Steroid receptor profile and receptor stability in subfractions of human prostatic tissues

Summary Androgen (AR), progesterone (PR), and estrogen (ER) receptor contents in cytosol and salt-extractable nuclear subcompartments from 6 normal, 39 benign hyperplastic (BPH), and 7 malignant prostatic tissue specimens were analyzed by radioligand-binding assay techniques. In addition, the temperature stability of AR and PR was measured in another three BPH specimens. Five punch-needle biopsy samples from prostate cancers were also analyzed for AR and PR content. All receptor data were calculated from saturation analyses. The highest AR content was found in the cytosol and nucleic from malignant prostatic tissues. The highest PR concentrations were found in BPH cytosol, whereas nuclei of all types of tissues were negative with regard to this receptor. Markedly lower concentrations of ER were found in cytosol and nuclei from BPH as compared with malignant and normal tissues. PR was the most temperature-stable receptor; a marked receptor loss at room temperature was not registered until after 12 h. AR was stable for 4–5 h in cytosol and for 8–9 h in nuclei. Needle-biopsy specimens from prostate cancer showed highly variable and confusing results for AR and PR content, indicating that microassay studies using biochemical techniques on small tissue samples are unreliable and should not be recommended.     

Immunohistochemistry and biochemistry in detection of androgen, progesterone, and estrogen receptors in benign and malignant human prostatic tissue.

The relative distribution of androgen (AR), progesterone (PR), and estrogen receptors (ER) was localized and estimated in human prostate tissue by immunohistochemistry in five normal tissue samples, in eight benign hyperplastic (BPH) samples, in nine primary cancers, and in seven prostate cancer metastases. Moreover, three prostatic cancer cell lines (LNCaP, DU 145, and PC 3) were analyzed. A comparison between the results obtained by radioligand binding assays and immunohistochemistry was performed for the AR and PR. Using immunohistochemistry, the AR was exclusively detected in the nuclei of both benign and malignant prostatic epithelial cells. The highest proportion of AR-positive cells was found in BPH and in prostate cancer metastases as compared with normal prostatic tissue. In a majority of the cases, the PR was only present in the nuclei of stromal cells. Benign hyperplastic prostates contained higher proportions of PR-positive cells as compared with primary carcinoma. PR was sparse in epithelial cells. ER-positive stromal cell nuclei were only detected in carcinomatous prostates. A few ER-positive epithelial cell nuclei were found in one sample each of a BPH and normal prostate. All cells from the androgen-dependent, LNCaP, cell line and a majority of the cells from the androgen-independent, DU 145, cell line were AR-positive. In contrast, the cells from the androgen-independent, PC 3, cell line were all AR-negative. All three cell lines were PR- and ER-negative. The radioligand binding technique detected the AR in extracts from both the cytosol and the nucleus. Again BPH contained higher amounts of AR as compared with normal prostatic tissue. The LNCaP cells contained high amounts of cytosolic AR while cells from the DU 145 and PC 3 cell lines lacked detectable AR as estimated by biochemical techniques. There seemed to be a discrepancy between biochemically measured and immunohistochemically estimated receptor content.     

Regional variation of cytosol androgen receptors throughout the diseased human prostate gland

The heterogeneous histology of the normal and diseased human prostate is well established. This study has investigated variations in cytosol androgen receptor (AR) content throughout the diseased gland to establish the most suitable sites to obtain tissue for AR assay. Only then can AR be investigated as a potential predictor of response to endocrine treatment. In a transverse slice of an enucleated prostate showing benign prostatic hyperplasia (BPH) AR levels in 1-g segments varied from 109 to 1,212 fmol/g tissue (mean 483 +/- 273; median 406), with no negative areas. Areas of higher receptor concentrations corresponded to the glandular regions of sections obtained from a slice taken in juxtaposition; areas of low receptor concentrations corresponded to the stromal regions. A significant correlation (P less than .02) was observed between AR concentration and the proportion of glandular components of each segment. Specimens were also obtained from each of three sites from 38 prostates; 45% of all specimens contained AR; however, distribution of receptor throughout the prostate was uneven. AR were significantly more likely to be measured in the peripheral zone (71% positive) than in periurethral tissue (39% positive) whilst only 24% of specimens taken from the limit of the resection possessed AR binding capacity. Similar distribution patterns were observed in both benign and malignant prostates, although 16% more specimens from carcinomatous prostates contained receptor than did those from benign glands; this difference was maintained at each site. In addition receptor levels were consistently lower in benign than in malignant specimens. It is therefore desirable to know the histological composition of specimens used for AR measurement.     

The effects of flutamide on total DHT and nuclear DHT levels in the human prostate

The effects of flutamide, an antiandrogen, on prostate tissue concentrations of total DHT, DHT present in both crude and purified nuclear fractions, prostatic acid phosphatase (PAP) and plasma testosterone were studied and compared to similar parameters in untreated benign prostatic hypertrophy (BPH). Flutamide was given to patients with BPH in a dosage of 750 mg per day by mouth for 10–14 days prior to transurethral resection of the prostate. Total prostate DHT was significantly decreased to 3.95 ng/g in 12 flutamide-treated patients compared to values of 6.61 ng/g in 12 patients with untreated BPH. However, no significant difference was noted in the concentration of DHT present in the crude nuclear fraction of flutamide-treated patients (646 pg/mg DNA, N = 5) and untreated BPH (882 pg/mg DNA, N = 10); nor was DHT in the purified nuclear fraction significantly different in drug versus untreated patients (251 pg/mg DNA for flutamide versus 353 pg/mg DNA for untreated controls). PAP concentration in BPH prostates was 7.11 S.U./mg wet weight and was significantly higher than 2.98 S.U. per mg wet weight noted in flutamide-treated patients. Plasma testosterone tended to rise in the flutamide-treated patients compared to the untreated BPH but this was not statistically significant. The decrease in total prostate DHT without changes in nuclear DHT was unexpected and difficult to explain in terms of current tenets regarding the mechanism of androgen action. The following hypotheses are offered: (a) Flutamide may decrease transport of testosterone into cells, thereby decreasing total prostate DHT. (b) Inhibitory effects of flutamide on receptor-bound DHT translocation to nuclei may be difficult to detect since 95% or more of nuclear DHT may not be bound to a salt extractable receptor. (c) The binding of DHT directly to putative nuclear matrix receptor sites may dilute the effects of flutamide on blocking translocation of receptor bound DHT, resulting in very small differences in DHT present in purified nuclei difficult to detect with current methodology.     

Apoptosis and hormonal milieu in ductal system of normal prostate and benign prostatic hyperplasia

Aim: To study the apoptotic rate (AR) and the androgen and estrogen milieu in the proximal and distal ductal sys-tems of prostate, in order to help exploring the effects of these factors on prostatic growth and the pathogenesis of be-nign prostatic hypertrophy (BPH). Methods: The proximal and distal ends of the ductal system were incised from20 normal prostate as well as the hypertrophic prostate tissue from 20 patients with BPH. The AR was determined bythe DNA end-labeling method and dihydrotestosterone (DHT) and estrodiol (E_2), by radioimmunoassay. Results:There was no significant difference in DHT and E_2 density between the proximal and distal ends of the ductal systems innormal prostate. E_2 appeared to be higher in BPH than in normal prostatic tissues, but the difference was statistically in-significant. In normal prostatic tissue, the AR was significantly higher in the distal than in the proximal ends of theductal system (P<0.05), while the AR of the proximal ends was significantly higher (P <0.     

Effect of stilboestrol and testosterone on the incorporation of 75selenomethionine by prostatic carcinoma cells

Controversy still exists as to whether oestrogens exert a direct effect on the prostatic cell. Incorporation of 75Selenomethionine (SeM) was used as a measure of protein synthesis by prostatic carcinoma cells in vitro to investigate the action of hormones on prostatic carcinoma cells in tissue culture. Stilboestrol (DES) and stilboestrol diphosphate (Honvan) inhibited protein synthesis in a proportion of patients, while testosterone was stimulatory. A similar effect was noted in cells from patients with benign hyperplasia (BPH). This work confirms that oestrogens have a direct inhibitory effect on prostatic cells at high concentrations which can be attained in patients given intravenous stilboestrol diphosphate.     

Steroid receptors and hormone responsiveness of human prostatic carcinoma

Androgen (AR), estrogen (ER), and progesterone (PR) receptors have been found in human prostatic hyperplasia (BPH) and prostatic carcinoma (PC) (AR mainly in the epithelial compartment and ER in the stromal compartment). These steroid receptors have been studied in the cytosol of 59 prostatic cancers in order to select patients to be treated with endocrine therapy (orchiectomy, administration of antiandrogens). Tissues were collected through transvesical resection and needle biopsy. Tritiated metribolone (R1881), 17b-estradiol and promegestone (R5020) with the use of exchange assay and a dextran-coated charcoal method have been usually employed. In some specimens receptor content was also analyzed in the nuclear fraction. AR was found in the cytosol of 47 of 59 (79.66%), ER in 14 of 26 (53.85%), and PR in 13 of 15 (86.67%) tumors examined. AR was detected in the nuclear fraction of 11 of 26, ER in nine of 13, and PR in two of four tumors examined. AR was found to correlate with the histological grade of the cancers, whereas no correlation could be demonstrated between the histological grade and cytoplasmic or nuclear ER. On the basis of receptor studies, hormonal treatment was started with orchiectomy followed by the administration of cyproterone acetate (CPA) 50 mg twice a day in 26 eligible out of 38 patients available for follow-up with tumors examined for cytoplasmic AR. The response to hormonal therapy depended strictly on the presence of AR. The endocrine treatment used seems, in our opinion, to be the most advisable form of therapy, and is capable of blocking the uptake of androgens of extratesticular origin by prostatic cells. Although the clinical advantages of progestational therapy for human PC are not yet established, the presence of PR in prostatic carcinoma leads us to believe that progestational compounds as well as the progestational activity of CPA may be effective on tumor growth of tissues that are positive for PR.     

Zinc and cadmium concentration in prostatic carcinoma of different histological grading in comparison to normal prostate tissue and adenofibromyomatosis (BPH)

Summary Zn and Cd concentrations in tissue of normal prostate gland, BPH and prostatic carcinoma of different histological grades have been determined by atomic absorption spectrometry before therapy. We found a distinct biological anatagonistic effect between Zn and Cd in the human prostate gland. We have seen an increasing amount of Zn in BPH, but a decrease in prostatic cancer. In contrast, we found a continuous increase of cadmium concentration from the normal prostate via BPH through carcinoma.     

Androgen receptors detected by autoradiography in prostatic carcinoma and benign prostatic hyperplastic tissue

Androgen receptor (AR) content in prostatic tissues from patients with either cancer or benign prostatic hyperplasia (BPH) is of interest from at least two standpoints: receptors may be a feature of the pathogenesis of these conditions, and they may be important to the management and prognosis of prostatic cancer patients. For these reasons, a quantitative autoradiographic assay for AR content in prostatic tissues has been developed. Application of autoradiography to rodent tissues yielded results that were highly correlated with those from biochemical assays. Thus, the autoradiographic analyses with human tissues reported in this paper were undertaken. Average AR content in 22 prostatic carcinomas was lower than that in tissues from 14 patients with BPH; the median values of the affinity index, the quantitative estimate of receptor content, were 7.0 and 12.0, respectively. For the cancer tissues, a trend of declining receptor content with advancing stage of disease appeared but was not statistically significant. No association between receptor content and degree of tumor aggressiveness as measured by Gleason score and MD Anderson score was evident. Patient age and race were not related to receptor content in either type of tissue.     

Clinical study on estramustine binding protein (EMBP) in human prostate

To elucidate the characteristics of estramustine binding protein (EMBP) in human prostate, tissue EMBP concentration was examined in 42 benign prostatic hypertrophy (BPH), 34 untreated prostatic carcinoma (PC), 8 hormone refractory PC (hr-PC), as well as 13 control prostate human tissue samples by RIA using rat-EMBP antibody, and the concentration thus obtained was compared with dihydrotestosterone (DHT), prostatic acid phosphatase (PAP), prostate-specific antigen (PSA), and zinc, indices exhibiting androgen dependency in the prostate. EMBP concentration correlated significantly with DHT and PSA levels in the control prostate and BPH, but not in untreated PC. In BPH, EMBP concentration increased significantly after administration of fluoxymesterone (4 mg/day for 2 weeks), whereas it decreased significantly after estramustine phosphate (280 mg/day for 2 weeks). The EMBP/DHT ratio in moderately and poorly differentiated, and the hr-PC was significantly higher than in controls, BPH, and well-differentiated PC. In addition, untreated PC with an EMBP/DHT ratio of more than 40 showed significantly lower progression-free probability as compared with PC with an EMBP/DHT of less than 40. These results suggest that (1) EMBP in BPH and well-differentiated PC preserves androgen dependency, but not in moderately and poorly differentiated, nor in hr-PCs, indicating that EMBP is a protein different from PAP and PSA, and (2) that the tissue EMBP/DHT ratio might be useful as a marker for predicting disease progression. © 1996 Wiley-Liss, Inc.     

Sex steroid receptors in normal and hyperplastic human prostate

The concentrations of sex steroid receptors (per unit DNA) were measured in normal periurethral and peripheral prostatic tissue samples from seven men (mean age 64 years; range 54-71 years) undergoing cystectomy for bladder cancer, and in hyperplastic nodules from 15 men with BPH (mean age 69 years; range 60-89). Occupied androgen (AR) and estrogen (ER) receptors were measured with an improved exchange procedure, where receptor-binding sites were stabilized by a combinatorial procedure involving careful washout of extracellular secretory products (including proteases) prior to homogenization, inclusion of 0.5 mM phenylmethyl sulfonylfluoride (PMSF) and 20 mM molybdate in the exchange medium, and long-term incubation at 0-4 degrees C. Bound radioligands were separated by a hydroxylapatite (HAP) batch adsorption procedure. Maximal specific exchange binding of 3H-R 1881 or 3H-estradiol in total homogenates of human prostate samples was achieved after incubation periods of about 72 h at 0-4 degrees C. In contrast, progestin receptors (PR) were readily available for binding 3H-R 5020; thus overnight binding at 0-4 degrees C was routinely used to measure PR. Binding specificities and equilibrium binding constants (calculated from 8-point Scatchard plots, correcting for nonsaturable binding) were found to be characteristic for AR, PR, and ER, respectively. The receptor results obtained in this study demonstrate that no significant differences existed in total AR per unit DNA between hyperplastic and either central or peripheral prostatic tissue samples; PR was present in both zones of normal prostatic tissue as often as in BPH samples, with PR concentrations significantly lower in hyperplastic samples; and ER was irregularly detected in both normal and hyperplastic tissue in low concentration relative to AR and PR; the frequency of ER detection was much lower in BPH than in normal prostate tissue. Studies of steroid receptor content relative to enzyme markers specific for epithelial and stromal cells in BPH samples showed a positive correlation between acid phosphatase activity (a specific marker for epithelial cells) and both AR and PR. No correlation was observed between AR or PR with either prolyl hydroxylase or myosin ATPase (specific markers for stromal cells). These observations suggest that PR, as well as AR, is primarily associated with the epithelial elements of prostate. Because of the relative infrequency of ER, similar correlation of ER with enzyme markers was not possible.     

Changes in the endocrine environment of the human prostate transition zone with aging: simultaneous quantitative analysis of prostatic sex steroids and comparison with human prostatic histological composition

BACKGROUND: It is well-known that the incidence of benign prostatic hyperplasia (BPH) increases with aging. The age-dependent changes in the ratio of serum sex steroid concentrations may play a role in BPH development. To clarify the relationship between the prostatic tissue concentrations of these steroids and age, we established a precise method of simultaneous quantitative analysis for prostatic sex steroids and used this method to investigate the tissue concentrations of three major sex steroids (testosterone, dihydrotestosterone, and estradiol) in the human prostate. METHODS: The methodology for the simultaneous quantitative analysis of prostatic sex steroids was established using castrated rat prostatic tissue, coupled with internal standards, for androgen-deprived medium, and the validation of the method was examined. Human prostatic tissues were collected during surgery and immediately frozen at -70 degrees C. Using our method, the steroidal fractions were extracted, purified, and quantified. The proportions of stroma, epithelium, and glandular lumen were measured on each histological specimen, using an image analyzer. RESULTS: The validation tests showed that our method of quantitative analysis was precise and sensitive enough for the quantification of testosterone, dihydrotestosterone, and estradiol in the prostate. In humans, the prostatic dihydrotestosterone concentration decreased with age, but the concentrations of testosterone and estradiol showed no relation with age. Therefore, the ratio of estradiol to dihydrotestosterone concentration (E2/DHT) in prostate increased with age. The E2/DHT ratio showed a significant positive correlation with the proportion of stroma. CONCLUSIONS: The age-dependent decrease in prostatic dihydrotestosterone and constant estradiol concentration lead to a relatively estrogen-dominant environment compared to that at younger ages. We assume that this relatively estrogen-dominant status induces stromal proliferation by some mechanism and leads to the development of BPH.     

前列腺肿瘤中雄激素受体基因突变的研究

为了探讨雄激素受体基因突变与前列腺癌和前列腺增生症发生发展的关系，采用多聚酶链反应单链构象多态分析技术对未经治疗的２０例前列腺癌和２０例前列腺增生症组织中雄激素受体基因第８外显子突变情况进行检测。结果：在２例晚期前列腺癌和１例前列腺增生症组织中检出雄激素受体基因第８外显子突变，２例晚期前列腺癌患者经内分泌治疗后均在９个月内复发。结果认为：前列腺癌中雄激素受体基因突变可能是不常见的，但雄激素受体基因突变与前列腺癌对内分泌治疗产生抵抗关系密切；对雄激素受体基因突变与前列腺增生症的关系有必要作进一步研究。     

前列腺癌、前列腺增生症患者血清骨保护素测定的临床价值

目的探讨前列腺癌（PCa）患者和前列腺增生（BPH）症患者血清中骨保护素（OPG）浓度的差异以及前列腺癌患者骨保护素浓度与血清前列腺特异性抗原（PSA）水平、前列腺体积是否具有相关性。方法采用双抗体夹心酶免法（ELISA）测定40例前列腺癌患者及40例前列腺增生患者血清OPG浓度,同时采集PCa患者的前列腺体积及PSA值。比较PCa患者及BPH患者血清OPG浓度的差异以及前列腺癌患者中OPG浓度与PSA、前列腺体积之间有无相关性。结果 PCa患者的血清OPG浓度平均值水平〔（14 900.19±5 168.65）pg/mL〕显著高于BPH组〔（10 457.87±4 786.29）pg/mL〕,差异有显著性意义（P〈0.01）。PCa患者血清OPG浓度与PSA值及前列腺体积之间均无明显相关性（r分别为=0.221、0.138,P均〉0.1）。结论血清OPG浓度对鉴别PCa和BPH有重要临床价值,PCa患者血清OPG浓度与血清PSA值及前列腺体积无明显相关性。     

Total protein and acid phosphatase concentrations in prostatic fluid from patients with BPH compared to carcinoma

Objective

To obtain evidence of metabolic changes in the human prostate associated with prostate pathology, in particular carcinoma of the prostate, by identifying and evaluating associated changes in prostatic secretory products.

Methods

Expressed prostatic fluid (EPF) from 36 patients with carcinoma, 128 with BPH histologically confirmed, and 148 with clinical BPH was subjected to determination of protein (Lowry; UV 280 nm absorption), enzymatic (DMA modified Row procedure) acid phosphatase (AcP), and immunologically identified (Tandem^®-PAP immunoenzymatic assay) prostatic acid phosphatase (PAP) concentration.

Resultsp

The important EPF findings are the following: (1) Protein concentrations (Lowry and UV determinations) are significantly increased in carcinoma as compared to histologic BPH, (2) AcP and PAP secretions remain stable in carcinoma versus BPH, and (3) AcP and PAP/Lowry protein ratios are significantly lower with carcinoma.

Conclusions

These findings of increased protein and the decreased relative secretions of AcP and PAP to total protein (ratio) in EPF from patients with carcinoma compared to BPH support and help to characterize the diffuse metabolic alteration in the prostate associated with prostate carcinoma. EPF observations identify potential metabolic changes occurring in prostate carcinoma that may have potential clinical and investigative relevance.     

Tamoxifen decreases human prostate CPK concentration

Creatine phosphokinase (CPK) was measured in whole prostate tissue of patients with benign prostatic hypertrophy (BPH) who were given Tamoxifen and/or megestrol acetate for seven days prior to transurethral resection of the prostate. Tamoxifen and Tamoxifen plus Megace®, but not Megace® alone, significantly decreased CPK concentrations in whole prostate tissue in comparison to untreated controls, suggesting that CPK is an estrogen-dependent enzyme in the prostate. CPK concentration was also measured separately in mechanically separated prostate stroma and epithelium. The levels of CPK were 2.19 times higher in stroma than epithelium. This study suggests that CPK may be a marker for estrogen stimulation of the stroma in BPH tissue.     

Determination of androgen receptors in human benign prostatic hypertrophy with two synthetic radiolabeled ligands

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.     

Studies on prostatism of males fifty years of age and older--analysis on results of a second mass screening for prostatic diseases in Tanno town

We conducted a second mass screening for prostatism on males 50 years of age and older in the Tanno town area, to determine the incidence of benign prostatic hypertrophy (BPH) and prostatic carcinoma. We also studied how often elderly males had symptoms related to prostatism according to their age. The incidence of BPH was 10.0% in these males who were received the first and/or second mass screening as well as those treated previously, when BPH was defined as the prostate with a moderately or greater enlarged size upon digital palpation. The incidence of BPH was 8.9% in males fifty years of age and older who lived in Tanno town. The figures tended to elevate with the advance of age. Two new patients with prostatic carcinoma were discovered during this second mass screening, which resulted in a total of 8 patients discovered in this area. Of these 8 patients, 6 had been found in the first screening. The incidence of prostatic carcinoma was 1.4% in those males who received the first or second mass screening as well as those treated previously, and that in all the males who lived in the town was 1.0%. Further efforts will be necessary to establish a more convenient system that can provide less costly screening procedures and more effective diagnostic procedures to detect localized prostatic carcinoma. With advance in age the male tended to have a higher incidence of prostate with a moderately or markedly enlarged size, and symptoms related to prostatism. In addition, the urinary flow rate tended to decrease, irrespective of the prostate size. The results supported the significance of mass screening for prostatism.     

Estrogen receptors and clinical correlations with human prostatic disease

Measurement of estrogen binding in human prostate using high-pressure liquid chromatography (HPLC) revealed the presence of cytosolic estrogen receptors (ER) both in benign prostatic hyperplasia (BPH) and adenocarcinoma. Receptor concentrations correlated with several histopathologic features in the specimens analyzed. Estrogen receptor levels generally were higher in BPH than in cancer specimens although there was a subgroup of patients with poorly differentiated carcinoma with levels higher than those of BPH, HPLC can be used for measuring ER in 50 microliters of cytosol, and thus needle biopsy specimens will be analyzed routinely for ER with this micromethod.