Biological properties of dehydrated human amnion/chorion composite graft: implications for chronic wound healing

Abstract

Human amnion/chorion tissue derived from the placenta is rich in cytokines and growth factors known to promote wound healing; however, preservation of the biological activities of therapeutic allografts during processing remains a challenge. In this study, PURION® (MiMedx, Marietta, GA) processed dehydrated human amnion/chorion tissue allografts (dHACM, EpiFix®, MiMedx) were evaluated for the presence of growth factors, interleukins (ILs) and tissue inhibitors of metalloproteinases (TIMPs). Enzyme-linked immunosorbent assays (ELISA) were performed on samples of dHACM and showed quantifiable levels of the following growth factors: platelet-derived growth factor-AA (PDGF-AA), PDGF-BB, transforming growth factor α (TGFα), TGFβ1, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), placental growth factor (PLGF) and granulocyte colony-stimulating factor (GCSF). The ELISA assays also confirmed the presence of IL-4, 6, 8 and 10, and TIMP 1, 2 and 4. Moreover, the relative elution of growth factors into saline from the allograft ranged from 4% to 62%, indicating that there are bound and unbound fractions of these compounds within the allograft. dHACM retained biological activities that cause human dermal fibroblast proliferation and migration of human mesenchymal stem cells (MSCs) in vitro. An in vivo mouse model showed that dHACM when tested in a skin flap model caused mesenchymal progenitor cell recruitment to the site of implantation. The results from both the in vitro and in vivo experiments clearly established that dHACM contains one or more soluble factors capable of stimulating MSC migration and recruitment. In summary, PURION® processed dHACM retains its biological activities related to wound healing, including the potential to positively affect four distinct and pivotal physiological processes intimately involved in wound healing: cell proliferation, inflammation, metalloproteinase activity and recruitment of progenitor cells. This suggests a paracrine mechanism of action for dHACM when used for wound healing applications.

Dehydrated human amnion/chorion membrane regulates stem cell activity in vitro

“…In prospective, randomized, controlled clinical trials, dHACM tissue has demonstrated the ability to resolve chronic inflammation and accelerate healing in a variety of refractory wounds;29, 30, 31, 48, 49, 50 however, the mechanisms of action remain somewhat unclear. Data from preclinical wound models in mice has additionally shown that dHACM implanted subcutaneously leads to the recruitment of circulating host MSCs and HSCs, suggesting that adult stem cells may contribute to the mechanism by which dHACM promotes healing 3, 33. The results from the current study contribute to an understanding of how different types of stem cells respond to dHACM in vitro and how dHACM‐treated stem cells may contribute to the mode of action for resolving chronic inflammation and ultimately promoting healing.…”

“…Extracts of dHACM contain protein factors that have been shown to stimulate cellular chemotaxis in vitro and in vivo ;3, 33 therefore, this study evaluated whether different stem cell types would respond to treatment with dHACM extract by populating a cell‐free zone using an in vitro cell closure model. Closure assays of ADSCs and BM‐MSCs, conducted by treating the cells with different concentrations of dHACM extract over 72 h, were evaluated based on percent closure of an initially acellular zone in each well.…”

“…As a nonadherent cell type, HSCs were not examined for cell migration using the Oris™ assay; however, previously published studies have shown that dHACM recruits HSCs to the site of a wound in vivo in a mouse subcutaneous implantation model 3. Over 50 soluble growth factors and cytokine proteins have been identified in soluble extracts of dHACM,32, 33 suggesting that dHACM contains soluble components that elute from the tissue and stimulate the growth and chemotactic migration of different types of stem cells, thus amplifying the number of stem cells locally. However, due to the complexity of this unique material, a causative effect cannot be assigned to any single regulatory protein or other signaling molecule either currently identified or yet to be identified in dHACM tissues.…”

“…Previously published animal dermal wound models have shown that dHACM promotes the recruitment of both hematopoietic and mesenchymal stem cells to the treated site of injury 3. Studies measured a significant increase in BM‐MSC recruitment by day 7 of dHACM treatment,33 as well as a ten‐fold increase of migrated HSCs to the wound site by day 14 in a mouse subcutaneous implantation model, accompanied by an increase in SDF‐1α expression 3. These in vivo results suggest that the combined impact of dHACM cytokine protein elution and regulation of stem cell activity at a wound site may be involved in enhancing tissue repair.…”

“…The dHACM allografts have been utilized in the treatment of chronic wounds, such as diabetic foot ulcers and venous ulcers 29, 30, 31. Over 50 growth factors, cytokines, chemokines, and tissue inhibitors of metalloproteinases (TIMPs) have been previously identified in dHACM tissues,32, 33 many of which are known to stimulate paracrine responses in cells involved in healing and tissue repair, such as human dermal fibroblasts, microvascular endothelial cells, and stem cells. Previously published animal dermal wound models have shown that dHACM promotes the recruitment of both hematopoietic and mesenchymal stem cells to the treated site of injury 3.…”