Indications for and Uses of Amniotic Membrane: Operating Room

The Amniotic Membrane: Histology, Biologic Properties, and Clinical Effects

The amniotic membrane (AM) is the innermost layer of the fetal membranes and is composed of three layers: epithelium, basement membrane (BM), and stroma. The epithelium is a monolayer of cuboidal cells attached to the basal lamina by hemidesmosomes. The BM is composed of collagen IV and VII, fibronectin, and laminin 1 and 5. The loose connective tissue of the stroma confers tensile strength. ^,

Studies on human AM preserved at -80°C for 1 month revealed the presence of growth factors and cytokines that maintain the microenvironment of the stem cells (SCs) of the corneal epithelium. The BM facilitates migration of epithelial cells, reinforces adhesion of basal epithelial cells, and prevents apoptosis. Finally, a component of the stroma suppresses signaling via transforming growth factor (TGF)-β, which modulates the proliferation of normal corneal, conjunctival, and limbal fibroblasts, and reduces subconjunctival fibrosis. The stroma also contains antiinflammatory and antiangiogenic proteins and protease inhibitors that reduce stromal inflammation and ulceration.

The biologic properties of AM are derived from its structure and composition. Clinical effects of AM include (1) enhanced epithelialization; (2) inhibition of inflammation, neovascularization, and scarring; and (3) antimicrobial properties.

This chapter will focus on the different uses and techniques of implantation in the operating room of AM from a very practical perspective.

Preservation Methods of Amniotic Membrane: Tissue Bank

Commercially available AM, the PROKERA (Prokera, BIO-Tissue Inc., Doral, FL) and the PURION Process (AmbioDry2, Ambio5, and AmbioDisk; IOP Ophthalmics, Costa Mesa, CA), have already been described in detail in the previous chapter. We will focus on AM prepared and delivered by most bank tissues to be used in the operating room.

Placentas from which AM is to be harvested should be obtained by elective C-section because bacterial contamination is lower. Only placentas with negative bacterial and fungal cultures and negative serologic results in the donor should be used.

The AM is prepared and preserved in the tissue bank using the method described by Tseng et al. The placenta is cleaned under laminar-flow using a balanced, sterile saline solution containing 50 μL/mL penicillin, 50 μL/mL streptomycin, 100 μL/mL neomycin, and 2.5 μL/mL amphotericin B. The amnion is separated from the chorion and is flattened onto nitrocellulose paper with the epithelial/BM up. The AM is then stored at -80°C in sterile vials containing cryoprotective agents (Dulbecco’s modified Eagle’s medium [Life Technologies Ltd., Paisley, UK] and glycerol or dimethylsulfoxide). The amount of active biologic factors decreases with preservation time: 50% after 2 months, being almost undetectable after 18 months.

Techniques for Amniotic Membrane Implantation in The Operating Room ( Video 148.1 )

Video 148.1 How to Know the Orientation of the Amniotic Membrane (AM). In case the membrane is supplied spread on a filter paper, the stromal side usually lies down onto the surface of the paper, and the epithelial side faces up; however, once it is separated from the filter paper, the membrane usually rolls itself. Both sides of the membrane are easily differentiated using a microsponge. The microsponge softly presses and dries the AM. The epithelial side of the membrane, with or without the amniotic epithelium, is smooth compared to the stromal (chorionic) side, which appears rough and irregular. Also, the stromal side sticks to the microsponge, whereas the epithelial side does not. Jose L. Güell, Oscar Gris, Daniel Elies, Felicidad Manero, Mercè Morral.

The technique of implantation of the AM will depend on the clinical effect desired. The AM may be used as a graft, as a patch, or a combination of both.

• Graft (epithelial-side up): in cases of both epithelial and stromal defects, AM replaces the missing stromal matrix and provides a BM for cells to grow on. The AM should fill the tissue defect but not overlap its limits. Multiple layers may be used in areas of corneal melt or thinning. Epithelialization occurs over the AM, which remains trapped within the stroma until reabsorbed.

• Patch (epithelial-side down): the AM covers an epithelial defect without stromal loss and is fixed with a running suture of Nylon 10/0 to the episclera near the limbus. It is either removed, reabsorbed, or falls off. The AM protects the ocular surface from external insults, reduces inflammation, and promotes epithelialization beneath the membrane.

• Combined approach: the inner membrane is sutured with the epithelial side up as a graft. The other, usually larger, is sutured on the top of the first, with the stromal side up. Dua et al. described that overlapping the edges of the second membrane with the peritomized conjunctiva may ensure that centripetally migrating epithelium from the conjunctiva will grow on the second membrane and not on the cornea.