Linoleic acid deficiency, interleukin 1, retinoids and androgens have been implicated as causative factors in the follicular hyperkeratinization seen in acne. The goal of this study was to test the hypothesis that more androgens are produced in follicles of acne subjects compared to subjects without acne. Thirty-four subjects (males and females with and without acne) were studied. The activity of 5 alpha-reductase (5 alpha-R) and 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) was determined in keratinocytes cultured from the infrainfundibulum and epidermis. Mean enzyme activities were slightly higher in the acne groups compared to the groups without acne, but differences were not statistically significant, perhaps due to limitations of this in vitro model. The activity of both 5 alpha-R and 17 beta-HSD was significantly greater in infrainfundibular keratinocytes compared to epidermal keratinocytes in all subject groups. 17 beta-HSD activity was 2.5- to 7-fold greater than the activity of 5 alpha-R in infrainfundibular keratinocytes. The regulation of 17 beta-HSD by endogenous factors may be important in determining the directional activity of 17 beta-HSD and hence the local concentration of testosterone within the infrainfundibulum. Additional studies of the effects of androgens on follicular keratinization are needed.