Stromal enlargement plays a key role in the development of benign prostatic hypertrophy in humans. Human prostatic fibroblasts were obtained from fetal and adult prostates and characterized as to their androgen and estrogen receptor status and growth in response to dihydrotestosterone (DHT), estradiol (E2), hydroxyflutamide (OH-FLU), hydrocortisone, basic fibroblast growth factor (bFGF), and epidermal growth factor (EGF). In addition, the ability of hormones and growth factors to induce the messenger RNA (mRNA) for the c-fos protooncogene was assessed as a measure of the early, direct effects of these compounds on cellular proliferation. Nuclear androgen receptors were demonstrable by immuno-cytochemistry in both fetal and adult cells. Nuclear estrogen receptor staining was negative. Neither E2nor hydrocortisone increased cellular proliferation. Both EGF and bFGF did in-crease cellular growth. DHT (10-8-10-7M) had a significant stimulatory effect on cell growth only in serum-free media. OH-FLU addition enhanced DHT induced proliferation. Changing the media during the course of the experiment obliterated the stimulatory effect of DHT. Both EGF (10 ng/ml) and bFGF (20 ng/ml) increased the mRNA for the c-fos protooncogene. DHT (10-7M) did not induce the mRNA for c-fos. We conclude that EGF, bFGF, and DHT (especially in combination with OH-FLU) increase the proliferation of human prostatic fetal and adult fibroblasts in vitro. E2has no effect on fibroblast proliferation. The stimulatory effects of EGF and bFGF are direct, whereas the effect of DHT appears to be indirect, possibly mediated via the increased production and/or secretion of growth factors.