Decellularization of human amniotic membrane and its impact on mesh bioscaffold properties

The human amniotic membrane (hAM) is a collagen-rich tissue increasingly used as a natural scaffold in tissue engineering. A key step in these applications is decellularization, which extracts the extracellular matrix (ECM) as a biomaterial. In this study, native hAM was processed using two different protocols: peptide bond hydrolysis with EDTA-assisted trypsin (En) and cell membrane disruption using a nonionic detergent combined with reversible alkaline swelling (SD), along with nuclease activity. The effects of each protocol on the composition, structure, and tensile behavior of the processed hAM were evaluated. Additionally, the repopulation efficiency of epithelial cells on both surfaces of the decellularized hAM was assessed. Comparisons between decellularized and digested hAM and native tissue revealed residual DNA contents ranging from 0.14 to 30% for the En method and from 1.7 to 10% for the SD method. Meanwhile, sulfated glycosaminoglycans (sGAG) content varied from 1.5 to 30% and from 1 to 18%, respectively. The En method substantially reduces fibronectin levels, while the SD method reduces lumican in the leached components of decellularized hAM. En-treated specimens exhibit altered elastic and rupture properties, negatively impacting their mechanical behavior. The hAM-derived ECM materials were repopulated by human vaginal epithelial cells, adopting morphologies that depend on the surface characteristics provided by the anatomical portion of the native tissue. Overall, these results suggest that native tissue variability influences the final composition of hAM-derived materials, that tensile properties are influenced by the used decellularization method, and that surface characteristics play a critical role in epithelial cell repopulation.