Short-Term In Vitro Culture of Human Ovarian Tissue: A Comparative Study of Serum Supplementation for Primordial Follicle Survival

Optimizing in vitro culture conditions for cryopreserved–thawed human ovarian cortical fragments (OCFs) represents a critical step in fertility preservation strategies. OCFs predominantly contain primordial follicles (PMFs), whose survival and integrity are essential for ex vivo folliculogenesis. This study aimed to evaluate the impact of different culture media supplementations on PMF survival and tissue morphology by comparing alpha-Minimum Essential Medium (αMEM) supplemented with Human Serum Albumin (HSA), Human Serum (HS), or Serum Substitute Supplement (SSS). Twenty-nine OCFs were cultured for three days, and follicular density and were morphology assessed. Generalized linear mixed model analysis showed that PMF density was significantly higher in OCFs cultured in medium supplemented with SSS (213 PMFs/mm3) compared to those cultured with HSA (107 PMFs/mm3) or HS (93 PMFs/mm3). Furthermore, SSS supplementation was associated with a significant increase in the number of PMFs showing healthy morphologies. These findings indicate that SSS supplementation to αMEM enhances the survival and preserves better morphologies of the human PMFs in short-term culture, highlighting its potential as a suitable culture supplement for ovarian tissue preservation.