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Defective clearance of the amyloid-β peptide (Aβ) from the brain is considered a strong promoter in Alzheimer's disease (AD) pathogenesis. Astrocytes and microglia are important mediators of Aβ clearance and Aβ aggregation state and the presence of amyloid associated proteins (AAPs), such as Apolipoproteins E and J (ApoE and ApoJ), may influence Aβ clearance by these cells. Here we set out to investigate whether astrocytes and microglia differ in uptake efficiency of Aβ oligomers (Aβoligo ) and Aβ fibrils (Aβfib ), and whether the Aβ aggregation state and/or presence of AAPs affect Aβ uptake in these cells in vitro. Adult human primary microglia and astrocytes, isolated from short delay post-mortem brain tissue, were exposed to either Aβoligo or Aβfib alone or combined with a panel of certain AAPs whereafter Aβ-positive cells were quantified using flow cytometry. Upon exposure to Aβ combined with ApoE, ApoJ, α1-antichymotrypsin (ACT) and a combination of serum amyloid P and complement C1q (SAP-C1q), a clear reduction in astrocytic but not microglial Aβoligo uptake, was observed. In contrast, Aβfib uptake was strongly reduced in the presence of AAPs in microglia, but not in astrocytes. These data provide the first evidence of distinct roles of microglia and astrocytes in Aβ clearance. More importantly we show that Aβ clearance by glial cells is negatively affected by AAPs like ApoE and ApoJ. Thus, targeting the association of Aβ with AAPs, such as ApoE and ApoJ, could serve as a therapeutic strategy to increase Aβ clearance by glial cells., (Copyright © 2014 Wiley Periodicals, Inc.)

Various C-terminally truncated amyloid beta peptides (Abeta) are linked to Alzheimer's disease (AD) pathogenesis. Cerebrospinal fluid (CSF) concentrations of Abeta38, Abeta40, and Abeta42 were measured by enzyme-linked immunosorbent assay in 30 patients with AD and 26 control subjects. CSF Abeta42 levels was decreased in patients with AD, whereas CSF Abeta38 and Abeta40 levels were similar in patients with AD and control subjects. All three Abeta peptides were interrelated, particularly CSF Abeta38 and Abeta40. Diagnostic accuracy of CSF Abeta42 concentrations was not improved by applying the ratios of CSF Abeta42 to Abeta38 or Abeta40.

Several effects of androgens on LNCaP-FGC prostate tumor cells showed a biphasic pattern. Stimulation of growth and inhibition of secretion of prostatic acid phosphatase (PAP) was observed at low androgen concentrations (below 1 nM of the synthetic androgen R1881), and inhibition of growth and stimulation of PAP secretion was observed at higher concentrations. In contrast, prostate specific antigen (PSA) secretion did not show this biphasic response pattern. Comparable effects were found for two sublines of the LNCaP-FGC cells: an early (passage 20, androgen-dependent) and relatively late (passage 70, androgen-sensitive) passage of the cells. Culturing of both sublines in the presence of a high concentration of androgens (10 nM R1881) resulted initially in a decrease in growth rate, but the cells started to proliferate within 3 weeks. These cells became less sensitive to androgens, lost their biphasic response pattern, and showed reduced androgen receptor levels. Three weeks after removal of the excess of androgens, the passage 70 cells regained a biphasic growth response to androgens. Culture in medium without steroids but with EGF resulted in a decrease of both androgen sensitivity and androgen receptor level. In conclusion, rapid changes of the androgen sensitivity and receptor level of the LNCaP cells occurred under the influence of culture conditions. These changes were partly reversible and, therefore, were most likely due to adaptation of the cells.

Nuclear androgen receptors in benign and malignant human prostatic tumors were estimated with a nuclear exchange assay to evaluate the applicability of the assay to biopsy size tissue samples. The mean nuclear androgen receptor content of six different prostate samples was not dependent on the amount of tissue used. In contrast, however, there was no significant correlation between the results obtained for individual prostates when large samples (500 mg) and samples weighing 100, 50(r = 0.38, n = 14), and 25 mg were compared. This lack of correlation could not be attributed to variations in the assay nor to differences in the percentage of epithelium in the samples. There was no effect of the presence of molybdate on the estimated nuclear androgen receptor level. We concluded that androgen receptors are distributed nonhomogeneously over prostatic tissue and that androgen receptor assays on multiple biopsies are required to obtain a proper estimate of the true androgen receptor content of the tissue.

Androgen receptors (ARn) were assayed in nuclear extracts of prostatic biopsies from 60 patients with benign prostatic hyperplasia (BPH) and 82 patients with prostatic cancer (PC), with an exchange assay using heparin extraction, labelling with 3H-R1881, and protamine sulphate precipitation. The content of ARn of BPH biopsies (38 +/- 34 fmol/mg protein [mean +/- SD]; n = 70) was not different from that of PC biopsies (39 +/- 32 fmol/mg protein; n = 115). Biopsies showing essentially normal prostatic tissue had a lower ARn content (12 +/- 13 fmol/mg protein; n = 6). The content of ARn was independent of the age of the patient and of the histological grade of the carcinomas. A considerable variation in ARn content within tumors of individual patients was found, indicating that ARn are not uniformly distributed over prostatic tissue; ie, cells with high and low receptor content may coexist in different proportions in different regions of the prostate. Therefore, assays on multiple biopsies may be required for a proper estimation of the mean receptor content. The question remains, however, whether the behavior of the tumor is adequately predicted by the mean receptor level or, for instance, by the region with the lowest receptor content.

The cooperative action of 17 beta-estradiol (E2) and polypeptide growth factors in stimulating proliferation of human breast cancer cells in vitro was investigated. To prevent background estrogenic stimulation, only phenol red-free media were used. When cultured in media supplemented with steroid-stripped serum in which all polypeptide growth factor activity had been chemically inactivated, MCF7 cells were unable to proliferate and became virtually quiescent. In the additional presence of insulin, epidermal growth factor (EGF), and E2, however, cells proliferated as rapidly as did cells cultured in media supplemented with fetal calf serum. Analysis by DNA flow cytometry showed that in the absence of external growth factors, MCF7 cells became arrested predominantly in the G1/G0 phase of the cell cycle. Upon addition of insulin in combination with EGF and E2, however, cells reentered the cell cycle with a high degree of synchrony. When added alone, E2 induced only slight mitogenic effects under these growth factor-defined conditions. In contrast, this steroid induced optimal proliferation in conventional steroid-stripped serum, which in itself contained considerable mitogenic activity. Insulin (at 10 micrograms/ml) was the most potent stimulator of MCF7 cell proliferation under growth factor-defined conditions, resulting in a more than sixfold increase in cell number after 96 hours. Other growth factors such as platelet-derived growth factor (PDGF), transforming growth factor beta (TGF beta), and EGF had little effect by themselves and only slightly influenced insulin-induced proliferation. At suboptimal concentrations of insulin (10-100 ng/ml), however, strong synergism was observed between E2 and insulin in inducing MCF7 proliferation. Using the CG5 cell line, a highly E2-sensitive MCF7 variant, synergism with E2 was already observed at 1 ng/ml insulin. It is concluded that MCF7 cells require insulin (or insulin-like growth factors) for proliferation. At suboptimal insulin concentrations, E2 acts synergistically with insulin, possibly by inducing autocrine production of polypeptide growth factors by these cells.

The rat ventral prostate contains prolactin receptors, and during sexual maturation prolactin stimulates the growth of this gland. The aim of the present investigation was to evaluate whether prolactin is involved in the regulation of the number of its own receptor sites in the rat ventral prostate. To this end, the content of prolactin receptors was estimated in prostate membranes of control and chronically hyperprolactinemic rats both before and after in vitro desaturation with 4 M MgCl2. Hyperprolactinemia resulted in a 40% increase in the number of available prolactin receptors (P less than 0.05). In vitro desaturation of receptors resulted in loss of 84% of protein and 36 +/- 6% and 52 +/- 6% of prolactin receptors from ventral prostate membranes of control and hyperprolactinemic rats respectively (P less than 0.05). We have concluded that the rat ventral prostate membranes are not suited to in vitro desaturation of prolactin receptors with MgCl2. From the increase in the number of available prolactin receptors after hyperprolactinemia we have concluded that prolactin is involved in the regulation of the number of its own receptors in the rat ventral prostate.

The nuclear androgen receptor (ARn) content of cancerous prostatic tissue has been investigated as a prognosticator for time to progression under endocrine therapy. In 1981 a prospective study was started to investigate whether the ARn content in biopsy specimens of patients with prostatic carcinoma predicts the duration of response following hormonal treatment. ARn was estimated by a microassay which involves extraction of nuclear pellets with a heparin-containing buffer, exchange labeling of the nuclear extract with 3H-R1881, and quantitation of the receptor with protamine sulphate precipitation. One hundred and fifteen patients with prostatic cancer entered this study; 47 patients had evidence of metastatic disease as proven by bone scan. Forty-two patients were treated by orchiectomy; 37 of these patients are evaluable with a minimal follow-up of 30 months. A relationship between the nuclear androgen receptor content and the time to progression following orchiectomy in these patients with metastatic disease of the prostate was not found. This could possibly be attributed to the heterogeneous nature of the prostatic tumor tissue with respect to the distribution of the ARn. We conclude that androgen receptor assay in needle biopsies, at least in this study, had no value for the prediction of the time to progression after orchiectomy.

Androgen receptor (AR) levels were measured in PC-82 tumor tissue grown in hormonally manipulated nude mice. In the nuclei of tumor tissue from intact male mice a relatively low concentration (mean 25 fmol/mg protein) of androgen receptors (ARn) was found, while no receptors for estrogens or progestins were detected. The total number of androgen receptors in the PC-82 tumor tissue (measured in the nuclei 1 h after injection of a single high dose of testosterone (T) was found to be 100 fmol/mg protein. The antiandrogen cyproterone acetate, administered in combination with the high dose of T, significantly lowered the amount of ARn in the tumor tissue. In the nuclei of tumor tissue from intact tumor-bearing male mice with T-containing Silastic implants, a 4-times higher amount of tightly associated AR was found. In addition, an increased growth rate of the tumor was observed following T implantation. This finding suggests that the increased growth rate of the PC-82 tumor is associated with a continuous occupancy of AR in the nuclei of the tumor tissue. Castration of tumor-bearing male mice, which arrests the growth of this tumor, did not affect the concentration of ARn in the tissue compared to that of tissue in the intact control situation. In addition, the total amount of AR measured after T injection was not affected by castration. Therefore, the availability of a sufficient and steady level of T in the plasma and consequently the duration of the presence of AR in the nucleus of the PC-82 tumor tissue, rather than the total concentration of AR, appear to be the limiting factors in the modulation of hormonal responses in this androgen target tissue.