Androgen uptake and binding in rat epididymal nuclei, in vivo.
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Animals↗
Biomedical subjects
D J Tindall
Publications and source records attributed to D J Tindall.
Metabolism of progesterone-4 14 C in porcine uterine endometrium.
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Animals↗
A comparison of the adrenal responses to hypoglycemia, metyrapone and ACTH.
The adrenal responses to insulin-induced hypoglycemia and the rapid adrenocorticotropic hormone (ACTH) stimulation test were compared in 24 healthy volunteers, 18 of whom also underwent a rapid oral metyrapone test. The cortisol levels after hypoglycemia (18.0-30.0 microgram/100 ml) were similar to and directly related to the levels after ACTH (21.0-31.0 microgram/100 ml). The levels after both stimuli were independent of age, sex, height, and weight. The 11-deoxycortisol response to the metyrapone test was less than the cortisol response to hypoglycemia and metyrapone administration was associated with more unpleasant side effects. In a group of 69 control subjects, the post-ACTH cortisol levels were 15.0 to 80.0 microgram/100 ml while in seven patients with Addison's disease they were less than 1-4.5 microgram/100 ml. In 44 control subjects, the posthypoglycemia cortisol levels were 18.0 to 30.0 microgram/100 ml compared with less than 1.0-9.0 microgram/100 ml in 22 patients with hypopituitarism. The absolute poststimulation cortisol levels provided better separation of control subjects from patients with adrenal or pituitary insufficiency than either the increment in cortisol levels or the 11-deoxycortisol response to metyrapone.
17-Hydroxycorticosteroids↗
Kininogenase activity of prostate-derived human glandular kallikrein (hK2) purified from seminal fluid.
Prostate-specific human glandular kallikrein (hK2) is an active enzyme in human seminal fluid. It is one of three serine proteases in the human kallikrein gene family, which includes hK1 (tissue kallikrein) and hK3 (prostate-specific antigen [PSA]). In order to examine kininogenase activity (i.e., production of kinin by these enzymes), we tested for bradykinin and/or Lys-bradykinin release upon incubation of hK2 and for other kallikreins with high-molecular weight kininogen (HMWK), which contains the nonapeptide bradykinin. Kinins are important regulatory peptides (especially for vascular permeability), and they may have a role in enhancing sperm motility. High-molecular weight kininogen is the substrate for plasma kallikrein (PKa potent kinin-generating enzyme circulating in blood, not of the same gene family) and for hK1. Glandular kallikrein and protein-C inhibitor (PCI)-hK2 complex, a serpin protease inhibitor that binds hK2, were purified to homogeneity by affinity and size-exclusion chromatography. About one-half of the hK2 is found in complex with PCI. The kallikrein enzymes were incubated with HMWK, and the resulting cleavage products were analyzed for kinin activity using enzyme immunoassay, high-performance liquid chromatography and mass spectrometry, and in vitro bioassay. Our results show that hK2 cleaves HMWK to produce bradykinin, not Lys-bradykinin (like hK1), and the resultant heavy (56-kDa) and light (42-kDa) chains of HMWK show similar electrophoretic mobility to those cleaved by PK. Prostate-specific antigen (hK3) had no kinin-generating activity. We also identified three other internal cleavage sites for hK2 in HMWK (Arg427, Arg437, and Arg457) that yielded two peptides, one of which is identical to a PK-cleaved peptide. Glandular kallikrein is about 500-fold less active than is PK or tissue kallikrein, but it may play a physiologically important role in bradykinin release in seminal fluid.
Amino Acid Sequence↗
Expression and androgenic regulation of human prostate-specific kallikreins.
In vitro studies suggest that the protease activity of PSA might play a functional role to facilitate the growth and spreading of cancerous prostatic cells. hK2 may have similar properties. Further investigation to prove their in vivo effects is required. Regulation of PSA and hKLK2 gene expression is mediated not only by androgens, but also by a number of autocrine and paracrine factors, suggesting that the control mechanisms for expression of these genes are complex and multifaceted. Such factors may also be integral for the growth and differentiation of prostate cells. Thus PSA and hKLK2 may serve as useful markers to study the regulation of gene expression during cell growth and differentiation of the prostate.
Androgens↗
Modulation of androgen receptor transcriptional activity by the estrogen receptor.
Estrogen/androgen receptor interactions in naturally occurring physiological systems and the effect of their respective steroid hormones on transcriptional activity remain undefined. In an attempt to delineate further the nature of the interaction between these two steroid hormone receptors we have examined the effect of cotransfection of androgen (AR) and estrogen receptor (ER) cDNAs on the expression of the mouse mammary tumor virus long terminal repeat region (MMTV-LTR) linked to the chloramphenicol-acetyltransferase (CAT) reporter gene. In QT6 cells, which contain neither AR nor ER, cotransfection of AR cDNA with the MMTV-LTR-CAT reporter, resulted in transactivation only in the presence of dihydrotestosterone (DHT). Treatment with 10(-8) M each of estradiol-17 beta (E2), dexamethasone, or progesterone did not enhance CAT activity, whereas treatment with the androgens DHT and mibolerone resulted in 87% and 89% CAT activity. Transfection of increasing concentrations of ER cDNA in the presence of 100 ng of AR cDNA and 10(-8) M each of DHT and E2 showed a dose-dependent decrease in CAT activity as compared to the response with DHT alone. Cotransfection of AR and ER cDNA in the presence of 10(-8) M DHT and increasing concentrations of E2 resulted in a dose-dependent decrease in CAT activity. When cells were treated with increasing concentrations of DHT with 10(-8) M E2 no significant increase in CAT activity was observed. In GC cells, which contain endogenous ER but no AR, cotransfection of AR cDNA and treatment with E2 and DHT, also reduced DHT-induced CAT activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Identification of human glandular kallikrein hK2 from LNCaP cells.
Based on studies indicating that human glandular kallikrein (hK2) mRNA is present in the prostate, we prepared a monoclonal antibody to a synthetic peptide corresponding to the 41-56 region of hK2 to try to identify the hK2 protein. Although prostate-specific antigen (PSA) and hK2 share 80% homology, the 41-56 amino acid sequence of hK2 is only 50% homologous with PSA. A monoclonal antibody, HK1A523, was identified that demonstrates high specificity for hK2. In western blot analysis, the antibody has a 1,000-fold greater sensitivity for the detection of hK2 than for PSA. The antibody was used to probe spent media from the prostate carcinoma cell line, LNCaP. An immunoreactive species was N-terminally sequenced and identified as mature hK2. HK1A523 was also utilized to probe prostate tumor cytosols and seminal fluid where putative forms of hK2 were also identified. The hK2 protein therefore is expressed and secreted from prostate carcinoma cells.
Antibodies, Monoclonal↗
Characterization of a TPA-response element in the 5'-flanking region of the androgen receptor gene.
The androgen receptor (AR) protein is an important transacting factor that is necessary for mediating gene expression of androgen-responsive genes. The expression of the AR gene is regulated by androgens and agents that utilize the calcium, protein kinase A, and protein kinase C pathways. Although the role of the calcium and protein kinase A pathways in the regulation of the AR gene has been investigated, the mechanism of regulation of AR through the protein kinase C pathway is not known. We have isolated the 5'-flanking region of the mouse AR gene and identified a consensus TPA (12-O-tetradecanoylphorbol 13-acetate)-response element (TRE). Transient transfection assays indicate that the TRE sequence is sufficient to confer TPA responsiveness to cells treated with TPA. Gel retardation assays and DNA footprint analysis demonstrated specific binding of the TRE and protection of the TRE sequence. Thus, these results describe a TRE in the 5'-flanking region of the AR gene and demonstrate that the TRE is responsive to TPA treatment.
Animals↗