BACKGROUND: Prostatic stroma affects both proliferation and differentiation of epithelial cells but the factors involved remain poorly understood. In order to identify and characterize potential paracrine mediators, we studied the effects of human prostate fibroblast-conditioned media (PFCM) in three bioassay systems. METHODS: The first bioassay uses transferrin secretion by cultured rat Sertoli cells as an endpoint for differentiating activity. Factors active in this (heterologous) assay were compared to PModS, a mediator of mesenchymal-epithelial interactions in the testis, also produced by rat prostate stromal cells. The two other (homologous) bioassays use LNCaP tumor cells or subcultured human prostate epithelial cells (PEC) as targets. Differentiation is evaluated by prostate-specific antigen (PSA) secretion and reverse transcriptase-polymerase chain reaction (RT-PCR) for a number of markers of epithelial function. Proliferation is assayed by measurements of DNA and thymidine incorporation. RESULTS: PFCM markedly stimulates transferrin production by Sertoli cells. The main factor(s) involved are acid stable and bind to heparin. However, both their size (approximately 37 kDa) and their behavior on reversed-phase chromatography differ from that of PModS. Although PFCM increases total RNA content of LNCaP, it does not increase or restore differentiated function of LNCaP or PEC. Proliferative effects are observed in LNCaP and these effects cannot be neutralized by an antiserum directed against basic fibroblast growth factor (bFGF). Antiproliferative effects are observed in PEC and these effects are largely due to transforming growth factor-beta (TGF-beta). CONCLUSIONS: PFCM provokes differentiating effects in a Sertoli cell bioassay, but unlike with rat stromal cells, the factor(s) involved differ from PModS. In the two homologous systems studied, differentiating effects could not be demonstrated and discordant effects were noted on proliferation. Various bioassay systems will be required to identify the spectrum of mediators present in PFCM.