mibolerone and Prostatic-Hyperplasia

The current study was undertaken, using cultures of prostatic epithelial and stromal cells, to determine the functional interactions between androgens, basic fibroblast growth factor (FGF2) and transforming growth factor-beta 1 (TGF beta 1) and their importance in maintaining stromal homeostasis. Treatment of stromal cells with TGF beta 1 significantly increased intracellular FGF2 and FGF2 sequestered to the extracellular matrix. FGF2 was also detected in stromal conditioned medium (SCM), but at levels 70-fold less than found in cell lysates. TGF beta 1 (0.1 ng/ml) treatment caused an initial increase of 86% in secreted FGF2 levels, but high concentrations of TGF beta 1 (5 ng/ml) decreased FGF2 levels by 38%, relative to the untreated control. Further studies showed that epithelial conditioned medium (ECM), androgen-treated, stromal conditioned medium (ASCM), but not SCM were mitogenic for stromal cells. Both ECM and ASCM caused a threefold increase in DNA synthesis. FGF2 may be the mediator of these interactions, since the mitogenic effect of both ECM and ASCM was significantly reduced by the addition of anti-FGF2 neutralising antibody. We hypothesise that the lack of response of stromal cells to SCM is due to TGF beta 1 blocking the mitogenic effect of FGF2. Thus down-regulation of TGF beta 1 synthesis, by androgens, results in stromal proliferation by ASCM.

Topics: Cell Division; Cell Separation; Cells, Cultured; DNA; Epithelium; Fibroblast Growth Factor 2; Flow Cytometry; Humans; Male; Nandrolone; Prostate; Prostatic Hyperplasia; Testosterone Congeners; Transforming Growth Factor beta

Stromal cells derived from collagenase-digested benign hyperplastic adult prostates were isolated and grown in culture. Androgen and oestrogen receptor status were determined and growth in response to mibolerone (a synthetic androgen) and oestradiol-17 beta was measured. In addition, the ability of oestrogens to regulate the androgen receptor in stromal cells was investigated. [3H]Thymidine incorporation into DNA was stimulated by mibolerone in primary and secondary cultures, but sensitivity was lost with subsequent passages. Androgen stimulation of [3H]thymidine incorporation was consistently inhibited by the anti-androgen cyproterone acetate. Oestradiol-17 beta also stimulated [3H]thymidine incorporation into DNA, and this effect was inhibited by the anti-oestrogen tamoxifen. Sensitivity to oestradiol was lost with subsequent passages. A combination of mibolerone and oestradiol was not synergistic in increasing [3H]thymidine incorporation into DNA, but maximal stimulation occurred at 100-fold lower concentrations of mibolerone and oestradiol when the two hormones were applied in combination. Specific high-affinity [3H]mibolerone- and [3H]oestradiol-binding sites were demonstrated by radioligand binding in intact cells. The affinity for oestradiol binding to its receptor exceeded that quantified for mibolerone binding to the androgen receptor, whilst the number of oestradiol-binding sites was approximately tenfold less than that quantified for mibolerone. Treatment with oestradiol down-regulated the number of [3H]mibolerone binding sites 1.7-fold (P < 0.005) as early as day 2 after oestradiol treatment. In conclusion, we successfully cultured stromal cells derived from hyperplastic prostates which retained sensitivity to androgen and oestrogen.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Cells, Cultured; Estradiol; Humans; Male; Nandrolone; Prostate; Prostatic Hyperplasia; Protein Binding; Receptors, Androgen; Stimulation, Chemical; Tamoxifen; Testosterone Congeners

This paper presents an approach for the assessment of the androgen receptor (AR) status in benign prostatic hyperplasia (BPH) and prostate cancer (PCa) tissues. Evaluation of AR was carried out in both soluble and nuclear fractions by a standard competition method, using tritiated mibolerone as radioligand. Based on our experience with breast and endometrial cancer, this approach focused on both type I (high affinity, low capacity) and type II (reduced affinity, higher capacity) binding sites, aiming mainly at establishing a putative "functional" receptor mechanism, i.e., the presence of type I AR in both cytosol and nucleus. Ancillary studies were carried out to exclude a potential overestimation of the AR content by interference with other steroid receptors, namely, progesterone (PgR) or glucocorticoid (GcR) receptors. Results showed that the interaction by PgR or GcR upon AR measurement was not relevant. The distribution of AR, namely the percent of positivity either in a single or in both cell compartments, was not significantly different in BPH (N = 32) or PCa (N = 24) tissues. For type I binding, the percent of positivity in both soluble and nuclear fractions (i.e., the "functional" AR status) was very close to that observed for other endocrine-related tumors, like breast cancer. Concentrations of type I AR appeared significantly higher in PCa than in BPH tissues; this was true for both soluble and nuclear fractions. In contrast, no significant difference was found in type II AR concentrations in either cell fraction.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aged; Binding Sites; Humans; Male; Nandrolone; Predictive Value of Tests; Prostate; Prostatic Hyperplasia; Prostatic Neoplasms; Radioligand Assay; Receptors, Androgen; Testosterone Congeners; Tritium

In rat uterus and prostate, 7 alpha, 17 alpha-dimethyl-19-nortestosterone (DMNT) binds to the androgen receptor specifically and with high affinity. However, this steroid does not bind to glucocorticoid receptors, since it does not displace binding of [3H]triamcinolone acetonide in calf thymus cytosol. In calf uterine and human breast tumor cytosols DMNT binds to the androgen and progesterone receptors, since binding of [3H] DMNT is displaced by unlabeled 16 alpha-ethyl-21-hydroxy-19-nor-4-pregnene-3, 20-dione triamcinolone acetonide, and 5 alpha-dihydrotestosterone (DHT). Conversely, binding of [3H]16 alpha-ethyl-21-hydroxy-19-nor-4-pregnene-3,20-dione is effectively competed for by unlabeled DMNT but not by DHT. The observed differences in binding of [3H]DMNT to rat and calf uterine cytosols suggest the species specificity of progesterone receptors. Unlike DHT, DMNT has no appreciable binding to human sex-steroid binding globulin. These findings suggest DMNT as a suitable ligand for measurement and characterization of androgen receptors in rat and human prostate.

Topics: Animals; Blood Proteins; Breast; Cattle; Cell Nucleus; Cytosol; Dihydrotestosterone; Estrenes; Female; Humans; Male; Metribolone; Nandrolone; Pregnenediones; Prostate; Prostatic Hyperplasia; Rats; Rats, Inbred Strains; Receptors, Androgen; Receptors, Progesterone; Substrate Specificity; Uterus

Accurate quantitation of androgen receptors requires a radioactive ligand which has affinity and specificity for the receptor and which is stable to metabolic enzymes. In this report, we have characterized the properties of 7 alpha,17 alpha-dimethyl-17 beta-hydroxy-4-estren-3-one (mibolerone) in human benign hyperplastic prostate cytosol and compared them to those of 17 beta-hydroxy-17 alpha-methyl-estra-4,9,11-trien-3-one (R1881). Mibolerone was found to have an affinity (Kd = 1.5 nM) greater than R1881. (Kd = 2.3 nM) for the androgen receptor in human prostate tissue. Surprisingly, mibolerone was found to bind with high affinity to the progesterone receptor in both human prostate (Kd = 5.9 nM) and rabbit uterus (Kd = 1.1 nM). However, binding to this receptor in both species could be blocked with a 500-fold excess of triamcinolone acetonide. [3H]Mibolerone binding to the androgen receptor was competed effectively with unlabeled dihydrotestosterone, R1881, and mibolerone but not by progesterone, diethylstilbestrol or R5020, in the presence of triamcinolone acetonide. Interestingly, mibolerone was more resistant to metabolism than R1881 in prostate cytosol when exposed to elevated temperatures (30 degrees C) for extended periods of time. However, when exposed to high-intensity ultraviolet irradiation, both compounds lost 50% of their binding ability in about 30 minutes. Mibolerone was found to have a very low affinity (Ki = 540 nM) for human sex steroid binding protein. These studies demonstrate that mibolerone is a useful ligand for androgen receptor assays. They also emphasize the need for including competitors of progesterone receptor binding in assays utilizing this steroid for androgen receptor measurements.

Topics: Animals; Chemical Phenomena; Chemistry; Dihydrotestosterone; Estrenes; Female; Humans; Male; Metribolone; Nandrolone; Promegestone; Prostate; Prostatic Hyperplasia; Rabbits; Receptors, Androgen; Receptors, Progesterone; Testosterone

Nuclear and cytosolic androgen receptor concentrations in tissues of human benign prostatic hypertrophy (BPH) were determined by use of methyltrienolone (R-1881) and 7 alpha,17 alpha-dimethyl-19-nortestosterone (DMNT) as radiolabeled ligands. Cytosolic R-1881-binding sites were 46.1 +/- 43 fmol/mg protein and nuclear R-1881-binding sites were 51.8 +/- 42 fmol/mg protein. DMNT-binding sites in cytosol were 44.3 +/- 38 fmol/mg protein and in nuclear extract 73.4 +/- 64 fmol/mg protein. No significant correlation was found between the number of R-1881- and DMNT-binding sites in either cytosol or nuclear extracts. Cytosolic or nuclear androgen receptor content was not significantly correlated with the percentage of epithelial or stromal cells as determined from the corresponding histological sections. In BPH tissue with marked cystic degeneration, very low androgen receptor levels were found.

Topics: Cell Nucleus; Cytoplasm; Cytosol; Epithelium; Estrenes; Humans; Male; Metribolone; Nandrolone; Promegestone; Prostate; Prostatic Hyperplasia; Radioligand Assay; Receptors, Androgen; Receptors, Progesterone; Testosterone Congeners

The binding nature of mibolerone in cytosols and nuclear extracts from hypertrophic human prostate was examined in comparison with that of R 1881. The binding of mibolerone in the cytosol and nuclear extract was single and of high affinity when evaluated by the method of Scatchard (1949). Binding of mibolerone with testosterone-binding globulin was not detected. The sedimentation coefficients of the binder for mibolerone in the cytosol and nuclear extract were 10.6 S and 3.6 S, respectively. When triamcinolone acetonide was induced in the binding medium, inhibition of mibolerone binding in the cytosol by testosterone and dihydrotestosterone was potentiated and this may imply that the binding observed in the presence of triamcinolone acetonide was responsible for the binding of the androgen receptor. In the nuclear extract, the binding was attributable mainly to the androgen receptor irrespective of the presence or absence of triamcinolone acetonide. These properties of the binding observed in the hypertrophic human prostate were almost same as those of the binding with R 1881. Although maximum binding sites measured using mibolerone were correlated with those using R 1881 in the cytosols as well as in the nuclear extracts, the values obtained with mibolerone were slightly greater than those with R 1881. Thus, mibolerone seems to be a suitable ligand for measuring the androgen receptor, but when compared with R 1881 no special merits in using mibolerone were detected.

Topics: Cell Nucleus; Cytosol; Estrenes; Humans; Male; Metribolone; Nandrolone; Prostatic Hyperplasia; Receptors, Androgen; Receptors, Steroid