Influence of short-term storage at 4 °C, -20 °C or -80 °C on the quality of decellularized porcine oviductal scaffolds

In reproductive medicine, decellularized scaffolds offer new opportunities for treating reproductive tract pathologies and improving assisted reproductive technologies. After obtaining them through decellularization procedures, these scaffolds are usually stored at 4°C, -20°C, or -80°C, although the effect of these temperatures on their quality is unknown. The objective of this study was to evaluate the short-term influence of these storage temperatures on the quality of the decellularized oviduct scaffolds derived from porcine species. Complete oviducts were obtained from prepubertal animals (Large White × Landrace) and decellularized following a previously described immersion–agitation protocol [1]. Subsequently, the scaffolds were stored at 4°C, -20°C, or -80°C for one month. Freshly decellularized oviducts were used as the control (n=4 per group). After storage, samples from each group underwent differential scanning calorimetry (DSC) and were tested for cytotoxicity by co-incubating them with porcine spermatozoa for 3 h at 38°C. Cytotoxicity analysis included the evaluation of sperm motility, kinematic parameters, viability, acrosomal integrity, and mitochondrial activity. The samples were also analyzed microbiologically, including counts of total aerobic bacteria (TAB), total coliforms (TC), and sulfite-reducing clostridia (SRC). The DSC results show no significant differences between the control group and the stored groups, indicating that storage temperature did not affect the thermal stability of the proteins (p > 0.05). No changes were observed in any sperm parameter evaluated after incubation of scaffolds with porcine spermatozoa (p > 0.05). However, storage temperature influenced TAB counts, which were significantly higher in oviducts stored at 4°C and -20°C compared to the control group (p < 0.05). Overall, these results demonstrate that decellularized oviductal scaffolds exhibit good stability after one month of storage. Nevertheless, bacterial growth can compromise scaffold quality, and storage at -80°C is the most effective way to prevent contamination.

[1] Martínez-López C et al. Theriogenology. 2025;231:36–51.