Amniotic Membrane Allografts in the Outpatient Wound Clinic: Current Practice Guidelines & Modalities

Wound care is a challenging and ever-changing field of medicine. Chronic wounds in particular are responsible for a large portion of time, effort, and cost in the medical community. A newly released economic evaluation of the impact, cost, and Medicare policy implications of nonhealing wounds determined that almost 15% of Medicare beneficiaries (8.2 million) were treated for a type of wound (surgical, arterial, venous, traumatic, pressure, or diabetic foot ulcer [DFU]) or wound-related infection in the 2014 calendar year.¹ The authors also noted that total Medicare spending estimates for all wound types was between $28.1 and $96.8 billion. The second most expensive wound type was the DFU,¹ and with the constant rise in the number of patients living with diabetes, DFUs are becoming more prevalent and are believed to affect roughly 25% of this patient population.^2-4 These wounds also have a high propensity to become chronic due to many factors, including neuropathy, hyperglycemia, infection, and other comorbid conditions.⁵ In their highly influential study, Sheehan et al defined chronic wounds as those that do not progress in a timely manner through the four phases of wound healing — hemostasis, inflammatory, proliferative, and remodeling — and determined progression of healing at four weeks to be a significant predictor of complete healing at three months.⁶ This research has had significant impacts on current treatment protocols as well as the ability to approve advanced treatment modalities with insurance providers. 

This article will focus on the discussion of amniotic membrane allografts (AMAs) as one of the many varied advanced treatment modalities available for providers to utilize in the outpatient wound care market today.

HISTORY & ANATOMICAL REVIEW

AMAs have been in use since the early 20th century for the treatment of burns, ulcers, traumatic wounds, surgical incisions, and ophthalmic conditions across an array of medical specialties.⁷ One area in which AMAs have gained significant traction is for their use in the healing of chronic ulcers. With the emergence of new preservation techniques and advancements in research, AMAs have become more readily used in the medical community, especially in the office and wound care center settings.^8-10 This has also expanded usage of the allografts to physicians who practice in nonsurgical specialties.

The human placenta is composed of two layers: the amnion and the chorion. The amniotic membrane is composed of five layers and is an avascular structure containing no lymph vessels, nerves, or muscles.^11,12 It serves to regulate metabolic activities via growth factors and cytokines, including epidermal growth factor, keratinocyte growth factor, transforming growth factor beta, vascular endothelial growth factor, and platelet-derived growth factor. These growth factors aid in activating cellular proliferation, differentiation, and cell migration.⁹ In a chronic wound, the cascade of wound healing becomes stalled in the inflammatory stage. This decreases growth factor production and creates an imbalance in enzymes, favoring degradative versus protective enzymes, which leads to breakdown of the extracellular matrix.^5,9 This is where AMAs can play a key role: overcoming the interruption in the complex cascade of wound healing. The amniotic membrane provides the key growth factors necessary to facilitate tissue growth and healing, and to overcome the stalled cycle of a chronic wound. It also has antimicrobial and anti-inflammatory properties.^13,14 The chorion layer is composed of three layers and is four times the thickness of the amniotic membrane. The chorion also contains growth factors, cytokines, and regulatory factors similar to the amnion. As such, the combination of the amnion/chorion layer contains five times the growth factors of the single amnion layer.¹⁵ The chorion is an avascular layer of dense collagen fibers, proteoglycans, and elastic fibers.¹⁴ Currently, there are AMAs on the market that contain either solely the amnion layer or both the amnion and chorion layers. With the influx of AMA products on the market, it is important to understand the different attributes of each product. The main distinguishing factors between the allograft products are the processing techniques and inclusion of amnion and/or chorion in the grafts. Processing techniques are very important in the preservation of the structural and biochemical composition and the maintenance of signaling molecules in the tissue.¹⁰ (See Table.)  

PROCESSING TECHNIQUES

As noted in the Table, each AMA features a patented processing technique for the production of the grafts. (For a listing of additional amniotic products and descriptions, see the sidebar in the online version of this article). Each technique is slightly different and aims to keep the most viable cells and growth factors within the tissues while creating a product that is easy to handle and store in an outpatient setting.¹⁰ Epifix is a product that has been on the market for many years and is heavily researched. The product is available in many sizes and is rehydrated with sterile saline. Additionally, it does not require suturing or stapling techniques to keep the graft in place. Most of the products on the market today are dehydrated or freeze-dried, allowing for a five-year shelf life. This has made AMAs readily accessible to a larger population of patients, including those being treated in private offices and wound care centers.⁸ There is one product on the market employing a processing technique so unique that it qualifies as the only fresh AMA: Affinity. The product is packaged in a small fluid-filled container that is shipped in a temperature-controlled environment and must be maintained at refrigerated temperature (1-10°C) prior to use. The graft floats within the liquid substance and has the consistency of a thin membrane, which is sticky on the stromal side. The novel processing technique and the makeup of the liquid in which the graft is stored remains undisclosed, protected information. This easy-to-apply graft contains the amnion layer and is said to maintain the most active growth factors and cytokines (as it is not dehydrated and does not require rehydration). Affinity has a 42-day shelf life from the date of processing. NuShield is a dehydrated AMA currently available in multiple sizes, including a 1.6 cm disc up to a 6-x-6 cm piece. NuShield comes ready to use or can be hydrated prior to application. 

Revita is one of the newer products on the market. This graft is freeze-dried; employs its own novel processing technique; and preserves the amnion, chorion, and the intermediate layers of tissue. The addition of the intermediate layer is important, as it adds collagen, hyaluronic acid, growth factors, glycosaminoglycans, and proteoglycans. The graft substance is thicker than many of its current peers, which sets it apart from other products.

FDA REGULATION PROTOCOL

AMAs are legally categorized as human cell, tissue, and cellular and tissue-based products (HCT/Ps) and are regulated under section 361 of the Public Health Service Act. Section 1271.10(a) outlines the requirements that products need to meet to be recognized solely as a “human cellular therapy product,” which allows for marketing to be conducted without prior approval or clearance from the U.S. Food & Drug Administration (FDA). The requirements are:

• The HCT/P is minimally manipulated.
• The HCT/P is intended for homologous use only, as reflected by the labeling, advertising, or other indications of the manufacturer’s objective intent. 
• The manufacture of the HCT/P does not involve the combination of the cells or tissues with another article, except water; crystalloids; or a sterilizing, preserving, or storage agent, (provided that the addition of water, crystalloids, or the sterilizing, preserving, or storing agent does not raise new clinical safety concerns with respect to the HCT/P); and either:

1. The HCT/P does not have a systemic effect and is not dependent upon the metabolic activity of living cells for its primary function;
2. The HCT/P has a systemic effect or is dependent upon the metabolic activity of living cells for its primary function, and is for autologous use, allogenic use in a first or second-degree relative, or reproductive use.¹⁶

HCT/Ps are also required to comply with FDA regulations, including donor screening and testing, current good tissue practice, adverse reaction reporting, establishment registration, and HCT/P deviation reporting and labelling. The tissue undergoes rigorous testing for communicable diseases at FDA-approved laboratories for donor testing. Infectious disease testing includes a minimum of human T-cell leukemia-lymphoma virus, HIV, cytomegalovirus, syphilis, hepatitis B and C, and West Nile virus.¹⁷ At this time, FDA approval of AMAs encompasses sheet grafts for use as an overlay (for example, in wound care); however, there is continued research for use of these products in micronized form in injection therapy. This would expand use across more specialties, allowing for the treatment of tendons, ligaments, and joint surfaces. The FDA has not approved the injectable version through HCT/P regulations, as it does not meet the criteria of minimal manipulation or homologous use. Although injection therapy is being used by physicians and is available for off-label use, it is not widely recognized by insurance companies, which has limited the reimbursement. Additional research is necessary to bring the micronized injectable version of AMA into practice.

SUMMARY

Chronic wounds pose a significant challenge to our healthcare system in terms of physician care and cost. The need for novel advanced treatments is apparent. AMAs are a viable option, especially with new preservation techniques that make them available across a wider variety of clinical settings and to surgical and nonsurgical specialties. Research continues to evolve, and FDA regulations and insurance reimbursement slowly will follow. 

Mallory Przybylski is board certified by the American Board of Podiatric Medicine and has a master’s degree in wound healing and tissue repair from Cardiff University. She is co-owner of Louisiana Foot and Ankle Specialists LLC, Lake Charles, LA. She does not disclose any financial conflicts of interest.

References 

1. Nussbaum SR, Carter MJ, Fife CE, et al. An economic evaluation of the impact, cost, and medicare policy implications of chronic nonhealing wounds. Value Health. 2018;21(1):27-32.

2. Zelen CM, Serena TE, Denoziere G, Fetterolf DE. A prospective randomised comparative parallel study of amniotic membrane wound graft in the management of diabetic foot ulcers. Int Wound J. 2013;10(5):502-7.

3. Boulton AJ, Armstrong DG, Albert SF, et al. Comprehensive foot examination and risk assessment: a report of the task force of the foot care interest group of the american diabetes association, with endorsement by the american association of clinical endocrinologists. Diabetes Care. 2008;31(8):1679-85.

4. Singh N, Armstrong DG, Lipsky BA. Preventing foot ulcers in patients with diabetes. JAMA. 2005;12(293):217-28.

5. Nanrhe A, Wu S. A guide to amniotic membrane modalities. Podiatry Today. 2016;29(9):40-8.

6. Sheehan P, Jones P, Caselli A, Giurini JM, Veves A. Percent change in wound are of diabetic foot ulcers over a 4-week period is a robust predictor of complete healing in a 12-week prospective trial. Diabetes Care. 2003;26(6):1879-82.

7. Garten A. A closer look at amniotic membrane allografts for wounds. Podiatry Today. Epub 11/12/14.

8. Sheikh ES, Fetterolf DE. Use of dehydrated human amniotic membrane allografts to promote healing in patients with refractory non healing wounds. Int Wound J. 2014;11(6):711-7.

9. Litwiniuk M, Grzela T. Amniotic membrane: new concepts for an old dressing. Wound Repair Regen. 2014; 22(4):451-6.

10. Cooke M, Tan EK, Mandrycky C, He H, O’Connell J, Tseng SC. Comparison of cryopreserved amniotic membrane and umbilical cord tissue with dehydrated amniotic membrane/chorion tissue. J Wound Care. 2014;23(10):465-74.

11. Mamede A, Carvalho MJ, Abrantes AM, Laranjo M, Maia CJ, Botelho MF. Amniotic membrane: from structure and functions to clinical applications. Cell Tissue Res. 2012. 349(2):447-58.

12. Werner S, Grose R. Regulation of wound healing by growth factors and cytokines. Physiol Rev. 2003;83(3):835-70.

13. Frykberg RG, Zgonis T, Armstrong DG, et al. Diabetic foot disorders. a clinical practice guideline (2006 revision). J Foot Ankle Surg. 2006.45(5 Suppl):S1-S66.

14. Niknejad H, Peirovi H, Jorjani M, Ahmadiani A, Ghanavi J, Seifalian AM. Properties of the amniotic membrane for potential use in tissue engineering. Eur Cell Mater. 2008;15:88-99.

15. Koob TJ, Lim JJ, Zabek N, Massee M. Cytokines in single layer amnion allografts compared to multilayer amnion/chorion allografts for wound healing. J Biomed Mater Res B Appl Biomater. 2015;103(5):1133-40.

16. Regulatory considerations for human cells, tissues, and cellular and tissue-based products: minimal manipulation and homologous use. guidance for industry and food and drug administration staff. U.S Department of Health and Human Services. 2017. Accessed online: www.fda.gov/downloads/biologicsbloodvaccines/guidancecomplianceregulatoryinformation/guidances/cellularandgenetherapy/ucm585403.pdf

17. Testing donors of human cells, tissues, and cellular and tissue-based products (HCT/P): specific requirements. FDA. 2018. Accessed online: www.fda.gov/biologicsbloodvaccines/safetyavailability/tissuesafety/ucm151757.htm

Clinician’s Report: Amniotic Wound Care Products

Neox^®Wound Allograft & CLARIX^®Regenerative Matrix
Company: Amniox^®Medical
Description: Amniox products are a cryopreserved form of human amniotic membrane and umbilical cord. Amniox Medical’s proprietary CRYOTEK^TMpreservation process retains the relevant natural structural and biologic characteristics of the tissue while devitalizing the cell activity to eliminate the possibility of graft rejection. It was first introduced in ocular application in 1997. The product was approved by the U.S. Food & Drug Administration in 2011 for foot and ankle use.

 There are two types of tissue that can be utilized to supplement healing. CLARIX is used as a surgical covering, wrap, or barrier. Physicians have described CLARIX as being used during foot and ankle surgery to decrease scar formation or augment the healing of tendon repairs. NEOX has been described as being used as a wound covering for dermal ulcers or defects. Both NEOX and CLARIX are available in multiple sizes and can be modified to fi t the wound or overlying area. The Amniox tissue can be sutured in place, if desired, with an absorbable tissue.

• NEOX

o   Indication: for use as a wound covering for dermal ulcers and defects

o  Composition: human umbilical cord and amniotic membrane preserved to maintain the innate physical and biological properties of these tissues

o   Launch: 2011

• CLARIX

o   Indication: for use as a surgical covering, wrap or barrier

o   Composition: modulates a local wound environment to simulate fetal scarless healing.   This bioactive matrix contains unique matrix proteins that regulate inflammation and prevent scar formation that may lead to limited functional recovery, poor cosmesis, and subsequent surgery or medical management. CLARIX is made from cryopreserved human amniotic membrane and umbilical cord, utilizing the patented CRYOTEK process.

o   Launch: 2011

• FLO variants

o   CLARIX FLO and NEOX FLO are the particulate forms of CLARIX and NEOX 100, and CORD 1K Regenerative and Wound Matrices.

AlloWrap^®DS
Company: AlloSource^®
Description: Amniotic tissue is naturally rich in growth factors to support the healing process and provides an immune-privileged barrier to support the patient’s body in preventing inflammation and scar tissue generation. AlloSource’s AlloWrap is a human amniotic membrane that is strong, pliable, and conforms to wound topography, serving as a barrier. AlloWrap has a double-sided epithelial layer that allows for easy placement, with no orientation necessary. The product is available in a moist, ready-to-use configuration (AlloWrap DS) or dry (AlloWrap Dry), both with room-temperature storage and a two-year shelf life.

AMNIOMATRIX^® & AMNIOEXCEL^®
Company: Integra Life Sciences
Description: AMNIOMATRIX is a cryopreserved suspension allograft derived from the amniotic membrane and components of the amniotic fluid. It is cryopreserved using the patented CryoPrime™ processing method that preserves the structural properties of the collagen, cytokines, growth factors, extracellular matrix (ECM), and viable cellular materials. The liquid-based suspension is especially suited to help repair wounds where membrane products might not be as effective (eg, tunneling or deep wounds). AMNIOMATRIX is intended for homologous use to help supplement the recipient’s tissue and aid in the closing of chronic wounds.

AMNIOEXCEL amniotic allograft membrane is a novel human placental-based tissue product. The membrane forms a protective covering over the wound while providing the key components found in human amnion, including an intact ECM, cytokines, and other growth factors. It easily integrates into the wound and helps provide the optimal environment to repair, reconstruct, and replace wound tissue. AMNIOEXCEL is a minimally manipulated amniotic membrane donated by pre-screened mothers during planned caesarean sections. The membrane is dehydrated using the proprietary DryFlex™ process, which keeps the tissue intact and malleable, gives it a five-year shelf life at room temperature, and retains the key components of human amnion. It is intended for homologous use as a wound covering to aid in closing chronic wounds.

Grafix^®
Company: Osiris Therapeutics Inc.
Description: Grafix is a cryopreserved placental membrane comprised of an extracellular matrix (ECM) that is rich in collagen, growth factors, fibroblasts, mesenchymal stem cells (MSCs), and epithelial cells native to the tissue. It is designed for application directly to acute and chronic wounds, is flexible, and is conforming cover that adheres to complex anatomies. Cryopreserved placental membranes have been shown to be beneficial for natural wound repair in the following ways.¹

Grafix is suitable for a variety of hard-to-treat acute and chronic wounds, including (but not limited to): diabetic foot ulcers, burns, deep tunneling wounds, pressure ulcers, pyoderma gangrenosum, surgical dehiscence, epidermolysis bullosa, surgical incisions, and venous leg ulcers. Grafix may be applied over both bone and tendon, naturally conforms to complex anatomies, and there is no need for sutures or Steri-Strips.^®

Reference

1. Niknejad H, Peirovi H, Jorjani M, Ahmadiani A, Ghanavi J, Seifalian A. Properties of the amniotic membrane for potential use in tissue engineering. Eur Cell Mater.2008;15:88-99.