Biomedical subjects

D J Horsfall

Publications and source records attributed to D J Horsfall.

At least 37 records · Page 2Linked to original sources

PubMed1995

Androgens induce divergent proliferative responses in human breast cancer cell lines.

Although the majority of primary human breast cancers express the androgen receptor (AR), the role of androgens in breast cancer growth and progression is poorly understood. We have investigated the effects of the naturally occurring androgen, dihydrotestosterone (DHT), and a synthetic non-metabolizable androgen, mibolerone, on the proliferation of six human breast cancer cell lines. The anti-proliferative and proliferative effects of androgens were only observed in cell lines that expressed the AR. Two of the AR-positive cell lines, T47-D and ZR-75-1 were growth inhibited in the presence of either DHT or mibolerone, while the proliferation of MCF-7 and MDA-MB-453 cells was increased by both androgens. Co-incubation of cultures with 1 nM DHT and a 100-fold excess of the androgen receptor antagonist, hydroxyflutamide, resulted in reversal of both inhibitory and stimulatory effects of DHT on T47-D, MCF-7 and MDA-MB-453 cell proliferation, indicating that DHT action is mediated by the AR in these lines. Hydroxyflutamide only partially reversed the DHT-induced growth inhibition of ZR-75-1 cultures, which suggests that growth inhibition of these cells may be mediated by non-AR pathways of DHT (or DHT metabolite) action. Mibolerone action on breast cancer cell growth was similar to that of DHT, with the exception that growth stimulation of MCF-7 and MDA-MB-453 cells was only partially reversed in the presence of a 100-fold excess of hydroxyflutamide. Anandron, another androgen receptor antagonist, was able to reverse all inhibitory and stimulatory actions of the androgens. AR antisense oligonucleotides reduced the level of immunoreactive AR expression in MDA-MB-453 and ZR-75-1 cells by more than 60%, but only reversed the growth inhibitory action of mibolerone in ZR-75-1 cultures. The results suggest that androgen action in breast cancer cell lines may not be solely mediated by binding of androgen to the AR. For example, metabolites of DHT with oestrogenic activity, or androgen binding to receptors other than the AR, may explain the divergent responses to androgens observed in different breast cancer cell lines.

Androgen Receptor Antagonists↗

PubMed1995

Differential expression of apolipoprotein-D and prostate specific antigen in benign and malignant prostate tissues.

PURPOSE: To investigate Apolipoprotein-D (Apo-D) and prostate specific antigen (PSA) immunohistochemical staining of nonmalignant and malignant human prostate tissues. MATERIALS AND METHODS: Apolipoprotein-D and PSA immunoreactivity were evaluated by video image analysis in nonmalignant prostates and in 30 stage D2 prostate cancers. RESULTS: Apolipoprotein-D was detected in all 30 tumors, and the level of staining was elevated in comparison to age-matched nonmalignant prostates (p < 0.05). In contrast, the level of PSA staining in tumors was less than that detected in nonmalignant prostates. CONCLUSIONS: Apolipoprotein-D is expressed in normal human prostate. Elevated Apo-D staining is associated with advanced prostate cancer.

Adolescent↗

PubMed1995

Medroxyprogesterone acetate therapy in advanced breast cancer: the predictive value of androgen receptor expression.

PURPOSE: To determine the predictive value of androgen receptor (AR) levels in primary tumors of women who undergo medroxyprogesterone acetate (MPA) therapy for advanced breast cancer after relapse on tamoxifen adjuvant therapy. METHODS: Between 1984 and 1987 at Flinders Medical Centre, South Australia, 136 postmenopausal women received adjuvant tamoxifen therapy for lymph node-positive breast cancer. Estrogen receptor (ER), progesterone receptor (PgR), and AR levels, tumor size, and degree of axillary node involvement were determined at the time of diagnosis. The median follow-up period was 81 months; 89 women developed metastatic disease, 83 of whom subsequently received MPA (500 mg/d). The objective response rate ([RR] ie, complete response [CR] and partial response [PR]) and progression-free interval (PFI) were assessed in response to MPA therapy. Associations between RR, PFI, and primary tumor characteristics including ER, PgR, and AR levels were examined using the Mann-Whitney U test, Kaplan-Meier product-limit estimator, and Cox proportional hazards regression, as appropriate. RESULTS: Thirty-two of 83 patients (38.6%) responded to MPA. RR was significantly associated with the presence of AR (P < .001), but not with other primary tumor characteristics or duration of tamoxifen therapy. After initiation of MPA treatment, PFI increased with increasing concentration of AR in the primary tumor. CONCLUSION: Response to MPA after adjuvant tamoxifen treatment for lymph node-positive breast cancer was positively associated with AR level in the primary tumor. This finding suggests that MPA action in breast cancer may be mediated in part by the AR.

Adult↗

PubMed1994

Detection of discrete androgen receptor epitopes in prostate cancer by immunostaining: measurement by color video image analysis.

To determine whether multiple features of immunohistochemical staining of the androgen receptor (AR) in prostate cancer could reliably predict androgen dependence, tumor biopsy specimens from 30 patients (stages A-D2) were stained using anti-peptide antibodies to the amino- and carboxyl-terminal of the AR. Measurements were made of the mean area and total amount (i.e., integrated optical density) of AR staining in at least 20 fields per section using a color video image analysis system, and the mean intensity of AR staining per cell and the percentage of AR positive tumor cells were derived. Video image analysis measurement identified quantitative differences in AR staining between the two antibodies, suggesting that this approach may provide a means of identifying receptor variants in prostate tumors. The AR staining measurements were analyzed by discriminant function analysis to assign individual cases to good and poor clinical outcome groups. AR staining features measured with a single antibody (e.g., amino-terminal) were sufficient to predict outcome following hormonal therapy in stage D2 patients (predictive value, 1.0), whereas all features of AR staining measured with both antibodies were required for the entire patient group (predictive value, 0.97). The principal discriminant in both patient groups contributing to the correct assignment of outcome was the mean intensity of AR staining per cell. These findings suggest that AR staining features measured by video image analysis have the potential to predict outcome in prostate cancer.

Aged↗

PubMed1994

The relative prognostic significance of total cathepsin D and HER-2/neu oncogene amplification in breast cancer. The South Australian Breast Cancer Study Group.

Total tumor cathepsin D (TCD) levels were determined prospectively by a radioimmunometric assay in tumor cytosol of 858 primary breast cancer patients diagnosed between 1989-1991. In 581 of these patients, tumor HER-2/neu oncogene amplification was simultaneously determined. In a "training-set" of 313 patients, "high" TCD was associated with significantly shorter disease-free survival (DFS). For the whole group, there was no correlation between TCD and pathologic stage, number of axillary nodes with tumor deposits, tumor size, histologic type and grade, or hormone receptor levels. In the node-positive group, high TCD level was associated with HER-2/neu amplification. After a median follow-up duration of 31 months, univariate analysis indicated that high TCD level was significantly associated with shorter DFS only in node-positive patients. The shorter DFS in association with high TCD levels was observed in both estrogen-receptor-positive and -negative patients. Cox multivariate analysis of DFS confirmed that high TCD level was predictive of shorter DFS in node-positive patients only. Because of the short duration of follow-up, the significance of TCD in overall survival was not determined. We conclude that high tumor TCD in node-positive patients is predictive of shorter DFS, and is often associated with HER-2/neu amplification. The possibility exists that high tumor TCD may act in combination with HER-2/neu amplification to promote dissemination of metastases.

Adult↗

PubMed1994

Nuclear shape and prognosis following orchiectomy in stage D2 prostate cancer.

In this study, we have examined whether tumor grade and morphometric nuclear features can predict the outcome of treatment by orchiectomy in patients with stage D2 prostate cancer. Two outcome groups based on duration of survival postorchiectomy were examined, a bad outcome group of 63 patients who died from prostate cancer within 12 months and a good outcome group of 34 patients who survived beyond 5 years. Tumors were histologically classified as well (17%), moderate (17%), or poorly differentiated (66%). Tumor grade and patient outcome were significantly associated (Mann-Whitney test; P < 0.005), with 76% of poorly differentiated tumors in the bad outcome group, and 65% of well-differentiated tumors in the good outcome group. Using discriminant function analysis, tumor grade correctly predicted outcome in 70% of cases. A statistically significant difference was also detected in nuclear shape values between the two outcome groups (P < 0.05) and histological grades (P < 0.05). Using discriminant function analysis, 51% of cases were correctly classified into outcome groups using nuclear shape factors, a figure which rose to 65% when all nuclear morphometric features were used. This demonstrates that nuclear morphometric features are of no clinical value in predicting the outcome of treatment in stage D2 disease. Furthermore, these evaluations cannot select patients who might be spared orchiectomy on the basis of a predicted poor response. However, nuclear shape and variance measurements of benign glandular epithelial cells within cancerous prostates were significantly different from those of malignant cells (P < 0.005). We conclude that, while video image analysis of prostatic nuclear shape can reliably discriminate between benign and malignant cells, nuclear morphometric features are of minimal prognostic value in men with stage D2 prostate cancer treated by androgen ablation.

Aged↗

PubMed1994

Glycosaminoglycans of guinea pig prostate fibromuscular stroma: influence of estrogen and androgen on levels and location of chondroitin sulfate.

The effects of aging and hormone manipulation on the glycosaminoglycan (GAG) content of prostatic stroma in guinea pigs were investigated. Total GAG and individual GAG classes (chondroitin, dermatan, and heparan sulfates, and hyaluronic acid) were measured biochemically in stromal extracts. Chondroitin sulfate was also measured and localized by video image analysis of immunocytochemically-stained tissue sections. The weight and total GAG (uronic acid) content of prostatic stroma increased between the ages of 2 weeks and 2 years by 7-8-fold and 4-5-fold respectively. GAG concentration per unit weight of stroma declined 4-fold during puberty and remained essentially unchanged thereafter. Similar results were obtained for each of the GAG classes. The decreases in GAG concentration were associated with a 3-fold increase in the size of the smooth muscle cells of the prostatic stroma during puberty. Hormonal control of GAG deposition in the prostatic stroma was investigated by steroid replacement in prepubertally-castrated animals. Administration of dihydrotestosterone (DHT) to castrate animals for 6 weeks resulted in significantly reduced concentrations of stromal uronic acid, compared with untreated castrate animals (P < 0.05). The GAG levels post-DHT treatment were similar to those observed after pubertal development in sham-operated control animals. Estradiol treatment had the opposite effect to that of DHT, resulting in a significantly increased concentration of uronic acid compared with castrate animals (P < 0.05). These steroid-induced changes in stromal GAG deposition were mostly contributed to by chondroitin and dermatan sulfates. Combined treatment with DHT and estradiol resulted in stromal uronic acid concentrations similar to those of animals receiving DHT alone, indicating that the effect of DHT on stromal GAG deposition is dominant over the effects of estradiol. Morphometric measurement, using computer-assisted video image analysis of a chondroitin sulfate epitope in prostatic sections stained with a monoclonal antibody (6C3), supported the biochemical data. Stereometric profiles across several sectioned glands demonstrated that chondroitin sulfate was confined to the periacinar basement membranes of the prostatic stroma in all groups except the estradiol-treated castrate animals, where the immunostaining extended from the periacinar basement membrane throughout the fibromuscular stroma. Treatment of castrate animals with estradiol alone also induced a physicochemical change in the chondroitin sulfate molecule, resulting in reduced electrophoretic mobility. In summary, this study identifies changes in the quantity, structure, and localization of chondroitin sulfate in the prostatic stroma of estradiol-treated guinea pigs. Furthermore, estradiol and DHT have opposing effects on the level of chondroitin and dermatan sulfate expression in the prostatic stroma.

Aging↗

PubMed1994

Effects of oestradiol-17 beta and 5 alpha-dihydrotestosterone on guinea-pig prostate smooth muscle cell proliferation and steroid receptor expression in vitro.

Smooth muscle cells (SMCs) are the major cellular component of the prostatic stroma. The aim of this study was to examine the effects of oestradiol-17 beta (OE2) and 5 alpha-dihydrotestosterone (DHT) on the proliferation of guinea-pig prostate SMCs in vitro. OE2 stimulated SMC DNA synthesis at all concentrations examined. At a plating density of 3.0 x 10(4) cells/cm2, maximal incorporation of [3H]thymidine (136% of control) was observed after 36 h of treatment with 1 nmol OE2/l. At the same plating density, DHT had an inhibitory effect on SMC DNA synthesis, with maximal effects (73% of control) being observed 24 h after treatment with 1 nmol DHT/l. These effects of OE2 and DHT were prevented by co-incubation with specific steroid receptor antagonists. At a threefold lower plating density (1.0 x 10(4) cells/cm2), the maximal stimulatory and inhibitory effects of OE2 and DHT were delayed by approximately 24 and 12 h respectively. At the lower plating density, a biphasic effect of DHT was observed on DNA synthesis; DHT was both inhibitory and stimulatory. Maximal inhibition (71% of control) and maximal stimulation (168% of control) were observed after 36 and 134 h treatment with DHT respectively. At the lower plating density, longer term treatment of SMC cultures with OE2 and DHT also resulted in an increase in cell number. After 7 days of treatment with OE2 and DHT, cell number increased by 13% and 12% respectively. When OE2 and DHT were added in combination, the short-term inhibitory effect of DHT on SMC DNA synthesis was dominant over the stimulatory effect of OE2. Treatment with DHT for 24 h significantly inhibited OE2-induced stimulation of [3H]thymidine incorporation, irrespective of the prior duration of OE2 treatment. At the lower plating density, OE2 also decreased oestrogen receptor (ER) mRNA levels to 38% of control levels after 24 h of treatment. ER mRNA levels remained repressed until 72 h after treatment with OE2, and returned to control values following 96 h of treatment. Both the androgen-induced inhibition and stimulation of DNA synthesis observed following treatment of SMCs with 1 nmol DHT/l were associated with a reduction in androgen receptor (AR) mRNA levels. At an intermediate time (i.e. 48 h after commencement of treatment with DHT) AR mRNA levels were increased more than twofold over control levels.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1994

Age-related changes in guinea pig prostatic stroma.

BACKGROUND: The histology of benign disease of the human prostate (benign prostatic hyperplasia) is heterogeneous. No other species demonstrates the same complexity, and current animal models do not appear to fully encompass the stromal and epithelial developmental changes involved in the human disease. This study describes age-related changes in the prostatic smooth muscle stroma of guinea pigs and humans, which may be pertinent to some aspects of the disease process in humans. EXPERIMENTAL DESIGN: Histologic and ultrastructural changes were examined and measured in the prostates of guinea pigs during aging (2 weeks to 31 months). Similar measurements were also made in human prostatic tissues during aging and the development of benign prostatic pathology. RESULTS: Morphometric analyses of prostates in guinea pigs and men demonstrated similar changes in stromal volume with age. The stromal volume proportion of the prostate in both species decreases at puberty due to the expansion of the epithelial cell compartment, and is followed by a progressive increase during adulthood until a maximum stromal content of approximately 75% of total tissue volume is reached at age 2 years in guinea pigs, and at age 70 years in men. The pathognomic feature of nodularity and the dramatic increase in gland size observed during the late stages of human benign prostatic disease did not occur in the guinea pig prostate. Ultrastructural analysis of guinea pig prostatic smooth muscle cells identified a progressive hypertrophy (approximately 10-fold) from prepuberty through to old age. Two-thirds of smooth muscle cells in the prostatic stroma of aging individuals of both species demonstrated perinuclear organelles (rough endoplasmic reticulum and free ribosomes) that were not present in younger individuals. CONCLUSIONS: The prominent histologic features of the guinea pig prostate during aging are increased stromal mass, significant stromal fibrosis, and occasional prostatitis. These features are frequently observed in men with clinical benign prostatic hyperplasia. The age-related increases in prostatic smooth muscle cell size and content of perinuclear organelles in the guinea pig suggest a re-activation of cellular synthetic activity. The similarity in some features of the prostatic smooth muscle stroma between aging men and guinea pigs suggests that there may be common pathophysiologic processes. We conclude that the guinea pig could be a useful model for examination of the age-related hypertrophy of the smooth muscle cell and the processes inducing reversion to a more synthetic smooth muscle cell phenotype.

Adolescent↗

PubMed1993

Clinical significance of HER-2/neu oncogene amplification in primary breast cancer. The South Australian Breast Cancer Study Group.

PURPOSE: To determine prospectively the prognostic significance of HER-2/neu oncogene amplification in the primary tumors of breast cancer patients. METHODS: HER-2/neu amplification in tumor DNA was determined by the slot-blot technique in 1,056 patients with breast cancer (stage I to III) diagnosed between 1987 and 1990. Parameters such as estrogen receptor (ER) and progesterone receptor (PgR) levels, tumor size, axillary nodal involvement, tumor grade, and time to relapse were prospectively obtained. RESULTS: HER-2/neu oncogene amplification, > or = 2, > or = 3, and > or = 5 copy number, was detected in 21%, 11%, and 7% of patients, respectively. In a test set of 529 patients, Cox multivariate analysis showed HER-2/neu copy number > or = 3 or > or = 5 was associated with shorter disease-free survival (DFS) duration. HER-2/neu copy number > or = 3 correlated significantly with pathologic stage of disease, number of axillary nodes with tumor, histologic type, and absence of ER and PgR. For all patients, after a median follow-up duration of 39 months, Kaplan-Meier univariate analysis indicated that tumor oncogene copy number > or = 3 correlated with shorter DFS in both node-negative and node-positive patients. In Cox multivariate analysis, HER-2/neu copy number > or = 3 was associated with shorter DFS, independent of nodal status, ER level, and tumor size. CONCLUSION: Although the follow-up duration of this study is relatively short, we conclude that HER-2/neu amplification is an independent predictor of shorter DFS in both node-negative and node-positive patients.

Adult↗

PubMed1992

Epidermal growth factor receptor in breast cancer: storage conditions affecting measurement, and relationship to steroid receptors.

This study investigates the effect of freezing and storage of tissue and subcellular fractions on the measurement of epidermal growth factor receptors (EGF-r); compares competition binding and single saturating dose assays (SSD) for quantitating EGF-r levels; investigates several tissues as potential quality control; and examines the relationship between EGF-r and hormone receptor expression in human breast cancers. Mouse and calf uterine cell membranes were preferred sources of quality control tissue with similar levels of high affinity EGF-r to human breast cancer tissue (less than 150-200 fmol/mg membrane protein). Studies using pooled mouse uterine tissues indicated a loss of 40% in EGF-r activity following a single-20 degrees C freeze/thaw cycle, while a breast cancer tissue showed a 75% loss, independent of storage temperature (liquid nitrogen, -70 degrees C, -20 degrees C). A single freeze/thaw cycle of mouse uterine broken cell pellets (nuclei plus membrane fraction) again indicated a loss of EGF-r irrespective of storage temperature (43% loss at -70 degrees C, 52% loss at -20 degrees C). In most cases irrespective of the tissue type or tissue fraction being stored, the length of storage had little impact on the extent of the loss in activity. A second freeze/thaw cycle of intact tissue, or freezing of broken cell pellets from a previously-frozen tissue, led to a further major or total loss of the remaining EGF-r. Overall these results are commensurate with the published effects of freezing and storage on estrogen receptor measurement. In addition, our studies suggest that the most suitable procedure for assaying frozen breast cancer specimens for EGF-r levels in conjunction with steroid receptor quantitation is to prepare and assay both cytosol and membrane fractions for their respective receptor content without further storage. A concordance of 86% was found in 44 breast cancers assayed for EGF-4 by saturation analysis and SSD. Statistically significant inverse relationships were found between EGF-r and estrogen and progesterone receptor levels in the study of approximately 350 breast cancer patients. No association was found with tumor stage or diameter, axillary node involvement, or patient age.

Animals↗

PubMed1991

Inhibition of T47D human breast cancer cell growth by the synthetic progestin R5020: effects of serum, estradiol, insulin, and EGF.

The mechanism of the antiproliferative effects of progestins on human breast cancer cells is not known. In view of the ability of estrogen to stimulate human breast cancer cell production of peptide growth factors, and since previous studies have suggested that the inhibitory action of progestins is dependent on estrogen-stimulated growth, the present study examined the interaction of growth factors and the synthetic progestin R5020 on the proliferation of T47D human breast cancer cells. In this study, the concentrations of estradiol, insulin, and EGF for optimal stimulation of T47D cell growth in 3% dextran-charcoal treated fetal bovine serum (DCC-FBS) were determined to be 1 nM, 100 nM, and 1 nM, respectively. Furthermore, incubation with these optimal concentrations of estradiol, insulin, and EGF in various combinations produced additive effects on T47D cell proliferation, suggesting that these agents act, at least in part, by different mechanisms. In contrast, in a chemically defined medium (DM), both estradiol and EGF were unable to stimulate T47D cell proliferation. In the case of estradiol, the inability to demonstrate stimulation of T47D cell growth in DM was not due to down-regulation of the estrogen receptor. R5020 inhibited the growth of T47D cells, although its effect was more marked in the presence of 3% DCC-FBS than in DM. Stimulation of T47D cell growth by either estradiol or insulin in 3% DCC-FBS was effectively inhibited by R5020. In contrast, growth of T47D cells stimulated by EGF in the absence of estradiol was not markedly inhibited by R5020, the growth being comparable to that of untreated control cells. These findings suggest that the inhibitory effect of R5020 on T47D cell proliferation is dominant over the action of some, but not all, breast cancer mitogens.

Animals↗

PubMed1991

The prognostic value of deoxyribonucleic acid flow cytometric analysis in stage D2 prostatic carcinoma.

This study was designed to compare the prognostic potential of tumor grade and ploidy status in patients with stage D2 prostate cancer. Two outcome groups were selected on the basis of survival after orchiectomy: a bad outcome group consisting of 66 patients who died of the disease within 12 months and a good outcome group comprising 37 patients who survived beyond 5 years. Tumors were classified histologically as well (17%), moderately (17%) or poorly (66%) differentiated. Tumor grade was a significant predictor of outcome, with 76% of poorly differentiated tumors in the bad outcome group and 65% of well differentiated tumors in the good outcome group (p less than 0.005). Deoxyribonucleic acid (DNA) ploidy analysis was performed on formalin fixed, paraffin embedded samples of the primary tumor to yield 97 final tracings that were classified using set criteria for DNA ploidy status. Over-all, 54% of the tumors were nondiploid (33% aneuploid and 21% tetraploid) and the remaining 46% were diploid. DNA ploidy status was a significant indicator of outcome (p less than 0.001), with 64% of diploid tumors in the good outcome group and 88% of the nondiploid tumors in the poor outcome group. Tetraploid tumors behaved no differently from other nondiploid tumors. We conclude that DNA ploidy status and tumor grading are significant independent predictors of outcome after orchiectomy and when combined yield important additional prognostic information.

Carcinoma↗

PubMed1989

The significance of oncogene amplification in primary breast cancer.

Alterations in the gene copy numbers of the proto-oncogenes HER2/neu and c-myc in primary human breast cancer investigated in 73 patients. We detected amplification of HER2/neu in 17 patient samples and amplification of c-myc in 11, while amplification of both was seen in 6 samples. There was no correlation of age, hormone receptor positivity or tumour size with amplification of either proto-oncogene. Amplification of HER2/neu was significantly correlated with the stage of the disease. HER2/neu amplification was observed in 18.5% and 38% of node-negative and node-positive patients, respectively; the association between HER2/neu amplification and advanced stage of the disease was statistically significant (p = 0.05). Since this is a prospective study, the clinical significance of oncogene amplification is not known. The relatively high frequency of HER2/neu amplification points to a functional role in human breast cancer, particularly in the progression of the disease. The method used in our study allows oncogene amplification to be studied in conjunction with hormone receptor determination and thus may be of value in large clinical trials to determine the significance of oncogene abnormalities in breast cancer.

Breast Neoplasms↗

PubMed1989

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.

Animals↗

PubMed1989

Development and characterization of primary cultures of smooth muscle cells from the fibromuscular stroma of the guinea pig prostate.

Primary cultures of smooth muscle cells (SMCs) were obtained by a two-step enzymatic digestion of guinea pig prostatic stroma. Ultrastructural morphology and growth characteristics of these cells conformed to those reported for SMCs isolated from vascular and visceral tissue sources. Electron microscopic examination indicated that the cells assumed modified myofibroblastoid features in culture. Microfilaments with associated dense bodies were markedly depleted in cultured smooth muscle cells, in comparison with those of the parent tissue. Cultured cells also possessed increased content of rough endoplasmic reticulum indicating the increased secretory or protein-synthetic capacity of the cells. Immunoperoxidase staining for cytoskeletal markers using monoclonal antibodies to desmin and vimentin supported the ultrastructural observations, suggesting a decline in desmin-staining intermediate filaments during "modulation" to the myofibroblastoid form. Despite this depletion of smooth muscle-specific differentiation markers and reversion to more general mesenchymal properties, the cells retained the ability to contract on challenge with norepinephrine, and grew in the characteristic "hill and valley" pattern on attaining confluence. Inasmuch as the estrogen and androgen receptor expression of the parent stromal tissue is also retained, these primary cell cultures should provide a useful model to study regulation of prostatic development.

Androgens↗

PubMed1989

Identification of the major sources of error in estrogen receptor measurements for individual laboratories using both tissue and cytosol samples.

The major sources of error between laboratories performing estrogen receptor measurements in tissue samples were identified for 17 participating laboratories in a trial conducted by the Australasian Quality Assurance programme. Both tissue and cytosol samples were provided, and the In-House assays were compared with the ER-EIA kit (Abbott Laboratories, U.S.A.) as a reference assay. For both the In-House and Abbott assays, tissue samples resulted in a between laboratory CV of about 55% and a within laboratory CV of about 30%. In contrast to tissue samples, the between laboratory CV for cytosol samples was reduced to 41% for the In-House assays and to 33% for the Abbott assay, whereas the within laboratory CV was reduced to 10% for both types of assay. The different methods of tissue homogenization by themselves were not found to be sources of error, and protein extraction efficiency from tissue was strongly correlated with protein measurement (P less than 0.0005). The major sources of error due to protein measurement, cytosol preparation, In-House and Abbott assays were evaluated for individual laboratories. The results indicated absence of any major sources of error for four laboratories, while one, two and three or more sources were indicated for seven, three and three laboratories respectively. The conclusion that about half the participants need to improve their ER assays was confirmed by three independent reviews. Furthermore, the trial demonstrated that tissue samples are essential as quality assurance material for a realistic assessment of ER assays in biopsy specimens.

Animals↗

PubMed1989

Sources of improvement in between laboratory variation in estrogen and progestin receptor measurement using tissue samples during the Australian Quality Assurance Programme.

Tissue and cytosol samples were compared as quality control material for assessment of between and within laboratory error in measurement of estrogen and progestin receptors (ER and PR) in a series of four trials for a total of 17 participating laboratories during the Australasian Quality Assurance Programme. For tissue samples, a substantial reduction in between laboratory CVs for both ER and PR from about 90 to 50% was achieved during the programme. In contrast, for cytosol samples a substantially lower between laboratory CV of about 30% was obtained. Tissue sample heterogeneity could be excluded as a major source of variation between laboratories. The likely source of the observed improvement in CV for tissue samples during the trials was due to a reduction of the initial under-estimation of receptor concentration in tissue samples by some of the participants. Although cytosol preparation from tissue samples was shown to be one major source of error, other sources of error such as the receptor assay itself and the associated protein measurements were identified. It is concluded that fragmented tissue samples are essential for a realistic assessment of between laboratory error in receptor measurements in biopsy material such as obtained from clinical breast cancer samples.

Animals↗