Dehydroepiandrosterone (DHEA) is used as a dietary supplement and can be metabolized to androgens and/or estrogens in the prostate. We investigated the hypothesis that DHEA metabolism may be increased in a reactive prostate stroma environment in the presence of proinflammatory cytokines such as transforming growth factor beta1 (TGFbeta1), and further, whether red clover extract, which contains a variety of compounds including isoflavones, can reverse this effect. LAPC-4 prostate cancer cells were grown in coculture with prostate stromal cells (6S) and treated with DHEA +/- TGFbeta1 or interleukin-6. Prostate-specific antigen (PSA) expression and testosterone secretion in LAPC-4/6S cocultures were compared with those in monocultured epithelial and stromal cells by real-time PCR and/or ELISA. Combined administration of TGFbeta1 + DHEA to cocultures increased PSA protein secretion two to four times, and PSA gene expression up to 50-fold. DHEA + TGFbeta1 also increased coculture production of testosterone over DHEA treatment alone. Red clover isoflavone treatment led to a dose-dependent decrease in PSA protein and gene expression and testosterone metabolism induced by TGFbeta1 + DHEA in prostate LAPC-4/6S cocultures. In this coculture model of endocrine-immune-paracrine interactions in the prostate, TGFbeta1 greatly increased stromal-mediated DHEA effects on testosterone production and epithelial cell PSA production, whereas red clover isoflavones reversed these effects.