Proliferation and phenotypic changes of stromal cells in response to varying estrogen/androgen levels in castrated rats

It is known that human benign prostatic hyperplasia might arise from an estrogen/androgen （E/T） imbalance. We studied the response of castrated rat prostate to different ratios of circulating E/T. The castrated male Wistar rats were randomly injected with E/T at different ratios for 4 weeks. The prostates of E/T （1：100） group showed a distinct prostatic hyperplasia response by prostatic index, hematoxylin and eosin staining, and quantitative immunohistochemical analysis of a-smooth muscle actin （SMA）. In this group, cells positive for Vimentin, non-muscle myosin heavy chain （NMMHC） and proliferating cell nuclear antigen （PCNA） increased in the stroma and epithelium. Furthermore, the mRNA levels of smooth muscle myosin heavy chain （SMMHC） and NMMHC increased. So E/T at a ratio of 1：100 can induce a stromal hyperplastic response in the prostate of castrated rats. The main change observed was an increase of smooth muscle cells, whereas some epithelial changes were also seen in the rat prostates.     

Androgen and prostatic stroma

Aim: To investigate the effect of androgen on the proliferation, differentiation and regression of canine prostatic stromal cells in vivo and human stromal cells in vitro. Methods: Twenty-two dogs, including 15 normal prostate dogs and 7 prostatic hyperplasia dogs, had their serum concentration of testosterone and estrodiol determined by radioimmunoassay before and after castration. The expression of androgen receptor (AR) and estrogen receptor (ER) in the prostate were analysed by immunohistochemistry and semi-quantitative RT-PCR before and after castration. Light microscopy, transmission electron microscopy and TUNEL assay were carried out successively before and after castration to evaluate the prostatic histomorphology. In vitro serum-free cell cultures from human prostatic stroma were established and exposed to dihydrotestosterone (DHT). The proliferation of the cell culture was detected by MTT assay. The expression of TGFβ, bFGF, AR, and smooth muscle cell (SMC) specific proteins (myosin and/or sm     

Antagonism of estrogen-mediated cell proliferation by raloxifene in prevention of ageing-related prostatic hyperplasia

Estrogen has important roles in the initiation and development of benign prostatic hyperplasia （BPH）. Regulators of the estrogen receptor （ER） are tissue- and cell-specific. We evaluated the effect of estrogen antagonist, raloxifene （Ral）, on the prevention and treatment of BPH by investigating its effect on the proliferation of two different prostate cell lines： a stromal cell line, WPMY-1, and a benign prostatic hyperplasia epithelial cell line, BPH-1. We additionally evaluated its effect on prostatic hyperplasia induced by estrogen and androgen in a rat model. The effect of Ral on the prevention of prostatic hyperplasia was analyzed by haematoxylin and eosin staining and quantitative immunohistochemistry （IHC） for proliferating cell nuclear antigen and α-smooth muscle actin. In vitro and in vivo, tamoxifen （Tam）, another anti-estrogen drug, and finasteride （Fin）, a drug for the clinical treatment of BPH, served as efficacy controls. The in vitro data showed that neither Ral nor Tam alone affected the proliferation of WPMY-1 and BPH-1, but both antagonized the effect of oestradiol in promoting the proliferation of the two cells. Results from the IHC staining of the rat prostates indicated that, similar to Tam and Fin, Ral inhibited the proliferation of stromal cells in vivo. Interestingly, in contrast to Tam, both Ral and Fin inhibited the proliferation of epithelial cells. Furthemore, Ral treatment much strongly decreased the number of prostatic acini and the surrounding layers of smooth muscle cells than Fin （P 〈 0.05）. Our data showed for the first time that Ral may have a role in the response of the rat prostate to selective ER modulators.     

Expression of estrogen receptor alpha and beta mRNAs in prostate cancers treated with leuprorelin acetate

OBJECTIVE: The discovery of a novel estrogen receptor (ER), ER-beta, has given rise to new possibilities regarding estrogen's roles in the prostate. Although ER-beta is reported to be expressed preferentially in the rat prostate, its expression in the human prostate and relationship to cancer development has not been investigated. Thus the purpose of the study was to examine mRNA levels of ER-alpha and ER-beta in benign prostatic hyperplasia and prostate carcinoma. METHODS: Samples of 15 prostate cancers obtained at radical prostatectomy were examined. All the patients had been maintained on androgen withdrawal therapy for at least 3 months. ER-alpha and ER-beta mRNAs were measured with a competitive PCR technique. RESULTS: Both ER-alpha and ER-beta mRNAs were detected in all of the prostate cancer tissues examined, as well as in PC3 and LNCap cells, although the levels varied among specimens. Interestingly, both types were significantly decreased in cases with lymph node metastasis. However, there was no correlation between ER mRNA levels and any other clinicopathological parameters. CONCLUSIONS: (1) Both ER-alpha and ER-beta mRNAs are expressed in prostate cancer and (2) expression of ER mRNA may not be related to cancer progression but may be negatively correlated with metastasis.     

Expression of sulfotransferase 1E1 in human prostate as studied by in situ hybridization and immunocytochemistry

BACKGROUND: Estrogen is recognized to play a role in the development and function of the prostate. Estrogen sulfotransferase (EST) 1E1 catalyzes the sulfoconjugation of estrogen and is thus involved in the metabolism of estrogen. We have recently shown that EST 1E1 is highly expressed in male mouse reproductive organs, including prostate. It appeared of interest to study the expression of EST 1E1 in human prostate. METHODS: EST 1E1 mRNA and protein expression was evaluated in benign prostatic hyperplasia (BPH) using in situ hybridization and immunocytochemistry, respectively. RESULTS: EST 1E1 mRNA and protein were found to be expressed in epithelial cells bordering alveola lumen (luminal cells) as well as stroma cells. CONCLUSION: The enzyme EST may play a physiological role in regulating local estrogen levels in human prostate.     

The role of tissue steroids in benign hyperplasia and prostate cancer

This paper presents a large body of data relating to benign prostatic hyperplasia, which have been derived from fundamental endocrinological research. For the urologist, the data open up interesting aspects of the pathomorphology of prostatic hyperplasia. The most interesting findings can be summed up as follows: 1. Testosterone is the circulating androgenic prohormone that mediates the intracellular message leading to androgen secretion, though by way of its metabolite dihydrotestosterone, which is really the active substance. 2. This metabolic conversion is catalyzed by 5 alpha-reductase, which is predominantly a stromal enzyme. 3. The estrogen metabolism in the stromal cells of the prostate may be associated with the abnormal growth of the prostate. 4. In the presence of benign prostatic hyperplasia dihydrotestosterone and 17 beta-estradiol accumulate in the nuclei of the stromal cells. 5. Adrenal androgens are also metabolized in the human prostate, yielding some substances with androgenic and some with estrogenic potency. 6. Changes in sex hormone binding globulins (SHBG) are found with age whether benign prostatic hyperplasia is present or not. It is therefore questionable whether it has any influence on the development of prostatic hyperplasia. 7. Although in some cases it is not yet possible to determine whether the findings presented in this paper have any causal significance, the data can be used as a rational basis for hormonal treatment of prostatic disease.     

Effect of pubertal development on estrogen receptor levels and stromal morphology in the guinea pig prostate

Using an immunocytochemical assay (ERICA) with a monoclonal antibody (H222Sp gamma) to the human estrogen receptor, we have demonstrated a stromal localization of the estrogen receptor in the dorsolateral prostate of the guinea pig. Specific staining of estrogen receptor in the guinea pig prostate was confined to the nuclei of periacinar and interacinar stromal cells. In comparison with prepubertal tissues, estrogen receptor staining intensity was markedly reduced in postpubertal prostatic tissues. No immunoreactive estrogen receptor was detected in the acinar epithelial cells irrespective of the developmental stage of the guinea pig prostate. Electron microscopic examination of the guinea pig prostate showed that the stromal component consists predominantly of smooth muscle cells, which, during pubertal development, undergo marked cytological changes and increase in size. These changes in the prostatic stroma were associated with a greater than fivefold reduction in levels of cytosolic and nuclear estrogen receptor determined by either a radioligand binding assay or an enzyme immunoassay (EREIA) and expressed relative to soluble protein. Morphometric analysis of the prostatic stromal cell density (SCD: nuclei/mm2 interacinar stroma), which is inversely proportional to stromal cell size, indicated that the SCD decreased approximately threefold during pubertal development. Furthermore, cytosolic estrogen receptor levels in mechanically separated prostatic stromal fractions were found to vary concordantly with the SCD during pubertal development. To determine whether estrogen influences normal development of the guinea pig prostate, the effect of various hormonal manipulations on stromal development was examined. Castration of prepubertal animals prevented the threefold decrease in SCD that is characteristic of pubertal development. Treatment of prepubertal castrates with estradiol and 5 alpha-dihydrotestosterone (DHT) in combination over a period equivalent to the transpubertal growth phase resulted in a stromal cell density similar to that seen in prostatic sections from intact postpubertal animals. In contrast, treatment of prepubertal castrates with either estradiol or DHT alone resulted in a prostatic stromal cell density intermediate between that observed in intact prepubertal and postpubertal animals. These findings suggest that both estrogen and androgen are required for the normal development of the guinea pig prostatic stroma.     

Induction of benign prostatic hyperplasia in intact dogs by near-physiological levels of 5α-dihydrotestosterone and 17β-estradiol

Benign prostatic hyperplasia was induced in mongrel dogs treated for 60 days with one silastic implant containing 17β-estradiol and four containing 5α-dihydrotestosterone. The condition was characterized by (1) a marked increase of the stromal elements, particularly the stromal septa between the individual glands, (2) a slight increase in prostatic volume, and (3) a morphology that resembled spontaneous complex benign prostatic hyperplasia in the dog. Other groups of animals that remained untreated or received only 17β-estradiol or only 5α-dihydrotestosterone did not develop this condition. Prostate volumes decreased by 14% in the estrogen-treated dogs, whereas they increased in the androgen-treated animals by 6% compared to pretreatment prostate volumes. The morphology of the epithelium of the prostates of androgen-treated animals was not different from that of controls despite the increase in prostate volume. The serum 17β-estradiol and 5α-dihydrotestosterone concentrations were increased from 25 ± 2 (mean ± SEM) and 256 ± 42 pg/mL, respectively, in control dogs to 52 ± 37 and 562 ± 37 pg/mL, respectively, in the dogs treated with the hormone combination. Thus, hormone concentrations were two- to three-fold higher than control values, and the ratio of estradiol-17β to 5α-dihydrotestosterone was increased by up to 19%. These data demonstrate that treatment of dogs with low levels of estrogen and androgen may be an excellent model for the study of spontaneous complex benign prostatic hyperplasia in aging men.     

Histological localization of estrogen receptors in normal and diseased human prostates by immunocytochemistry.

The role of estrogens and estrogen receptors (ER) in the human prostate remains unresolved. In this study we have used the monoclonal ER antibody H222 to investigate the histological localization of ER in normal and diseased human prostates by immunocytochemistry. Prostate tissue was obtained from 3 young organ donors (Group I-normal prostate), from 14 prostates removed by radical prostatectomy or radical cystoprostatectomy, which had caused no or only mild obstructive symptoms (Group II-non-obstructive prostate), and from 11 prostates removed by suprapubic prostatectomy, which had caused severe obstructive symptoms due to a large benign prostatic hyperplasia (BPH) (Group III-obstructive prostate). In prostates of all groups ER were found to be in nuclei of the prostatic urethra and of the periurethral prostatic duct. In striking contrast, ER in the interglandular prostatic stroma was not as homogeneous among the different groups. We observed a low concentration of ER in the stroma of normal prostates, the highest concentration in non-malignant stroma of non-obstructive prostates, and no ER at all in stroma of obstructive prostates. Based on the immunocytochemical localization of ER in normal and diseased human prostate, our results indicate that stromal growth in obstructive BPH may not be mediated via ER. However, we cannot exclude that an increase of stromal ER concentration (as observed in non-obstructive prostates) is directly involved in induction of BPH, leading further prostate growth thereafter into an estrogen independent state.     

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.     

Estrogen receptor in human benign prostatic hyperplasia

Estrogens have been proposed as a major etiological factor in the pathogenesis of benign prostatic hyperplasia in man. The presence of estrogen receptor in benign prostatic hyperplasia would support this concept. Using the receptor stabilizer, sodium molybdate, and a hydroxylapatite assay we assayed human benign prostatic hyperplasia for the presence of cytosolic estrogen receptor. For comparison, we assayed estrogen receptor in cytosols of prostatic cancer and normal tissue, and we also measured androgen receptor and progesterone receptor concentrations in the 3 tissue types. Estrogen receptor was present in 8 of 15 benign prostatic hyperplasia specimens at a mean concentration of 9.2 fmol./mg. protein for the estrogen-receptor-positive samples. Sucrose gradient analysis of the estrogen receptor of benign prostatic hyperplasia revealed that it sedimented in the region of 8S, and steroid specificity studies confirmed that the binding to estrogen receptor was estrogen-specific. Estrogen receptor was also found in normal (3 of 3) and malignant (4 of 6) tissues, and all tissues were positive for androgen receptor. The presence of estrogen receptor in human benign prostatic hyperplasia supports the proposal that circulating estrogens may have a role in the pathogenesis of this disorder.     

前列腺组织中EGF、bFGF的表达

目的：研究EGF、bFGF在前列腺组织中的表达。方法：应用mRNA斑点杂交、原位杂交、免疫组织化学及原位杂交与免疫组织化学双染法检测6例正常前列腺（NP）、27例良性前列腺增生症（BPH）前列腺组织中EGF及bFGF的表达。结果：BPH前列腺组织和NP组织中无均无EGF mRNA表达,EGF蛋白表达呈弱阳性,两组间差异无显著性意义（P＞0.05）;NP组织上皮细胞有较多bFGF mRNA表达,但无bFGF翻译,基底基质细胞有少量mRNA及蛋白表达,二者表达水平基本一致;BPH前列腺组织上皮细胞无bFGF mRNA表达,但局灶性增殖上皮细胞细胞膜上有bFGF,基底基质细胞有大量bFGF mRNA转录及蛋白质翻译,以局灶性增殖区最为明显。结论：NP及BPH的前列腺组织中无EGF分泌细胞;bFGF在前列腺基底基质细胞过度表达,以自分泌、旁分泌方式促进了基质和上皮的非均一性增殖。     

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     

Immunocytochemical localization of estrogen receptors in spontaneous and experimentally induced canine benign prostatic hyperplasia

Estrogens are believed to play a critical role in the etiology of canine benign prostatic hyperplasia (BPH); however, the mechanism has not been elucidated. To gain insight into this problem, we investigated the immunocytochemical localization of estrogen receptors (ER) in normal prostates, spontaneous BPH, and experimentally induced BPH by using a monoclonal ER antibody (H222). In all canine prostates the majority of ER was localized in nuclei of the same histological components: (1) transitional epithelium and subjacent stroma of the prostatic urethra, (2) periurethral prostatic ductal epithelium, and (3) prostatic stroma. ER content in the stroma was highest in the periurethral region of the prostate. Among the different groups of dogs, differences in ER location were seen only in the glandular epithelium. No ER was found in the glandular epithelium of normal prostates of young untreated dogs. In striking contrast, glandular epithelium of spontaneous BPH contained specific nuclear ER staining, though this staining was heterogeneous and was observed in only a minority (less than 10%) of the acinar epithelial cells. ER-positive acini in BPH were located predominantly in the periurethral region. These data demonstrate anatomical and biochemical heterogeneity of prostatic components and indicate that the estrogen sensitivity of prostatic cells is heterogeneous. If estrogen does play a role in BPH, it appears to act selectively rather than uniformly throughout the prostate. We reasoned that if glandular epithelial ER are involved in the development of spontaneous BPH, one might expect to find the same location of ER in BPH that was induced experimentally by specific types of treatment with androgens +/- estradiol. However, among hormone-treated dogs the presence of ER-positive prostatic glandular epithelium varied with the type of hormonal treatment but did not correlate with the experimental induction of glandular BPH. Some treatment groups with induced BPH had ER-positive prostatic glandular epithelial nuclei (with the same extent and pattern of ER localization as in spontaneous BPH); however, other treatment groups with induced BPH had ER-negative glandular epithelium. These data indicate either that glandular epithelial ER may not be involved in the pathogenesis of canine BPH or that there may be different types of BPH that have different etiologies. Possible mechanisms by which estrogen may affect the canine prostate are discussed in light of these new data on ER location.     

Resistance to Apoptosis and Up Regulation of Bcl-2 In Benign Prostatic Hyperplasia After Androgen Deprivation

Purpose

Benign prostatic hyperplasia (BPH) is related to advancing age and the presence of androgens and occurs in virtually all older men. BPH causes morbidity, most often by urinary obstruction, in a substantial fraction of men over sixty. Both finasteride and androgen ablation induce partial diminution in BPH that occurs over weeks to months. This is in contrast to the often rapid involution seen in both normal prostatic epithelium and prostatic carcinoma in response to androgen withdrawal. This study was performed to analyze the response of prostatic cells, and in particular BPH, to acute androgen ablation.

Materials and Methods

We subjected a cohort of 26 men to androgen ablation with goserelin, a gonadotrophin releasing hormone agonist, for 3-4 weeks prior to radical prostatectomy for prostate cancer. Preablation biopsy specimens and prostatectomy specimens were immunohistochemically stained for apoptotic cells and for expression of apoptosis regulatory proteins Bcl-2, Bax, Bcl-x, and Bak.

Results

Normal prostatic epithelial cells and prostate cancer responded to hormone deprivation by undergoing apoptosis, but in 19/26 specimens prostatic hyperplasia had a total absence of apoptosis. In all 26 specimens, benign prostatic hyperplasia demonstrated increased expression of the Bcl-2 protein, but no change in the expression of Bax, Bcl-x, and Bak. In contrast, adjacent normal and malignant prostatic epithelium showed positive staining for apoptosis and did not alter Bcl-2 expression in response to androgen ablation.

Conclusions

BPH demonstrated increased staining for Bcl-2 after androgen deprivation that may render hyperplastic epithelium relatively resistant to apoptosis induced acutely by androgen withdrawal.     

前列腺癌和癌旁组织中雌激素受体α和β的表达及意义

目的 通过测定前列腺癌中癌组织、癌旁组织和良性前列腺增生组织中雌激素受体(estrogen receptor,ER)α和β的表达水平,探讨ER在前列腺腺癌发生、发展中的变化和作用机制.方法 采用超高敏链酶亲合素-过氧化物酶法检测28例前列腺腺癌标本中癌和癌旁组织以及29例良性前列腺增生组织中ERα和ERβ的表达水平,对比其在各组之间表达的差异,分析ERα和ERβ的表达水平与前列腺癌患者的Gleason评分、临床分期、年龄和TPSA的关系.结果 前列腺癌组织中ERα主要在间质细胞表达.ERα在前列腺癌组织、癌旁组织和良性前列腺增生组织上皮细胞的表达阳性率分别为0%、14%、24%,间质细胞阳性率分别为57%、68%、31%,组间比较差异有统计学意义(P＜0.05).前列腺癌上皮细胞和间质细胞均有ERβ的表达且差异无统计学意义(P＞0.05).ERβ在前列腺癌组织、癌旁组织和良性前列腺增生组织上皮细胞的表达阳性率分别为39%、64%、29%;间质细胞阳性率分别为50%、75%、79%,组间比较差异有统计学意义(P＜0.05).ERβ在不同Gleason评分前列腺癌组织中表达差异有统计学意义(P＜0.05).结论 前列腺癌中ERα主要在间质细胞表达;ERβ在上皮细胞和间质细胞均有表达.ERα和ERβ的表达水平在前列腺癌组织、癌旁组织和良性前列腺增生组织均存在差异.ER口与前列腺癌变发生和恶性程度相关.

Abstract：

Objective To investigate the expression of estrogen receptor (ER) α and β in human prostate cancer (PC), peri-cancer tissue and benign prostatic hyperplasia (BPH) tissue, and to discuss the role of estrogen receptor in prostate cancer. Methods The expression of ERα and ERβ in PC (n=28), peri-cancer tissue (n=28) and BPH (n=29) were detected by immunohistochemistry with En vision method. The ERα and ERβ expression were compared among different tissues by chisquare. The relationship between ER expression and related clinicopathologic features was statistically analyzed by spearman rank collection. Results ERα was localized dominantly in the stromal cell of PC. There were significant differences of the expression of ERα in PC, peri-cancer tissue and BPH tissue (epithelial cell 0%, 14%, 24%, P＜0. 05; stromal cell 57%, 68%, 31%,P＜0. 05). ERβ was localized in both epithelial and stromal cell of PC. There were significant differences of the expression of ERβ in PC, peri-cancer tissue and BPH tissue (epithelial cell 39%, 64%, 29%, P＜0.01; stromal cell 50%, 75%, 79%, P＜0.05). There was a significant difference of the expression of ERβ in different Gleason scores of PC tissue. Conclusions ERα is localized in the stromal cell of PC tissue.ERβ is localized in both epithelial and stromal cell of PC tissue. The ERβ might be related to the tumor differentiation of PC.     

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.