The effects of cryopreservation on angiogenesis modulation activity of human amniotic membrane

Amniotic membrane (AM), as the innermost layer of placenta, has side dependent effects on the angiogenesis. Cryopreservation is a necessary process to avoid the challenging problems of fresh tissues; a procedure which makes the AM ready-to-use. Since the cryopreservation can influence the AM characteristics for experimental and clinical purposes, in this study the effects of cryopreservation were evaluated on angiogenesis modulation activity of the AM compared to fresh tissues in an animal model. The AM was implanted mesenchymal side up or epithelial side up in a rat dorsal skinfold chamber. The length and number of branches of formed capillaries were measured via intravital microscopy after 7 days. The amount of IL-8 (interleukin-8) and TIMP-2 (Tissue Inhibitor of Matrix Metalloproteinase-2) as two factors in amniotic cells which have great impacts on angiogenesis were evaluated using ELISA assay. The epithelial surface of cryopreserved AM had inhibitory effects on vessel formation. The cryopreserved amniotic mesenchymal side increased the vessel length and sprout. The result of cryopreserved AM on angiogenesis was similar to that of fresh tissues. The levels of IL-8 and TIMP-2 in cryopreserved samples were significantly less than fresh AMs which shows that angio-modulatory properties are not limited to the effects of amnion epithelial and mesenchymal stem cells and the other components such as extracellular matrix may contribute in angio-modulatory effects. These promising results show that inducing and inhibitory effects of the AM, which make it an appropriate candidate for different clinical situations, were maintained after cryopreservation.