Testicular CNR1 and CNR2 expression in stray dogs from Campania Region, Italy: insights from a One Health sentinel model

In Campania Region (Italy), illicit drug residues, including cocaine, have been detected in surface waters, highlighting the occurrence of emerging anthropogenic contaminants. Within a One Health perspective, stray dogs represent useful sentinels for tracking environmental risks. In this context, the endocannabinoid system contributes to male reproductive homeostasis and inflammatory regulation, making cannabinoid receptor 1 (CNR1) and cannabinoid receptor 2 (CNR2) relevant candidates for investigating testicular remodelling. This study investigated changes in CNR1 and CNR2 expression during progressive spermatogenic impairment in canine testes. To this aim, twenty male stray dogs from the Campania Region were enrolled and classified according to spermatogenic status as follows: normal spermatogenesis (Group A), altered spermatogenesis (Group B), severely impaired spermatogenesis (Group C), and intratubular seminoma (Group D). CNR1 and CNR2 were assessed by immunohistochemistry and Western blot. CNR1 showed weak to moderate diffuse cytoplasmic immunopositivity in spermatogonia and primary spermatocytes in Group A. Group B displayed reduced positivity confined to scattered residual germ cells and peritubular areas. Group C exhibited the highest immunoreactivity across all groups, with markedly increased staining in residual germ cells and peritubular compartments. Group D showed heterogeneous staining in tumour and stromal cells. CNR2 displayed scattered positivity in spermatogonia, primary spermatocytes and interstitial cells in Group A, reduced germ cell signal in Groups B and C, and stromal positivity in Group D, including cells morphologically compatible with macrophage-like elements and mesenchymal components. Western blot analysis showed reduced CNR1/CNR2 protein signal in Group B compared with Group A, followed by increased expression in Groups C and D. Overall, these findings suggest that CNR1 and CNR2 expression changes during progressive impairment of spermatogenesis and neoplastic remodelling. However, further studies integrating environmental exposure assessment and cell specific co-localization are needed to clarify the biological significance of these receptors in canine testicular damage.