Issue: January 15, 2006

ByLauren Wolkoff

January 15, 2006

11 min read

Save

This article is more than 5 years old. Information may no longer be current.

Cryopreserved amniotic membrane transplantation offers wealth of healing properties

This “relatively simple” procedure continues to grow in popularity as studies show the efficacy of treatment.

Issue: January 15, 2006

ByLauren Wolkoff

Researchers are tapping into the potential for the application of human amniotic membrane to ocular surface disorders.

Scheffer C.G. Tseng, MD, PhD [photo]
Scheffer C.G. Tseng

Today, cryopreserved amniotic membrane is used – albeit not yet routinely – to address a variety of corneal, epithelial and conjunctival disorders, including neurotrophic ulcers, persistent epithelial defects, microbial keratitis, band keratopathy and chemical burns. It has also been used to speed wound healing and promote the regeneration of stem cells in unhealthy corneas, as well as to prevent scarring after a variety of procedures, including PRK.

While this field of research is not a new one – dating back more than a half century to 1940 – it was not until the mid-1990s when researchers brought the amniotic membrane into the mainstream of corneal research with studies using non-cryopreserved human membrane as a xenograft in rabbit eyes.

Numerous applications

With many exciting applications in full swing, it is impossible to pick just one reason to celebrate the advent of amniotic membrane transplantation (AMT), according to Scheffer C.G. Tseng, MD, PhD, a pioneer researcher in the field who helped resurrect its study for ocular use.

“Amniotic membrane can be used for so many different things – and that’s what is so exciting about it,” Dr. Tseng said in an interview with Ocular Surgery News. “It has truly revolutionized what we used to do.”

For example, in certain cases, AMT can avert the need for a corneal transplant or delay the transplant to a less urgent time when conditions are more favorable for success.

Dr. Tseng said he now routinely uses amniotic membrane grafts to begin to heal or regenerate traumatized or diseased corneas before proceeding with further treatment.

“It has become my first-line approach to quiet the eye and restore the [ocular] environment,” he said. The hope is to eliminate the need for more invasive surgery or at least create less antagonistic conditions for surgery.

Dr. Tseng is one of the developers of AmnioGraft, the trade name for cryopreserved amniotic membrane used specifically for ocular surface reconstruction. He is director of the research and development department of TissueTech Inc., which holds the patent for AmnioGraft. AmnioGraft is procured, processed and distributed by Bio-Tissue Inc., which licenses the patent and is a subsidiary of Tissue Tech.

AmnioGraft is the only amniotic membrane approved by the Food and Drug Administration for ocular surface reconstruction. Dr. Tseng noted that the FDA has approved the cryopreserved tissue to deliver anti-inflammatory, anti-scarring and anti-angiogenic actions and to promote epithelial healing.

He said that the cryopreserved AmnioGraft is also useful to “give an extra edge of security” to a healthy donor eye from which a limbal graft is removed for transplantation to the fellow eye. Dr. Tseng said that his research has shown that applying amniotic membrane speeds up the cell regeneration process.

Mechanism of action

Amniotic membrane consists of three layers: the epithelium, basement membrane and stroma. Its efficacy in ocular surgery stems from its ability to promote epithelialization, inhibit fibrosis, protect against inflammation and angiogenesis, decrease the risk of infection and conduct fluids, according to a review of existing research published by Merle Fernandes, MS, and co-authors in the August 2005 issue of Cornea.

In an editorial in the same issue of Cornea, Kenneth R. Kenyon, MD, wrote that “the remarkable mix of growth factors, neurotrophins and cytokines enmeshed within a cornea-like basement membrane-coated extracellular matrix offers a potent ‘soup’ plus ersatz stroma with anti-inflammatory, anti-angiogenic and anti-microbial activities in the near absence of immunogenicity.”

“It is this multipotency that facilitates amnion’s multiplicity of clinical applications,” Dr. Kenyon said.

Yet Fernandes and colleagues point out that “most of the reports described in literature are case series, either retrospective or prospective, and there are not any randomized, controlled trials comparing this modality of treatment with the existing ones. Hence, it is difficult to draw conclusions based on existing knowledge.”

Worldwide research

AMT has perhaps demonstrated the greatest benefit in repair of epithelial defects following chemical injury and for chronic persistent epithelial defects requiring an amniotic membrane overlay or inlay, according to Dr. Kenyon.

Dr. Kenyon has experimented with AMT for repairing persistent epithelial defects, including neurotrophic keratitis. He said that performing keratoplasty alone under these circumstances might lead to surface failure.

In his editorial, Dr. Kenyon noted that he attained a 90% success rate by combining penetrating or lamellar keratoplasty with a temporary AMT graft and minimal lateral tarsorrhaphy.

Researchers at the Tokyo Dental College in Chiba, Japan, have been studying the use of amniotic membrane for ocular surface disorders such as corneal ulcer, persistent epithelial problems and recurrent pterygium for about 10 years, according to Jun Shimazaki, MD.

Much of their research has focused on using amniotic membrane as a substrate for cultivated epithelial sheets. They have used the technique to treat Stevens-Johnson syndrome, ocular cicatricial pemphigoid, chemical and thermal burns and other severe ocular surface disorders.

An eye with band keratopathy and persistent corneal epithelial defect (a, b). Following superficial keratectomy, AmnioGraft was adhered to the cornea by fibrin glue and protected by ProKera (not shown). The eye was healed with a stable surface and little scarring 75 days later (c, d).

Images: Tseng SCG

“The ocular surface is the area where regenerative medicine has been advanced,” Dr. Shimazaki said. “Transplantation of the cultivated epithelial sheet is one of such applications, and the method has already more than 5 years’ history for clinical use.”

For cell sources, Dr. Shimazaki and his colleagues use both limbal epithelium – autologous, allogenic or from a living relative – and autologous oral epithelium.

Another group of Japanese researchers, led by Shigeru Kinoshita, MD, PhD, of the Kyoto Prefectural University of Medicine, has also conducted research on using cultivated epithelial stem cell sheets.

Dr. Kinoshita presented an update on his group’s research at the European Society of Cataract and Refractive Surgeons meeting in Lisbon. The presentation focused on using cultivated stem cells to repair inflamed tissues in Stevens-Johnson syndrome or following chemical trauma. He described how the cells are cultivated on a layer of amniotic membrane for up to 2 weeks and then cut into grafts using a 19-mm trephine.

According to Dr. Kinoshita, ophthalmologists might be able to avoid limbal transplantation failure by using cultivated stem cell sheets. He said that applying cultivated surgical grafts at the acutely inflamed stage of corneal damage may help cure surface damage in the underlying corneal stroma.

Maria Teresa Rodriguez-Ares, MD, and colleagues at the University Hospital in Santiago de Compostela, Spain, published a study in Cornea in 2004 showing that corneal perforations less than 1.5 mm in diameter were treated successfully with multilayer amniotic membrane transplantation.

They used two layers of amniotic membrane for corneas with microperforations and three to four layers for corneas with larger perforations. The grafts were trimmed to the fit the ulcer size and sutured into place. The procedure was deemed successful in 73% of the treated eyes, with the failed cases involving perforations larger than 3 mm in diameter.

Dr. Rodriguez-Ares’ study results on multilayered AMT are supported by other studies, including a 2001 study published by Dr. Shimazaki’s research group in the American Journal of Ophthalmology.

Nonpreserved amniotic membrane

Nonpreserved amnion has also been used experimentally, though it has a much shorter shelf life than cryopreserved amnion and has not been approved for clinical use by the Food and Drug Administration.



A patient suffered from corneal perforation (2 mm) and flat chamber for 2 weeks (a). AmnioGraft was glued and sutured, and ProKera was inserted with a sutured tarsorrhaphy (b). Corneal thickness (arrow) and anterior chamber depth were restored 5 weeks later (c,d).

Kaan Gündüz, MD, and colleagues at Ankara University in Turkey reported in the April 2005 issue of Eye on their experience using nonpreserved AM in ocular surface reconstruction after excision of extensive ocular surface neoplasia.

They found that 10 eyes of 10 patients with extensive ocular surface neoplasia achieved a satisfactory result with a wet, stable conjunctiva and rapid healing after tumor excision followed by placement of AM. The investigators observed no inflammation or rejection reaction to the nonpreserved AM.

Dr. Gündüz and co-authors speculate that the potential advantages of nonpreserved AMT include increased potency because of the effect of growth factors and cytokines. In contrast, cryopreserved amniotic membrane contains no or few viable cells.

Potential drawbacks include the risk of HIV infection, despite seronegativity of the donor when the membrane was harvested.

“Although rare, a risk exists that the donor could become infected shortly before harvesting, which could not be elicited in the serologic studies performed before surgery,” Dr. Gündüz and colleagues said.

To prevent the transmission of disease, extreme care must be taken in the processing of nonpreserved amniotic membrane, they noted.

It should be noted that the FDA has not approved AMT to be performed with live cells.

AMT for recurrent pterygium

Among its other properties, amniotic membrane can suppress normal conjunctival and pterygium body fibroblasts, numerous studies have shown. Dr. Shimazaki’s group has also published studies evaluating AMT techniques for recurrent pterygium, as well as pterygium associated with symblepharon.

In the literature review published by Fernandes and colleagues, AMT was recommended as first-line management for primary pterygium, and in particular for double-headed pterygium to cover a large conjunctival defect.

However, Dr. Kenyon noted that AMT appears to be only moderately advantageous for advanced and recurrent pterygia.

While agreeing that amniotic membrane can be useful to cover exposed sclera when a particularly large dissection is made, he added that the conventional conjunctival autograft techniques used for the past 25 years are also adequate.

“I am certain that conjunctival grafts as large as 15 by 15 mm can be harvested without risk of graft ischemia or compromise of the donor site,” Dr. Kenyon said in his Cornea editorial.

“The rationale that [amniotic membrane] spares conjunctiva in patients who might potentially require glaucoma filtration surgery is tenuous at best,” he said.

For primary pterygia, Dr. Kenyon said, conjunctival autograft transplantation remains “the procedure of choice against which all alternatives must be measured.” He said the 95% rate of this approach is better than that with current AMT techniques, and that operating times are the same.

Dr. Tseng noted that he uses Amnio-Graft for primary and recurrent pterygium together with intraoperative application of mitomycin C. “I find this approach is far better than conjunctival autograft,” he said.

Limitations

Among the limitations of AMT is that it will not work in conditions of extreme dry eye or if the host tissue cells are damaged to the point where the eye cannot blink, close or tear, according to Dr. Tseng. Other limitations include severe inflammation and ischemia if not corrected.

However, these issues can be addressed with adjunctive measures, then AMT can be successful, he added.

Dr. Kenyon noted that extensive limbal stem cell deficits must be first managed by limbal auto- or allograft, sometimes accompanied by amniotic membrane overlay.



A patient suffered from an acute chemical burn presenting with pain and a large corneal limbal epithelial defect (a, b). ProKera was inserted the next day (c), and the pain was markedly reduced. Healing was completed in three days (d).

He also pointed out that AMT should not be considered in cases when microbial keratitis is present, unless it is needed urgently to prevent corneal perforation. In these cases, the AMT should ideally be postponed until the infection has been controlled, he said.

As for the availability of amniotic membrane – in light of the potential for increased demand in the near future – Dr. Kenyon said he does not foresee major problems.

In an e-mail interview, Dr. Kenyon said that procuring placentas by cesarean section is a straightforward process, “albeit somewhat tedious with respect to testing for transmittable diseases.”

“Given the ability of a single placenta to provide amnion for perhaps a hundred or more individual procedures, I would not expect a donor demand limitation as we have always experienced, for example, with corneas,” Dr. Kenyon told Ocular Surgery News. “Also, as there are no significant antigenicity concerns, any amnion will suffice for application.”

Current and future innovations

One recent innovation by Dr. Tseng is the development of a sutureless amniotic graft, known as ProKera (also from Bio-Tissue). This device consists of a circular piece of AmnioGraft in a dual ring system that conforms to the cornea much like a contact lens.

ProKera is a class 2 medical device that was approved by the FDA in December 2003. It has been approved for up to 8 weeks of wear, but Dr. Tseng said patients typically do not need it for longer than 3 weeks.

It is used as a temporary overlay in indications such as acute chemical or thermal burns, acute Stevens-Johnson syndrome and chronic recalcitrant keratitis caused by herpes zoster ophthalmicus, herpes simplex virus or vernal keratitis.

It can also be used to resolve persistent or recurrent epithelial defects, to reduce complications of high-risk corneal grafts, or to prevent rubbing from the lid or lashes in conjunction with socket or fornix reconstruction, according to Dr. Tseng.

Most literature on AMT focuses on cryopreserved membrane, but a dehydrated form exists as well. Dried amniotic membrane grafts, commercially known as AmbioDry, are available from Okto Ophtho Inc., of Costa Mesa, Calif. This dry form is not approved by the FDA as a surgical graft.

Research is limited on the effectiveness of dehydrated membrane, and no head-to-head studies exist comparing the cryopreserved to dehydrated forms, Dr. Kenyon said.

For Your Information:

Scheffer C.G. Tseng, MD, PhD, is the director of the Ocular Surface Center in Miami, Fla. He can be reached at 7000 S.W. 97th Ave., Suite 213, Miami, FL 33173; 305-274-1299; fax: 305-274-1297; e-mail: stseng@ocularsurface.com. Dr. Tseng has a financial interest in TissueTech Inc., which owns the U.S. patents for AmnioGraft.
Merle Fernandes, MS, can be reached at the L.V. Prasad Eye Institute, L.V. Prasad Maarg, Banjara Hills, Hyderabad 500 034, India; e-mail: fernandes@lvpei.org.
Kenneth R. Kenyon, MD, is OSN Cornea/External Disease Section Editor. He can be reached at Eye Health Vision Centers, 51 State Road, North Dartmouth, MA 02747; 508-994-1400; fax: 508-992-7701; e-mail: kenrkenyon@cs.com. Dr. Kenyon has no direct financial interest in the products mentioned in this article, nor is he a paid consultant for any companies mentioned.
Jun Shimazaki, MD, is associate professor of ophthalmology at the Tokyo Dental College in Chiba, Japan. He can be reached at Department of Ophthalmology, 5-11-13 Sugano Ishikawa 272-8513, Japan; 81-47-322-0151; fax: 81-47-325-4456; jun@eyebank.or.jp. Dr. Shimazaki has no direct financial interest in the products mentioned in this article, nor is he a paid consultant for any companies mentioned.
Shigeru Kinoshita, MD, PhD, can be reached at 465 Kajii-cyo, Agaru, Hirokoji, Kawaramachi, Kamigyo-ku, Kyoto 602-0841, Japan; 81-75-251-5577; fax: 81-75-251-5663; e-mail: skinoshi@ophth.kpu-m.ac.jp.
Maria Teresa Rodriguez-Ares, MD, can be reached in the Anterior Segment Unit, Department of Ophthalmology, Hospital de Conxo-Complejo Hospitalario Universitario de Santiago, Universidad de Compostela, Spain; e-mail: trares@usc.es.
Kaan Gündüz, MD, can be reached at Mesa Koza Plaza 3, Blok 24/18, GOP 06700, Ankara, Turkey; 90-532-6337997; fax: 90-312-2291812; e-mail: eyemd@ada.net.tr.

References:

Fernandes M, Sridhar M, et al. Amniotic Membrane Transplantation for Ocular Surface Reconstruction. Cornea. 2005;24(6):639-642.
Gündüz K, Uçakhan Ö, et al. Nonpreserved human amniotic membrane transplantation for conjunctival reconstruction after excision of extensive ocular surface neoplasia. Eye [advance online publication]. April 29, 2005.
Hanada K, Shimazaki J, et al. Multilayered amniotic membrane transplantation for severe ulceration of the cornea and sclera. Am J Ophthalmol. 2001;131(3):324-331.
Kenyon K. Amniotic membrane: Mother’s own remedy for ocular surface disease. Cornea. 2005;24(6):639-642.
Rodrigues-Ares MT, Tourino R, et al. Multilayer amniotic membrane translplantation in the treatment of corneal perforations. Cornea. 2004;23(6):577-583.
Bio-Tissue Inc., maker of AmnioGraft and ProKera, can be reached at 7000 SW 97th Ave, Suite 211, Miami, FL 33173; 305-412-4430; Fax: 305-412-4429; Web site: www.biotissue.com.
OktoOphtho Inc., maker of AmbioDry, can be reached at 3184-B Airway Ave., Costa Mesa, CA 92626, 1-800-658-6411; Web site: www.oktoophtho.com.
BioTeck, maker of AMX, can be reached at Via E. Fermi, 49, 36057 Arcugnano ; +39-0444-289366; Fax: +39-0444-285068; Web site: www.bioteck.com.
Lauren Wolkoff is Managing Editor of OSN Latin America and Japan Editions.