<p>The blood-testis barrier (BTB), a unique structure established through intercellular connections of Sertoli cells, establishes a protective microenvironment for spermatogenesis and male fertility. Cadmium (Cd), known for its toxicity, is ubiquitously present in the environment. Here, our findings revealed that Cd exposure compromises BTB integrity, as demonstrated by decreased expression of BTB-associated proteins and elevated D_signal/D_radius values. Mechanistically, we demonstrated that activation of the CXCL2/CXCR2 axis contributes to Cd-induced BTB impairment, as evidenced by experiments using a CXCR2 inhibition model. As key BTB components, Sertoli cells rely on autophagy to maintain their physiological functions. However, the specific role and mechanism of Sertoli cell autophagy in Cd-induced BTB damage remain unknown. Notably, our results showed that autophagy inhibition aggravated the Cd-induced BTB disruption and testicular CXCL2/CXCR2 axis activation in mice, whereas autophagy activation alleviates Cd-evoked BTB disruption and testicular CXCL2/CXCR2 axis activation. Further verification by Sertoli cell specific <i>Atg5</i> knockout mouse model showed that the autophagy suppression exacerbated Cd-induced BTB disruption and upregulated the expression of CXCL2. Collectively, our finding points out that ATG5-dependent autophagy in Sertoli cells protects against Cd-induced BTB disruption via perturbing CXCL2/CXCR2 axis. Our study not only reveals a novel molecular mechanism underlying Cd-induced reproductive toxicity but also provides potential therapeutic targets for male infertility intervention.</p>