Integrated histological, biochemical and molecular analysis of grass carp (Ctenopharyngodon idella) responses to acute hypoxic stress.

Hypoxia critically impacts grass carp (Ctenopharyngodon idella) aquaculture, causing oxidative damage, mortality and economic losses. This study revealed the physiological and genetic response mechanisms of grass carp under hypoxic stress by integrating histology, biochemistry and miRNA-mRNA network analysis. The experimental individuals were confined in sealed containers with pre-aerated water, allowing dissolved oxygen (DO) to decline naturally, and three groups were established: Normoxia (DO: 7.99 ± 0.43 mg∙L-1; control), Hypoxia (DO: 0.88 ± 0.16 mg∙L-1; upward swimming and gulping), and Asphyxia (DO: 0.29 ± 0.04 mg∙L-1; loss of equilibrium), each with three replicates. Hypoxia and Asphyxia groups exhibited increased hepatic lactate concentration, as well as reduced interlamellar cell mass area, hepatic glucose and glycogen concentrations. Hepatic malondialdehyde concentration, catalase activity and pyruvate content increased as hypoxia intensified, while superoxide dismutase activity decreased. Transcriptomic analysis identified 2661, 2834 and 1908 differentially expressed genes (DEGs) in Hypoxia vs. Normoxia, Asphyxia vs. Normoxia and Hypoxia vs. Asphyxia comparison, respectively, revealing enrichment in the cell cycle, glycolysis and biosynthesis of unsaturated fatty acids as DO declined. The miRNA profiling detected 71, 47 and 51 differentially expressed miRNAs (DEMs) in the three comparisons, respectively. These DEMs mediated the regulation of pathways involving gill cytoskeletal dynamics, antioxidant defenses and glycolipid metabolism through targeting key genes [e.g., uncoupling protein 2 (ucp2) and krüppel-like factor 15 (klf15)]. The results demonstrated the miRNA-mRNA regulatory mechanism related to grass carp physiological response to hypoxia, providing a novel theoretical foundation for understanding hypoxia tolerance in this species.