Browsing Scholarly Projects 2012 by Authors
The Cryptic Peptides, Prepro-Thyrotropin Releasing Hormone 186-199 and 194-199, Suppress Anterior Pituitary Prolactin Secretion in vivo and in vitroShortridge, Emily; The University of Arizona College of Medicine - Phoenix; Handa, Robert (The University of Arizona., 2012-05-02)Prepro-thyrotropin releasing hormone (ppTRH)-176-199 is one of several peptide fragments cleaved during TRH synthesis and has been implicated as a regulator of neuroendocrine function. ppTRH 176-199 has been shown to acutely inhibit the stress-induced rise in ACTH, corticosterone (CORT), and prolactin (PRL) in the rat. The receptor for ppTRH 176-199 currently remains unknown. In this study we sought to characterize the active domain of ppTRH 176-199 and, using in vivo and in vitro approaches, determine its role in regulating anterior pituitary secretion of PRL. The 186-199, 194-199, and 186-191 amino acid fragments of ppTRH were administered I.P. to adult male Sprague-Dawley rats 15 min. prior to a 20 min restraint stress to determine the peptide’s active moiety in regulating prolactin secretion. Animals were euthanized and plasma was saved for assay of circulating PRL using enzyme immunoassay (EIA). ppTRH 186-199 significantly attenuated the stress-induced rise in prolactin in male rats in a dose-responsive fashion. This effect was mimicked by ppTRH 194-199 but not by ppTRH 186-191. At the highest dose (10 mg/kg BW), ppTRH 194-199 also reduced the stress-induced rise in plasma CORT. Additionally, in vitro studies were performed using the rat growth hormone (GH)/PRL –secreting MMQ cell line. MMQ cells were treated with ppTRH 186-199 and media was assayed for PRL levels. Cells were harvested and examined for changes in PRL mRNA. Within 30 minutes following treatment of estradiol-stimulated MMQ cells with ppTRH 186-199 there was a decrease in media levels of PRL compared to vehicle. Furthermore, in MMQ cells that were primed with 10nM estradiol for 48 hours there was an increase in media PRL levels, which was reduced following ppTRH 186-199 treatment. After 4 hrs of treatment, the inhibitory effect of ppTRH 186-199 on PRL secretion from MMQ cells was only seen on estradiol-stimulated cells. There were no effects of ppTRH 186-199 when examined after 24 hrs of treatment. There were no effects of ppTRH 186-199 or 194-199 of PRL mRNA levels. These data suggest that the carboxy terminal fragment of preproTRH 178-199 contains all the activity of this ppTRH cryptic peptide for regulation of PRL and corticosterone secretion. This suggests a potential moiety responsible for interaction with the peptide’s receptor. The inhibitory effect of ppTRH 186-199 and 194-199 on media PRL levels and not on mRNA synthesis implicates it as an effector of hormone secretion rather than protein synthesis. The short-lived duration of its effects supports a role as 6 an acute effector of the PRL system. The target receptor of the ppTRH 178-199 fragment remains uncertain. However the use of ppTRH 194-199 as a peptide bait may prove useful in identifying the receptor.
A Role for Estrogen Receptor β in the Inhibition of Prostate Cancer Cell GrowthIbragimov, Angela; The University of Arizona College of Medicine - Phoenix; Handa, Robert (The University of Arizona., 2012-04-30)Prostate cancer (PC) and benign prostatic hypertrophy (BPH) are highly prevalent neoplasms. Studies have demonstrated the androgen-dependent nature of benign and pathologic growth of prostate cells. Although Androgen Receptors (AR) have strong proliferative activity in the prostate, recent studies have implicated an anti-proliferative role for Estrogen Receptor Beta (ERβ). This study investigates the effects of ERβ stimulation on normal prostate growth in vivo as well as on PC cell growth in vitro to better elucidate a mechanism for the proposed anti-proliferative actions of ERβ. We also study the interplay between concurrent androgen and ERβ stimulation on PC cell proliferation in vitro. Our hypothesis is that ERβ activation will decrease cell growth and increase cell death in PC cells. Three different ERβ-simulating compounds were analyzed; the selective ERβ agonist diarylpropionitrile (DPN), the dihydrotestosterone (DHT) metabolite 5 alpha androstane-3 β 17b diol (3β-diol), and the isoflavone metabolite, equol, a daidzein-derived compound with phytoestrogen properties. DPN (2mg/Kg) treatment of adult male EZC3 mice for 21 days caused a significant decrease in dorsolateral lobe weight as 4 compared to control (P=.0002). Equol has the same effect on the dorsolateral lobe weight of Sprague-Dawley rats. Furthermore, DPN treatment of human Lymph Node Carcinoma of the Prostate cells (LNCaP) decreased cell proliferation, an effect that was overcome by concurrent treatment with DHT. Interestingly, equol also showed an anti-proliferative effect in cells when used alone as well as in the presence of DHT. 3β-diol, however, did not alter cell growth. Prostate specific antigen (PSA) levels measured from treated LNCaP cells as a measure of androgen stimulation demonstrated that DPN does not interfere with the ability of DHT to stimulate the AR. Furthermore, in vitro data strongly suggest an antagonistic action of equol on the effects of DHT not seen by DPN or 3β-diol. Our data suggest an anti-proliferative role of some ERβ agonists, notably DPN and equol. Although these agonists are ligands of the same receptor, it appears that they activate different molecular pathways and have varying effects on androgen stimulation by DHT. The effects of ERβ agonists are of paramount importance in modulating hormone-induced PC cell proliferation and may have future clinical implication in this widely-prevalent disease condition.