Growth Hormone mRNA Expression Research Articles

Malnutrition during lactation changes growth hormone mRNA expression in offspring at weaning and in adulthood

Mothers' nutrition during lactation programs growth in their offspring. We studied the contribution of the growth hormone (GH) for this programming, evaluating GH mRNA expression. Lactating dams were grouped as follows: C, control diet with 23% protein; PR, 8% protein-restricted diet; and ER, energy-restricted diet, receiving the control diet in restricted quantities of the PR group's ingestion. Some pups were killed at weaning; the others received the control diet until they were sacrificed as adults. Pituitary GH mRNA was analyzed by Northern blot analysis. At weaning, the ER and PR animals had lower GH mRNA levels (−29% and −18%, respectively) and lower length as well as body weight. Ninety-day-old PR offspring showed a lower body length (−5%), whereas ER offspring showed a higher one (+5%); however, at 180 days, the lengths were not different. Both 90- and 180-day-old animals showed body weight differences against control animals, with PR offspring showing a lower (−10%) and ER offspring showing a higher (+12%) body weight. GH mRNA was higher in ER offspring at 90 and 180 days (+19% and +22%, respectively); it was lower in PR offspring at 90 and 180 days (−19% and −17%, respectively). Thus, we showed a direct relation between GH mRNA expression and length as well as body weight. We suggest that malnutrition during lactation may program GH mRNA expression patterns in adulthood and that these changes could be responsible for differences in growth patterns.

CDNA cloning and mRNA expression of growth hormone in Belontiidae (Anabantoidei suborder)

Complete cDNA encoding for the growth hormone (GH) of the Belontiidae fish family (Anabantoidei suborder) was cloned by RACE PCR using several sets of degenerate oligonu- cleotides. GH cDNA of the pearl gourami (Trichogaster leeri; pgGH), cloned from the pituitary, included the 5’ and 3’ noncoding sequences of 44 bp and 181 bp, respectively, and consisted of 840 bp that encoded for a prehormone of 204 amino acid (aa) residues. GH cDNA of the blue gourami (T. trichopterus; bgGH) and its deduced aa sequence had the same lengths as those of the pgGH, with nucleotide and aa identities of 97% and 99%, respectively. GH cDNA cloned from the dwarf gourami (Colisa lalia; dgGH) differed from both pgGH and bgGH. The identity of the dgGH cDNA nucleotides was 88%, compared to pgGH and bgGH. However, the identity of the deduced dgGH aa sequence was 97% when compared to bgGH and 96% when compared to pgGH. Nucleotides of GH cDNA of the fighting fish (Betta splendens; ffGH) had an identity of 82% to those of pgGH and bgGH, and 81% to the dwarf gourami. Higher identity was found among the aa sequences than among the nucleotide sequences. The identity of the cloned aa ffGH compared to bgGH, pgGH, and dgGH was 93%, 92%, and 91%, respectively. Higher levels of GH mRNA were found in females in immature, vitellogenic, and mature stages than in males in various stages of gonadal development. No significant differences in the GH transcription level were found between immature and mature females and males. However, the mRNA level decreased significantly during sexual behavior in males. GH sequence and expression may be used as systematic markers for Belontiidae fish and possibly other fish.

Thyroid hormone-dependent gene expression as a biomarker of short-term 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) exposure in European common frog (Rana temporaria) tadpoles

The effects on thyroid hormone-dependent gene biomarker responses of the persistent organochlorine pesticide metabolite 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) were investigated after exposure of 4-week-old European common frog (Rana temporaria) (stage 36) tadpoles to two (0.001 and 0.01 ppm) DDE concentrations. Total body weight, total length, and tail length and width increased after 3-day exposure to DDE. Expression patterns of genes encoding for growth hormone, thyroid-stimulating hormone (TSHβ) and thyroid hormone receptor (TRα and TRβ) isoforms were evaluated in the head, body and tail regions using a validated real-time polymerase chain reaction (PCR) method. The mRNA expression of growth hormone in the body, and TSHβ in the head showed significant DDE concentration-dependent decreases. While DDE caused variable effects on TRα mRNA steady-state, the expression of TRβ was significantly decreased in the tail by DDE in a concentration-specific manner. The effect of DDE exposure on TRβ mRNA expression showed a negative correlation with tail length and width during the exposure period. The unique pattern of a DDE-induced decrease of tail TRβ expression probably reflects the significant role of this thyroid hormone receptor isoform in tail re-absorption and overall metamorphosis in anuran species. Therefore, the present study shows that the evaluation of thyroid hormone-dependent genes may represent quantitative biomarkers of acute exposure to organochlorine pesticides in anuran species during critical developmental periods such as metamorphosis. Given the widespread environmental levels of DDT and its metabolites, these pollutants will remain a subject of concern and their effects on anuran species should be studied in more detail.

Independent differentiation of mammotropes and somatotropes in the chicken embryonic pituitary gland

It has been reported that mammotropes in a rodent pituitary gland are derived from somatotropes via somatomammotropes (SMTs), cells that produce both growth hormone (GH) and prolactin (Prl). However, no studies have been done on the transdifferentiation of somatotropes in the chicken pituitary gland. In this study, in order to determine the origin of mammotropes, we studied detail property of appearance of chicken somatotropes, mammotropes and pit-1 cells and then evaluated the existence of SMTs in the chicken embryonic pituitary gland. Immunohistochemical analysis revealed that GH-immunopositive (GH-ip) cells appeared on embryonic day (E) 14 and were mainly distributed in the caudal lobe, while Prl-immunopositive (Prl-ip) cells appeared in the cephalic lobe of the pituitary gland on E16. In situ hybridization (ISH) and RT-PCR analysis showed that expression of GH and Prl mRNA starts at E12 in the caudal lobe and at E14 in the cephalic lobe respectively. Pit-1 mRNA was first detected on E5 by RT-PCR, and pit-1 mRNA-expressing cells were found in the cephalic lobe on E8. Then with the ontogeny of the chicken, these cells spread into both lobes. Using a double staining method with ISH and immunohistochemistry, we could not detect the existence of SMTs in the chicken embryonic pituitary gland even in the marginal region of either lobe. These results suggest that chicken somatotropes and mammotropes independently appear in different lobes of pituitary gland and that transdifferentiation from somatotropes to mammotropes is not the central route for differentiation of mammotropes in the embryonic chicken pituitary gland.

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) as a Growth Hormone (GH)-Releasing Factor in Grass Carp. I. Functional Coupling of Cyclic Adenosine 3′,5′-Monophosphate and Ca2+/Calmodulin-Dependent Signaling Pathways in PACAP-Induced GH Secretion and GH Gene Expression in Grass Carp Pituitary Cells

Pituitary adenylate cyclase-activating polypeptide (PACAP), a member of the glucagon/secretin peptide family, has been recently proposed to be the ancestral GH-releasing factor. Using grass carp as a model for bony fish, we examined the mechanisms for PACAP regulation of GH synthesis and secretion at the pituitary level. Nerve fibers with PACAP immunoreactivity were identified in the grass carp pituitary overlapping with the distribution of somatotrophs. At the somatotroph level, PACAP was shown to induce cAMP synthesis and Ca(2+) entry through voltage-sensitive Ca(2+) channels (VSCC). In carp pituitary cells, PACAP but not vasoactive intestinal polypeptide increased GH release, GH content, total GH production, and steady-state GH mRNA levels. PACAP also enhanced GH mRNA stability, GH promoter activity, and nuclear expression of GH primary transcripts. Increasing cAMP levels, induction of Ca(2+) entry, and activation of VSCC were all effective in elevating GH secretion and GH mRNA levels. PACAP-induced GH secretion and GH mRNA expression, however, were abolished by inhibiting adenylate cyclase and protein kinase A, removing extracellular Ca(2+) or VSCC blockade, or inactivating calmodulin (CaM)-dependent protein kinase II (CaM kinase II). Similar sensitivity to VSCC and CaM kinase II blockade was also observed by activating cAMP production as a trigger for GH release and GH gene expression. These results suggest that PACAP stimulates GH synthesis and secretion in grass carp pituitary cells through PAC(1) receptors. These stimulatory actions probably are mediated by the adenylate cyclase/cAMP/protein kinase A pathway coupled to Ca(2+) entry via VSCC and subsequent activation of CaM/CaM kinase II cascades.

Induction of GH, PRL, and TSHβ mRNA by transfection of Pit-1 in a human pituitary adenoma-derived cell line

The functional development of pituitary cells depends on the expression of a combination of transcription factors and co-factors. Pituitary-specific transcription factor-1 (Pit-1) is required for the expression of growth hormone (GH), prolactin (PRL), and the thyroid-stimulating hormone beta subunit (TSH beta) and acts synergistically with the estrogen receptor (ER) and GATA-binding protein 2 (GATA-2) to induce PRL and TSH beta expression, respectively. The glycoprotein hormone alpha subunit (alpha SU) is the first hormone to be expressed during pituitary development. In addition to being expressed in follicle-stimulating hormone, luteinizing hormone (LH), and TSH cells, alpha SU is reported to co-localize with GH in pituitary cells. These findings have led to the suggestion that the expression of Pit-1 in cells of the alpha SU-based gonadotropin cell lineage might also lead to the expression of GH. In this study, we transfected HP 75 cells (derived from a human non-functioning pituitary adenoma that expressed alpha SU and LH beta) with Pit-1 by using an adenovirus FLAG-Pit-1 construct. Most of the transfected cells expressed GH mRNA, with fewer cells expressing PRL and TSH beta mRNA. The HP 75 cells expressed the genes for ER and GATA-2, thus allowing their expression of GH, PRL, and TSH beta mRNA in response to Pit-1. These results support the hypothesis that GH can be induced in cells that possess an active alpha SU gene and shed light on the basic molecular mechanism that drives the development of GH, PRL, and TSH beta expression in the alpha SU-based gonadotroph lineage.

Molecular cloning, tissue distribution, and ontogeny of mRNA expression of growth hormone in orange-spotted grouper ( Epinephelus coioides)

A full-length cDNA encoding for growth hormone (GH) was cloned from orange-spotted grouper ( Epinephelus coioides) pituitary using reverse transcription and rapid amplification of cDNA ends (RACE). The GH precursor cDNA consists of 956 bp in size with a 85 bp 5′-untranslated region and 259 bp 3′-untranslated region. The 612 bp open reading frame encodes a 204 amino acid (aa) protein, which represents the precursor of grouper GH composed of a 17 aa signal peptide followed by a 187 aa mature GH polypeptide. The sequence of grouper GH shares 95% aa sequence homology with GH reported in gilthead sea bream ( Sparus aurata), and it also exhibits structural features highly homologous to GH reported in other fish species in the domains representing conserved motifs of GH polypeptides. A single GH transcript of 0.93 kb in size has been detected with Northern blot in the pituitary. Using semi-quantitative PCR approach, dominant PCR products were observed in grouper pituitary, while less PCR products were detected in the brain, spleen, and ovary. The expression of GH mRNA could be detected in 1 dph larvae, after that a significant increase in PCR products was found in 5-day-old fish larvae followed by a drop to very low levels in 15-day-old fish larvae. A second rise was then observed in 25-day-old grouper larvae. These findings suggest that in grouper GH mRNA expression can be detected in day 1 post-hatching larvae, and the GH present in eggs and larvae may play a key role in early development of grouper, especially during the process of metamorphosis of fish larva.

Oestrogen Replacement in vivo Rescues the Dysfunction of Pituitary Somatotropes in Ovariectomised Aromatase Knockout Mice

The mechanism of regulation of growth hormone (GH) secretion by oestrogens and androgens is still controversial. Available data on the action of oestrogens on GH expression and secretion in somatotropes is poorly understood. We previously reported that the aromatase knockout (ArKO) mouse with oestrogen deficiency and excessive androgen levels had dysfunctional somatotropes. In order to clarify the influence of androgen and oestrogen, we investigated the in vivo treatment of ovariectomised (OVX) ArKO mice with exogenous oestradiol (E₂) on the mRNA expression of GH, GH-secretagogue receptor (GHS-R), GH-releasing hormone receptor (GHRH-R), pituitary-specific transcription factor (Pit-1), and somatostatin receptors (sst1–5) in pituitary glands. Circulating plasma GH levels were also evaluated. The results showed that ArKO/OVX mice have a low expression of pituitary GH, GHRH-R, GHS-R and Pit-1, and significantly reduced GH levels. Treatment of female ArKO/OVX (E₂-deficient without excessive androgen) mice with E₂ for 21 days enhanced expression of pituitary GHRH-R and Pit-1 to 151 and 168%, respectively, of that in mice without treatment. E₂ treatment increased GH expression and plasma levels in ArKO/OVX mice to levels comparable with those in wild-type female mice. We conclude therefore that long-term E₂ replacement rescues the dysfunction of somatotropes in ArKO/OVX mice through increases in expression of GH, GHRH-R, and Pit-1 in the pituitary somatotropes, whereas the level of androgen in this oestrogen-deficient female mouse does not significantly influence the function of somatotropes.

Paracrine regulation of growth hormone gene expression by gonadotrophin release in grass carp pituitary cells: functional implications, molecular mechanisms and signal transduction

Growth hormone (GH) is known to stimulate luteinizing hormone (LH) release via paracrine interactions between somatotrophs and gonadotrophs. However, it is unclear if LH can exert a reciprocal effect to modulate somatotroph functions. Here we examined the paracrine effects of LH on GH gene expression using grass carp pituitary cells as a cell model. LH receptors were identified in grass carp somatotrophs and their activation by human chorionic gonadotropin (hCG) increased 'steady-state' GH mRNA levels. Removal of endogenous LH by immunoneutralization using LH antiserum inhibited GH release and GH mRNA expression. GH secretagogues, including gonadotrophin releasing hormone (GnRH), pituitary adenylate cyclase-activating polypeptide (PACAP) and apomorphine, were effective in elevating GH mRNA levels but these stimulatory actions were blocked by LH antiserum. In pituitary cells pretreated with actinomycin D, the half-life of GH mRNA was not affected by hCG but was enhanced by LH immunoneutralization. Treatment with LH antiserum also suppressed basal levels of mature GH mRNA and primary transcripts. hCG increased cAMP synthesis in carp pituitary cells and hCG-induced GH mRNA expression was mimicked by forskolin but suppressed by inhibiting adenylate cyclase and protein kinase A. Similarly, the stimulatory actions of hCG and forskolin on GH mRNA expression were blocked by inhibiting Janus kinase 2 (JAK2) and MAP kinase (MAPK), including P42/44(MAPK) and P38 (MAPK). These results suggest that LH is essential for the maintenance of GH release, GH gene expression, and somatotroph responsiveness to GH-releasing factors. The paracrine actions of LH on GH mRNA expression are mediated by a concurrent increase in GH gene transcription and GH mRNA turnover, probably through JAK2/MAPK coupled to the cAMP-dependent pathway.

Effects of Different Culture Conditions and Leptin on GH mRNA Expression and GH Secretion by Pig Pituitary Cells

Growth hormone (GH) is enhanced in malnutrition; physiological increments in GH secretion seem to play an important role in regulating metabolism during fasting. Leptin has also been shown to play a role, amongst others, in modulating the somatotropic axis. In this study, we investigated how the composition of culture media could influence basal and leptin-stimulated GH secretion and expression in pig pituitary cells. Pituitary cells from 8-month-old sows were incubated for 48 h in presence and absence of 10% fetal calf serum, either in DMEM/Ham's F12, in arginine-free DMEM/Ham's F-12, or in DMEM/Ham's F12 Salts. Cells were then treated for 24 h with GHRH or recombinant human leptin (rhLep) individually or in association with GHRH; cell proliferation, nitric oxide (NO) production and GH expression and secretion were determined. The absence of nutritional factors induced a decrease in cell proliferation, but stimulated both GH secretion and expression. Furthermore, rhLep significantly increased GH expression and secretion irrespective of culture conditions. NO production was only significantly enhanced by leptin under DMEM/Ham's F12 culture conditions. These observations lead us to hypothesize that the adaptive capabilities of pituitary cells may overcome the negative effects of undernutrition; in this context, leptin does not seem to depend on NO pathways in modulating GH secretion.

Effects of exogenous cortisol on the GH/IGF-I/IGFBP network in channel catfish

Glucocorticoids are known to hinder somatic growth in a number of vertebrate species. In order to better understand the mechanisms through which they may act in channel catfish, we examined the effects of feeding cortisol on the growth hormone (GH)/insulin-like growth factor-I (IGF-I)/IGF-binding protein (IGFBP) network. Fish (30.6 ± 3.0 g) were fed once daily for 4 weeks and treatments included: (1) High-cortisol (dietary cortisol provided at 400 mg/kg feed), (2) Low-cortisol (dietary cortisol provided at 200 mg/kg feed), and (3) Control (commercial catfish feed). Fish fed diets with cortisol weighed approximately 50% less than Controls. Feed intake was reduced by approximately 30% in both treatments of cortisol fed fish compared to Controls. A ∼20-kDa IGFBP was observed in plasma from High- and Low-treated fish while it was not detected in Control fish plasma. High-cortisol treatment increased pituitary GH mRNA expression approximately 10-fold while liver IGF-I mRNA expression was not different between cortisol-treated fish and Controls. Cortisol treatments decreased plasma levels of IGF-I. These data indicate that feeding cortisol for 4 weeks reduces weight gain, feed intake, and plasma levels of IGF-I and induces a ∼20-kDa IGFBP. One mechanism through which cortisol may impede growth of catfish is through an increase in a low molecular weight IGFBP which may lead to inhibitory effects on the action of IGF-I.

Rescue from Dwarfism by Thyroid Function Compensation in rdw Rats

The rdw rat was initially reported as having hereditary dwarfism caused by pituitary dysfunction. Subsequent studies on the rdw rat, however, have demonstrated that the primary cause of rdw dwarfism is present in the thyroid gland but not in the pituitary gland. The primary cause of rdw rat disorders is a missense mutation of the thyroglobulin (Tg) gene by a one-point mutation. In the present study, we attempted to rescue the dwarfism of the rdw rats using a diet supplemented with thyroid powder (T-powder) and a thyroid graft (T-graft). The infants of the rdw rat were successfully raised to a mature stage body weight, accompanied by elevation of serum growth hormone (GH) and prolactin (PRL), by the T-powder. Furthermore, the T-graft successfully increased the body weight with fertility. The serum GH and PRL levels in the T-graft rdw rat significantly increased. The serum thyroid-stimulating hormone (TSH) levels in the T-graft rdw rat were significantly decreased but were significantly higher than those in the control rat. The GH and PRL mRNA expression in the rdw rat with the T-graft was virtually the same as that of the control, but the TSH beta mRNA differed from that of the control rats. Thus, the dwarfism in the rdw rat is rescued by thyroid function compensation, such as that afforded by T-powder and T-graft.

Establishment of a time-resolved fluoroimmunoassay for measuring plasma insulin-like growth factor I (IGF-I) in fish: effect of fasting on plasma concentrations and tissue mRNA expression of IGF-I and growth hormone (GH) in channel catfish ( Ictalurus punctatus)

A time-resolved fluoroimmunoassay (TR-FIA) was established and validated that allows for the determination of plasma concentrations of insulin-like growth factor I (IGF-I) in three domestically cultured fishes: channel catfish ( Ictalurus punctatus), hybrid striped bass ( Morone chrysops × M. saxatilis), and rainbow trout ( Oncorhynchus mykiss). Sensitivity of the assay was 0.20 ng/ml. Intra- and inter-assay coefficients of variation (CV) were <7 and <12%, respectively. Serial dilutions of plasma from each species were parallel to the standard curve. Recovery of IGF-I from spiked plasma samples was >90% for all three species of fishes. The IGF-I TR-FIA was biologically validated via its use to determine the effect of fasting on circulating IGF-I levels in channel catfish. Fasting-induced changes in plasma growth hormone (GH), hepatic IGF-I mRNA expression, and pituitary GH mRNA expression were also determined. Fasted channel catfish lost 5.6 and 15.6% body mass after 2 and 4 weeks of fasting, respectively. Plasma IGF-I concentrations were depressed ( P < 0.05) relative to fed controls following 2 and 4 weeks of fasting. Plasma GH concentrations were not different ( P > 0.05) in fasted fish after 2 weeks, but significantly increased ( P < 0.05) by 4 weeks of fasting. Hepatic IGF-I mRNA expression after 2 and 4 weeks of fasting was reduced ( P < 0.05) relative to fed controls. Pituitary GH mRNA expression was similar ( P > 0.05) between 2-week-fasted catfish and fed controls, but was increased ( P < 0.05) in 4-week-fasted catfish. The IGF-I TR-FIA was sensitive, accurate, and precise for all three species of fishes, and provided a low-cost, and non-radioisotopic method for quantifying plasma IGF-I levels in fed and fasted channel catfish.

Novel aspects of growth hormone (GH) autoregulation: GH-induced GH gene expression in grass carp pituitary cells through autocrine/paracrine mechanisms.

GH feedback on its own secretion at the pituitary level has been previously reported, but the mechanisms involved have not been elucidated. Here we examined the autocrine/paracrine effects of GH on GH synthesis using grass carp pituitary cells as a cell model. GH receptors were identified in carp somatotrophs, and their activation by exogenous GH increased steady-state GH mRNA levels and GH production. Removal of endogenous GH by immunoneutralization using GH antiserum inhibited basal as well as stimulated GH mRNA expression induced by GH-releasing factors in fish, including GnRH, apomorphine, and pituitary adenylate cyclase-activating polypeptide-38. Cytosolic mature GH mRNA levels were elevated by GH treatment and reduced by GH antiserum, whereas nuclear GH primary transcripts were almost undetectable after GH immunoneutralization. Inhibition of Janus kinase-2 (JAK2), phosphoinositide 3-kinase, and MAPK also abolished GH-induced steady-state GH mRNA expression. GH immunoneutralization in pituitary cells pretreated with actinomycin D induced a marked decrease in the half-life of GH mRNA, indicating that the clearance of GH transcripts could be enhanced by removing endogenous GH. These results provide evidence that GH can serve as a novel intrapituitary autocrine/paracrine factor maintaining GH gene expression in somatotrophs, and this action is mediated by JAK2/MAPK and JAK2/phosphoinositide 3-kinase cascades coupled to GH receptors.

In vitro and in vivo effects of ghrelin on luteinizing hormone and growth hormone release in goldfish.

We studied the in vitro and in vivo effects of octanoylated goldfish ghrelin peptides (gGRL-19 and gGRL-12) on luteinizing hormone (LH) and growth hormone (GH) release in goldfish. gGRL-19 and gGRL-12 at picomolar doses stimulated LH and GH release from dispersed goldfish pituitary cells in perifusion and static incubation. Incubation of pituitary cells for 2 h with 10 nM gGRL-12 and 1 or 10 nM gGRL-19 increased LH-beta mRNA expression, whereas only 10 nM gGRL-19 increased GH mRNA expression. Somatostatin-14 abolished the stimulatory effects of ghrelin on GH release from dispersed pituitary cells in perifusion and static culture. The GH secretagogue receptor antagonist d-Lys(3)-GHRP-6 inhibited the ghrelin-induced LH release, whereas no effects were found on stimulation of GH release by ghrelin. Intracerebroventricular injection of 1 ng/g body wt of gGRL-19 or intraperitoneal injection of 100 ng/g body wt of gGRL-19 increased serum LH levels at 60 min after injection, whereas significant increases in GH levels were found at 15 and 30 min after these treatments. Our results indicate that, in addition to its potent stimulatory actions on GH release, goldfish ghrelin peptides have the novel function of stimulating LH release in goldfish.

Effect of genotype–nutrition interaction on growth and somatotropic gene expression in the chicken

The neuroendocrine system integrates genotype with external factors such as nutrition to regulate animal growth. To investigate the role of somatotropic axis in the interaction of genotype and nutrition, two series of experiments were conducted using broiler and layer chickens as a model. In the first experiment, both strains of chickens were raised on their respective standard diets and the mRNA expressions of somatotropic genes were investigated on day 5 (D5), D21, and D42 after hatching as composites of genotype and nutrition. The hypothalamic somatostatin (SS) and pituitary growth hormone (GH) mRNA expression as well as the plasma GH levels were higher in layer chickens while the opposite was true for hepatic GH receptor (GHR) mRNA. Regulation of GHR mRNA expression was found to be tissue-specific. Hepatic GHR mRNA content increased with age whereas in the muscle, the peak levels of expression were observed at D5 with significantly higher abundance ratio in the layer. To evaluate genotype–diet interaction on growth and the patterns of gene expression of both layer and broiler chickens, layers were fed broiler diet and vice versa from D1 to 42 in the second experiment. The D42 body weight of layer chickens increased by 35% when fed with broiler diet, whereas that of broiler chickens decreased by 51% when fed with layer food. The diet exchange completely reversed the patterns of hypothalamic SS and pituitary GH mRNA expression and the strain differences vanished when the comparison was made on the same diet basis. The hepatic GHR mRNA decreased by 46.1% in broilers fed with layer food, but increased by 45.6% in the layer fed with broiler diet. The strain differences were diminished but did not completely disappear on the same diet basis for hepatic GH receptor mRNA. In contrast, however, the muscle GHR mRNA expression was not affected by diet exchange and thus, was more genotype-specific. The results suggest that genes of the somatotropic axis respond to nutrition differently at the level of transcription.

Arginine Increases Growth Hormone Gene Expression in Rat Pituitary and GH3 Cells

The effect of arginine (Arg) and Ornitargin^® (OT) [a compound containing the aminoacids Arg, citrulline (Cit) and ornithine (Orn)] administration upon growth hormone (GH) gene expression was studied both in vivo and in vitro (hemipituitaries and GH3 cells) by Northern blot analysis. For in vivo studies, adult male Wistar rats were anesthetized, subjected to i.v. infusion of 200 µl of 150 mM NaCl (control group), Arg (15 or 150 mg) or OT (15 mg of Arg, 1 mg of Cit and 4 mg of Orn) at a rate of 20 µl/min, and killed 50 min thereafter. For the in vitro studies, hemipituitaries or GH3 cells were incubated in 1 ml of appropriate medium containing Arg (15 or 150 mg) or OT (15 mg of Arg, 1 mg of Cit and 4 mg of Orn) for 60 min. The pituitaries of the in vivo and in vitro studies and GH3 cells were subsequently processed for RNA extraction. Total RNA was subjected to electrophoresis in agarose (1%)/formaldehyde gel, transferred to a nylon membrane and subjected to hybridization with a rat GH ³²P-cDNA, and ³²P-18S rRNA probe to correct for the variability in RNA loading. After autoradiography of the membrane, the abundance of GH mRNA and 18S rRNA bands was quantified by densitometry. The in vivo study demonstrated that Arg and OT infusion induced a 2.3-fold increase in GH mRNA expression, which could result from the Arg-mediated inhibition of somatostatin release. In addition, in vitro Arg, but not OT, induced GH gene expression in hemipituitaries and GH3 cells, indicating that the aminoacid can act per se at the pituitary somatotrope level. In conclusion, our data show for the first time that arginine stimulates GH gene expression in parallel to its recognized GH-releasing activity.

The inhibition of inducible nitric oxide synthase reverts arthritic-induced decrease in pituitary growth hormone mRNA but not in liver insulin-like growth factor I mRNA expression.

Experimental arthritis induced by Freund-adjuvant administration is a model of chronic inflammation and rheumatoid arthritis associated with a decrease in pituitary growth hormone (GH) and hepatic insulin-like growth factor I (IGF-I) gene expression. Excessive nitric oxide (NO) synthesis by inducible NO synthase (iNOS) has been implicated in the pathogenesis of inflammatory illness. Moreover, NO participates in the regulation of GH secretion at both the hypothalamus and the pituitary. We have examined the role of iNOS activation in producing the changes in the GH-IGF-I axis in arthritic rats. Adult male Wistar rats received aminoguanidine or vehicle from day 20, after adjuvant or vehicle injection, until day 28. Two hours and 30 min after the last aminoguanidine injection, all rats were killed by decapitation. Arthritis increased hypothalamic expression of somatostatin mRNA while it decreased pituitary GH mRNA expression, and both effects were prevented by aminoguanidine administration. In arthritic rats, the parallel decrease in serum IGF-I, and in hepatic IGF-I content and mRNA expression, correlates with the decrease in circulating GH concentrations. Aminoguanidine administration to arthritic rats did not modify either serum GH or serum IGF-I concentrations, or hepatic IGF-I mRNA expression. However, aminoguanidine administration to control rats resulted in a decrease in serum GH concentrations and in a decrease in both hepatic IGF-I mRNA expression and serum IGF-I concentrations. These data suggest that NO mediates the arthritis-induced decrease in GH mRNA expression by acting at a hypothalamic level, but it is not involved in the decrease in hepatic IGF-I mRNA expression.

Targeted Ablation of Gonadotrophs in Transgenic Mice Depresses Prolactin but Not Growth Hormone Gene Expression at Birth as Measured by Quantitative mRNA Detection

We previously reported that transgenic ablation of gonadotrophs results in impaired development of cells immunostainable for prolactin (PRL) but not of cells immunostainable for growth hormone (GH) or pro-opiomelanocortin (POMC) in pituitary of newborn mice. The question remained whether this reduction in PRL protein is a reflection of reduced PRL mRNA expression, or whether this regulation is only situated at the translational level. We therefore generated a new series of transgenic mice in which gonadotrophs were ablated by diphtheria toxin A targeting, and analyzed hormone mRNA levels instead of hormone protein around the day of birth. Pituitary mRNA expression levels of luteinizing hormone-β (LHβ), PRL and GH were quantified using real-time TaqMan RT-PCR. Of the 13 transgenic mice obtained, 8 showed a clear-cut reduction (ranging from 62 to 98%) in LHβ mRNA levels. PRL mRNA values were significantly reduced in the transgenic mice (p = 0.0034), while GH mRNA expression was unaffected (p = 0.93). An additional observation was that female newborn mice produce 5 times more LHβ mRNA than male mice whereas no sex difference was observed for expression levels of PRL and GH mRNA. Moreover, in the wild-type mice, LHβ mRNA expression was 20-fold higher than GH mRNA expression which in turn was 500- to 1,000-fold higher than PRL mRNA expression, suggesting a low expression level of the PRL gene at birth. In conclusion, the present data support the hypothesis that embryonic development of PRL gene expression is stimulated by gonadotrophs.

Relationships between performance traits and the expression of growth hormone, insulin‐like growth factor‐I, and insulin in pigs selected for growth or leanness

SummarySelection for growth rate or backfat thickness (BFT) in pigs is related with changes in the blood plasma concentrations of growth hormone (GH) and insulin‐like growth factor‐I (IGF‐I). Two experimental pig selection lines based upon a common commercial Large White (LW) selection line were selected for increased growth rate (F‐line), or decreased BFT (L‐line), respectively. We report here on the statistical evaluation of the phenotypic relationships between growth rate and BFT on the one hand, and (1) mRNA expression levels of GH and its transcription factor pituitary‐specific transcription factor‐1 (Pit‐1), (2) GH protein plasma pulse pattern components and (3) IGF‐I and insulin plasma concentrations on the other hand. Residual regression coefficients (RRC) and residual correlation coefficients (RCC) were used to evaluate the phenotypic residual relations between the traits and the hormone levels. Growth hormone mRNA levels, and to a lesser extent Pit‐1 mRNA levels, were inversely related with BFT and growth rate when these traits were fitted separately in the model suggesting that GH is more closely related to the traits than Pit‐1. Within line evaluation showed that GH mRNA levels had significant RRC with the performance traits in both lines, while Pit‐1 mRNA levels were only significantly related to BFT in the L‐line in both analyses. When growth rate and BFT were fitted together in the model GH mRNA levels had also significantly RRC with growth rate and BFT. Growth hormone mRNA levels seem to be more closely related with BFT than with growth rate. Growth rate was inversely related with the GH plasma baseline value in both lines. Growth hormone baseline concentration, and to a lesser extent GH area under the curve, were inversely related to BFT. However, GH level‐line‐specific effects were only found for the baseline concentration in the F‐line. A positive relationship between growth rate and plasma IGF‐I concentrations was found in the L‐line. These results suggest a relationship between GH mRNA expression and performance traits in pig lines selected for growth rate or against BFT. No such relationship could be found for its pituitary‐specific transcription factor. Furthermore, specific components of the GH blood plasma pulse pattern were related to specific performance traits in these selection lines. The IGF‐I blood plasma concentrations were related to growth rate in these selection lines.