Involvement of estrogen receptors in prostatic diseases

Accumulating evidence shows that estrogens participate in the pathogenesis and development of benign prostatic hyperplasia and prostate cancer by activating estrogen receptor α. In contrast, estrogen receptor β is involved in the differentiation and maturation of prostatic epithelial cells, and thus possesses antitumor effects in prostate cancer. However, the natural ligands of estrogen receptor β are not fully understood, and its mode of action according to its ligands and the binding sites located in the promoter regions of downstream genes remains to be elucidated. Here, we review recent experimental investigations of estrogen receptors and their urological relevance. Estrogen receptor‐mediated signaling in the prostate is essential together with the androgen receptor‐mediated pathway, providing a new therapeutic target for prostatic diseases.     

Expression of estrogen receptor in diseased human prostate assessed by non-radioactive in situ hybridization and immunohistochemistry

To understand the role of estrogen in the pathogenesis of benign prostatic hyperplasia, expressions of estrogen receptor (ER) mRNA and ER protein by in situ hybridization and by immunohistochemistry, respectively, were investigated in human prostatic tissues. In non-malignant region, ER mRNA and ER protein were found in cytoplasm and nucleus, respectively, of stromal cells, but not in glandular epithelial and basal cells. In benign regions, ER mRNA/ER protein positive cells were found in fibromyoadenomatous and myoadenomatous hyperplasia, but not in adenomatous hyperplasia. A striking feature was periacinar arrangement of ER mRNA/ER protein positive stromal cells in all prostate carcinoma treated with androgen withdrawal. The ER mRNA/ER protein positive cells were immunohistochemically identified as fibroblasts, myoblasts, and smooth muscle cells. These results indicate that stromal cells are the primary target of estrogen in prostate, and that androgen withdrawal upregulates the expression of ER gene. © 1995 Wiley-Liss, Inc.     

Androgen binding sites on nuclear matrix of normal and hyperplastic human prostate

To further characterize human prostatic androgen receptor, nuclei were isolated from normal prostate (no. = 3) and benign prostatic hyperplasia specimens (no. = 10). High ionic strength (0.6 M KCl) treatment of nuclei released nuclear extractable androgen receptor and DNase I digestion then yielded nuclear matrices. Androgen receptor was quantified in the nuclear extract and nuclear matrix preparations by Scatchard analysis of specific R1881 binding. Only 1 of the 3 normal tissues had extractable androgen receptor (113 fmol. per gm. of tissue) while the mean concentration of extractable androgen receptor for BPH was 189 fmol. per gm. of tissue. The mean concentrations of matrix-bound androgen receptor were 325 fmol. per gm. of tissue and 548 fmol. per gm. of tissue for normal and hyperplastic prostate, respectively. The androgen binding sites on nuclear matrix may represent the functional intranuclear androgen receptor and a characterization of these sites may provide an understanding of the etiology of BPH.     

Comparison of the R-3327H rat prostatic adenocarcinoma to human benign prostatic hyperplasia and metastatic carcinoma of the prostate with regard to steroid hormone receptors

Quantitative analyses of cytosolic steroid hormone receptors were performed on nine tumors from the transplantable rat prostatic adenocarcinoma R-3327H. Androgen receptors and estrogen receptors were found in eight of nine and five of six tumors, respectively. None of the tumours analyzed contained detectable progestin or glucocorticoid receptors (four and seven tumors, respectively). The apparent equilibrium dissociation constants for the androgen and estrogen receptors were 0.7-4.3 nM and 0.6-1.8 nM, respectively. The apparent equilibrium Bmax values (maximum number of binding sites) were 1,500-25,000 fmoles/gm tissue for the androgen receptor and 640 to 5,800 fmoles/gm tissue for the estrogen receptor. A comparison between the receptor contents of the R-3327H rat tumor and human benign prostatic hyperplasia and metastatic carcinoma of the prostate showed that the rat tumor was different from the human tissues in several respects. Hence, the search for an optimal animal model for prostatic carcinoma in man must be continued.     

Ornithine decarboxylase activity and its gene expression are increased in benign hyperplastic prostate

BACKGROUND: Ornithine decarboxylase (ODC) is the first key enzyme in the polyamine biosynthesis pathway. Polyamine is believed to participate in cellular proliferation and differentiation. To study the relationship between ODC and the pathogenesis of benign prostatic hyperplasia (BPH), the polyamine levels, ODC activities, and expression of ODC mRNA in benign hyperplastic and normal human prostates were assayed. METHODS: Polyamine contents and ODC activities in tissue extracts were determined by reverse-phase high-performance liquid chromatography and spectrophotometric procedures, respectively. The ODC mRNA levels were assayed by Northern blot analysis. RESULTS: The contents of putrescine, spermidine, and spermine in BPH tissues were 2.2, 3.4, and 6.0 times higher than those in normal tissues, respectively; the ODC activity of BPH tissue was about 3.2 times higher than in normal tissue; the expression level of ODC mRNA in the BPH tissues was greater than that of normal tissues. CONCLUSIONS: The findings imply that 1) the increased ODC activity and polyamine content in prostatic tissue may correlate with the pathogenesis of BPH, and 2) the high level of ODC activity is induced by the overexpression of ODC mRNA.     

Creatine phosphokinase isoenzymes in human prostatic tissues: a comparison between benign hyperplasia and adenocarcinoma

The objective of this study was to determine the distribution of creatine phosphokinase (CPK) into its three isoenzymes, MM, MB, and BB, in human prostatic tissue, in patients with benign hyperplasia (BPH) and adenocarcinoma. Specimens were obtained from 23 patients with adenocarcinoma of the prostate and 25 patients with benign hyperplasia. We also had the opportunity to analyze the CPK content in two normal prostates, the first from a 16 1/2-year-old boy and the second from a 9 1/2-year-old child. Our results showed prostate tissue to contain almost exclusively the BB isoenzyme with traces of the MB and MM dimers in both cancer and BPH as well as the specimen of normal prostate from the 16 1/2-year-old boy. As for the 9 1/2-year-old child, we found the following distribution: 39% MM, 21% MB, and 40% BB dimer. A comparison of the CPK-BB content in benign hyperplasia and adenocarcinoma revealed no significant difference between the two groups. Furthermore, we tried to correlate prostatic tissue CPK-BB levels with another possible tumor marker of the prostate, prostatic acid phosphatase (PAP) measured in the cytosol. No correlation was found between these two markers. We also studied the relationship of CPK-BB and PAP content in prostatic tissue to nuclear and cytosolic androgen receptor content in human prostatic tissue. We found some correlation between CPK-BB and androgen cytosolic receptors as well as between PAP content and androgen cytosolic receptors in patients with benign hyperplasia. No such correlation was found in the group with adenocarcinoma. In conclusion, this study does not show that the measurement of CPK-BB in the prostatic tissue could be used as an index of tissue malignancy.     

Rationale for using aromatase inhibitors to manage benign prostatic hyperplasia. Experimental studies.

Today, human benign prostatic hyperplasia (BPH) is considered primarily to be a disease of the stroma, in which estrogens are thought to play a considerable causative or permissive role. The growing incidence of BPH with increasing age coincides with a shift in the androgen:estrogen ratio in favor of estrogens, not only in terms of serum hormone values, but also in the prostate itself. Furthermore, evidence has been provided for a preferential accumulation of estrogens in the stroma of human hyperplastic tissue, and the presence of an estrogen receptor satisfying the classical criteria of high affinity and low capacity has been demonstrated. Also, animal studies have emphasized the potential role of estrogens in the pathogenesis of BPH. Experimentally, stimulation of the stroma, particularly of smooth muscle, can be induced by aromatizable substrates, such as androstenedione, in the prostates of beagles and cynomolgus monkeys. These effects can be antagonized by aromatase inhibitors, such as atamestane. In addition, the increase in intraprostatic estrogen concentrations and immunohistochemically detectable estrogen receptor content induced by androstenedione in intact dogs is completely reversed by simultaneous treatment with atamestane. In conclusion, clinical data, as well as that from animal models, emphasize an important role for estrogens in the development of BPH. Estrogen deprivation might, therefore, represent a useful treatment for human BPH.     

Mitogenic factors in prostatic tissue and expressed prostatic secretion

Expressed prostatic secretions and extracts of benign prostatic hyperplasia tissue contain a polypeptide growth factor(s) that stimulates the uptake of tritium-labeled thymidine by cultured 3T3 fibroblasts. Mitogenic activity was present in expressed prostatic secretions and extracts of benign prostatic hyperplasia tissue. The apparent molecular weights of the mitogenic fractions were estimated to be 300,000, 150,000 and 60,000 daltons for prostatic tissue extracts, and 30,000 daltons for expressed prostatic secretions. Bioassays yielded a mean of 27 units of mitogenic activity per mg. protein in expressed prostatic secretions obtained from men with normal and enlarged prostate glands. There was no difference in bioassayable mitogenic activity in the expressed prostatic secretions from normal and benign prostatic hyperplasia samples but gel filtration studies revealed a high molecular weight component present only in samples from men with prostatic enlargement. A dialyzable low molecular weight inhibitor of fibroblast growth was found in the prostatic tissues and expressed prostatic secretions. We report the characterization studies and discuss the possible roles of growth factors in the pathogenesis of benign prostatic hyperplasia.     

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.     

雌激素及其受体在良性前列腺增生中的作用

雌激素是前列腺间质细胞强有力的生长调节剂 ,它与其特异性受体结合后发挥作用。大量研究表明 ,雌激素及其受体参与了老年男性良性前列腺增生的发生、发展全过程。本文主要综述了雌激素及其受体在良性前列腺增生发病过程中作用的研究进展     

Altered androgen metabolism in metastatic prostate cancer.

Admixture of androgen-sensitive elements from normal or hyperplastic prostatic tissue interferes with biochemical studies of prostate cancer in its primary site. Heterogeneity of cancer tissues, varying in stromal and epithelial elements, also complicates interpretation of data relating to androgen metabolism. Accordingly, we have compared metastatic deposits composed of epithelial cancer cells to the primary biopsies of 4 patients in respect to uptake of 3H-testosterone and its conversion to 5-alpha-dihydrotestosterone during in vitro incubation. 3H-testosterone uptake was similar for both tissue sites but 3H-dihydrotestosterone formation was reduced by 76% in the metastases compared to primary tissues. This group was not large enough to show statistical significance, whereas a total of 11 such primary studies compared to 6 metastatic specimens was significant. When either primary or secondary tissue results were compared to 12 cases of benign prostatic hyperplasia similarly studied the differences were highly significant. These results demonstrate a major impairment in the formation of dihydrotestosterone by metastatic prostatic cancer and a similar but less evident alteration in the primary site. This abnormality in testosterone metabolism is of major importance in the attempt to obtain effective hormonal control of human prostatic cancer.     

Estrogen formation in human prostatic tissue from patients with and without benign prostatic hyperplasia

Prostatic tissue removed at the time of cystoprostatectomy was separated into periurethral and peripheral zones. Homogenized tissue was incubated with [1,2,6,7(3)H] androstenedione in the presence or absence of an aromatase inhibitor, 4-hydroxyandrostenedione (4-OHAD) and a 5 alpha-reductase inhibitor 4-MA (N,N-diethyl-4-methyl-3-oxo-4-aza-5 alpha-androstane 17 beta-carboxamide). Estrogen formation was determined by reverse isotope dilution of [3H] estrone and [3H] estradiol and crystallization to constant specific activity. Control incubations were carried out in parallel utilizing heated prostatic tissue. Total estrogens produced in the periurethral zone in patients with benign prostatic hyperplasia (BPH) was 223 fmol/mg protein/hr (SE +/- 57) compared to 102 fmol (SE +/- 17) in patients without BPH. Estrogen formation in the peripheral zone was 175 fmol (SE +/- 69) and 105 fmol (SE +/- 26) in patients with and without BPH, respectively. The prostatic aromatase exhibits Michaelis-Menton kinetics with an apparent Km of 90 nM. 4-OHAD inhibited aromatization in the prostatic tissue by 57-93%. Aromatization was also strongly inhibited by 4-MA, indicating that 4-MA is a potent aromatase as well as a 5 alpha-reductase inhibitor in this tissue. These results suggest that aromatization of androgens to estrogens in the human prostate proceeds at a substantial rate and that local estrogen formation could preexist and be a factor in the etiology of BPH and prostatic cancer.     

Increased recovery of androgen receptors from human prostate through the use of mersalyl: evidence for androgen regulation of the binding sites in carcinomatous prostate

With a technique adapted for needle biopsies from human prostate, androgen receptors have been quantitated in normal, hyperplastic, and carcinomatous samples. An important improvement in the yield of cytosol specific binding sites was obtained when samples were pre-incubated with the mercurial reagent mersalyl, which dissociates endogenously bound hormone receptor complexes, before the binding assay with 3H-methyltrienolone (3H-R1881). Androgen receptors in normal prostate tissue were found to be highest (7.81 +/- 1.12 pmoles/g tissue), and significantly different from hyperplastic prostate (2.02 +/- 0.55 pmoles/g tissue, p less than 0.025), but not from carcinomatous samples (4.47 +/- 0.79 pmoles/g tissue). Mean values for hyperplastic and carcinoma were not statistically distinguishable (p less than 0.1). The clinical response to hormone therapy in 85% of 13 patients with prostatic adenocarcinoma reflected the prostatic androgen receptor content. Orchidectomy followed by estrogen administration for several months leads to a dramatic fall (8-fold) in total androgen receptors in carcinomatous prostate, while estrogen alone did not seem to produce a significant effect. These preliminary data suggest that androgen could directly regulate its binding sites, as demonstrated earlier for other animal target organs.     

Partial purification and characterization of a growth factor from human hyperplastic prostatic tissues

Summary A growth factor capable of stimulating DNA synthesis of Balb/c 3T3 cells was purified by heparin-Sepharose column chromatography about 1900-fold from the cytosol of human prostatic tissues obtained at autopsy or open prostatectomy. This growth factor bound to heparin-Sepharose in the presence of 0.5 mol/l NaCl and was eluted by 1.0–1.55 mol/l NaCl. Its molecular weight was estimated to be 68000 by SDS-polyacrylamide gel electrophoresis. The amino acid composition was determined and compared with the data of other growth factors, which revealed no striking conformity. Distribution of growth factor activity was investigated in mechanically separated prostatic tissues of benign prostatic hyperplasia. The separation scheme provided two fractions: the stromal fraction consisting mainly of fibroblasts, fibers and smooth muscle, and the epithelial fraction consisting of epithelial cells. The specific growth-stimulating activity in the stromal fraction was about 2-fold that in the epithelial fraction. Referred to the total activity of whole tissue, about 74% of the activity could be detected in the stromal fraction, while only about 5% was detectable in the epithelial fraction. This study demonstrates the existence of a growth factor in human benign hyperplastic prostatic tissues, showing a remarkable distribution of growth factor activity, which may play a role in the pathogenesis of benign prostatic hyperplasia.     

DNA synthesis in the canine prostate: effects of androgen and estrogen treatment

Canine prostatic DNA synthesis was evaluated by measuring [3H]thymidine incorporation into DNA of tissue slices in vitro. Among untreated beagles, prostatic DNA synthesis rates in young dogs with normal prostates, young dogs with spontaneous benign prostatic hyperplasia (BPH), and old dogs with BPH were 676 +/- 186, 1,220 +/- 156, and 641 +/- 88 cpm/100 micrograms DNA/hr, respectively. Among 81 young beagles (intact or castrated) that had been treated for 4 months with various steroids, rates of DNA synthesis varied according to the type of hormonal treatment. Prostatic DNA synthesis (cpm/100 micrograms DNA/hr) was significantly different (P less than 0.001) for dogs treated with estradiol alone (1,658 +/- 221 cpm/100 micrograms DNA/hr; n = 10 dogs), androgen alone (testosterone, 5 alpha-dihydrotestosterone, or 5 alpha-androstane-3 alpha,17 beta-diol; 1,000 +/- 61 cpm/100 micrograms DNA/hr; n = 31 dogs). However, there was no correlation between prostate size and rate of DNA synthesis (cpm/100 micrograms DNA/hr). Although treatment with estrogen alone resulted in the highest rate of DNA synthesis, it produced squamous metaplasia and the smallest prostates; these results are indicative of a high rate of cell turnover. Comparing prostates that reached the same size following 4 months of treatment with androgen alone or androgen plus estrogen, the rate of prostatic cell turnover was lower in the androgen plus estrogen group. These results are interpreted to indicate an inhibitory effect of estradiol on the rate of cell death in the presence of androgens.     

Quantitation of cytosolic and nuclear estrogen and progesterone receptor in benign, untreated, and treated malignant human prostatic tissue by radioligand binding and enzyme-immunoassays.

Estrogen receptor (ER) and progesterone receptor (PgR) were quantitated in benign and malignant human prostatic tissue by using radioligand binding assays (RBA) and enzyme-immunoassays (EIA). Using a hydroxylapatite exchange method for ER, little or no nuclear ER (ERN) could be detected, but with the EIA both cytosolic (ERC) and ERN were detected in almost all specimens, although in meager concentrations. Tissue from patients with carcinoma had significantly higher ERC concentrations than tissue from patients with benign disease. Specimens from estrogen-treated patients had significantly lower ERC:ERN ratios than those from untreated patients. Progesterone receptor was detected in virtually all specimens by both methods, at concentrations higher than those of ER. Carcinoma tissue with a high malignant involvement contained significantly less PgR than benign tissue. Using either method, the highest concentrations of PgR were observed in tissue from carcinoma patients treated with estrogen. Overall, the data suggest that although human prostatic tissue contains only modest amounts of ER, this is active in that treatment with estrogen promotes association of this receptor with the nuclear fraction and increases PgR content.     

The prostate cancer-up-regulated long noncoding RNA PlncRNA-1 modulates apoptosis and proliferation through reciprocal regulation of androgen receptor

ObjectiveEmerging evidences implicate long noncoding RNAs (lncRNAs) are deregulated in cancer development. The purpose of the current study is to investigate the role of new lncRNA, named PlncRNA-1, in prostate cancer (CaP) pathogenesis.Materials and methodsIn this study, real-time q-PCR was used to demonstrate the expression of PlncRNA-1 in 16 pairs CaP tissues and matched normal tissues, 14 pairs CaP tissues and BPH tissues, 4 CaP cell lines, including LNCaP, LNCaP-AI, PC3, and C4-2, and 2 normal prostate epithelial cell lines RWPE-1 and PWR-1E. After PlncRNA-1 was suppressed by siRNA in LNCaP and LNCaP-AI cell lines, cell proliferation and apoptosis were assessed using CCK-8 and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). After PlncRNA-1 and AR was suppressed by siRNA in LNCaP and LNCaP-AI cell lines, real-time q-PCR and Western blotting were used to measure reciprocal regulation of PlncRNA-1 and AR.ResultsWe showed that expression PlncRNA-1, was significantly higher in CaP cells relative to normal prostate epithelial cells, as well as higher in human CaPs compared with normal tissues and benign prostatic hyperplasia (BPH). Silencing of PlncRNA-1 significantly reduced cell proliferation and induced apoptosis in CaP cell lines LNCaP and LNCaP-AI. Mechanistically, PlncRNA-1 suppression by siRNA resulted in a decrease of androgen receptor (AR) mRNA, protein and AR downstream target. Of note, blockade of AR signaling with siRNA also resulted in a suppression of PlncRNA-1 expression in CaP cell lines.ConclusionsOur study suggests reciprocal regulation of PlncRNA-1 and androgen receptor contribute to CaP pathogenesis and that PlncRNA-1 is a potential therapy target.     

A chicken chorioallantoic membrane assay for the evaluation of the androgen responsiveness of prostatic tissue

A biological assay using the chorioallantoic membrane (CAM) of the chicken egg as an in vitro culture system has been developed and standardized in order to assess the androgen responsiveness of normal and abnormal prostatic tissue. In this assay, test tissue is implanted onto the CAM of chicken eggs in the presence or absence of exogenous testosterone treatment, with subsequent serum levels of 45 ng./dl. and greater than 5,000 ng./dl., respectively. The responsiveness of test tissue to low versus high androgen levels was evaluated in this CAM assay using both cellular morphology and mitotic index as response criteria. Both androgen responsive and unresponsive tissues from the rat were grown on the CAM and demonstrated appropriate morphologic and proliferative responses to the presence and absence of testosterone supplementation. Human prostatic adenocarcinoma and benign hyperplastic tissues were also grown successfully on the CAM. Documentation of human tissue survival was performed by immunocytochemical analysis for prostate specific antigen and prostate specific acid phosphatase. The human prostatic adenocarcinomas studied demonstrated a two to four-fold increase in mitotic index with androgen augmentation. In conclusion, the ability of this assay to determine androgen responsiveness as measured by morphologic and proliferative criteria has been documented in rat tissues. Human prostatic adenocarcinoma has been grown consistently on the chick chorioallantoic membrane for the first time using newly developed techniques. Therefore, this technique shows promise in the clinical application for the determination of the androgen responsiveness of human prostatic carcinomas.     

人良性前列腺增生组织中性激素结合位点与细胞外基质的研究

用免疫组织化学方法（ＡＢＣ法）在４９例良性前列腺增生（ＢＰＨ）组织及１０例正常前列腺组织中对性激素结合位点、纤维粘连蛋白、胶原Ⅳ型蛋白，基膜粘连蛋白进行免疫定位半定量表达。结果提示：ＢＰＨ组织中性激素结合位点表达增强，雄激素的作用部位在上皮及基质细胞，雌激素的作用部位主要在基质细胞；另外ＢＰＨ组织中细胞外基质显著增加，后者与性激素的调节密切相关。