Preserved Human Amniotic Membrane Research Articles

Effects of different doses of gamma irradiation on oxygen and water vapour transmission rate of preserved human amniotic membrane

Preserved human amniotic membrane either air dried or glycerol preserved has been used effectively to treat superficial and partial thickness wounds without leaving any obvious hypertrophic scar. The preserved amnion, sterilised by ionising radiation, is known as an effective barrier for heat, fluid and protein loss while adheres nicely on wound. Air drying slightly reduced the oxygen transmission rate (OTR) of the amnion and the value significantly dropped after 15 kGy (p < 0.05). Glycerol preservation significantly reduced (p < 0.05) the OTR indicating less oxygen transmitted through the well structured cells of the amnion. Increase in the OTR with the increasing radiation doses up to 35 kGy possibly due to direct effects of radiation that resulted in large intercellular gaps. Both preservation methods significantly increased (p < 0.05) the water vapour transmission rate (WVTR). However, the low WVTR in the air dried amnion at 15 and 25 kGy was postulated due to cross-linking of collagen. Changes in the biophysical properties can be linked to direct and indirect effects of radiation on collagen bundles. The radiation dose of 25 kGy caused no adverse effect on biophysical properties hence it is still acceptable to sterilize both the air dried and the glycerol preserved amnions.

Two separate methods of stem cell transplantation in both eyes of a single patient presented with bilateral stem cell deficiency following exposure to Euphorbia latex

Limbus is the most dynamic area of ocular surface which is the “home” of stem cells especially in the region of palisades of Vogt. Stem cell deficiency is manifested by ingrowth of vessels into the cornea with resultant corneal haze that results in decreased vision [1]. Stem cell deficiencies are treated by transplantation of stem cells. The methods are varied and includes transplantation from cadaveric tissue, from living related donor (in bilateral cases) and from fellow eye (in unilateral cases). In vitro cultured stem cells in human amniotic membrane (HAM) base may also be utilised. We report here the successful outcome of stem cell implantation in a patient with end stage limbal stem cell deficiency following exposure to the latex of Euphorbia. The Euphorbiaceae family includes trees, succulents and herbaceous plants. Different species of Euphorbia grow all over the world. The milky latex or sap is toxic and may cause intense inflammation of the skin and the eye [2, 3]. Ocular toxic reaction varies from mild conjunctivitis to keratouveitis to severe corneal vascularisation in neglected cases. Ocular toxicity can be caused by three different species of Euphorbia, namely, Euphorbia trigona (African milk tree), Euphorbia neriifolia (Indian Spurge tree) and Euphorbia milii (crown-of-thorns houseplant). In this present case, the patient (female, aged 45 years) with bilateral stem cell deficiency was treated by transplantation of stem cell from living related donor(mother) in the left eye and in vitro cultured stem cell on HAM base in the right eye. A normotensive, non-diabetic, patient (female, aged 45 years) presented to our clinic with loss of vision in both eyes. She gave a history of exposure to Euphorbia latex 5 months back in her both eyes. She neglected the condition and there was gradual decrease in his vision following the brief acute episode. Visual acuity in RE was 1/60, in LE it was finger counting at 2 m. There was no improvement in vision with refractive correction. Schirmer test values in both eyes measured 15 mm. Corneal sensation in both eyes were normal. Intraocular pressure in BE were 18 mmHg and USG (A+B) scans in both eyes were normal. Anterior segment of both eyes revealed 360° limbal vascularisation with diffuse stromal haze up to the extent that anterior chamber details including the pupil is barely visible (Fig. 1). Fig. 1 Right eye showing limbal vascularisation with diffuse stromal haze up to the extent that anterior chamber details including the pupil is barely visible A decision for transplantation of stem cells was made. She was put on systemic steroids (1 mg/kg/day) 3 days prior to admission. From the mother, two separate conjuctivolimbal lenticules were prepared [4 × 2 mm each], one from each eye to reduce chances of development of iatrogenic stem cell loss. The recipient bed in LE was prepared by marking two separate areas: one centred around 6 o’clock hours and the other around 12 o’clock region of the limbus. Peritomy was done and bleeding controlled by cauterisation. The donor lenticules were sutured to the recipient bed with interrupted 10-0 monofilament nylon sutures. In the post-operative period, the patient was put on systemic steroids (1 mg/kg/day) in a tapering dose, topical steroid (pred acetate 1 %), topical antibiotic (topical moxifloxacin) and artificial tears (carboxymethyl cellulose). Both topical and systemic steroids were tapered slowly over a period of 6 weeks and topical antibiotics were withdrawn after the first 2 weeks. The patient was advised weekly visit at the cornea clinic. Four months after the first operation, her RE was operated. Recipient bed was prepared as described before and in vitro cultured stem cells on HAM base were sutured at around 360° of limbus with interrupted 10-0 monofilament nylon sutures. The whole of cornea including transplanted discs were covered by preserved HAM and sutured to adjacent conjunctiva with interrupted 10-0 monofilament nylon. Patient was placed on same post-operative regimen as before. The patient also underwent cataract surgery in both eyes 5 months after her last operation in RE.

Scanning electron microscopic assessment on surface morphology of preserved human amniotic membrane after gamma sterilisation

Human amniotic membrane that has been processed and sterilised by gamma irradiation is widely used as a biological dressing in surgical applications. The morphological structure of human amniotic membrane was studied under scanning electron microscopy (SEM) to assess effects of gamma radiation on human amniotic membrane following different preservation methods. The amniotic membrane was preserved by either air drying or submerged in glycerol before gamma irradiated at 15, 25 and 35 kGy. Fresh human amniotic membrane, neither preserved nor irradiated was used as the control. The surface morphology of glycerol preserved amnion was found comparable to the fresh amniotic membrane. The cells of the glycerol preserved was beautifully arranged, homogonous in size and tended to round up. The cell structure in the air dried preserved amnion seemed to be flattened and dehydrated. The effects of dehydration on intercellular channels and the microvilli on the cell surface were clearly seen at higher magnifications (10,000×). SEM revealed that the changes of the cell morphology of the glycerol preserved amnion were visible at 35 kGy while the air dried already changed at 25 kGy. Glycerol preservation method is recommended for human amniotic membrane as the cell morphological structure is maintained and radiation doses lower than 25 kGy for sterilization did not affect the appearance of the preserved amnion.

Ocular Surface Reconstruction using Amniotic Membrane following Excision of Conjunctival and Limbal Tumors

To report the clinical results of patients treated by preserved human amniotic membrane transplantation (AMT) following the removal of conjunctival and limbal tumors. Retrospective noncomparative interventional case series of 9 patients (9 eyes) who underwent AMT after removal of conjunctival and limbal tumors with lesion-free margins and perilesional cryotherapy. The excised tumors were histopathologically examined and included 2 squamous cell carcinomas, 2 papillomas, and 5 nevi. Bulbar conjunctiva was involved in all of the eyes, limbus and cornea in 7 and 3 eyes, respectively. The mean extent of the limbal involvement was 4 clock hours (range 2-9, SD 2.4); the average diameter of the base of the tumor was 12.8 mm (range 10-20, SD 4.4). The mean follow-up time was 38 months (range 13-60, SD 15). No surgical or early postoperative complications were observed. In all eyes, complete healing of the tissue defect occurred, resulting in a stable, wet, and noninflamed epithelium. All eyes demonstrated a smooth ocular surface except one with a clinically insignificant symblepharon after the excision of a squamous cell carcinoma. Superficial peripheral corneal vascularization and opacification as a sign of partial limbal stem cell deficiency developed in 2 eyes. In one case, a recurrence of conjunctival papilloma was diagnosed after a 3-year follow-up. Amniotic membrane transplantation is an effective method of reconstruction following a conjunctival and limbal tumor excision and cryotherapy of surgical wound margins. In most cases, complete healing of an ocular surface can be achieved without any clinically significant complications.

Treatment of Primary Pterygium

To compare preserved human amniotic membrane transplantation and conjunctival autografts for treatment of primary pterygium. Thirty-seven eyes of 35 patients with primary pterygium underwent surgical excision. In group A, 25 eyes were treated with conjunctival autografts. In group B, 12 eyes were treated with preserved human amniotic membrane transplantation. These two groups were compared in terms of complications, recurrence, and final appearance, prospectively. We observed two cases of recurrence in group A and five cases of recurrence in group B. The onset of recurrence was about 5.5 months in both groups. We observed two eyes with Dellen ulcer and one case with a Tenon's cyst in the superior temporal quadrant in group A and one case of Dellen ulcer and granulation tissue of one eye in group B. We suggest that free conjunctival autografting is a more effective technique in primary pterygium in terms of lowering recurrences compared with the amniotic membrane transplantation technique.

Mitomycin C, amniotic membrane transplantation and limbal conjunctival autograft for treating multirecurrent pterygia with symblepharon and motility restriction

Treatment of eyes with multirecurrent pterygia associated with severe symblepharon and motility restriction is challenging. A combined surgical procedure of intraoperative mitomycin C, amniotic membrane transplantation and conjunctival limbal autograft was applied for treating such eyes. Seven eyes of seven patients who had previously undergone an average of four operations for pterygial removal and who manifested recurrent pterygia associated with severe symblepharon and motility restriction were involved in this retrospective study. The surgical procedures involved clearing fibrovascular membrane over the cornea, extensive excision of epibulbar fibrovascular tissue to the bare sclera, application of 0.02% mytomycin C onto the bare sclera for 5 min and transplantation of preserved human amniotic membrane and conjunctival limbal autograft. Postoperatively, all seven eyes showed rapid epithelialization on the corneal surface in 3-5 days and, on the conjunctival surface, in 10-18 days. For a mean follow-up period of 22.4+/-6.1 months, six eyes recovered deep fornices, smooth and stable ocular surface and full ocular motility without recurrence. One eye showed regrowth of fibrovascular tissue and motility restriction but less severe than before surgery. No complication was noted due to mitomycin C. Combined intraoperative mitomycin C, amniotic membrane graft and limbal conjunctival autograft are successful approaches for treating multirecurrent pterygia with severe symblepharon to restore the ocular surface integrity and prevent recurrence.

Ophthalmic Applications of Preserved Human Amniotic Membrane: A Review of Current Indications

Preserved human amniotic membrane (AM) is currently being used for a wide spectrum of ocular surface disorders. The AM has a basement membrane, which promotes epithelial cell migration and adhesion. The presence of a unique avascular stromal matrix reduces inflammation, neovascularization and fibrosis. The basic tenets of amniotic membrane transplantation (AMT) are to promote re-epithelialization, to reconstruct the ocular surface and to provide symptomatic relief from surface aberrations. AMT is a useful technique for reconstruction of surface defects resulting from removal of surface tumors and symblephara. AMT has effectively restored a stable corneal epithelium in eyes with, persistent epithelial defects and corneal ulcers. In the setting of acute ocular burns and SJS, AMT has satisfactorily reduced scarring and inflammation. AMT alone may be an effective alternative for partial limbal stem cell deficiency. However remarkable improvements in surface stability have resulted from concurrent AMT and limbal stem cell transplantation, wherein the limbal grafts are obtained from the normal fellow eye, living relative or cadaveric eye. In severe or bilateral cases, well being of the donor eye is a major concern. Currently, the most unique application of preserved human AM in ophthalmology is its use as a substrate for ex-vivo expansion of corneal and conjunctival epithelium. In this novel technique of tissue engineering, epithelial stem cells can be safely harvested and expanded on denuded AM. The resultant composite cultured tissue has been successfully transplanted to restore vision, as well as the structure and function of damaged ocular surfaces.

Amniotic membrane transplantation to reconstruct the conjunctival surface in cases of chemical burn

Purpose: To evaluate the efficiency of preserved human amniotic membrane transplantation for conjunctival surface reconstruction in cases of chemical burn. Methods: Preserved amniotic membrane transplantation was performed in 6 patients (6 eyes) having symblepharon and fornix insufficiency due to chemical burn. Amniotic membrane was sutured to the intact conjunctiva after the fibrotic tissue was excised. The fornix was reconstructed in cases having fornix insufficiency. These cases were followed up for 4–24 months. Results: During the mean follow-up period (10±7.37 months) adequate bulbar conjunctiva and fornix depth was achieved in 5 patients without recurrence and with mild fibrosis. In 1 patient who did not have a healthy conjunctiva preoperatively, conjunctival fibrosis and symblepharon recurred. Conclusions: Preserved human amniotic membrane transplantation is a good alternative treatment method for conjunctival surface reconstruction in those cases with some healthy peripheral conjunctival tissue.

Single and multilayer amniotic membrane transplantation for persistent corneal epithelial defect with and without stromal thinning and perforation

AIMSTo evaluate the efficacy of amniotic membrane transplantation (AMT) in persistent corneal epithelial defect with or without stromal thinning and corneal perforation.METHODS28 patients (28 eyes) with persistent corneal epithelial defect...

Epiphora caused by conjunctivochalasis: treatment with transplantation of preserved human amniotic membrane.

To present our experience with the use of preserved human amniotic membrane on patients with epiphora caused by conjunctivochalasis. Twelve patients, seven women and five men, ages ranging from 56 to 72 years (mean, 61 years) were referred to our Cornea Service with chronic epiphora. In all patients, no punctal ectropion was present, the lacrimal pathway was patent, and the dye disappearance test was abnormal. All patients had already undergone various therapies including multiple irrigations of the lacrimal system, antibiotic drops, steroid drops, and artificial tear drops. In all patients, conjunctivochalasis, which was not previously diagnosed, was evident on slit-lamp examination. After surgical removal of the excess conjunctiva, preserved human amniotic membrane was placed over and sutured with 10-0 nylon continuous suture to the free conjunctival edges. During the postoperative period, artificial tear drops and steroid/antibiotic drops were applied. Improvement of the epiphora was evident from the first postoperative day. After removal of the suture 10 to 15 days (mean, 12 days) after surgery, no patient complained of epiphora. The dye disappearance test was normal. During the follow-up period, which ranged from 6 to 11 months (mean, 8 months), no patient complained of epiphora and no conjunctivochalasis was detected in the area in which human amniotic membrane was transplanted. In our experience, transplantation of preserved human amniotic membrane greatly improved symptoms of epiphora caused by conjunctivochalasis. Continued education of the general ophthalmologists concerning this condition is required.

Reduction in corneal haze and apoptosis by amniotic membrane matrix in excimer laser photoablation in rabbits

Purpose To determine whether preserved human amniotic membrane can reduce corneal haze and keratocyte apoptosis induced by excimer laser photoablation in rabbit corneas. Methods Excimer photoablation was performed bilaterally in 30 rabbits with a 6.0 mm ablation zone and 120 μm depth using the VISX Star laser with the phototherapeutic keratectomy (PTK) mode. One eye was randomly covered by preserved human amniotic membrane secured with 4 interrupted 10-0 nylon sutures, and the other eye served as the control. The amniotic membranes were removed at 1 week, and corneal haze was graded with slitlamp biomicroscopy by 3 masked corneal specialists biweekly for the ensuing 12 weeks until the rabbits were killed. Another 18 rabbits were divided into 4 subgroups and received PTK alone, PTK with membrane, PTK with sham sutures, or PTK with tarsorrhaphy. All eyes were studied histologically, and 3 eyes in each group were studied by in situ terminal deoxynucleotidyl transferase deoxy-UTP-nick end labeling assay at 1, 3, and 7 days and 12 weeks, respectively. Results A consistent grading of differences in corneal haze scoring between the control corneas and the amniotic-membrane-covered corneas was noted among the 3 masked observers. Organized reticular post-PTK corneal haze peaked at 7 weeks in both groups, and the corneal haze score in the amniotic-membrane-covered group was significantly less than in the control group from 7 to 12 weeks (all P < .001). Compared to the control corneas, the amniotic-membrane-covered corneas had less inflammatory response at 1 and 3 days, less keratocyte apoptosis in the ablated anterior corneal stroma at 1, 3, and 7 days ( P < .001), and less stromal fibroblast cellularity and epithelial hyperplasia at 12 weeks. Conclusions Amniotic membrane matrix introduced at an early stage of the corneal wound healing process effectively reduced corneal haze induced by excimer laser photoablation in rabbits. Studies linking suppression of apoptosis in the acute wound-healing process with reduction of subsequent corneal scarring may have useful clinical applications.

Preserved Amniotic Membrane Transplantation for Conjunctival Surface Reconstruction.

Objective. To study the efficacy of preserved human amniotic membrane in the reconstruction of conjunctival defect created during surgical removal of conjunctival lesions or symblepharon lysis.Methods. Preserved human amniotic membrane transplantation was performed in 93 eyes of 85 patients for reconstruction of various conjunctival surface problems. The indications for surgery were (1) pterygium excision (54 eyes), (2) conjunctival tumors excision (23 eyes), lysis of symblepharon (13 eyes), and covering a scleral graft (three eyes).Results. Success was noted in 69.9% (65/93) eyes, partially success in 22.6% (21/93) eyes, and failure in 7.5% (7/93) eyes with a mean follow-up of 8.9 months (1-28 months). In pterygium, conjunctival tumor, symblepharon, and scleral graft group, the success rate in each group was 70.3%, 78.3%, 53.8%, and 66.7% respectively. No serious immediate post-operative complications or graft rejection occurred.Conclusion. Amniotic membrane transplantation can be considered an alternative treatment for difficult ocular surface problems, and is effective in promoting epithelial healing, and reducing inflammation and scarring.

Laminin-5 is a component of preserved amniotic membrane

Purpose. To determine if laminin-5 is retained in the matrix of cryopreserved human amniotic membrane tissue prepared for ocular surgeries. Methods. Amniotic membrane was solubilized in urea/SDS buffer. Constituent proteins were resolved by SDS-PAGE and laminin-5 content was determined by Western blot analysis using a panel of antibodies directed against the a3, ß3 or ?2 chains of the molecule. Human corneal epithelial cells were seeded on amniotic membrane and cultured in the presence or absence of EGF. The cell-membrane construct was examined for laminin-5 content using Western blot analysis and immunofluorescence microscopy. Results. In preserved amniotic membrane the laminin-5 a3 chain is present in both the unprocessed (190-kDa) and processed (160-kDa) forms. The ß3 chain is found in the 145-kDa form. The ?2 chain appears to be predominantly in the processed (105-kDa) form. Very little of the unprocessed form of the ?2 chain (155-kDa) could be detected using immunoblot analysis. A similar distribution of laminin-5 was also present in extracts of corneal epithelial cells cultured on amniotic membrane. Immunofluorescence analysis of cells cultured on the membrane demonstrated polarization of laminin-5 at the cell-membrane interface. Conclusions. The presence of both the unprocessed and processed forms of laminin-5 a3 and ?2 chains in preserved human amniotic membrane suggests that when used as a substrate in ocular surgeries, this membrane may be capable of promoting corneal epithelial cell motility and adhesion. Regulation of the motile or adhesive function may lie with factors secreted by the corneal epithelium that populates the membrane following surgery.

Use of nonpreserved human amniotic membrane for the reconstruction of the ocular surface.

To describe the use of nonpreserved human amniotic membrane (NP-AMT) as an alternative to preserved human amniotic membrane (AMT) for the reconstruction of the ocular surface in several diseases. NP-AMT was used in the treatment of five patients with the following diseases: noninvasive conjunctival squamous cell carcinoma, corneal persistent epithelial defect, severe alkali burn, near total limbal deficiency secondary to multiple surgeries, and ocular cicatricial pemphigoid. In some cases, a limbal autograft or allograft was employed simultaneously, sutured on top of the NP-AMT. All sutures were made with 10-0 Nylon and were removed at two weeks. Ocular surface was satisfactorily reconstructed, eyes were quiet, and patients were comfortable despite prolonged deepithelialization in some cases. There was a case of a limbal autograft ischemia--in the burned patient--that caused partial corneal conjunctivalization. Initially, the NP-AMT looks thickened but thins around the fifth day and looks similar to AMT. Results using NP-AMT are similar to those of AMT. It is a good alternative and it is easily obtained in places were AMT is not available or is too expensive to procure.

Growth factor mRNA and protein in preserved human amniotic membrane

Growth factor mRNA and protein in preserved human amniotic membrane

Growth factor mRNA and protein in preserved human amniotic membrane

Purpose. To investigate the expression of growth factor mRNA and the level of growth factor protein in preserved human amniotic membrane (AM). Methods. RT-PCR was used to examine the expression of mRNA for eight growth factors (EGF, TGF-a, KGF, HGF, bFGF, TGF-ß1, -ß2, -ß3) and two growth factor receptors (KGFR and HGFR) in human AM preserved at -80°C for one month. In addition, ELISAs were used to measure the protein concentrations of seven growth factors (EGF, TGF-a, KGF, HGF, bFGF, TGF-ß1, -ß2) in preserved human corneas and in AM both with and without amniotic epithelium. Results. RT-PCR revealed that human AM expresses mRNA for EGF, TGF-a, KGF, HGF, bFGF, TGF-ß1, -ß2, -ß3, KGFR and HGFR, while ELISAs showed that it contains EGF, TGF-a, KGF, HGF, bFGF, TGF-ß1, -ß2. AM without amniotic epithelium also contains all seven growth factors examined, however, in this tissue the protein levels of EGF, KGF, HGF and bFGF were found to be significantly lower than in native AM. Conclusions. Preserved human AM expresses mRNAs for a number of growth factors and contains several growth factor proteins that might benefit epithelialization after AM transplantation. High levels of EGF, KGF, HGF and bFGF in AM with amniotic epithelium as compared to AM without amniotic epithelium suggest an epithelial origin for these growth factors. We feel that EGF, KGF and HGF in particular might play important roles in ocular surface wound healing after AM transplantation.

Application of Preserved Human Amniotic Membrane for Corneal Surface Reconstruction.

Objective: To evaluate the efficacy of preserved human amniotic membrane transplantation for reconstruction of the corneal surface diseases.Methods: Preserved human amniotic membrane transplantations were performed in 84 eyes of 78 patients for corneal surface reconstruction. The indications were limbal stem cell deficiency from Steven-Johnson syndrome, chemical burn and herpes keratitis (27 eyes), bullous keratopathy (26 eyes), persistent epithelial defect and dellen (17 eyes), band keratopathy (11 eyes), preparing for prosthesis (1 eye), corneal ulcer (1 eye) and acute chemical burn (1 eye).Results: Success was noted in 83.3% (70/84) eyes, partial success in 13.1% (11/84) eyes, and failure in 3.6% (3/84) eyes for an average follow-up of 10.5 months (3 - 29 months). No patient developed major immediate post-operative complications.Conclusion: Amniotic membrane transplantation can reduce inflammation, promote corneal epithelial healing, and decrease irritation in corneal surface problems.

Techniques for reconstruction of the corneal surface by transplantation of preserved human amniotic membrane

Persistent defects of the ocular surface remain a therapeutic challenge, which often requires surgical intervention. Recently, transplantation of human amniotic membrane has been suggested for reconstruction of the corneal surface. We describe two techniques using cryo-preserved amniotic membrane. Transplantation of several layers of amniotic membrane can successfully restore deep corneal ulcers associated with persistent epithelial defects. When such defects are not associated with stromal ulcers, they might benefit from amniotic membrane grafts sutured to cover the entire corneal surface. Both methods may serve as valuable tools for ocular surface reconstruction, especially in the context of impaired epithelial function.

Suppression of transforming growth factor-beta isoforms, TGF-beta receptor type II, and myofibroblast differentiation in cultured human corneal and limbal fibroblasts by amniotic membrane matrix.

Down-regulation of the transforming growth factor-beta (TGF-beta) signaling system is a strategy for preventing scarring during wound healing. Human corneal and limbal fibroblasts were cultured on the stromal matrix side of preserved human amniotic membrane. The levels of TGF-beta1, beta2, and beta3 and TGF-beta type II receptor transcripts and TGF-beta1 and beta2 proteins were suppressed as early as 8 hr and more dramatically at 24 hr after contact with an amniotic membrane. This suppressive effect was accompanied by down-regulation of alpha-smooth muscle actin, EDA spliced form of fibronectin, and integrin alpha5. It persisted even when challenged by 10 ng/ml TGF-beta1. In contrast with their counterparts grown on plastic or in collagen gel, such suppression in amniotic membrane cultures remained complete after 1 week of culturing. Cells cultured on amniotic membrane showed significantly reduced [3H]-thymidine incorporation compared to cells cultured on plastic and displayed no DNA fragmentation. These results reveal a novel mechanism by which the TGF-beta signaling system, DNA synthesis, and subsequent myofibroblast differentiation can be suppressed by an amnionic membrane matrix. This action explains in part the antiscarring results of amniotic membrane transplantation used for ocular surface reconstruction, a surgical technique applicable to other subspecialties. It may also explain in part why fetal wound healing is scarless.

Amniotic Membrane Transplantation for Ocular Surface Reconstruction in Patients with Chemical and Thermal Burns

The purpose of the study is to examine the usefulness of preserved human amniotic membrane transplantation in patients with chemical and thermal burns. The study design was a nonrandomized clinical trial. Seven eyes of six patients with severe chemical (n = 5) and thermal (n = 2) burns were studied. Eyes were treated with excision of cicatricial tissues followed by a placement of amniotic membrane on the sclera. Transplantation of limbal grafts from an opposite eye (n = 4) or from donor eyes preserved at -80 degrees C (n = 2) was performed simultaneously. Reconstruction of ocular surface epithelia and visual acuity were measured. With the mean observation period of 53.3 weeks, central corneal epithelium was reconstructed successfully in all eyes. Neither amniotic membrane nor limbal grafts were rejected. A persistent epithelial defect developed in one eye, which was treated successfully by tarsorrhaphy. After surgery, the corneal epithelium showed normal arrangements on specular microscopy, and its barrier function recovered to seminormal. Corrected visual acuity markedly improved in each eye. Regenerated conjunctiva on the amniotic membrane was stable and uninflammed with minimum-to-mild scarring. Slight recurrence of conjunctivalization was noted in three eyes. However, because these eyes were stable and central cornea was clear, no further surgery was needed. Amniotic membrane transplantation promotes normal conjunctival epithelialization while suppressing fibrosis formation. The procedure, especially when performed with limbal autograft transplantation, appears to be effective for the treatment of chemical or thermal burns of the ocular surface.