Inductive stimuli from mesenchymal cells direct the growth and phenotypic differentiation of epithelium in prostate gland development. This process is androgen regulated, yet specific mechanisms are unknown. To address mechanisms, we have characterized paracrine acting factors from fetal rat urogenital sinus (UGS) organ cultures and derived mesenchymal cell lines. UGS mesenchymal cells secrete a factor (UGIF) which alters epithelial cell phenotype, stimulates secretory protein synthesis, and inhibits proliferation, suggesting this protein functions as a differentiation-inducing factor. Physicochemical, biological, and immunological properties have been determined and the UGIF protein purified to homogeneity. UGIF inhibits cell proliferation, stimulates protein secretion, and alters phenotype in PC-3 and LNCaP human prostatic carcinoma cell lines, suggesting a tumor suppressor function. A variant, phenotypically transformed UGS mesenchymal cell strain showed decreased UGIF expression coordinate with a stimulated secretion of activated TGF-B2 not observed in the parent cell line. Mutations or alterations in UGIF expression coupled with alterations in other growth factors may produce a growth factor imbalance, potentially relevant to the initiation and progression of prostatic cancer. The following studies are proposed: To develop antibody and oligonucleotide probes to UGIF; To determine the spatial and temporal expression of UGIF and TGF-B isoforms in human normal prostate and in clinical stages of human prostatic cancer; To determine the primary structure of human UGIF cDNA and characterize mutational defects in prostate cancer by analysis of PCR amplified cDNA products encoding UGIF and; To characterize the in vitro and in vivo responses of PC-3 and LNCaP human prostatic adenocarcinoma cells to UGIF. Antibody and cRNA probes will be used for immunocytochemistry and in situ hybridization. The mixed oligonucleotide primed amplification procedure (MOPAC) will used to analyze human UGIF cDNA, generate probes, and clone full length cDNA. UGIF cDNA from stages of human prostatic carcinoma will be analyzed by PCR/GC-clamp DGGE methods and sequenced to map putative mutations. PC-3 and LNCaP cell exposed to purified rat UGIF +/- androgens will be analyzed for growth and marker protein expression in vitro to correlate with in vivo effects. The athymic nude mouse host system will be used with PC-3 and LNCaP cells inoculated with UGIF secreting cells or UGIF containing Alzet Micro-Osmotic pumps to assess in vivo effects. Long range goals are to understand the specific mechanisms of UGIF action in prostate neoplasia and the potential therapeutic value of UGIF as a prostate tumor suppressor.
Showing the most recent 10 out of 36 publications
