0.1% Peracetic Acid Treatment Is the Optimal Process for Ovarian Decellularized Extracellular Matrix

Abstract

This research aimed to find the optimal sterilization process for ovarian decellularized extracellular matrix (OV-dECM). OV-dECM was prepared. Four different sterilization processes, such as ultraviolet ray, 1 mg/L CuCl₂ + 0.5% H₂O₂, 70% ethanol, and 0.1% peracetic acid (PAA), were evaluated. The sterilization efficiency and the in vitro and in vivo biocompatibility were assessed. Hematoxylin and eosin, 4′,6-diamidino-2-phenylindole staining, and DNA quantitation indicated the successful production of OV-dECM. 0.1% PAA and 70% ethanol achieved better sterilization. The sterilized OV-dECM in the 70% ethanol and 0.1% PAA groups could integrate with human umbilical cord mesenchymal stem cell (between them: 7 days: p = 0.187; 10 days: p = 0.293; both p > 0.05) and keep the cell viability (CCK-8, cell counting kit-8) and cell morphology (F-actin staining). The sterilized OV-dECM in both 70% ethanol and 0.1% PAA groups had good CD31 (a neovascularization maker) fusion indexes (between them: p = 0.288; p > 0.05), while α-smooth muscle actin (another neovascularization maker) in the 0.1% PAA group was higher than that in the 70% ethanol group (between them: p = 0.012; p < 0.05). The assessment of CD68, CD86, and CD206 (macrophage phenotype makers) demonstrated that the 0.1% PAA group had better chance to promote the M1–M2 transformation of macrophage than 70% ethanol after the implantation of OV-dECM in mice. In conclusion, 0.1% PAA is the optimal sterilization process for OV-dECM with better sterilization capacity and biocompatibility.

Impact Statement

This study identifies 0.1% peracetic acid as the optimal sterilization method for ovarian decellularized extracellular matrix (OV-dECM), resolving a critical barrier to artificial ovary (AO) development. It ensures OV-dECM sterility while preserving biocompatibility, promoting macrophage M1-to-M2 polarization to reduce inflammation. This advances AO translational potential, offering hope for fertility preservation in patients with cancer/leukemia and guiding standardized OV-dECM sterilization in regenerative medicine.