Amniotic membrane is an increasingly utilized tool for physicians to spur healing in chronic diabetic foot ulcers. These authors take a closer look at the research and potential of both cryopreserved and dehydrated amniotic membrane.
The prevalence of diabetes continues to rise year after year. In 2011, an estimated 366 million people, 7 percent of the world’s population, had diabetes.1 It is expected that the number will increase to 552 million adults by 2030.1
Complications due to diabetes are also an expanding public health issue. Diabetic foot ulcers (DFUs) represent one of the most common complications of diabetes. Approximately 25 percent of patients with diabetes will develop a lower extremity ulceration in their lifetime.2 Diabetic foot ulcers can have a profound impact on the morbidity, mortality and quality of life of patients. For example, patients who develop DFUs have a higher risk of myocardial infarction and fatal stroke than those who have never had a wound.3
The treatment of DFUs can be difficult as they are often slow to heal and frequently recur. The poor prognosis of these wounds is often attributed to other complications of diabetes such as peripheral neuropathy, peripheral vascular disease and persistent hyperglycemia. Unfortunately, the prolonged healing of ulcers frequently leads to wound infection and subsequent amputation. Clearly, lower extremity ulcerations pose a significant problem to patients suffering from diabetes mellitus as well as the physicians who treat them.
A Pertinent Primer On The Anatomy And Physiology Of Amniotic Tissue
Luckily, there are many advanced therapies that the wound specialist can employ in order to facilitate healing when traditional approaches fail. Amniotic tissue represents one of the most promising advanced therapies available for achieving wound closure. However, the relatively high cost of using advanced wound care products often poses an obstacle to their use. Factors such as tissue storage as well as the time and skill required to apply amniotic membranes also represent challenges inherent to these products. Therefore, we must closely examine the effectiveness of these products in comparison to more affordable methods before employing them regularly.
Before examining the characteristics of individual products, it is important to understand the anatomy and physiology of amniotic tissue in general.
The amniotic membrane derives from the placental sac and supports the fetus by forming the inner lining of the amniotic cavity. Functions of the amniotic membrane include the exchange of water-soluble molecules and production of cytokines and growth factors.5 By regulating metabolic processes, it is the responsibility of the amniotic membrane to facilitate the development of the fetus. The anatomic makeup of the amniotic membrane dictates its functionality.
There are two layers that make up the amniotic membrane: an inner amnion that is adjacent to the fetus as well as an outer chorion that lies next to the uterus. The amnion is made up of a thin layer of epithelial cells connected to a basement membrane. By comparison, the chorion is much thicker and consists of densely packed collagen fibers, forming a protective outer layer between the fetus and uterus. Additionally, the chorion has powerful anti-inflammatory and immunologic properties that shield the fetus from harm.
Perhaps the most striking characteristic of amniotic membrane is its ability to produce a wide variety of regenerative growth factors that facilitate fetal development. Examples of growth factors within amniotic membrane include basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), keratinocyte growth factor (KGF), nerve growth factor (NGF), transforming growth factors (TGF) and vascular endothelial growth factor (VEGF).5–7 Both amnion and chorion contain growth factors but researchers have determined that chorion contains nearly five times as many growth factors as amnion.5 Therefore, it stands to reason that an in vitro study comparing bilayer allografts (amnion/chorion) to single layer amnion allografts identified a greater number of growth factors in the bilayered product.8
These growth factors in combination with various other cytokines have great potential benefits in wound healing. These benefits include creating a structural scaffold for tissue proliferation, modulating the immune response, inhibiting matrix metalloproteinase production, reducing inflammation, stimulating angiogenesis and facilitating tissue remodeling.
What The Evidence Says About Dehydrated Human Amniotic Membrane
One of the available dehydrated human amniotic membrane (dHAM) products is AmnioExcel (Integra LifeSciences). This product consists of an intact extracellular matrix intended to assist in tissue regeneration. According to the manufacturer, the dehydration process removes the need for thawing, making AmnioExcel available for use at room temperature with a shelf life of five years.9 Furthermore, the allograft immediately adheres to a moisturized wound bed due to its hydrophilic nature.10 These traits make the dHAM relatively easy to store and use.
Several recently published studies have looked at the clinical effectiveness of this dHAM. Snyder and colleagues published a randomized controlled trial comparing the efficacy of the dehydrated amniotic membrane allograft to the standard of care in healing chronic diabetic foot ulcers.10 Results of this study demonstrated that one-third of patients receiving dHAM in addition to standard of care had complete wound closure by six weeks. Comparatively, none of the 14 patients in the standard of care cohort achieved wound closure in the same period of time. Although the sample size in this study was small, the results were promising for the potential healing benefits of dHAM.
Another study conducted by Lintzeris and coworkers consisted of a retrospective analysis on the use of dehydrated amniotic membrane for wounds with delayed healing.9 This study included a total of nine wounds that had failed to have greater than 50 percent healing after the use of both traditional and advanced wound care modalities. The wounds were present for an average of 11 weeks before the first application of dHAM. All wounds healed in an average of 5.7 weeks after a mean of 2.7 applications. The authors noted no adverse events. Although the sample size was quite small, the authors’ findings suggest that further clinical trials are worth conducting in order to evaluate the utility of dHAM in treating complex, non-healing wounds.
EpiFix (MiMedx Group) represents another promising product and consists of dehydrated amnion/chorion membrane. EpiFix contains angiopoietin (ANG‐2), EGF, bFGF, heparin-binding epidermal growth factor (HB-EGF), hepatocyte growth factor (HGF), platelet-derived growth factor BB (PDGF-BB), placental growth factor (PlGF) and VEGF.11 In theory, these factors give EpiFix the potential to promote revascularization of chronic wounds.
In 2017, Bianchi and colleagues conducted a randomized controlled, multicenter trial evaluating the utility of EpiFix in treating venous leg ulcers.12 The study included 52 patients receiving EpiFix plus multilayer compression and 57 patients receiving multilayer compression alone. The authors found that “participants receiving weekly application of EpiFix and compression were significantly more likely to experience complete wound healing (60 percent) than those receiving standard wound care and compression (35 percent)” at 12 weeks. These results confirm the theory that dehydrated amniotic/chorionic membrane allograft can improve the wound healing environment in chronic venous leg ulcers.
Key Pearls On The Potential Of Cryopreserved Human Amniotic Membrane
Cryopreservation represents another modality of processing and delivering amniotic membranes. There are several cryopreserved amniotic products currently on the market. Grafix and Stravix (Osiris Therapeutics) are two such products indicated for use in treating diabetic foot ulcers.
Grafix is a bioengineered tissue allograft consisting of a cryopreserved placental membrane matrix. Grafix processing occurs in a manner that maintains the integrity of the extracellular matrix, growth factors, fibroblasts and mesenchymal stem cells of the tissue.
Touting Grafix as the first product to have a randomized clinical trial that included viable stem cells, Lavery and colleagues conducted a study comparing diabetic foot ulcers treated with Grafix to those treated with high-quality standard of care alone.13 The authors defined standard of care as “surgical wound debridement, high‐quality offloading devices and non-adherent dressings that were provided uniformly to all patients in both treatment groups.” The results of this study demonstrated a higher proportion of ulcers achieving complete closure at 12 weeks in the Grafix group (62 percent) in comparison to 21 percent in the control group. Furthermore, the authors noted that 82 percent of Grafix-healed ulcers remained closed at a 12-week follow-up in comparison to 70 percent in the control group. Additionally, the authors noted a lower incidence of wound infection during the 12-week period in the Grafix cohort (18 percent) versus 36.2 percent in the control group.
Overall, this randomized clinical study demonstrated that weekly application of Grafix increased the proportion of wounds that healed, decreased the incidence of wound infection and accelerated the time to 100 percent reepithelialization.13 Based on their findings, the authors claim that Grafix is a viable treatment option in the effective treatment of chronic diabetic foot ulcers.
Similar to Grafix, Stravix is a cryopreserved placental tissue composed of umbilical amnion containing extracellular matrix, growth factors and endogenous mesenchymal cells.14 What makes Stravix unique is the fact that it is combined with Wharton’s jelly, thus allowing it to conform to various surfaces such as bone, muscle, tendon, nerve, capsule and hardware. Stravix essentially confers the advanced wound healing properties of Grafix in a more pliable texture.
Comparing The Processing Of Dehydrated And Cryopreserved Amniotic Membranes
Clearly, there is literature to suggest that both dehydrated and cryopreserved amniotic membranes provide significant benefits in treating complex, stalled wounds. However, it is important for the wound specialist to be familiar with the processing methods of each set of products.
In a double-blind peer-reviewed study, Cooke and colleagues compared the Cryotek (cryopreservation) method with the Purion (dehydration) processing method.15 The aim of the study was to determine if different processing methods affected tissue integrity and therapeutic efficacy. The authors noted that cryopreservation retains the natural architecture of the amniotic membrane extracellular matrix whereas dehydration can “compact” the tissue and potentially alter it. Furthermore, the authors’ results suggested that cryopreservation maintained the integrity of biological signaling molecules such heavy-chain hyaluronic acid/pentraxin 3 complex, a compound with potent anti-inflammatory and anti-scarring properties.16
Although these results suggest that cryoprocessing preserves the amniotic membrane on a molecular level better than dehydration, it is worth noting that there is lack of head-to-head studies comparing the two processes in the clinical setting.
Amniotic tissue products are powerful options for physicians to keep in their wound care arsenal. These products offer the potential for accelerating wound healing, reducing the incidence of infection and preventing reulceration. Yet, despite the published advantages of using amniotic membrane in treating DFUs, it is not appropriate to apply them to every wound. Advanced tissue products are not a replacement for the traditional mainstays of wound care such as aggressive sharp debridement, adequate offloading and the application of sterile dressings.
Amniotic membrane, like most biologic tissue products, requires significant processing and therefore carries a high price tag. On average, amniotic tissue products cost $500 to $1,000 per application. This may seem astronomically high at first glance but the treatment and complications of DFUs carry an approximate cost of $28,000 per patient per year.17 Therefore, when clinicians use amniotic membrane products appropriately, the modalities may result in sizeable cost savings for our health care system.
Furthermore, in the setting of treating patients who are at a high risk for amputation, financial considerations must take a back seat to limb preservation efforts. Further studies are needed to determine whether improved healing rates achieved with these products correlate with a reduced rate of amputation and mortality.
Dr. Sullivan is a first-year resident within the Division of Podiatric Medicine and Surgery at Beth Israel Deaconess Medical Center in Boston.
Dr. Rosenblum is an Assistant Clinical Professor of Surgery at Harvard Medical School and the Associate Chief of the Division of Podiatric Medicine and Surgery at the Beth Israel Deaconess Medical Center in Boston. He is a Fellow of the American College of Foot and Ankle Surgeons.
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