Abstract: 【Objective】 This study aimed to investigate the molecular mechanisms underlying heat stress-induced apoptosis of skeletal muscle cells in Hu sheep， thereby providing a theoretical support for elucidating the inhibitory role of heat stress on mutton production performance of Hu sheep.【Method】 A heat stress model was established by treating skeletal muscle cells in Hu sheep at 41.5 ℃. The cell viability was assessed using the CCK-8 kit to determine the appropriate heat treatment duration. The reactive oxygen species （ROS） level， superoxide dismutase （SOD） activity， and malondialdehyde （MDA） content in the cells were measured using kits. The relative expressions of heat shock proteins， apoptosis-related genes， and antioxidant-related genes， along with their corresponding encoded proteins， were determined using real-time fluorescence quantitative PCR and Western blotting.【Result】 The treatment at 41.5 ℃ could be used to build a heat stress model for skeletal muscle cells in Hu sheep. In skeletal muscle cells in Hu sheep under heat stress， ROS level extremely significantly increased （P<0.01， the same below）， MDA content significantly increased （P<0.05，the same below）， and SOD activity extremely significantly increased； a large amount of nuclear condensation and strongly stained blue fluore-scent spots were observed in the cells， with an extremely significant increase in cell apoptosis rate； the relative expressions of pro-apoptotic genes BAX and Caspase8 were significantly and extremely significantly increased， while the relative expressions of anti-apoptotic genes BCL-2 and BCL-XL were significantly and extremely significantly decreased. In skeletal muscle cells in Hu sheep under heat stress， the relative expression of the NOX2 gene and protein was significantly and extremely significantly increased. After intervention with the inhibitor Apocynin， the relative expression of NOX2 protein was significantly reduced， the ROS level and MDA content extremely significantly decreased， SOD activity significantly increased； the number of apoptotic cells with nuclear condensation and strong staining obviously decreased， resulting in an extremely reduction in the cell apoptosis rate. In skeletal muscle cells in Hu sheep under heat stress， the relative expression of p38 protein extremely significantly increased， while the relative expression of phosphorylated protein p-p38 significantly increased. After intervention with the inhibitor Apocynin， the relative expression of p-p38 protein was extremely significantly decreased.【Conclusion】 Heat stress induces a cascade of oxidative stress in the skeletal muscle cells of Hu sheep. By up-regulating the expression of the NOX2 gene and its encoded protein， heat stress leads to ROS over-accumulation that activates the p38 MAPK signaling pathway that regulates the expression of apoptotic genes， thus mediating apoptosis in Hu sheep skeletal muscle cells.