In Vivo Evaluation of Angiogenic Properties of a Dehydrated Amnion Chorion Membrane

Angiogenesis is an essential part of wound healing, and placental membranes have been shown to have angiogenic properties both in vitro and in vivo. 

The purpose of this study was to evaluate the angiogenic properties of a commercially available dehydrated Amnion/Chorion membrane (dACM) º in vivo.  Gelatin sponges soaked with conditioned media (CM) from dACM were implanted subcutaneously into Sprague Dawley rats and retrieved at 7 and 14 days and evaluated via histology and gene expression.  CM was obtained by incubating dACM grafts in basal media for 5 days at 4°C at a ratio of 1 cm2 dACM to 1 mL media. 

The protocol for this study was reviewed and approved by the Bridge PTS IACUC (protocol 17-02).  Control groups consisted of sponges soaked with basal media. At 7- and 14-days, implants were retrieved with half placed in 10% neutral buffered formalin for histological evaluation, and half placed in RNAzol for PCR evaluation for angiogenesis related targets.  Fixed samples were then embedded and sections were then stained with CD31 and _SMA with hematoxylin counterstaining.   By 7 days there were significant increases in several pro-angiogenic genes in the dACM CM group including FN1, EFNA1, TGFB3, VEGFC, TYMP, THBS1, and SERPINE1.  ANGPT2, an antagonist of angiopoietin 1, was significantly downregulated at 7 days. 

At 7 days SMA quantification showed small increases in staining within the dACM implant compared to negative controls, but no differences in _SMA staining on the outside of the implant.  By 14 days, _SMA staining outside the implant was increased in dACM CM groups compared to controls.  These results demonstrate that dACM contains a number of growth factors and cytokines that promote a pro-angiogenic response in vivo.