Compromised extracellular vesicle-mediated communication impairs bull spermatozoa function

Oviduct-derived extracellular vesicles (EVs) modulate spermatozoa function through bioactive cargo transfer; however, how stress-induced alterations in EVs affect spermatozoa function remains poorly understood. In particular, this study aimed to assess whether EVs exert differential, potentially uncoupled effects on distinct sperm functions. In this study, bovine oviductal epithelial cells (BOECs) were cultured with or without exposure to cobalt chloride (CoCl₂) to induce oxidative stress (OS). Isolated EVs were characterized for physicochemical properties and proteomic composition, and their functional effects on bovine spermatozoa were assessed using fluorescence staining and microscopy. EVs derived from stressed BOECs exhibited altered physicochemical characteristics and a distinct proteomic profile compared with unstressed EVs. Using unstressed EVs, a dose-dependent enhancement of spermatozoa viability, capacitation, and acrosome reaction was observed, with maximal effects at 1 × 10⁸ EVs (p < 0.001). The minimum effective concentration of 1 × 10⁶ EVs was selected for subsequent experiments. At this concentration, EVs from stressed BOECs failed to improve spermatozoa viability or acrosome reaction, although capacitation levels remained comparable to those induced by unstressed EVs, which was significantly higher than the control group (p < 0.001). These findings demonstrate that OS reshapes the molecular cargo and bioactivity of oviductal EVs, selectively impairing their ability to support the acrosome reaction while preserving capacitation, highlighting their role as stress-sensitive modulators of reproductive success.