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J F Wilber

Publications and source records attributed to J F Wilber.

At least 19 recordsLinked to original sources
PubMed2000

Interactions between the prohormone convertase 2 promoter and the thyroid hormone receptor.

The majority of prohormones are cleaved at paired basic residues to generate bioactive hormones by prohormone convertases (PCs). As PC1 and PC2, two neuroendocrine-specific PCs, appear to be the key enzymes capable of processing a variety of prohormones, alterations of PC2 and/or PC1 levels will probably have a profound effect on hormonal homeostasis. We investigated the regulation of PC2 messenger RNA (mRNA) by thyroid hormone using GH3 cells to demonstrate that T3 negatively regulated PC2 mRNA levels in a dose- and time-dependent fashion. Functional analysis of progressive 5'-deletions of the human (h) PC2 promoter luciferase constructs in GH3 cells demonstrated that the regulation probably occurs at the transcriptional level, and that putative negative thyroid hormone response elements were located within the region from -44 to + 137 bp relative to the transcriptional start site. Transient transfections in JEG-3 cells and COS-1 cells showed that the suppressive effect of T3 was equally mediated by the thyroid hormone receptor (TR) isoforms TRalpha1 and TRbeta1. Electrophoretic mobility shift assays using purified TRal and retinoid X receptor-beta protein as well as GH3 nuclear extracts showed that regions from +51 to +71 bp and from +118 to +137 bp of the hPC2 promoter bind to TRalpha1 as both a monomer and a homodimer and with TRalpha1/retinoid X receptor-beta as a heterodimer. Finally, the in vivo regulation of pituitary PC2 mRNA by thyroid status was demonstrated in rats. These results demonstrate that T3 negatively regulates PC2 expression at the transcriptional level and that functional negative thyroid hormone response elements exist in the hPC2 promoter. We postulate that the alterations of PC2 activity may mediate some of the pathophysiological consequences of hypo- or hyperthyroidism.

Animals↗
PubMed1999

Synthetic retinoid CD437 induces S-phase arrest and apoptosis in human prostate cancer cells LNCaP and PC-3.

BACKGROUND: Exposure of prostate carcinoma cell lines to retinoids, which function through the classical retinoic acid nuclear receptor, (RARs) or retinoid X receptors (RXRs), results in minimal cytostatic inhibition of cell proliferation. METHODS: Growth inhibition and various regulatory responses were investigated in two human prostate carcinoma cell lines (LNCaP and PC-3) treated with or without a synthetic retinoid, CD 437. RESULTS: Incubation of prostate carcinoma cell lines with a novel retinoid CD437 resulted in the marked inhibition of proliferation. LNCaP and PC-3 possessed IC50 values for CD437 of 375 nM and 550 nM, respectively. Incubation with 1 microM CD437 for 24 hr resulted in 100% and 60% inhibition of growth in LNCaP and PC-3 cells, respectively. Simultaneously, cell flow cytometric analyses revealed a dramatic increase of the cell population in S phase, in both LNCaP (from 38.6% up to 86.7%) and PC-3 (27.9% to 55.7%), and a decreased proportion of cells in G2 phase, in LNCaP (from 23.7% down to 1.2%) and PC-3 (14.9% to 2.2%), indicating a significant S-phase arrest. The cell growth inhibition and S-phase arrest in these cells were followed by apoptosis, as revealed by the acquisition of the characteristic cell morphology including the appearance of apoptotic bodies, and further confirmed by cellular DNA fragmentation. CD437-induced-S phase arrest was associated with upregulated mRNA levels of p21waf1/cip1/sdi1 in both LNCaP (p53+/+) and PC-3 (53-/-) cells. CONCLUSIONS: CD437 represents a unique retinoid that induces S-phase arrest and apoptosis in both androgen-dependent (LNCaP) and -independent (PC-3) human prostate cancer cells, suggesting a potential role of CD437 in the treatment of human prostate cancer.

Antineoplastic Agents↗
PubMed1999

D-tagatose, a novel hexose: acute effects on carbohydrate tolerance in subjects with and without type 2 diabetes.

AIM: D-Tagatose (D-tag), a hexose bulk sweetener, does not affect plasma glucose levels when orally administered to rodents. Additionally, D-tag attenuates the rise in plasma glucose after mice are administered oral sucrose. The current study was undertaken to investigate the acute glycaemic effects of oral D-tag alone or in combination with oral glucose in human subjects with and without type 2 diabetes mellitus. Glycaemic responses to D-tag also were investigated in subjects after oral sucrose to examine whether the glucose-lowering effects of D-tag in rodents may result from a direct inhibition of intestinal disaccharidases. METHODS: Eight normal and eight subjects with diabetes mellitus were administered 75 g of glucose, 75 g of D-tag, or 75 g of D-tag 30 min prior to a 75 g oral glucose tolerance test (OGTT). Five patients with diabetes mellitus were challenged with a 75 g oral sucrose tolerance test (OSTT) with and without oral pre-treatment with 75 g of D-tag. Patients with diabetes mellitus also received separate 0, 10, 15, 20 and 30 g of D-tag 30 min prior to a 75 g OGTT. RESULTS: Oral loading with D-tag alone led to no changes in glucose or insulin levels in either normal patients or those with diabetes mellitus. Pre-OGTT treatment with 75 g D-tag, however, attenuated the rise in glucose levels in patients with diabetes mellitus (p < 0.02 at 60 and 180 min, and p < 0.01 at 120 min). The glucose area under the curve (AUC) was reduced significantly also by pre-treatment with D-tag in a dose-dependent manner in patients with diabetes mellitus (p < 0.05 for 10 g D-tag, p < 0.001 for 20 g D-tag, and p = 0.0001 for 30 g D-tag). In patients with diabetes mellitus 75 g D-tag similarly attenuated the rise in glucose following an OSTT (p < 0.01 at 30 min, and p < 0.02 at 60 min). Pre-treatment with 75 g D-tag also tended to blunt the rise in insulin following an OGTT in normal patients (p = 0.07 for insulin AUC) but not patients with diabetes mellitus (p = 0.66). Following 75 g of oral D-tag in four normal patients, plasma D-tag levels rose to a mean peak level of 3.6 mg/dl at 90 min. The administration of 75 g D-tag led to diarrhoea, nausea and/or flatulence in 100% of subjects. When D-tag was administered at lower doses ranging from 10 g to 30 g, only three of 10 patients with diabetes mellitus had gastrointestinal symptoms which were much more mild than those evoked by 75 g D-tag. CONCLUSIONS: These results show that oral D-tag significantly blunts the rise in plasma glucose seen after oral glucose in patients with diabetes mellitus in a dose-dependent manner without significantly affecting insulin levels. The minimal elevation of plasma D-tag levels in normal patients and the adverse gastrointestinal effects seen following larger doses of D-tag support poor absorption of this hexose and suggest that D-tag may act by attenuating glucose absorption in the intestine. D-tag may be a useful therapeutic adjunct in the management of type 2 diabetes mellitus.

Adult↗
PubMed1998

The thyrotropin-releasing hormone gene 1998: cloning, characterization, and transcriptional regulation in the central nervous system, heart, and testis.

Mechanisms of triiodothyronine (T3) negative regulation of the human thyrotropin-releasing hormone (TRH) gene were investigated with a chimeric construct of the 5' flanking region fused to a luciferase reporter gene, transfected into human neuroblastoma cells (HTB-11). Maximum negative regulation was achieved with constructs containing bases -242 to +54. Four sequences in this region exhibited homology with half sites of thyroid hormone response elements (TRE) (AGGTCA). The most important site was a sequence with an overlapping TRE/CRE, involving bases -53 to -60 (TGACCTCA). Potential combinatorial interactions of thyroid hormone receptors and CREB at this site were explored. Modest promoter stimulation was achieved with dibutyryl cyclic adenosine monophosphate (cAMP) (10(-3) M) plus IBMX (0.5 mM). Stimulation was greatly enhanced (+820%) by cotransfection of a constitutively activated protein kinase A (pPKA) construct. Cotransfection with pCREB increased stimulation further to 1350% above control. Stimulation of pPKA and pCREB interfered with stimulation by unliganded TRbeta1, and co-transfected pPKA and pCREB blocked T3 negative inhibition by TRbeta1-T3 complexes. When this site was mutated by polymerase chain reaction (PCR) mutagenesis, the mutant construct failed to respond to unliganded TRbeta1, and stimulation by pPKA and/or pCREB was inhibited markedly, from 12.5- to 2.1-fold, p < 0.001. Moreover, TRbeta1-T3 complexes failed to show any inhibition of the mutated promoter. These results suggest that negative regulation is achieved by inhibition of CREB stimulation of the TRH promoter at this overlapping TRE/CRE site. The two cosuppressors, NCoR and SMRT, were able to augment stimulation of the TRH promoter by unliganded TRbeta1 and enhance the magnitude of T3 inhibition. The potential role of the TRH gene and the pathophysiology of thyroid hormone resistance was investigated with three mutant TRbeta1 constructs. Thyroid hormone resistance was found to be expressed at the level of TRH gene regulation, due to lowered inhibition by mutant TRbeta1-T3 complexes and by their dominant negative effects on wild-type TRbeta1-T3 inhibition. TRH gene expression has been identified in the heart. Cardiac TRH mRNA was not regulated by T3, in contrast to HTB-11 cells, but cardiac TRH mRNA density could be augmented by glucocorticoids and by testosterone. TRH receptors were identified using Scatchard blots that showed a kilodalton of 1.4 nM and a bmax of 10 pmol/mg protein. TRH-R mRNA was identified also by reverse transcription polymerase chain reaction (RT-PCR). Enhanced ventricular contractility by TRH was demonstrated in both an open-chested dog preparation and in ex vivo ventricular myocytes, using video edge cinematography. Under controlled conditions, myocyte shortening was 13.3%, and TRH (10(-6) M) caused muscle shortening to increase 140%, (p < 0.005). TRH gene expression was demonstrated exclusively in Leydig cells of the testis. High affinity binding sites were identified in testicular membranes with a kilodalton of 1.6 x 10(-6) M. TRH was able to inhibit LH and HCG-activated testosterone secretion significantly. Thus, one paracrine role of TRH in the testis may be to serve as inhibitory modulator of gonadotropin-stimulated testosterone secretion.

Animals↗
PubMed1997

Serotonin (5-HT) stimulates thyrotropin-releasing hormone (TRH) gene transcription in rat embryonic cardiomyocytes.

Thyrotropin-releasing hormone (TRH) and its mRNA have been identified in the rat heart, and TRH can enhance cardiomyocyte contractility in vivo. At present, little is known about cardiac TRH gene transcriptional regulation in the heart. Hormones and neurotransmitters, including thyroid hormone (T3), glucocorticoids, testosterone, and 5-HT initiate effects not only in the cardiovascular system, but also in the regulation of hypothalamic TRH. To clarify the potential roles of these modulators upon the cardiac TRH gene transcription, rat TRH promoter activity was assessed in rat embryonic myocyte cells (H9C2) by transient transfection assays. TRH promoter activity was stimulated significantly by dexamethasone (10(-4) M) and testosterone (10(-5) M), and was inhibited by T3 (10(-7) M). Interestingly, the neurotransmitter 5-HT stimulated TRH promoter activity in H9C2 cells, but not in HTB-11 cells. To further clarify this selective role of 5-HT on TRH promoter transcriptional activity in cardiac cells, 5-HT receptor antagonists and agonists were tested. A selective 5-HT2 receptor antagonist blocked 5-HT stimulation, whereas 5-HT agonist analogs caused augmentative effects when combined with 5-HT. Neither 5-HT nor any antagonists or agonists influenced H9C2 cell growth or morphology. These data suggest that 5-HT is an important transcriptional regulator of the cardiac TRH gene.

Animals↗
PubMed1997

Postnatal ontogeny of the thyrotropin-releasing hormone receptor messenger ribonucleic acids in the rat forebrain.

Postnatal developmental change of the thyrotropin-releasing hormone receptors (TRHR) in the rat forebrain was investigated using TRH binding assays and Northern blot analyses from postnatal day 8 to the age of 2 years. TRH binding assays, with [3H]MeHisTRH as the radioactive ligand, demonstrated that the binding capacity in the forebrain was lowest at postnatal day 8 and increased to a maximum level at postnatal day 20. The TRH binding significantly decreased to adult levels between days 20 and 35, and no significant change was observed thereafter. Northern blot analysis, with a 32P-labeled TRHR cRNA probe, revealed that expression of the TRHR gene in the forebrain was not detectable on day 8 after birth, whereas apparent gene expression could be detected in the anterior pituitary. In contrast to the binding capacities, TRHR mRNA levels were very low until postnatal day 20, and increased significantly between days 20 and 35. No significant alteration in mRNA levels was observed after day 35. These results indicated that: (1) TRH binding capacities in the forebrain increased to a maximum levels between the second and third postnatal week and thereafter decreased to adult level, (2) the levels of TRHR mRNA and the TRH-binding capacities did not correlate in that period, suggesting that the TRHR number in the immature forebrain might be regulated by a posttranscriptional mechanism, and (3) expression of the TRHR gene in the forebrain and pituitary seemed to be regulated differentially during development.

Aging↗
PubMed1997

The detection of thyrotropin-releasing hormone (TRH) and TRH receptor gene expression in Siberian hamster testes.

Thyrotropin-releasing hormone (TRH) from the hypothalamus is the major regulator of TSH synthesis and secretion. Most recently, TRH and TRH receptors (TRH-R), as well as their mRNAs, have been identified in rat testis. To expand our knowledge on the testicular TRH and TRH receptor gene expression in different species, in the present study the mRNA levels of testicular TRH and TRH-R were investigated in Siberian hamsters. To further localize the cellular sites of the gene expression, the animal model was treated with a single injection of ethylene dimethane sulfonate (EDS) (i.p., 80 mg/kg body weight), a compound known as to specifically eliminate testicular Leydig cells. The elimination of Leydig cells induced by EDS treatment was confirmed by histological studies of the testis sections and by serum hormonal analyses, which showed a dramatic reduction of serum testosterone (T) levels and significantly elevated serum LH concentrations. Messenger RNA levels of TRH and TRH-R in the testes were determined by Northern blot analyses quantitated with densitometry scanning. The results showed that specific TRH-R mRNA, 3.8 kb in size, was identified in Siberian hamster testes and the mRNA levels were significantly elevated in the EDS-treated testes compared to the controls (p < 0.01). Testicular TRH mRNA was also detected; however, no significant differences in TRH mRNA levels were found between EDS-treated and control groups. The size of TRH mRNA was characterized as about 1.2 kb in hamster testes, which was smaller than that observed in the rat hypothalamus (1.6 kb) and in the rat testis (2.0 kb). Further studies by RNase H digestion revealed the presence of smaller TRH transcripts in the hamster testes than those in the rat testis. No hybridization signal for TRH mRNA was detected by RNase protection assay, when a rat TRH riboprobe was applied to hamster testis RNA, suggesting the limited homology of TRH gene sequences between these two species. Our results demonstrate that both TRH and TRH-R genes are expressed in Siberian hamster testes, and a significant increase of TRH-R mRNA levels occurs in the Leydig cell eliminated hamster testes. Unlike the rat testicular TRH mRNA mainly detected in Leydig cells, in hamster TRH mRNA could also be detected in other testicular compartment.

Animals↗
PubMed1997

Regulation of the human TRH (hTRH) gene by human thyroid hormone receptor beta 1 (hTR beta 1) mutants.

TRH is negatively regulated by T3 both in the hypothalamic paraventricular nucleus and transient transfection models. Mutations in hTR beta 1 genes are associated with the syndrome of generalized resistance to thyroid hormone. To investigate potential effects of mutant TRs on T3 regulation of the hTRH gene, transient gene expression assays were performed in human neuroblastoma (HTB-11) cells with an hTRH promoter-luciferase construct, wild type (WT) hTR beta 1, and three qualitatively distinct hTR beta 1 mutant forms (ED, OK and PV). In the presence of T3 (10(-9) M), liganded WT-hTR beta 1 inhibited hTRH promoter activity significantly (40%). Cotransfection of each of the two mutants (ED and OK) achieved similar levels of inhibition only at 10 to 100 fold increased T3 concentrations. Of interest, a 10x excess of mutant ED or OK could also exert dominant negative effects upon WT hTR beta 1-T3 mediated inhibitory actions on the hTRH promoter. In contrast, mutant TR-PV exerted neither inhibitory nor dominant negative effects at even higher concentrations of T3. Moreover, all three unliganded mutant forms stimulated TRH promoter activity significantly in the absence of T3, despite their different mutations in the ligand-binding domain (LBD). These data demonstrate that thyroid hormone resistance at the level of TRH gene regulation, due to reduced inhibitory actions of mutant TR-T3 complexes, as well as dominant negative effects upon WT hTR beta 1 mediated inhibition, likely contribute to elevated TSH values observed in the syndrome of thyroid hormone resistance.

Dose-Response Relationship, Drug↗
PubMed1996

Reversal of TR-T3 inhibition of the hTRH gene by excess TR ligand-binding domain: evidence for novel accessory protein.

The thyrotropin-releasing hormone (TRH) gene is regulated negatively at the transcriptional level by thyroid hormone (T3). T3 positive regulatory effects on other target genes, such as the growth hormone gene, are mediated through heterodimerization of thyroid hormone receptors (TRs) with RXR or other auxiliary nuclear protein(s). To explore whether an accessory co-suppressor protein(s) may be involved in T3 inhibitory regulation of human TRH gene transcription, transient gene expression studies have been carried out using a hTRH-luciferase (TRH-Luc) chimetric reporter construct, an hTR beta 1 expression construct, and pABgal-hTR beta 1 ligand-binding domain (LBD) fusion constructs, cotransfected into a human neuroblastoma cell line (HTB-11,ATCC). Results herein indicate that T3-dependent inhibitory regulation (48-60% of control) of the hTRH gene promoter by hTR beta 1-T3 complexes could be abrogated completely by cotransfection of a 10 x excess of hTR beta 1-LBD (TR 168-456 aa) in a pABgal94 vector. In striking contrast, cotransfection of a 10 x excess of highly truncated hTR beta 1-LBD (TR 452-456 aa) failed to reverse T3-mediated TRH promoter inhibition. This squelching effect by excessive intact TR-LBD, moreover, could not be reversed by raising T3 concentration 100-fold (from 10(-8) to 10(-6) M), thus excluding a squelching effect of T3 itself by excess LBD. These results suggest that negative regulation of the hTRH gene promoter activity by TR beta 1-T3 complexes involves interactions with an accessory co-suppressor protein, which may bridge DNA-bound TR beta 1-T3 complexes to the transcriptional initiation complex.

Binding Sites↗
PubMed1996

Localization of thyrotropin-releasing hormone mRNA expression in the rat heart by in situ hybridization histochemistry.

Our laboratory has recently detected mRNA of thyrotropin-releasing hormone (TRH) in the rat heart. The density of mRNA for TRH is five-fold higher in the atria than in the left and right ventricle. We also found TRH receptor mRNA and 3H-TRH-binding sites in both ventricles. Cardiac contractility was stimulated after intracoronary administration of TRH. This study was performed to investigate the localization of TRH in the heart. We utilized in situ hybridization histochemistry (ISHH) to localize TRH mRNA expression in the rat heart. ISHH was performed on fresh frozen heart tissue sections which were hybridized with a specific 35S-TRH oligo probe and subsequently processed by autoradiography. The autoradiographic signals corresponding to TRH mRNA were analyzed with an image program. For positive controls TRH mRNA was identified in the hypothalamic paraventricular nucleus. This test confirms the specificity of the TRH oligo probe. Cardiac hybridization signals were observed predominantly in the atria and localized preferentially in atrial connective tissues, vascular adventitia and atrial cardiomyocytes. No hybridization signals were found in ventricular cardiomyocytes. These observations suggest that TRH is synthesized in atrial myocytes and atrial vascular structures. Based on studies which show synthesis of the TRH receptors in ventricular cardiomyocytes, we hypothesize that atrial TRH is an endocrine source for the stimulation of ventricular contractility and that endothelial and adventitial TRH may play a role(s) in the regulation of the growth and/or vasomotor tome of the cardiac vascular system.

Animals↗
PubMed1995

Thyroid hormone enhancement of estradiol stimulation of breast carcinoma proliferation.

Thyroid hormone (T3) and estradiol (Est) modulate biological processes by binding to nuclear receptor proteins that, through interactions with specific response elements in the regulatory regions of genes, modulate gene transcription. Est stimulation of estrogen receptor (ER)-positive breast carcinoma cell growth occurs through its ability to bind to the ER and activate gene transcription. We now report that physiological concentrations of T3 significantly enhance Est stimulation of growth of a number of human breast carcinoma cell lines. The effect of T3 is specific for Est stimulation of growth and has no effect on insulin-like growth factor-I stimulation of growth. The effect of T3 on enhancing Est-mediated growth was specifically blocked by the addition of ligands inducing retinoid X receptor (RXR) homodimer receptor formation, suggesting that RXR-thyroid nuclear receptor (TR) heterodimer formation is required for the T3-mediated effect on estradiol-stimulated growth. Four thyroid nuclear receptors have been described in tissues, TR alpha 1, alpha 2, beta 1, and beta 2. Breast carcinoma cells were found to express TR beta 1 and TR alpha 2 mRNA and very low levels of TR alpha 1 mRNA. T3 did not increase ER mRNA or protein levels and did not enhance Est-mediated increases in gene transcription of a number of genes, i.e., transforming growth factor-alpha and pS2 which contain estrogen-response elements (EREs) in their regulatory regions. However, T3 enhanced Est-stimulated ERE-TK-CAT activity. Thus significant cross-talk appears to occur between the TRs and ER and T3 appears to enhance Est-mediated gene transcription.

Blotting, Northern↗
PubMed1994

Ligand (T3) dependent and independent effects of thyroid hormone receptors upon human TRH gene transcription in neuroblastoma cells.

Thyrotropin releasing hormone (TRH) gene is regulated negatively at the transcriptional level by thyroid hormone (T3) in rat anterior hypothalamus. The actions of T3 upon other target genes are known to be mediated through the thyroid hormone receptors (TR), TR alpha and TR beta. To explore whether the inhibitory regulation of human (h) TRH gene transcription by T3 is TR isoform specific and whether TRH gene transcription can be modulated as well by unliganded TR isoforms, transient gene expression studies have been carried out using hTRH-luciferase (TRH-Luc) chimeric constructs and TR expression constructs, co-transfected into a human neuroblastoma cell line (HTB-11). Data herein demonstrate T3-dependent inhibitory regulation of the hTRH gene promoter by TR-T3 complexes. Moreover, significant inhibition (39%-60%) could be achieved by T3 bound to either hTR alpha 1, hTR beta 1, or rTR beta 1, beta 2 and was comparable quantitatively, indicating an absence of TR isoform specificity for T3 inhibition. Conversely, basal promoter activity of the hTRH gene could be activated significantly by unliganded hTR alpha 1, beta 1, rTR beta 1, and beta 2 (150% to 334%), but not by hTR alpha 2. Thus, TRs appear to exert opposite effects on hTRH gene transcription, depending on the presence or absence of ligand (T3). These dual effects of TR suggest that the addition of the T3 ligand effects conformational changes that can abrogate the initiation of transcription.

Animals↗
PubMed1994

Identification of thyrotropin-releasing hormone receptor in the rat testis.

We have recently documented the expression of preprothyrotropin-releasing hormone (TRH) gene in murine, human and rat testis. Moreover, we have localized TRH to rat Leydig cells immunohistochemically, and found that both prepro TRH mRNA and TRH levels are developmentally regulated in the rat testis. To investigate the potential roles of TRH in testicular function, characterization of TRH receptors (TRH-R) in this tissue was undertaken. Recently, a cDNA encoding murine TRH-R has been isolated, making possible cloning of a rat TRH-R cDNA from the anterior pituitary gland. This cDNA was used for detection of TRH-R gene expression in the rat testis by Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR). TRH receptor assays were also performed with (3H)MeHisTRH as the radioactive ligand. In Northern blot analysis, a single and specific hybridization band, approximately 3.8 kb in size, was identified in whole testis RNA, identical in size with that found in the anterior pituitary gland. The concentration of TRH-R mRNA in the testis was approximately 10% of that in the pituitary. TRH-R mRNA was also detected by RT-PCR in Metrizamide gradient-purified Leydig cells. TRH receptor binding assays revealed the presence of specific, high affinity binding sites with a Kd of 1.6 x 10(-8) M in the testis. Such TRH binding was inhibited by chlordiazepoxide, a specific antagonist of TRH receptor binding. We conclude that TRH may exert local, probably autocrine, actions in the testis via a transmembrane receptor very similar or identical to that in pituitary.

Animals↗
PubMed1993

Identification, localization and developmental studies of rat prepro thyrotropin-releasing hormone mRNA in the testis.

Thyrotropin-releasing hormone (TRH) plays the central regulatory role in the hypothalamic-pituitary-thyroid axis, but is also present in many extra-hypothalamic loci. The adult rat testis has been identified previously as a source of hypothalamic neuropeptides including TRH. To investigate whether the TRH gene is transcribed in testis, the identification and localization of prepro(pp) TRH mRNA and TRH were studied. Northern blot analyses of ppTRH mRNA in the adult rat testis showed a 2.0 kb band, hybridized with a ppTRH cRNA probe. This band was 0.4 kb greater than the 1.6 kb hypothalamic band. The concentration of ppTRH mRNA in the adult testis was approximately 13% of that found in the hypothalamus. Developmental studies of testicular ppTRH mRNA revealed that no ppTRH mRNA could be detected at the earliest stage (day 8). However, hybridization signals were detected on day 20 and increased progressively on days 35, 45 and 70 by 5.8, 6.4, and 9.8-fold, respectively. In addition, ppTRH mRNA was determined in Leydig cells by Northern analyses of elutriated testicular cell fractions. TRH was also measured in the rat testes at different developmental stages by RIA. TRH concentrations paralleled ppTRH mRNA during development. TRH was localized to Leydig cells by immunohistochemistry. These results indicate that ppTRH mRNA and TRH are present in the rat testis, especially in the Leydig cells. The changes of ppTRH gene expression and the concentration of TRH in the rat testis are developmentally dependent. TRH may function as a new paracrine or autocrine regulator of testicular function.

Animals↗
PubMed1993

A truncated isoform of the thyrotropin-releasing hormone receptor is expressed in the rat central nervous system as well as in the pituitary gland.

Using the reverse transcription-polymerase chain reaction (RT-PCR), a cDNA encoding the entire rat thyrotropin-releasing hormone receptor (TRH-R) was isolated from normal rat pituitary gland mRNA. In addition, a novel truncated isoform of TRH-R which lacks 52 base pairs (bp) in the carboxyl (C-) terminal tail was isolated. This truncation, probably generated by alternate splicing, causes a frame-shift and results in a truncated TRH-R 25 amino acids shorter and with a different C-terminal amino acid sequence than the longer type receptor. This truncated TRH-R mRNA, along with the longer receptor form, was found to be expressed throughout the rat pituitary gland and brain.

Amino Acid Sequence↗