Properties of androphilic proteins in cytosols of human benign prostatic hypertrophy.

Androphilic proteins in the cytosol from the human benign prostatic hypertrophy are separated into two fractions by Sephadex chromatography; void volume fraction and IgG fraction which was eluted near the site of hIgG. In the present study, properties of these two androphilic proteins were compared. Association constants of these proteins were in the order of 10(9) M-1. However, the binding capacity of the former was smaller than that of the latter. These two androphilic proteins well bound to nuclei, and the high-affinity and saturable binding to nuclei was observed in the 3H-dihydrotestosterone-IgG fraction complex, while binding of 3H-dihydrotestosterone-void volume fraction complex to nuclei was low affinity and unsaturable. The binding of the complexes to chromatin seems to be of low affinity and nonsaturable. These androphilic proteins did not bind to calf thymus DNA. Salt extractability of the bound void volume fraction after incubation with nuclei was not different from that of the bound IgG fraction. It was observed that the chromatographic behavior of the androphilic protein in IgG fraction was changed after incubation with nuclei.     

Binding to R 1881 (methyltrienolone) of proteins from human benign prostatic hypertrophy

Cytosol of human benign prostatic hypertrophy bound to R 1881 in a high affinity manner. Most of the protein which bound to R 1881 was recovered in the precipitate of a 0-30% saturation of ammonium sulfate, and was eluted in the void volume on a Sephadex G-200 column. The binding of cytosol to R 1881 was more inhibited by progestins than by dihydrotestosterone and estradiol-17 beta. The binder therefore seemed to be different from dihydrotestosterone-binding protein. The R 1881-binding component extracted from the nuclei by 0.4M KCl bound also to dihydrotestosterone in a high affinity manner. Cytosol prelabeled with R 1881 was bound to the nuclei in a nonsaturable process, and the extraction pattern of R 1881 by 0.4M KCl from the nuclei was almost identical to that in the case of dihydrotestosterone as the ligand. These results suggested that a part of the cytosolic protein which bound to R 1881 entered the nuclei where it bound to nuclear such components as dihydrotestosterone-binding protein.     

Serum of patients with prostatic cancer or benign prostatic hypertrophy contains nonpolar testosterone.

We have previously described a nonpolar form of radioimmunoassayable serum testosterone (NPT) not measured by available antitestosterone antibodies. It is detected by mild alkaline hydrolysis of the petroleum ether extract of serum and subsequent radioimmunoassay. The properties of NPT are consistent with that of a fatty acid ester of testosterone or dihydrotestosterone. The serum of young males contains 1 to 3 ng/ml NPT, but it is not detected in female serum. We measured serum testosterone and NPT levels in 36 men between 58 and 87 years of age. Seventeen subjects with advanced prostatic cancer (NPT 1.70 +/- 1.44 ng/ml) were compared with a control group consisting of six patients with benign prostatic hypertrophy (BPH) and 13 patients with no prostatic disease (NPT 0.72 +/- 0.46, P = 0.017). There was no significant difference between BPH patients and patients with no prostatic disease; the results were pooled. The concentration of NPT in prostatic cancer patients but not in controls was inversely correlated with that of testosterone. Immunoassayable testosterone was present in the serum of two orchiectomized patients and, therefore, cannot derive solely from the testes.     

Inhibition by various steroids on dihydrotestosterone binding to androphilic protein in human benign prostatic hypertrophy.

The effect of various steroidal compounds on the binding of the androphilic protein to dihydrotestosterone in the cytosol of tissues of human benign prostatic hypertrophy was examined. Androgens, as well as estrogens and gestagens showed an inhibitory effect on the binding. Non-steroidal anti-androgens were revealed to be weak inhibitors on the binding. Two androphilic proteins were observed in Sephadex G-200 chromatography in fractions eluted at the void volume and in fractions appearing at the site of IgG, and the rate of inhibition on the binding of both fractions by various steroids was compared. The rate of inhibition by various compounds was generally higher in the IgG fraction than in the void volume fraction. When the ligand and inhibitors were incubated with the cytosol prior to fractionation by Sephadex chromatography, rate of inhibition was lower than that obtained when the ligand and inhibitors were reacted with fractions after chromatography. Implications of the difference observed in these two experiments are not clear at this moment. The results obtained by the protamine precipitation experiment were almost the same as those by the experiment using the extract of the unfractionated acetone-dried cytosol, therefore, the protamine procedure does not seem to precipitate the tissue specific androphilic protein specifically.     

Steroid receptors in the human prostate. 1. Estradiol - 17beta binding in benign prostatic hypertrophy.

Agar gel electrophoresis and ultracentrifugation on continuous sucrose gradients revealed the presence of a 4S estradiol 'receptor' in cytosols of samples of human benign hyperplastic prostate tissue. High affinity (charcoal stability) and saturability (disappearance with excess competitor) binding characteristics of the molecular species concerned facilitated its clear distinction from similarly migrating serum albumin-steroid complexes. Our data, obtained with human serum, purified human albumin and albumin-enriched cytosol strongly suggest that agar gel electrophoresis, when used alone, may lack specificity for the quantification of estrogen 'receptors'. Radioligand binding to these molecules may be obscured by similarly migrating albumin-steroid complexes which fail to dissociate during electrophoresis. We advocate the inclusion of competitor experiments to improve the specificity of the method.     

Clinical experience with alpha-adrenoceptor antagonists in benign prostatic hypertrophy

The symptomatic treatment of benign prostatic hyperplasia with alpha-adrenoceptor antagonists is based on the concept of a dynamic component in prostatic obstruction, dependent on the sympathetically controlled tone of smooth muscle in the prostate and its capsule. Blockade of the alpha adrenoceptors minimizes this tone, hence the degree of obstruction, and also relieves the irritative symptoms. A review of the published clinical experience reveals convincing evidence of a beneficial action on the irritative symptoms of diurnal and nocturnal frequency, and on the objective parameters of uroflow, residual urine, urethral closure pressure, and cystometrographic evidence of detrusor instability. It is also of value in the prophylaxis or treatment of certain cases of acute retention in these patients. The greatest amount of experience has been with phenoxybenzamine, which appears to be the most effective drug in practice, but other drugs are being assessed for this purpose. It is possible that a combined alpha 1 and alpha 2 antagonist may be more effective than a pure alpha 1 antagonist.     

Steroid receptors in the human prostate. 2. Some properties of the estrophilic molecule of benign prostatic hypertrophy.

Some physicochemical properties of the estrophilic ‘receptor’ of human benign prostatic hypertrophy were examined by agar gel electrophoresis. 1) Competition analyses revealed the high selectivity of the molecule for the naturally occurring estrogens but not for representatives of other classes of steroid hormones (androgens, corticosteroids, progesterone). This, coupled with the failure of an estrogen ‘receptor’-rich extract to exhibit detectable tissue specific binding of (³H) 5α-dihydrotestosterone suggests that prostatic androgen and estrogen receptors may have separate identities. 2) The molecule proved highly resistant to enzyme attack, a stability conferred by estradiol-17β rather than by the thiol reagent dithiothreitol. Its proteinaceous nature was finally demonstrated when extract was exposed to enzymes at 0°C prior to steroid addition. 3) Initial complex formation between estrogen and its ‘receptor’ protein was rapid and reached a plateau after 4 hours. Binding was greater at 0°C than 37°C.     

Binding characteristics of naftopidil and alpha 1-adrenoceptor antagonists to prostatic alpha-adrenoceptors in benign prostatic hypertrophy.

Binding properties of naftopidil and alpha 1-adrenoceptor antagonists to alpha-adrenoceptors in prostates from benign prostatic hypertrophy (BPH) were characterized by radioreceptor assays using [3H]prazosin and [3H]rauwolscine. Specific binding of [3H]prazosin and [3H]rauwolscine in human prostatic membranes was saturable and of high affinity, and it showed a pharmacological specificity which characterized alpha 1 and alpha 2-adrenoceptors, respectively. Naftopidil and several alpha 1 antagonists competed for prostatic [3H]prazosin binding in order: R-(-)-YM-12617 greater than prazosin greater than bunazosin greater than terazosin greater than naftopidil greater than urapidil, and the inhibitory effect (Ki = 11.6 nM) of naftopidil was 10 to 45 times less potent than quinazoline derivatives such as prazosin, bunazosin and terazosin. The potencies of these antagonists in competing for [3H]prazosin binding sites in human prostates correlated well with their pharmacological potencies (pA2). Scatchard analysis indicated that the decrease of prostatic [3H]prazosin binding by naftopidil was due to a marked increase in the Kd value without a change in the Bmax value. The inhibition of prostatic [3H]prazosin binding by naftopidil was reversible. Naftopidil also inhibited prostatic [3H]rauwolscine binding (Ki = 70.0 nM). Thus, it is suggested that naftopidil antagonizes alpha 1-adrenoceptors in human prostates in a competitive and reversible manner.     

Estradiol- and estriol-binding in human benign prostatic hyperplasia.

Binding of the natural estrogens, estradiol and estriol, was investigated, in 34 samples of human benign prostatic hypertrophy (BPH) tissue, using Scatchard analysis and agar gel electrophoresis. Saturation binding analysis using a wide range of concentrations of both ligands resulted in curvilinear Scatchard plots. This confirmed the presence of two binding forms for estradiol: a true estrogen receptor, and a protein with lower affinity and higher capacity. Both binding species were also demonstrated and quantified with estriol. The electrophoretic process, after incubation at low and high ligand concentrations also resulted in separation, for both estrogens, of two binding peaks. They are probably two distinct forms of the low affinity, high capacity binding measured by Scatchard. The procedure used in our laboratory was not able to provide accurate determination of the concentrations of these binding forms. Possible modifications to alleviate these drawbacks are discussed.     

Plasma steroids in benign prostatic hypertrophy and carcinoma of the prostate

Cortisol, 17-hydroxyprogesterone (17-OH-P), progesterone, testosterone, 5 alpha-dihydrotestosterone (DHT), estrone (E1) and estradiol (E2) were determined by RIA after chromatographic separation of steroids on Sephadex LH-20 columns, in 54 hospitalized patients with benign prostatic hyperplasia (BPH) and in 32 hospitalized patients with prostatic carcinoma (PCA) (T34, N01, M01). The patients' values were compared with those of 63 age-matched controls. Increased cortisol and DHT levels, subnormal estrogen and 17-OH-P values and normal progesterone level were found in both benign and malignant groups. Higher mean values for testosterone, and T/DHT ratio and lower mean values for E2/T ratio were found in PCA, as compared with those in BPH. An age invariance of cortisol, testosterone, T/DHT ratio and estradiol was found in both BPH and PCA, instead of the age dependence found in normal subjects. The normal relations between testosterone and its precursor (17-OH-P) and its peripheral metabolite (E2), respectively, and the normal relation between estrone and 17-OH-P were not evident in either BPH or PCA group. The normal direct relation between testosterone and DHT has been found in both patient groups.     

Transurethral microwave treatment for benign prostatic hypertrophy: a randomised controlled clinical trial.

OBJECTIVES--To determine whether transurethral microwave treatment for patients with benign prostatic hypertrophy provides significant symptomatic relief, a reduction in residual urine volumes, and improvements in flow rates compared with sham treatment. DESIGN--Prospective double blind randomised study with follow up at three months. SETTING--Department of Urology in a London teaching hospital. PATIENTS--40 men completed the study: 22 received microwave treatment and 18 received sham treatment. Entry criteria were symptoms of prostatism of at least six months'' duration, a total symptom score > 14, and a peak urine flow rate < 15 ml/s or a residual urine volume > 50 ml. Exclusion criteria were prostatic cancer, a residual urine volume > 200 ml, a very large prostate, an obstructing middle lobe, acute urinary retention, impaired renal function, coexisting urinary tract disease, and previous prostatic surgery. INTERVENTIONS--A single 90 minute transurethral microwave treatment or sham treatment. OUTCOME MEASURES--Patients'' symptoms (including daytime frequency and nocturia) recorded in a self assessment symptom score questionnaire, peak urinary flow rates, and residual urine volumes. RESULTS--The mean total symptom scores of the patients who received microwave treatment fell from 30 to 11 compared with a fall from 31 to 26 for patients who received sham treatment (p < 0.001). Among patients who received microwave treatment daytime frequency fell from 9.4 to 5.5 voids a day and night time frequency from 3.5 to 1.6 voids a night; residual urine volumes fell from 104 ml to 52 ml; and peak urine flow rates increased by 2.3 ml/s. In the control group there was no improvement in any of these features. Treatment preserved sexual function and antegrade ejaculation. CONCLUSIONS--For selected patients with prostatism microwave treatment is effective and has few side effects.     

Normal and benign human prostatic epithelium in culture. I. Isolation.

Isolation of normal human glandular epithelia and their growth and maintenance in vitro have been major problems. The primary objective of studies presented here was to isolate postpubertal, normal human, viable prostatic epithelium for in vitro cultivation. The long-term objective of these investigations was to develop an in vitro human cell model system for studies on prostatic carcinogenesis. A method for isolation of viable, normal and benign human prostatic epithelium, using collagenase for tissue dissociation, is described. Intact acini were isolated, which, on plating gave rise to vigorously growing monolayer cultures of epithelial cells. The purity of epithelial cultures partly depended upon the source of tissue. Specimens of normal prostate and those of benign tissue derived from open prostatectomies provided primarily pure epithelial cultures with occasional fibroblast colonies in some cultures, which could be removed. Cultures from some specimens of transurethral resection of the prostate (TURP) contained many fibroblast colonies due to incomplete separation of acini from the stroma. This resulted from incomplete digestion of denatured tissue caused by electrocauterization during surgery. Cultures established in this manner are being used to study the effects of hormones, vitamins and other growth regulators in order to establish growth requirements of these cells in vitro, which would facilitate their long-term maintenance.     

Evaluation of the effectiveness of prazosin in conservative treatment of benign prostatic hypertrophy]

Prazosin--a selective blocker of alpha 1-adrenergic receptors--was administered to 30 patients with benign prostatic hypertrophy. Twenty four patients (80%) reported an improvement in voiding and observed more potent urinary stream after the treatment. Average and maximum flow rates increased in 18 patients (60%). Therapy had to be discontinued in 2 patients because of the adverse reactions (hypotension and syncope in one and exacerbation of the coronary disease symptoms in another patient).     

Transforming growth factor-alpha expression in benign and malignant human prostatic disease.

Investigation of biological variables in prostatic disease may not only prevent patients with a good prognosis being overtreated, but allow better selection of appropriate therapy, and may identify potential targets for novel therapies. This study investigates the growth factor transforming growth factor-alpha (TGF alpha) expression in benign and malignant prostatic biopsies using both radioimmunoassay and immunohistochemistry, considering its role in malignant epithelial transformation and as a prognostic indicator. Biochemical methods were less satisfactory than the more selective immunohistochemical methods, due to the heterogeneity of prostatic tissue. Seventy-one percent of benign biopsies (range 0-18.62ng/mg DNA) and 69% of malignant biopsies (range 0-11.1ng/mg DNA) had detectable levels of TGF alpha using radioimmunoassay. Immunohistochemical staining for TGF alpha identified expression in 15% of benign (4 out of 27) and 53% malignant biopsies (18 out of 34). Positive staining was also identified in premalignant lesions and within stromal elements, thus implying the factor's role in autocrine/paracrine growth and/or malignant transformation. Immunostaining for TGF alpha may enhance detection of premalignant lesions and small foci of malignant glands which are otherwise difficult to identify using standard histopathological techniques.     

Ki-67 antibody immunostaining in benign and malignant human prostatic disease.

Indices of mitotic potential may improve prognostic discrimination in patients with malignant disease. Ki-67 is a monoclonal antibody directed against an unknown proliferation antigen which has been shown to be a measure of mitotic potential. Sixty-four benign and eighty malignant prostatic biopsies were stained with the Ki-67 antibody. Nuclear and cytoplasmic staining was identified in benign and malignant biopsies using immunoalkaline phosphatase and immunoperoxidase staining reactions. Nuclear staining was identified in 14 benign and 44 malignant biopsies. Nuclear staining for Ki-67 was seen in 36% of biopsies with Gleason histological score (GHS) 2-4, 71% with GHS 5-7 and 62% with GHS 8-10. Nuclear staining was associated with advanced local disease stage, but not with metastatic disease stage. Clinical follow-up is required to establish the value of Ki-67 immunostaining as a prognostic determinant in prostatic cancer.     

Measurement of androgen receptor in cytosols from normal, benign hypertrophic and cancerous human prostates

The binding of R 1881 in cytosols from the normal, benign hypertrophic and cancerous human prostates was examined in the presence of molybdate and triamcinolone acetonide. The addition of 10 mM Na2MoO4 resulted in an increase in the stability of the binder without any change in Kd. Triamcinolone acetonide added to the incubation of cytosol with R 1881 modified the inhibition pattern by other steroids, and the binding in the presence of triamcinolone acetonide exhibited the characteristics of androgen receptor. The complex of cytosol and R 1881 formed in the incubation in the presence of triamcinolone acetonide was sedimented at 8.5S. Kd of the binding to R 1881 of the normal and pathological prostates was almost identical, but maximum binding sites of the androgen receptor were larger in benign hypertrophic and cancerous prostates than in normal tissues. For the assay of binding capacity in needle biopsy specimens, one point determinations were performed using 2.5 nM R 1881 as the ligand. The number of binding sites obtained by this method were well correlated with those obtained by the Scatchard plot in various prostatic tissues.