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Human Amniotic Membrane Transplantation
For The Treatment of Ocular Surface Disease

Brian L. Schwam, M.D.
Brian L. Schwan, M.D. is a Senior Associate Consultant,
Department of Ophthalmology, Mayo Clinic Jacksonville and
Assistant Professor of Ophthalmology, Mayo School of Medicine.
The first reported use of fetal membranes in skin transplantation was by Davis in 1910.1 In 1913, Sabella used amniotic membrane on burned and ulcerated skin surfaces and observed lack of infection, marked decrease in pain, and increased rate of re-epithelialization of traumatized skin surface.2 Others have demonstrated the use of amniotic membrane as a biological dressing for open wounds including burns and chronic ulceration of the legs.3

The first use of amniotic membrane transplantation (AMT) in ophthalmology was by De Rotth in 1940 who reported partial success in the treatment of conjunctival epithelial defects after symblepharon4 (scarring and adhesions between palpebral and bulbar conjunctiva). Sorsby and Symons in 1946 found that patients with caustic burns of the conjunctiva with corneal involvement could be treated successfully using amniotic membrane.5 Little else regarding AMT appeared in the ophthalmic literature until 1995 when Kim and Tseng used AMT for ocular surface reconstruction of severely damaged corneas in a rabbit model.6 Since that experimental study, AMT has been used for persistent corneal epithelial defects,7 neurotrophic corneal ulcers, 8,9 leaking filtering blebs after glaucoma surgery, 10 pterygium surgery 11,12 conjunctival surface reconstruction, 13 bullous keratopathy, 14,15 chemical or thermal burns, 16,17,18 ocular surface reconstruction with or without limbal stem cell grafting, 19,20 and in patients with ocular cicatricial pemphigoid or Stevens-Johnson syndrome. 21

The normal ocular surface is covered by corneal, limbal and conjunctival epithelial cells. The limbal stem cells are felt to give rise to the corneal epithelium and therefore are especially important for maintenance of a smooth, clear corneal surface. These cells, together with a stable tear film, maintain the health of the ocular surface and therefore allow for good visual acuity. Damage to these cells from certain systemic inflammatory diseases or primary ocular diseases or trauma (Tables 1 and 2) may lead to state of limbal stem cell deficiency. The result is breakdown of the ocular surface and corneal epithelial defects that may become chronic if the normal epithelialization process fails. Conjunctival epithelium may replace corneal epithelium resulting in loss of corneal transparency. Chronic inflammation may then occur characterized by neovascularization, corneal scarring and opacification, corneal thinning, and possible corneal perforation, all of which may lead to loss of visual acuity.
 

Table 1: Systemic Diseases Which
May Cause Ocular Surface Damage:

Table 2: Ocular Diseases/Injuries Which
May Cause Ocular Surface Damage:

  • Stevens-Johnson syndrome
  • Ocular cicatricial pemphigoid
  • Rheumatoid arthritis
  • Sjogren's syndrome
  • Toxic epidermal necrolysis
  • Herpes zoster ophthalmicus
  • Surgically induced neurotrophic keratitis
  • Chemical or thermal injuries
  • Keratoconjunctivitis sicca
  • Severe microbial infection
  • Multiple ocular surgeries

Treatment of chronic epithelial defect includes limiting or removing its causes. Failure of the normal epithelialization process may be exacerbated by the friction generated from the mechanical movement of the eyelid over the chronic defect, especially when the palpebral conjunctiva is abnormal. The use of frequent topical lubrication with preservative-free artificial tears is advised. Removal of ocular medications as tolerated, especially those with preservative which may be toxic to the corneal epithelium, is recommended as well. Punctal occlusion may be beneficial. Therapeutic contact lenses have been shown in some cases to be effective therapy for persistent epithelial defect but carry the increased risk of infection. More invasive surgical therapies include temporary or permanent tarsorrhaphy (sutures that are used to close the palpebral fissure at least partially) but its efficacy is limited by the ability of the corneal wound to heal. Use of human amniotic membrane for transplantation may be an alternative or adjunctive therapy.

Human amniotic membrane is derived from the fetal membranes which consist of the inner amniotic membrane made of single layer of amnion cells fixed to collagen-rich mesenchyme 6 to 8 cells thick loosely attached to chorion. It is composed of three layers: a single epithelial layer, thick basement membrane, and avascular stroma. A detailed discussion of the embryology and histology can be found elsewhere.22 Human amniotic membrane has been shown to contain collagen types III and V. It also contains collagen types IV and VII similar to corneal epithelial basement membrane as well as fibronectin and laminin.23, 24 Additionally, it contains fibroblast and other growth factors. 25, 26

Human amniotic membrane is believed to be nonimmunogenic. Antibodies or cell-mediated immune response to amniotic membrane have not been demonstrated, suggesting low antigenicity. Therefore, the use of systemic immunosuppressives in AMT is not required. In contrast, chorion provokes neovascularization and typical rejection phenomenon.22 The amnion surface epithelial cells do not express HLA A, B, C, or DR or beta2-microglobulin.27, 28 However, others have found that amnion epithelial cells do express HLA class I molecules under certain conditions.29 Gabler, et al. used repeated amniotic membrane transplantation to heal a sterile, non-healing ulcer related to rheumatoid arthritis.30 They found sterile hypopyon after the second and third procedures and proposed this was due to an immunologic reaction, possibly caused by sensitization to unknown antigens as the amniotic membrane tissue used each time was from the same donor placenta. The authors therefore recommended using amniotic membrane from different donor placenta in cases of repeated amniotic membrane transplantation. Others have found possible immunologic reactions after repeated exposure using amniotic membrane for non-ophthalmic indications. 31

To date, most clinical experiences with human amniotic membrane transplantation have been with tissue preserved using the method described by Tseng and colleagues. 19 The fragments of tissue are preserved at 80 degrees Celsius in a part glycerol solution and are flattened onto nitrocellulose paper with the epithelium side up. Kruse, et al. suggest that amniotic membrane epithelial cells are not viable after preservation and it is unclear whether the growth factors survive cryopreservation.32 Several authors have described the use of fresh or non-preserved human amniotic membrane for transplantation. Mejia, et al. successfully used amniotic membrane for symptomatic management of bullous keratopathy in eyes with poor visual potential.15 Ucakhan et al evaluated safety and efficacy of non-preserved AMT with or without limbal autograft transplantation in acute and chronic chemical eye injuries.18 They found that AMT promoted epithelial healing, reduced inflammation, increased comfort, and decreased severity of vascularization. They did not find any infectious, inflammatory, or toxic reactions. Although the use of fresh tissue may be associated with a higher risk of blood-borne diseases33, the authors felt non-preserved tissue may be important for ocular surface reconstruction in developing countries where cost and availability may prohibit use of preserved tissue.

The exact mechanisms by which amniotic membrane delivers its beneficial effects on the ocular surface are still being investigated. Amniotic membrane modulates levels of cytokines and growth factors and has also been shown to have unique properties, including pain reducing, fibrosis suppressing, antibacterial, and wound protecting. 22, 34-36 It is unclear whether amniotic membrane promotes limbal stem cell proliferation. 16, 19 Many studies have shown that amniotic membrane promotes epithelialization of the corneal surface. Its structure and the presence of growth factors favor the growth of epithelium and thus help to restore the ocular surface. 25 The mechanism by which it facilitates epithelial healing may vary depending on the technique used at the time of amniotic membrane transplant surgery: inlay vs. overlay technique. AM may also be used in a "filling" technique.37
 

Figure 1. Amniotic membrane grafting:
Inlay technique
 

 The inlay technique (Figure 1) involves placement of the amniotic membrane graft into the corneal ulcer secured into place by interrupted sutures without extending beyond the edge of the epithelial defect. The amniotic membrane thus acts as a basement membrane to which epithelialization may take place over it from the surrounding epithelium. Therefore, persistent corneal defects or ulceration may be treated prior to 7, 9 or after 37 perforation. Mejia, et al. successfully used larger AMT for treatment of symptomatic bullous keratopathy by leaving peripheral cornea uncovered to promote epithelialization over the graft.15 They suggested that AM used in this fashion may be an alternative to conjunctival flap in corneas with poor visual potential. In using this technique, however, the amniotic membrane graft becomes trapped under the healed corneal epithelium and may limit corneal transparency for several months or more affecting vision.

 

Figure 2. Amniotic membrane grafting:
Overlay technique

 In the overlay technique (Figure 2), the entire corneal surface including the limbus is covered with the amniotic membrane graft. Here the amniotic membrane functions primarily as a biological contact lens.38 The graft protects regenerating epithelium from the frictional forces of the eyelid and palpebral conjunctiva while at the same time appears to allow adequate oxygen permeability and moisture to the epithelium. Corneal transparency remains when the graft eventually detaches or dissolves. This method has been used successfully in cases of stem cell deficiency of various causes 19 and in cases of persistent epithelial defect unresponsive to medical therapy and surgically induced epithelial defects.39 Gris, et al. propose that this method may be a safe alternative that some patients may prefer over tarsorrhaphy when medical treatment has failed.39 Both the inlay and overlay techniques may be used together.

Limbal stem cell transplantation using allograft or autograft may be used in conjunction with amniotic membrane transplantation.20 While amniotic membrane transplantation alone may be best in patients with partial limbal stem cell deficiency, the combined use of these procedures is necessary in patients with total limbal stem cell deficiency.18,19

Letko, et al. compared inlay versus overlay amniotic membrane grafting techniques in ability to heal persistent corneal epithelial defects and found no difference in terms of healing time and recurrence rate.38 However, they did note lower success rates in healing epithelial defects than previously reported and higher incidence of recurrences of epithelial defects. They concluded that amniotic membrane transplants for persistent epithelial defects should not be used as first or second-line therapy and should be reserved for use when all other measures have failed.

Amniotic membrane may be used as a "filler" for closing corneal perforations.37, 40 This method may be combined with the above techniques. Hanada, et al. used multilayered amniotic membrane transplantation to heal ulceration of the cornea and sclera of various causes.37 In their study, amniotic membrane was used as a "filler" (providing a substitute for collagens), graft (providing basement membrane for epithelialization), and patch (providing protection for the wound).

Two cases of using AMT combined with a sealant for small corneal perforations have been reported. Su reported the use of cyanoacrylate glue overlying amniotic membrane placed into the anterior chamber in a patient with corneal melting and perforation following alkali injury.40 The glue subsequently dislodged with the perforation healed below. Duchesne used human fibrin glue combined with AMT to seal corneal perforations approximately 2mm in diameter.41 The authors proposed that this method might be a good alternative to delay penetrating keratoplasty for treating corneal perforations when acute graft rejection risk is high.40, 41

Few intraoperative or postoperative complications have been observed. However, there has been variation in the reported success rates of healing persistent epithelial defects using AMT. Lee and Tseng reported an initial success rate of 90% (10/11).7 Letko, et al. using both inlay and overlay techniques reported a success rate of only 70% (21/30) of eyes after the first AMT.38 They also reported a higher rate (29%) of recurrence of epithelial defects. Gris, et al. using the overlay technique found complete epithelialization in 3 of 4 cases when the amniotic membrane graft remained in place for 2 or more weeks but in no cases when the graft remained in place for less than 1 week.39 Early detachment of the amniotic membrane graft occurred despite multiple fixation sutures. Hanada, et al. noted a success rate of 73% (8 of 11 eyes) in eyes with severe ulceration of the cornea and/or sclera and concluded that eyes with total limbal dysfunction or autoimmune disorder may not be cured by AMT alone.37 Using the overlay technique, Azuara-Blanco, et al. reported that AMT failed to stabilize the cornea in 5 eyes with impending or recent corneal perforation.42 It can be postulated that differences in surgical technique, causes of persistent epithelial defects, severity of ocular surface disease, and small numbers of patients in these and other studies may explain these differences in outcomes.

The ophthalmic uses of human amniotic membrane for transplantation are many and its rediscovery has greatly improved our ability to treat debilitating ocular surface disease. Its rebirth for ophthalmic use has been likened to the in vivo studies and new technology used for the development of a method to mass-produce penicillin during World War II.43 While results from many studies are encouraging, others show less enthusiastic results. It does appear that amniotic membrane transplantation is effective in healing persistent epithelial defects and sterile ulceration when medical therapy fails. Future studies directly comparing AMT to other methods of treatment of persistent epithelial defects would help to better define the role of amniotic membrane transplantation in ocular surface disease and perhaps further elucidate the mechanisms by which this therapy seems to work.

REFERENCES

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  8. Chen HJ, Pires RTF, Tseng SCG. Amniotic membrane transplantation for severe neurotrophic ulcers. Br J Ophthalmol. 2000;84:826-833.
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  37. Hanada K, Shimazaki J, Shimmura S, et al. Multilayered amniotic membrane transplantation for severe ulceration of the cornea and sclera. Am J Ophthalmol. 2001;131:324-331.
  38. Letko E, Stechschulte SU, Kenyon KR, et al. Amniotic membrane inlay and overlay grafting for corneal epithelial defects and stromal ulcers. Arch Ophthalmol. 2001;119:659-663.
  39. Gris O, del Campo Z, Wolley-Dod C et al. Amniotic membrane implantation as a therapeutic contact lens for the treatment of epithelial disorders. Cornea 2002;21:22-27.
  40. Su CY, Lin CP. Combined use of an amniotic membrane and tissue adhesive in treating corneal perforation: A case report. Ophthalmic Surg Lasers. 2000;31:151-154.
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  42. Azuara-Blanco A, Pillai CT, Dua HS. Amniotic membrane transplantation for ocular surface reconstruction. Br J Ophthalmol. 1999;83:399-402.
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August-September 2002 / Jacksonville Medicine

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