Optimizing Decellularization of Bovine Ovarian Tissue: Toward a Transplantable Artificial Ovary Scaffold with Minimized Residual Toxicity and Preserved Extracellular Matrix Morphology

Introduction: The decellularized extracellular matrix (dECM) from ovarian tissue could be the best scaffold for the development of a transplantable artificial ovary. Typically, dECM from ovarian tissue has been obtained using sodium dodecyl sulfate (SDS), at a concentration of 1% for 24 h. However, SDS can leave residues in the tissue, which may be toxic to the seeded cells. This study aimed to obtain dECM from bovine ovarian tissue using SDS and NaOH at a minimum concentration in the shortest incubation time. Methods: The respective SDS and NaOH concentrations investigated were 1% and 0.2 <sc>m</sc>; 0.5% and 0.1 <sc>m</sc>; 0.1% and 0.02 <sc>m</sc>; and 0.05% and 0.01 <sc>m</sc>, with 24-, 12-, and 6-h incubation periods. After the incubation time, the tissue was washed in 50 mL of distilled water for 6 h. Results: Histological analysis confirmed decellularization and showed the conservation of collagen fibers in all samples following treatment. Furthermore, the lowest SDS and NaOH concentrations that showed no DNA remaining during electrophoresis analysis were 0.1% and 0.02 <sc>m</sc> when incubated for 24 and 12 h. DNA quantification resulted in &lt;0.2 ng DNA/mg ovarian tissue using these protocols. Additionally, the coculture of dECM (obtained by 0.1% SDS and 0.02 <sc>m</sc> NaOH for 12 h) with ovarian cells showed that there was no toxic effect for the cells for up to 72 h. Conclusion: The protocol involving 0.1% SDS and 0.02 <sc>m</sc> NaOH for 12-h incubation decellularizes bovine ovarian tissue, generating a dECM that preserves the native ECM morphology and is nontoxic to ovarian cells.