This study employed a ground-based microgravity analog system to assess mouse oocyte meiotic progression and developmental competence, providing mechanistic insights into space environment-induced defects during oocyte maturation. Germinal vesicle (GV)-stage mouse oocytes were encapsulated in polydimethylsiloxane (PDMS) chip chamber and subjected to simulated microgravity (SMG) culture using a random positioning machine (RPM). Meiotic dynamics were systematically analyzed at five key stages: GV (0 h), GV breakdown (GVBD, 2 h), pro-metaphase I (Pro-MⅠ, 5 h), metaphase I (MⅠ, 8 h), and metaphase II (MⅡ, 16 h). Mitochondrial distribution, spindle morphology, and chromosome alignment were quantified through confocal laser microscopy coupled with fluorescent probes. The results showed that SMG exposure reduced oocyte maturation rates by 32.75% compared to normal gravity (NG) controls (p<0.01). Mitochondrial dynamics exhibited stage-specific perturbations: perinuclear clustering at MI (70.00% vs 41.18% in NG) and disorganized aggregation patterns in 71.88% of MII oocytes. Spindle assembly and chromosome alignment were also disrupted: multipolar spindles during MⅠ caused disordered chromosome segregation. At MⅡ, SMG oocytes displayed exacerbated spindle defects (57.58% abnormality rate vs 22.32% in NG, p<0.05) and widened equatorial plates (15.63 μm vs 7.55 μm, p<0.0001). These findings suggest that SMG exposure compromises meiosis and oocyte quality through tripartite disruption of mitochondrial-spindle-chromosomal coordination. These results provide important insights into how mechanical perturbations regulate subcellular structure interaction networks to affect oocyte quality.
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