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Mitochondrial Dysfunction and Oxidative Stress in Spermatogonia: Mechanisms of Heat Stress-Induced Spermatogenic Impairment in the Testis

CHEN Siyu^1,2,3, LU Jiawei^1,2,3, WEI Ke^1,2,3, ZHOU Lixin⁴, XUE Jia⁴, SHEN Linyuan^1,2,3, ZHU Li^1,2,3, GAN Mailin^1,2,3*

(¹National Key Laboratory of Swine and Poultry Breeding Industry, College of Animal Science and Technology, Sichuan Agricultural University, Chengdu 611130, China; ²Key Laboratory of Livestock and Poultry Biomics, Ministry of Agriculture and Rural Affairs, College of Animal Science and Technology, Sichuan Agricultural University, Chengdu 611130, China; ³Sichuan Provincial Key Laboratory of Livestock and Poultry Genetic Resources and Breeding, College of Animal Science and Technology, Sichuan Agricultural University, Chengdu 611130, China; ⁴Chengdu Animal Disease Prevention and Control Center, Chengdu 611130, China)

Abstract:

To investigate the regulatory effects of heat stress on mitochondrial function, oxidative stress, and cell fate in spermatogonia, in vivo experiments were conducted to establish a mouse testicular heat stress mod el. Testicular coefficient, sperm density, and motility were analyzed. Structural alterations in seminiferous tubules were examined by hematoxylin-eosin staining with measurements of their diameter and area, and mitochondrial ultrastructure was observed by transmission electron microscopy. Additionally, testicular transcriptome sequencing was performed to screen for differentially expressed pathways. In spermatogonia, mitochondrial membrane poten tial was examined using JC-1 probe, ROS (reactive oxygen species) levels were measured with DCFH-DA fluores cent probe, and lipid metabolism-related gene expression was quantified by real-time PCR. Cell proliferation and apoptosis were assessed by EdU and PI fluorescence staining, respectively. The results demonstrated that heat stress significantly reduced testicular coefficient, sperm density, and motility, and induced disorganization of seminifer ous tubules. Transcriptome analysis revealed significant enrichment of pathways related to mitochondrial function, TNF-mediated response, metabolic processes and oxidative stress. TEM (transmission electron microscopy) of tes ticular tissue revealed ultrastructural damage to mitochondria, while spermatogonia exhibited decreased mitochon drial membrane potential and disruption of the mitochondrial network, with upregulation of lipid metabolism gene ACSL4 and downregulation of SLC7A11. Cell proliferation was significantly inhibited, whereas apoptosis rate was markedly increased. Collectively, heat stress induced mitochondrial dysfunction, an oxidative stress elevation, and lipid metabolic reprogramming, thereby suppressing spermatogonial proliferation and promoting apoptosis. Mito chondrial dysfunction and oxidative stress may represent key early events mediating heat stress-induced reproduc tive impairment.

CSTR: 32200.14.cjcb.2026.03.0003