Characterization of the Amnion, Chorionic Membrane, and Trophoblast Layers of a Decellularized, Freeze-Dried Placental Membrane for Wound Care Application

Introduction: Placental membranes have been successfully used for chronic wound treatment. Placental membrane contains three major, distinct layers: the amniotic membrane (AM), the chorionic membrane (CM), and a trophoblast (Troph) layer; the latter two layers together form the chorion. Recently, a full-thickness (AM+CM+Troph), decellularized, lyophilized, and terminally sterilized placental membrane product* has been introduced as a novel wound dressing material.

The objective of this study was to determine the quantities of major biological factors in each of the three layers of the placental membrane and assess the related anti-inflammatory activity.

Methods: Placental membranes from 3 authorized donors were decellularized, and each layer was separated, lyophilized, and then extracted to quantify five biological factors using ELISA or multiplex: basic fibroblast growth factor (bFGF), interleukin-1 receptor antagonist (IL-1RA), tissue inhibitors of metalloproteinases (TIMP-2), hyaluronic acid, and fibronectin. Biological factors were quantitated and reported as a percentage of total quantity in all three layers for each factor tested. DNA quantification and histological analysis were also performed to confirm the decellularization and successful separation of each layer. In addition, an anti-inflammatory assay was performed for each layer.

Results: Successful decellularization was demonstrated from DNA quantification. Histological analysis confirmed the complete separation of each layer and trophoblast layer is the thickest. The trophoblast layer contained the highest percentage of bFGF (60%), IL-1RA (67.8%), TIMP-2 (54.8%), and fibronectin (79.4%). The AM and CM contained higher percentages of hyaluronic acid than the trophoblast layer. The trophoblast layer also demonstrated the highest anti-inflammatory activity of all three layers.

Discussion: This study demonstrated that the trophoblast layer can provide more than 50% of endogenous biological factors and ECM components, with the exception of hyaluronic acid, and suggests the inclusion of the intact trophoblast layer in this placental membrane product* is beneficial for wound healing.