Induction of apoptosis by cannabidiol and its main metabolites in human Leydig cells.

Cannabidiol (CBD) is one of the most prevalent and abundant cannabinoids extracted from the plant Cannabis sativa. CBD has been reported to induce male reproductive toxicity in animal models. In this study, we examined the effects of CBD and its main metabolites, 7-carboxy-CBD and 7-hydroxy-CBD, on primary human Leydig cells, which play a crucial role in male reproductive health. Our results showed that CBD, at concentrations below the Bayesian benchmark dose (BMD), inhibited the growth of human Leydig cells by arresting the cell cycle at G1/S transition, disrupting cell cycle regulators, and decreasing DNA synthesis. Concentration-response transcriptomic profiling identified that apoptosis was one of the top biological processes significantly affected by treatment with CBD for 24 h. The occurrence of apoptosis was confirmed by increased activation of caspase-3/7 and an increased proportion of annexin V and propidium iodide (PI)-positive cells. Similar to CBD, both 7-carboxy-CBD and 7-hydroxy-CBD decreased cell viability and induced apoptosis after treatment for 24 h. 7-Hydroxy-CBD and 7-carboxy-CBD showed lower cytotoxicity than CBD, and 7-carboxy-CBD had the lowest cytotoxicity among the three compounds. Our findings revealed that CBD and its main metabolites can cause adverse effects on primary human Leydig cells.