Culture medium modulates nucleolar architecture and disrupts ribosomal RNA levels in preimplantation mouse embryos

Embryonic culture media support preimplantation development in vitro, yet their broader effects on the molecular organization have not been fully characterized. In mammalian embryos, the nucleolus is a prominent and highly dynamic organelle that orchestrates ribosome biogenesis through RNA polymerase I-dependent ribosomal DNA (rDNA) transcription. However, the effect of culture conditions on nucleolar organization and rRNA dynamics during early development remains poorly understood. In this study, we investigated the effects of two compositionally distinct culture media on the nucleolar architecture and rRNA levels in mouse preimplantation embryos cultured in vitro. In vitro fertilized embryos were cultured in either Chatot-Ziomek-Bavister (CZB) or α-minimal essential medium (α-MEM). The nucleolar morphology was assessed by analyzing the localization of the nucleolar protein NOPP140 and rRNA levels were quantified by immunostaining the embryos with the Y10b antibody clone. Quantitative analyses showed that embryos cultured in α-MEM exhibited a significantly higher proportion of single and enlarged nucleoli per blastomere than those cultured in CZB (P < 0.05). However, α-MEM cultured embryos displayed a significant reduction in Y10b signal intensity relative to CZB-cultured embryos (P < 0.05). Although developmental rates were similar between the culture conditions, embryos cultured in α-MEM displayed altered cell lineage allocation compared to those cultured in CZB. Together, these findings indicate that culture medium regulates nucleolar organization and rRNA abundance in mouse preimplantation embryos, with nucleolar plasticity serving as a sensitive molecular indicator of in vitro culture-dependent developmental effects.