Ex vivo method for assessing androgen synthesis and histological changes in rat testicular tissue slices treated with azole compounds.

As part of toxicity assessment in the development of pharmaceuticals and pesticides, ex vivo method using testicular tissue slices is suggested for evaluating the impact and underlying mechanisms of action of compounds on androgen synthesis. However, relevant comprehensive assessments, such as comprehensive profiling of androgens and related hormones and histological examination of tissue slices, remain limited. We evaluated an ex vivo method using testicular tissue slices by determining whether the amounts of each hormone synthesized in successive metabolic steps could be measured to reflect compound-induced effects. First, rat testicular tissue slices were incubated with ketoconazole (Ket) solutions (a representative azole fungicide that inhibits androgen synthesis) for 3 h; hormone assays of the supernatant and histological examination of the slices, including hematoxylin and eosin staining, ultrastructural examination, and immunohistochemistry, were conducted. Changes in androgen and precursor hormone levels were consistent with cytochrome P450 (CYP) 17A1 inhibition by Ket, together with morphological observations in Leydig cells that generally reflected the altered hormone profiles. Subsequently, eight azoles (hexaconazole, flusilazole, triticonazole, imazalil, penconazole, myclobutanil, triadimefon, and fluconazole) with inhibitory effects on CYP17A1 were examined. Hormone assays revealed changes suggestive of CYP17A1 inhibition for all the compounds, along with apparent differences in their inhibitory potency. Histological examination provided supportive morphological observations that were generally consistent with differences in hormone synthesis among the compounds. These results demonstrate that this ex vivo method can detect azole-induced disruptions in androgen synthesis and associated histopathological changes with fewer resources, thus offering a valuable tool for mechanistic toxicity studies.