Elevated non-esterified fatty acid (NEFA) levels are closely associated with metabolic disorders in dairy cattle, yet their direct effects on adipocyte physiology remain poorly understood. In this study, we demonstrate that high NEFA concentrations significantly impair bovine adipocyte function by simultaneously inhibiting proliferation/differentiation (p < 0.01) and promoting pathological lipid deposition. Through integrated transcriptomic and functional analyses, we identified NDUFAB1 as a central metabolic regulator that counteracts NEFA-induced adipocyte dysfunction. Mechanistically, NDUFAB1 activation attenuates the cytotoxic effects of excessive NEFA exposure. These findings provide both fundamental insights into energy metabolism regulation and a potential therapeutic target (NDUFAB1) for preventing bovine metabolic diseases.

Low-temperature environments in cold regions pose a significant threat to cattle farming. Bovine mammary epithelial cells (BMECs) are highly sensitive to cold stress, and acute cold stress can induce apoptosis, adversely affecting lactation performance and health. To explore the mechanism of acute cold stress-induced apoptosis in BMECs, we established an in vitro acute cold stress model. Results showed that mRNA levels of HSP90 increased significantly in a time-dependent manner after 2 h of cold stress, confirming successful model establishment. Following 4 h of cold stress, pro-apoptotic genes (Caspase-3, Bax) exhibited significantly elevated mRNA levels, while the anti-apoptotic gene (BCL-2) showed significantly reduced mRNA levels. Concurrently, the apoptosis rate increased significantly, indicating that acute cold stress induces apoptosis and suggesting the 4 h mark may represent a critical transition point. Integrated transcriptomic and functional analyses identified ENO1 as a core metabolic regulator counteracting acute cold stress-induced apoptosis in BMECs. As a multifunctional protein, ENO1 (alpha-enolase) acts as a central enzyme in glycolysis while exerting additional roles in cellular signaling and apoptotic processes, thereby participating in various pathophysiological regulations. Both mRNA and protein levels of ENO1 were significantly elevated in cold-stressed cells compared to untreated controls. Importantly, interference with ENO1 expression aggravated the extent of cold stress-induced apoptosis, demonstrating the regulatory role of ENO1 in this process. To our knowledge, this is the first report elucidating the core regulatory function of ENO1 in acute cold stress-induced apoptosis in BMECs. These findings provide a theoretical basis for understanding apoptotic mechanisms under stress.

FTO expression correlates with adipose tissue development in rabbits, yet its regulatory role in preadipocyte differentiation remains poorly understood. Therefore, we employed transcriptome sequencing to identify 422 differentially expressed genes (DEGs) between the FTO overexpression group and the FTO-negative control group. Multi-omics evidence from proliferation assays (CCK-8), protein verification (WB), gene quantification (qPCR), and lipid staining (Oil Red O) established FTO as a positive regulator of rabbit preadipocyte development with gain-of-function enhancing and loss-of-function impeding differentiation. In conclusion, FTO regulates the proliferation and differentiation of rabbit preadipocytes, providing deeper insights into livestock energy metabolism and physiological functions. These findings hold significant implications for healthy animal farming, high-quality livestock product production, and the prevention and control of metabolic diseases in animals.