Isolation and characterization of rabbit intramuscular preadipocytes: Establishing a foundation for studies of intramuscular fat metabolism

Intramuscular fat (IMF) significantly influences meat quality, affecting flavor, tenderness, and juiciness. Despite extensive research on livestock such as cattle and pigs, the regulation of IMF in rabbits remains poorly understood. This study establishes a standardized in vitro method for isolating and characterizing intramuscular preadipocytes from the longissimus dorsi muscle of 3-day-old Hycole rabbits using enzymatic digestion and differential centrifugation. Three-day-old neonates were selected for initial protocol development due to their higher progenitor cell abundance and tissue accessibility, which facilitates method optimization and ensures consistent cell yields. Isolated cells displayed typical preadipocyte morphology and expressed key markers, including PREF-1, PPARγ, and Leptin, confirmed by RT-qPCR and immunofluorescence. Upon adipogenic induction, cells exhibited progressive lipid accumulation, as indicated by Oil Red O staining, and showed upregulated expression of PPARG, C/EBPa, and FABP4, confirming their differentiation potential. This study establishes a comprehensive protocol for isolating rabbit intramuscular preadipocytes, providing a foundational tool for investigating the molecular mechanisms of adipogenesis in lagomorphs. The model supports future studies on genetic and nutritional strategies to enhance rabbit meat quality while preserving its lean characteristics.