited text no. 29
Falcinelli G, Falcinelli B, Taloni M, Rigby H, Fullwood NJ, Kinoshita S. Modified osteo-odonto keratoprosthesis for treatment of cornealblindness: Long term anatomical and functional outcomes in 181 cases. Arch Ophthalmol 2005;123:1319-29.  Back to cited text no. 30
Chan CC, Holland EJ. Infectious keratitis after Boston type 1 keratoprosthesis implantation. Cornea 2012;31:1128-34.  Back to cited text no. 31


  [Figure 1], [Figure 2], [Figure 3], [Figure 4]

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