The thyrotrophin-releasing hormone (TRH)-like peptides in rat prostate are not formed by expression of the TRH gene but are suppressed by thyroid hormone

1992 ◽  
Vol 132 (2) ◽  
pp. 177-184 ◽  
Author(s):  
R. Bilek ◽  
P. J. Gkonos ◽  
M. A. Tavianini ◽  
D. G. Smyth ◽  
B. A. Roos
Keyword(s):  
High Performance ◽  
Polymerase Chain Reaction Amplification ◽  
Exchange Chromatography ◽  
Adult Rats ◽  
Thyrotrophin Releasing Hormone ◽  
Basic Group ◽  
Releasing Hormone ◽  
Significant Difference ◽  
Reaction Amplification ◽  
Rat Prostate

ABSTRACT Thyrotrophin-releasing hormone (TRH)-immunoreactive peptides were extracted from rat prostate and divided into two groups by mini-column cation exchange chromatography. The amounts of the peptides in each group were determined by radioimmunoassay with a TRH antiserum. The unretained peptides which lacked a basic group and the retained peptides which possessed a basic group were further purified by high-performance liquid chromatography. The unretained fraction was found to contain a series of TRH-immunoreactive peptides, one of which corresponded chromatographically to synthetic pGlu-Glu-Pro amide and another to pGlu-Phe-Pro amide. None of the TRH-immunoreactive peptides in either fraction exhibited the chromatographic behaviour of TRH. Additional evidence for the absence of TRH gene expression in the prostate was obtained by Northern blot analysis and by application of polymerase chain reaction amplification, which failed to reveal TRH mRNA. Furthermore the preproTRH-derived peptide, preproTRH(53–74), could not be detected by radioimmunoassay. The influence of thyroid status was investigated on the levels of the TRH-like peptides in the prostate. Adult rats were treated chronically with thyroxine (T4) or propylthiouracil (PTU) and the concentrations of the TRH-immunoreactive peptides were determined by chromatography and radioimmunoassay. Treatment with T4 caused the levels of the neutral and acidic TRH-like peptides to fall to approximately one-third of the levels in the controls. No significant difference from the controls was seen in the concentrations of the peptides in the prostates of rats rendered hypothyroid by administration of PTU. The results demonstrate that rat prostate contains TRH-immunoreactive peptides which are not derived from the TRH gene. It is concluded that the TRH-like peptides arise from one or more genes which are structurally distinct from that which codes for the TRH preprohormone. Since these peptides are amidated and their levels are sensitive to hormone administration, it is likely that they fulfil a biological function. Journal of Endocrinology (1992) 132, 177–184

ABSENCE OF PYROGLUTAMYL-N3im-METHYL-HISTIDYL-PROLINEAMIDE (METHYL-THYROTROPHIN RELEASING HORMONE) IN THE RAT HYPOTHALAMUS

1978 ◽  
Vol 77 (3) ◽  
pp. 405-408 ◽  
Author(s):  
A. E. PEKARY ◽  
J. E. MORLEY ◽  
J. M. HERSHMAN
Keyword(s):  
Cation Exchange ◽  
Exchange Chromatography ◽  
Mammalian Brain ◽  
Synthetic Analogue ◽  
Cation Exchange Chromatography ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Rat Hypothalamus

Pyroglutamyl-N3im-methyl-histidyl-prolineamide (methyl-thyrotrophin releasing hormone, methyl-TRH) is a potent synthetic analogue of TRH. N3im-Methyl-histidine is present in mammalian brain and it has been suggested that methyl-TRH is a physiological releasing hormone normally present in the hypothalamus. A non-gradient cation-exchange chromatography system that uses SP-Sephadex C-25 and completely resolves methyl-TRH and TRH has been developed. Because methyl-TRH cross-reacts in the immunoassay for TRH, this assay was used to measure TRH and methyl-TRH in the chromatographic fractions. By this means it has been demonstrated that the amount of methyl-TRH present in the rat is less than 0·025 ng/hypothalamus.

Expression, purification and structural analysis of recombinant rBdh-2His6, a spermadhesin from buck (Capra hircus) seminal plasma

2012 ◽  
Vol 24 (4) ◽  
pp. 580 ◽  
Author(s):  
Antônia Sâmia F. Nascimento ◽  
João B. Cajazeiras ◽  
Kyria S. Nascimento ◽  
Sara Monalisa S. Nogueira ◽  
Bruno L. Sousa ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Structural Changes ◽  
Polymerase Chain Reaction Amplification ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Exchange Chromatography ◽  
Capra Hircus ◽  
Secretory Proteins ◽  
Reaction Amplification ◽  
Polymerase Chain

Spermadhesins, a family of secretory proteins from the male genital tract of ungulate species, belong to the group of animal lectins. Spermadhesins have a prominent role in different aspects of fertilisation, such as spermatozoid capacitation, acrosomal stabilisation, sperm–oviduct interaction and during sperm–oocyte fusion. Proteins (spermadhesins) in buck seminal plasma were described. In the present study, bodhesin Bdh-2 cDNA present in buck seminal plasma was subcloned with the expression plasmid pTrcHis TOPO used to transform Escherichia coli Top10 One shot cells. The recombinant clones were selected by growth in 50 µg mL–1 ampicillin-containing LB broth and polymerase chain reaction amplification. Recombinant rBdh-2His6 synthesis was monitored by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and followed by immunoblotting using monoclonal anti-His antibody. Production of rBdh-2 using low temperatures was not satisfactory. Greater production of rBdh-2 occurred with 1.5 mM isopropyl β-d-thiogalactoside after 2 h of induction. The method used to purify rBdh-2 was affinity chromatography on a His-Trap column following ion-exchange chromatography on a DEAE-Sephacel column. The secondary structure of the rBdh-2His6 was evaluated by spectral profile circular dichroism (CD). The prevalence of secondary structures like β-sheets, with fewer unfolded structures and α-helices, was confirmed. The structure of rBdh-2His6 remained stable up to 35°C. However, significant structural changes were observed at temperatures higher than 40°C related to a distortion of the CD spectrum.

Levels of dopamine and thyrotrophin-releasing hormone in hypophysial stalk blood during an oestrogen-stimulated surge of prolactin in the ovariectomized rat

1985 ◽  
Vol 105 (1) ◽  
pp. 107-112 ◽  
Author(s):  
W. J. de Greef ◽  
W. Klootwijk ◽  
B. Karels ◽  
T. J. Visser
Keyword(s):  
High Performance ◽  
Vascular System ◽  
Ovariectomized Rats ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Methanolic Extracts ◽  
Oestradiol Benzoate ◽  
Hypothalamic Dopamine ◽  
Reliable Increase ◽  
Hypophysial Portal

ABSTRACT The changes in hypothalamic release of dopamine and thyrotrophin-releasing hormone (TRH) into the hypophysial portal vascular system during an oestrogen-stimulated surge of prolactin in ovariectomized rats were investigated. A single injection of 5 μg oestradiol benzoate resulted in a reliable increase in the plasma levels of prolactin during the afternoon 3 days later. Anaesthesia did not block this afternoon surge of prolactin, although its magnitude was only half of that of unanaesthetized rats. Before and during this surge, hypophysial stalk blood was collected into methanol to analyse the hypothalamic release of dopamine and TRH. Immunoreactive TRH in these methanolic extracts eluted as a single peak with the same retention time as authentic TRH on reverse-phase high performance liquid chromatography. In comparison to the morning values, levels of dopamine decreased and those of TRH increased in hypophysial stalk blood by 50 and 240% respectively. These data indicate that hypothalamic dopamine and TRH may be involved in the afternoon surge of prolactin. Daily treatment with parachlorophenylalanine, an inhibitor of serotonin synthesis, reduced the hypothalamic release of TRH by 50%, but did not prevent the afternoon surge of prolactin and TRH induced by oestradiol benzoate. J. Endocr. (1985) 105, 107–112

INTRAVENOUS AND PERORAL TRH STIMULATION IN SPORADIC ATOXIC GOITRE

1977 ◽  
Vol 85 (3) ◽  
pp. 508-514 ◽  
Author(s):  
C. Kirkegaard ◽  
J. Faber ◽  
T. Friis ◽  
U. Birk Lauridsen ◽  
P. Rogowski ◽  
...  
Keyword(s):  
High Frequency ◽  
Diagnostic Value ◽  
Stimulation Test ◽  
Trh Test ◽  
Thyrotrophin Releasing Hormone ◽  
Normal Thyroid ◽  
Releasing Hormone ◽  
Trh Stimulation ◽  
Significant Difference ◽  
Bovine Tsh

ABSTRACT Thyrotrophin releasing hormone (TRH) stimulation test with 200 μg iv was performed in 35 patients with atoxic sporadic goitre. In 23 patients with diffuse goitre 7 showed a lack of increase in serum thyrotrophin (TSH) at a significantly increased frequency compared to controls (P = 0.0028). In 4 patients with solitary nodules 2 showed no significant response to TRH (negative), while 3 of the 8 patients with multinodular goitres had negative TRH test. Only 6 of the 12 TRH negative patients also had non-suppressible 131I uptake following T3. No significant difference in age and thyroid parameters was found between the TRH negative and TRH positive patients. In 7 TRH negative patients the test was repeated with 400 μg TRH but all remained negative. Five of these patients were given TRH perorally 80 mg daily for 2 weeks resulting in a significant increase in serum T4 and T3. No detectable increase in TSH was found. The response to iv bovine TSH in 4 TRH negative patients was found to be normal, suggesting that there was normal thyroid sensitivity to TSH. Our findings suggest that patients with TRH negative atoxic goitre can release biological active TSH following prolonged TRH stimulation. The high frequency of a negative standard TRH test in atoxic goitre seems to diminish the diagnostic value of the standard TRH test.

DEVELOPMENTAL CHANGES IN THE DEGRADATION OF THYROTROPHIN RELEASING HORMONE BY THE SERUM AND BRAIN TISSUES OF THE MALE RAT

1977 ◽  
Vol 74 (2) ◽  
pp. 339-340 ◽  
Author(s):  
C. OLIVER ◽  
C. R. PARKER ◽  
J. C. PORTER
Keyword(s):  
Medical School ◽  
Developmental Changes ◽  
Male Rats ◽  
Male Rat ◽  
Obstetrics And Gynecology ◽  
Adult Rats ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Old Rats ◽  
Phosphate Buffered Saline

*Laboratoire de Médecine Expérimentale, UER Médecine Nord, Boulevard Pierre Dramard, 13326 Marseille Cedex 3, France and †Department of Obstetrics and Gynecology, Southwestern Medical School, 5323 Harry Hines Boulevard, Dallas, Texas 75235, U.S.A. (Received 15 March 1977) Thyrotrophin releasing hormone (TRH) is rapidly degraded when incubated at 37 °C with plasma (Redding & Schally, 1969) or brain homogenates (Bassiri & Utiger, 1974) from adult rats. However, immunoreactive TRH is stable in serum obtained from rats less than 2 weeks old (Oliver, Taurog & Porter, 1974). No loss of biological or immunological TRH activity occurs during incubation with serum from 4- or 16-day-old rats (Neary, Kieffer, Federico, Mover, Maloof & Soodak, 1976). In this report, we have determined the TRH degrading activity of brain homogenates and serum obtained from male rats at various stages of development after birth. Synthetic TRH (1 ng, Beckman Instruments, Inc.) diluted in 50 μl phosphate-buffered saline (0·01

A Double-Blind Trial of Intravenous Thyrotrophin-Releasing Hormone in the Treatment of Reactive Depression

1975 ◽  
Vol 127 (3) ◽  
pp. 227-230 ◽  
Author(s):  
L. E. J. Evans ◽  
P. Hunter ◽  
R. Hall ◽  
Moira Johnston ◽  
V. Mathew Roy
Keyword(s):  
Double Blind Trial ◽  
Double Blind ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Blind Trial ◽  
Reactive Depression ◽  
Antidepressant Effects ◽  
Significant Difference

SummaryIn a double-blind trial 600μg. of thyrotrophin-releasing hormone (TRH) was compared with placebo given daily for four days to two groups of ten patients. There was no significant difference between the antidepressant effects of TRH and placebo.

Effect of prolonged intermittent thyrotropin-releasing hormone administration to fetal and neonatal rats

1979 ◽  
Vol 57 (1) ◽  
pp. 31-34 ◽  
Author(s):  
P. Walker ◽  
J. H. Dussault
Keyword(s):  
Repeated Administration ◽  
Perinatal Period ◽  
Thyroid Stimulating Hormone ◽  
Thyrotropin Releasing Hormone ◽  
Neonatal Rats ◽  
Adult Rats ◽  
Subcutaneous Injections ◽  
Releasing Hormone ◽  
Significant Difference ◽  
The Mean

Fetal and neonatal rats received daily subcutaneous injections of 10 μg thyrotropin-releasing hormone (TRH) until 7 or 14 days postnatally. At 70 days the pups were challenged with 1 μg TRH intravenously via an indwelling jugular cannula. Basal serum thyroxine, triiodothyronine, and thyroid-stimulating hormone (TSH) concentrations did not differ among the three groups. The mean TSH responses as determined by the mean peak TSH concentration and the total TSH response as determined by planimetry were not significantly different, and there was no significant difference in pituitary TSH content following the TRH challenge among the three groups. This study suggests that the integrity of the hypothalamo–pituitary axis in adult rats cannot be affected by the repeated administration of pharmacologic doses of TRH during the perinatal period.

Postnatal developmental pattern of thyrotrophin releasing hormone-degrading activity in rat plasma, hypothalamus and liver: role of tri-iodothyronine

1983 ◽  
Vol 97 (3) ◽  
pp. 409-418 ◽  
Author(s):  
Sonia Aratan-Spire ◽  
Kari Moilanen ◽  
Paul Czernichow
Keyword(s):  
Degradation Products ◽  
Rat Plasma ◽  
Developmental Pattern ◽  
Adult Rats ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Degrading Enzyme ◽  
Liver And Kidney ◽  
Layer Chromatography

Thyrotrophin releasing hormone-degrading activity (TRH-DA) is present in plasma, hypothalamus, pituitary gland, liver and kidney of adults of several species. Each of these tissues contains more than one TRH-degrading enzyme but only one, pyroglutamate aminopeptidase, isolated from the blood, is a TRH-specific enzyme. The aim of this study was to describe the developmental pattern of TRH-DA in the plasma, hypothalamus and liver and the role of tri-iodothyronine (T3) in the development of TRH-DA in the rat. Based on the hypothesis that thyroid hormones stimulate plasma TRH-DA in adult rats, degradation of TRH was studied in hypo- or hyperthyroid rats induced by 6-n-propyl-2-thiouracil or T3 respectively. Tritiated TRH was incubated with plasma and homogenates of hypothalamus or liver. After separation of degradation products by thin-layer chromatography, the amount of degraded [3H]TRH (pmol/50 μl plasma or homogenate) was taken as a comparative index of TRH-DA. Plasma TRH-DA was not detectable before day 9 while hypothalamic and hepatic TRH-DA was already active at birth. Furthermore, the maturation pattern of total TRH-DA was different in plasma compared with other tissues and T3 appeared to play a significant role in its development. The absence of plasma TRH-DA in the neonatal period, its special thyroid-dependent developmental pattern and the presence of a specific TRH-degrading enzyme in adult blood suggest a physiological regulatory role for this activity.

STUDIES ON THE NATURE OF MAMMALIAN HYPOTHALAMIC THYROTROPHIN RELEASING HORMONE USING IMMUNOCHEMICAL, CHROMATOGRAPHIC AND ENZYMIC TECHNIQUES

1975 ◽  
Vol 65 (1) ◽  
pp. 83-90 ◽  
Author(s):  
S. L. JEFFCOATE ◽  
N. WHITE
Keyword(s):  
Ion Exchange ◽  
Human Plasma ◽  
Carboxymethyl Cellulose ◽  
Mammalian Species ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Normal Human Plasma ◽  
Thyrotrophin Releasing Hormone ◽  
Releasing Hormone ◽  
Normal Human

SUMMARY Hypothalamic extracts from three mammalian species (rat, rabbit and sheep) were found to contain several ng of immunoreactive thyrotrophin releasing hormone (TRH)-like activity. This substance chromatographed on ion exchange chromatography (carboxymethyl cellulose) as a single peak that was indistinguishable from synthetic TRH. Hypothalamic TRH was also inactivated by normal human plasma at a rate (1·21–1·46%/μl plasma/h and 1·59–1·77%/50 μl plasma/min) similar to that of synthetic TRH (1·42%/μl plasma/h and 1·73%/50 μl plasma/min). This combination of chromatographic and enzymic techniques can be applied to the identification of immunoreactive TRH in body fluids.

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