Simple limbal epithelial transplantation another option for ocular surface disease
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The regenerative capacity of stem cells has been of great interest in every field of medicine.
The additional role of a pericorneal structure contributing to epithelial regeneration was first postulated in 1971. Detailed evaluation of the limbal palisades of Vogt hypothesized the presence of repository cells in the interpalisadal region. The XYZ hypothesis further elaborated the role of centripetal movement of peripheral cells (Y) along with proliferation of basal cells (X) to replace epithelial cell loss (Z) in the cornea. A recent report by Dua and colleagues suggests the existence of distinct “limbal epithelial crypts” that provide a niche to maintain a favorable microenvironment for ocular stem cells.
Limbal stem cell transplantation
Limbus stem cell transplantation has evolved over time as a useful technique in management of limbal stem cell deficiency (LSCD). The availability of multiple techniques led to the development of a separate nomenclature system by the Cornea Society for ocular surface rehabilitative procedures.
Keratolimbal allografts were the earliest form of ocular stem cell regeneration, using 360° cadaveric donor ring segments (periphery cornea and limbus) sutured to the corneal periphery of the diseased host eye. This procedure was combined with penetrating/lamellar keratoplasty with the cornea of the same donor corneoscleral rim. The role of sectoral/partial conjunctival limbal autograft was first published in 1989 by Keivyon and Tseng. This was followed by the development of cultivated limbal epithelial transplant (CLET), which used donor limbal cells from the contralateral eye cultured in vitro to treat such patients.
Simple limbal epithelial transplant
Simple limbal epithelial transplant (SLET) by Dr. Virendra Sangwan and colleagues offered a low-cost, single-step alternative to CLET in absence of sophisticated laboratories. The donor graft (limbal epithelial cells) can be harvested from the other eye of the patient (autologous SLET) or a live related/cadaveric donor eye (allogeneic SLET), with the risk for rejection being significant in the latter case.
Preoperative evaluation
The primary indication for this procedure is epithelial healing disorders in secondary LSCD due to ocular surface diseases such as chemical burns, Stevens-Johnson syndrome (SJS), mucous membrane pemphigoid (MMP), allergic conjunctivitis or post-excision of ocular surface squamous neoplasia/large pterygium. A multidisciplinary approach is needed for proper management of systemic features and serological parameters in case of SJS or MMP. This is done in collaboration with an internist. Detailed preoperative examination is performed with special attention to surface abnormalities. Lid abnormalities such as ectropion, entropion and lagophthalmos, if present, need to be corrected surgically before or along with SLET to ensure minimal adnexal disturbance. Proper evaluation of associated dry eye syndrome with lubricants, autologous serum or punctal occlusion is a must to ensure good tear film stability. Ocular surface pathologies such as pterygium, pseudopterygium, pannus and symblepharon are excised or released to provide a smooth non-inflamed bed for good graft adhesion. It is necessary to evert the lids and look for small symblepharon that may be missed on cursory examination.
Diagrammatic representation and photographic documentation of all ocular findings, such as corneal vascularization (clock hours, depth into cornea, superficial/deep) and corneal opacity (size, location, depth), are a must. This aids in monitoring the progression of disease and response after surgery.
Detailed high-resolution OCT images are captured to analyze stromal integrity, measure corneal thickness and decipher structural organization of the anterior segment in cases with a limited view. The best surgical outcomes are experienced in wet ocular surfaces with resolved inflammation, minimal/no symblepharon and clear underlying stroma. In case of autologous SLET, it is important to ensure that informed consent is obtained for both the donor and recipient eye. This is imperative due to the rare but existent possibility of focal LSCD developing at the donor site.
Surgical technique
Preoperative instillation of brimonidine eye drops 0.15% and phenylephrine 5% three times every 5 to 10 minutes in the donor and recipient eye help to reduce intraoperative bleeding.
Harvesting donor limbal tissue
Superior limbal palisades are preferred (Figure 1) to harvest donor limbal tissue. In case of allogeneic SLET, the harvested limbal cells should be less than 48 hours old, from a live-related or cadaveric donor younger than 60 years and seronegative for HIV, hepatitis B surface antigen, hepatitis C virus and VDRL. Local anesthesia (peribulbar/topical) is administered in case of autologous or allogeneic SLET. Under sterile precautions in the operating room, a small limbus-based conjunctival flap is created 2 mm away from the limbus. Subconjunctival dissection is continued with a crescent blade up to 1 mm of superficial clear cornea. Adjacent 1 to 2 clock hours of limbus stem cells are then excised using Vannas scissors and preserved in balanced salt solution. The conjunctival flap is repositioned using fibrin glue or sutures.
Source: Amar Agarwal, MS, FRCS, FRCOphth, Shana Sood, DNB, FICO, MRCS, Soosan Jacob, MS, FRCS, DNB, and Dhivya Ashok Kumar, MD, FRCS, FICO, FAICO
SLET
In SLET, local peribulbar anesthesia is administered. Under sterile precautions, 360° conjunctival peritomy is performed with complete removal of vascularized pannus. Superficial fibrotic scar, conjunctivalized tissue and hypertrophied epithelium are gently dissected (Figure 2). The surface is kept moist to ensure smooth movement of the instrument. This is followed by complete drying with a Weck-Cel spear and application of a thin, uniform layer of fibrin glue. The amniotic membrane graft (AMG) is stretched taut and applied basement membrane (non-sticky) side up with the ends firmly tucked under the peritomy margins. A blunt instrument is used to iron out excess glue and folds in the AMG. This is crucial as lumpy, bumpy glue application or folds in amniotic membrane can interfere with graft adhesion and anchoring. The donor rim of limbal epithelial cells, cut in small bits, is placed epithelial side up (identified by pigmented palisades) in the mid-periphery of the cornea (2 mm to 3 mm away from limbal vessels to prevent rejection) and secured in place by a drop of fibrin glue over each bit. After the glue polymerizes, a soft bandage contact lens is applied and acts as a protective bandage.
Postoperative care
Moxifloxacin 0.5% is instilled topically four times daily for a period of 2 weeks or until the epithelium heals in the donor and recipient eye. Prednisolone 1% or dexamethasone 0.1% are instilled four times a day for a period of 6 weeks in case of autogenous SLET. In patients with allogeneic SLET, topical steroids are continued in a tapering dose, followed by one to two drops a day for many years after surgery. Topical cyclosporine 2% can also be administered for long-term immunosuppression. Lubricants in the form of drops or gel are continued to maintain the ocular surface. It is imperative to counsel the patient to report immediately in case of redness, photophobia, pain or decrease in vision. These may herald the onset of rejection (in allogeneic SLET) or sterile/microbial keratitis, which is common in patients with compromised ocular surfaces. Rejection in cases of allogenic SLET is treated with a combination of systemic and topical steroids.
Re-epithelialization and healing
Re-epithelialization (Figure 3) of the ocular surface takes place within 2 weeks of the surgery, after which the bandage contact lens can be removed. Fluorescein staining patterns help track the growth of the epithelium nest from each donor bit. It is this epithelial nest that further anchors the donor bit to the AMG with time.
Inadvertent growth of conjunctival epithelium from adjacent peritomy margins poses a significant threat to proper re-epithelialization of the cornea by limbal stem cells. The conjunctivalized corneal surface is characterized by goblet cells and vascularization and prone to recurrent erosions. Hence, regular denuding of encroaching conjunctival cells is necessary until the limbal stem cells cover the complete corneal surface (10 to 14 days). This can be achieved by sequential sectoral conjunctival epitheliectomy (SSCE) using a brush or gentle scraping under topical anesthesia. However, subconjunctival hemorrhage may result from damage to encroaching conjunctival vessels, and patients experience ocular discomfort on most sittings. Additionally, SSCE is contraindicated in eyes with thin stroma, total LSCD and dry, anesthetic corneas. An amnion-assisted conjunctival epithelial redirection (ACER) technique by Dua and colleagues described the role of an additional AMG overlay (stromal side down) on donor limbal stem cells as a barrier shield (Figure 4). This prevents contamination of the corneal surface by conjunctival cells, thereby ensuring complete epithelialization of the cornea by limbal stem cells alone.
At 3 months after surgery, the donor limbal bits are taken up completely. A successful outcome is defined as a stable, transparent, epithelialized and avascular surface of the cornea.
Summary
The procedure of SLET has undergone multiple modifications to achieve its present form. It is a low-cost, single-step procedure with a relatively easy learning curve. The use of AMG as an in vivo culture medium eliminates the need for cell culture laboratories and plays a role in resolution of associated corneal haze. It is much more effective in pediatric patients as compared with CLET. Reported efficacy of autologous SLET after 1 year is 70% to 84% in patients with unilateral LSCD due to chemical burns. The majority of patients achieved an improvement in best corrected visual acuity of two lines after surgery. SJS and MMP are the most common indications for allogeneic SLET. Stromal haze may interfere with the visual outcome of the procedure, warranting a lamellar/penetrating keratoplasty or keratoprosthesis for complete rehabilitation of the patient.
- References:
- Basu S, et al. Ophthalmology. 2016;doi:10.1016/j.ophtha.2015.12.042.
- Dua HS, et al. Br J Ophthalmol. 2017;doi:10.1136/bjophthalmol-2015-307935.
- Gupta N, et al. Indian J Ophthalmol. 2018;doi:10.4103/ijo.IJO_602_17.
- Sangwan VS, et al. Br J Ophthalmol. 2012;doi:10.1136/bjophthalmol-2011-301164.
- Vazirani J, et al. Br J Ophthalmol. 2016;doi:10.1136/bjophthalmol-2015-307348.
- For more information:
- Amar Agarwal, MS, FRCS, FRCOphth, is director of Dr. Agarwal’s Eye Hospital and Eye Research Centre. Agarwal is the author of several books published by SLACK Incorporated, publisher of Ocular Surgery News, including Phaco Nightmares: Conquering Cataract Catastrophes, Bimanual Phaco: Mastering the Phakonit/MICS Technique, Dry Eye: A Practical Guide to Ocular Surface Disorders and Stem Cell Surgery and Presbyopia: A Surgical Textbook. He can be reached at 19 Cathedral Road, Chennai 600 086, India; email: aehl19c@gmail.com; website: www.dragarwal.com.
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