Liver GH Receptor mRNA Research Articles

Growth Hormone Secretion Patterns in German Landrace (DL) Fetuses and Piglets Compared to DL Piglets with Inherited 1,25-Dihydroxyvitamin D3 Deficiency.

The regulation of growth hormone (GH) release during prenatal development and during early postnatal life is not entirely clarified. In this study plasma GH concentrations in pigs with inherited pseudo vitamin D deficiency type I (PDDR-I), which regularly show growth retardation, were compared during ontogeny with unaffected pigs of the same breed (German Landrace, DL) as control. Plasma GH concentrations were measured in plasma of chronically catheterized fetuses (beginning on day 101 after mating or after artificial insemination) and in piglets (day 37 postpartum (p.p.)—day 42 p.p.) of both lines. A growth curve beginning at day 7 p.p. was recorded for both lines. The relative amount of GH receptor (GHR) mRNA in liver was quantified by competitive reverse transcription polymerase chain reaction in piglets at day 42 p.p. A trend for higher GH concentrations was observed in PDDR-I fetuses (p < 0.1). In PDDR-I piglets compared to DL piglets higher plasma GH values (p < 0.01), were observed despite lower body weight. The relative quantity of GHR mRNA in liver was not significantly different between the two lines. Piglets with an inherited defect of vitamin D synthesis showed higher GH concentrations. A hormonal imprinting by low 1,25(OH)2D3 could be one reason for our observations and should be analysed in detail in future.

Esterase 1 is a novel transcriptional repressor of growth hormone receptor gene expression: a unique noncatalytic role for a carboxyesterase protein.

The pleiotropic actions of GH result from its engagement with the GH receptor (GHR). GHR expression is regulated by free fatty acids (FFA). A cDNA phage expression library was screened to identify a phage clone expressing esterase 1 (ES1) binding to the FFA-response element (FARE), L2-D1, in the murine GHR promoter. Ectopically expressed ES1 inhibited GHR promoter activity via effects at two FARE, L2-D1 and L2-A2. Chromatin immunoprecipitation experiments demonstrated specific association of ES1 with the FARE. Catalytically inactive ES1 retained inhibitory activity on the GHR promoter and excluded the possibility that the effect on the GHR promoter was an indirect effect secondary to ES1's actions on the intracellular metabolism of FFA. Ectopically expressed ES1 inhibited the endogenous GHR mRNA and protein expression in 3T3-F442A preadipocytes. Subcellular fractionation and confocal microscopy established that ES1 localizes both to the cytoplasm and the nucleus. Experiments demonstrated chromosome region maintenance 1-dependent nuclear export and the presence of a functional nuclear export signal in ES1. The domain of ES1 responsible for the effect on the GHR promoter was localized to the C-terminal portion of the protein. The in vivo significance of ES1's effect on GHR expression was suggested by decreased liver GHR mRNA expression in mice on a high-fat diet correlating with increased steady-state abundance of liver ES1 mRNA. Our results identify and characterize ES1 as a novel transcriptional regulator of GHR gene expression, thereby establishing a unique nonenzymatic role for a carboxyesterase and expanding the potential biological roles of this protein superfamily.

Effects of a lone star tick ( Amblyomma americanum) burden on performance and metabolic indicators in growing beef steers

We examined the response of selected animal performance, endocrine, immune, and metabolic factors from 13 steers (254 ± 6.1 kg) with and without a lone star ( Amblyomma americanum) tick burden during progressive days of the tick feeding cycle. Steers were randomly assigned to either non-treated controls or treated with 300 adult pair of A. americanum per animal. Animals were weighed and blood sampled on days −7, 0, 10, 14, 21, 28, and 35 relative to tick treatment. Tick treatment did not affect ( P < 0.1) animal performance. Experimental day did ( P < 0.05) affect body weight gain and dry matter intake. Tick treatment did not affect ( P < 0.1) metabolic indicators. Experimental day affected ( P < 0.05) IGF1 and lactate, tended to affect cortisol ( P < 0.07), but did not affect ( P < 0.1) glucose concentrations. Tick treatment did not significantly ( P < 0.1) affect growth hormone receptor (GHR) mRNA in liver, but liver tissue from treated animals had numerically lower GHR mRNA than did tissue from control animals. Day had a significant ( P < 0.05) effect on liver GHR mRNA. There was a significant treatment by day interaction ( P < 0.05) for liver IGF1 gene expression, as IGF1 mRNA was reduced in tick-treated cattle versus control cattle on day 35. Overall, liver IGF1 gene expression was lower ( P < 0.05) in tick than in control animals while there was no effect ( P > 0.1) due to day. Within the tick-treated group, correlations were found between quantitative female tick feeding characteristics and host metabolic indicators. Feeding by adult female lone star ticks did cause acute stress in growing beef steers on a moderate plane of nutrition as indicated by some physiologic indicators. In particular there may be longer term effects on the somatotrophic axis in the liver which could affect subsequent (i.e. feedlot) performance. It is not known how these observed effects would be manifest under a lower plane of nutrition, as is common and may become more so within the current native range of A. americanum. Other acute effects due to tick feeding may have been masked by the effects of handling and invasive sampling. Non-invasive experimental procedures are called for in order to study the effects of a stressor such as arthropod infestation on grazing animals. Future research efforts will be aimed at non-invasively elucidating the effects of tick stress on grazing animals under various nutritional environments.

Uterine and Hepatic Gene Expression in Relation to Days Postpartum, Estrus, and Pregnancy in Postpartum Dairy Cows

The somatotropic axis consisting of growth hormone, the growth hormone receptor (GHR) insulin-like growth factor (IGF)-I, and IGF binding proteins changes with the stage of lactation and nutrition of the cow and may be 1 mechanism through which lactation and nutrition affect the establishment of pregnancy. The objective of this study was to quantify GHR, IGF-I, and IGF binding protein-2 (IGFBP-2) mRNA in liver and uterine endometrial tissue at 4 stages of lactation (40, 80, 120, and 160 days in milk) and around the time of artificial insemination. Estrus was synchronized with GnRH and PGF2α, and cows were inseminated 12h after estrus. Uterine biopsies were collected immediately before the second injection of PGF2α (before estrus), at the initiation of standing estrus, and 4 d after estrus. Liver biopsies were collected once on 4 d after estrus. The abundance of GHR, IGF-I, and IGFBP-2 mRNA in liver and uterus was determined by real-time quantitative PCR. The amount of liver IGF-I mRNA was positively correlated with plasma IGF-I concentrations. Cows that became pregnant after AI had more GHR and IGFBP-2 mRNA in their liver than cows that did not become pregnant. There was no effect of DIM or pregnancy status on abundance of uterine mRNA; however, uterine GHR and IGF-I mRNA was most abundant at estrus. In summary, cows at different stages of lactation or with different pregnancy statuses had similar quantities of uterine mRNA. In contrast, liver quantities of mRNA differed relative to pregnancy status. These data provide evidence that liver indices of metabolic state may be indicative of pregnancy success.

Response of the salmon somatotropic axis to growth hormone administration under two different salinities

We compared the response of plasma insulin-like growth factor-I (IGF-I) to growth hormone (GH) administration under two different salinities to test the hypothesis that environmental salinity alters the “activity” of the GH–IGF-I axis. In July, postsmolt coho salmon reared in fresh water (FW) were transferred to either FW or half seawater (1/2 SW) (15 ppt) tank. During the experiment, water temperature was maintained at 10 °C for both salinities; photoperiod was adjusted to that of Seattle (48°N), and fish were not fed. Two days after transfer, fish were injected once with porcine GH (pGH) at a dose of 2 or 8 μg/g body weight. Liver and blood samples were collected 1, 2 and 3 days after injection. Liver GH receptor (GHR) mRNA expression was analyzed by quantitative real-time RT-PCR, and plasma IGF-I, 41-kDa IGF-binding protein (main carrier of IGF-I) and pGH were quantified by radioimmunoassays. Transfer to 1/2 SW resulted in transient increases in basal levels of liver GHR mRNA and 41 kDa IGF-binding protein (IGFBP) but not IGF-I. The GH injection increased liver GHR mRNA, plasma IGF-I and 41-kDa IGFBP in fish in both FW and 1/2 SW. However, the time course and magnitude of the response differed between salinities. Fish in FW receiving 8 μg/g pGH had the highest IGF-I levels (63.7±6.8 ng/ml) one day after injection, whereas fish in 1/2 SW showed a peak (88.8±14.3 ng/ml) two days after injection of the same dose. It is speculated that the prolonged response to GH by fish in 1/2 SW may be due to slower disappearance of pGH from the circulation in fish in 1/2 SW. The transient increase in basal liver GHR mRNA may also contribute to a greater response for fish in 1/2 SW. These results suggest that salinity is capable of altering the “activity” of the GH–IGF-I axis in salmon.

Growth Hormone Stimulates Hepatic Expression of Bovine Growth Hormone Receptor Messenger Ribonucleic Acid through Signal Transducer and Activator of Transcription 5 Activation of a Major Growth Hormone Receptor Gene Promoter

The objective of this study was to determine whether and how GH regulates hepatic expression of GH receptor (GHR) mRNA in cattle. Ribonuclease protection assays revealed that injection of GH in a slow-release formula increased both hepatic GHR and IGF-I mRNAs 1 wk after the injection. The increases in GHR and IGF-I mRNAs were highly correlated. Western blot analysis showed that the injection also increased liver GHR protein level. In cattle and other mammals, hepatic GHR mRNA is expressed as variants that differ in the 5'-untranslated region due to the use of different promoters in transcription and/or alternative splicing. We found that GH increased the expression of the liver-specific GHR mRNA variant GHR1A without affecting the other two major GHR mRNA variants in the bovine liver, GHR1B and GHR1C. In transient transfection analyses, GH could robustly activate reporter gene expression from a 2.7-kb GHR1A promoter, suggesting that GH augmentation of GHR1A mRNA expression in the liver is at least partially mediated at the transcriptional level. Additional transfection analyses of serially 5'-truncated fragments of this promoter narrowed the GH-responsive sequence element down to a 210-bp region that contained a putative signal transducer and activator of transcription 5 (STAT5) binding site. EMSAs demonstrated that this putative STAT5 binding site was able to bind to STAT5b protein. In cotransfection assays, deletion of this putative STAT5 binding site abolished most of the GH response of the GHR1A promoter. Like 1-wk GH action, 6-h (i.e. short-term) GH action also increased liver expression of GHR1A and total GHR mRNAs in cattle. These observations together suggest that GH directly stimulates the expression of one GHR mRNA variant, GHR1A, through binding STAT5 to its promoter, thereby increasing GHR mRNA and protein expression in the bovine liver.

Pituitary mRNA Expression of the Growth Hormone Axis in the 1‐Year‐Old Intrauterine Growth Restricted Rat

Intrauterine growth restriction (IUGR) is one of the major causes of short stature in childhood. Abnormalities in the growth hormone (GH) axis have frequently been observed in children who are born intrauterine growth restricted and GH treatment is effective to improve final height. However, the way that the GH axis is involved is not fully understood. Previously, when investigating the effect of IUGR on the central somatotrophic axis, a hypothalamic effect was discovered with elevated somatostatin and decreased neuropeptide Y mRNA expression levels, whereas serum GH and insulin-like growth factor I (IGFI) were unaltered. These findings were thought to indicate a hypothalamic alteration of the GH axis due to IUGR, probably to compensate pituitary output, thereby normalising peripheral values of GH and IGFI. Therefore, the present study aimed to evaluate the effect of IUGR on the pituitary GH axis in this rat model. Pups from rats that underwent bilateral uterine artery ligation at day 17 of pregnancy were studied. Pituitary glands were collected from 1-year-old offspring for quantitative measurements of GH, GH-receptor (GH-R), GH-releasing hormone receptor (GHRH-R), somatostatin receptor subtype 2 and 5, IGFI and IGFI receptor mRNA levels using a real-time reverse transcriptase-polymerase chain reaction. In addition, liver GH-R and IGFI mRNA expression levels were measured and a radioimmunoassay was performed to determine serum IGFI levels. In the IUGR rat, levels of pituitary GH, GH-R and GHRH-R relative gene expression (RGE) were increased. No differences were found in the RGE level of all other pituitary growth factors, liver GH-R and IGFI, and serum IGFI concentration between IUGR and control rats. The present data show that intrauterine growth failure leads to changes in the pituitary that might counterbalance the effects found previously in the hypothalamus.

Diurnal Variation in Growth Hormone Receptor Messenger Ribonucleic Acid in Liver and Skeletal Muscle of lit/+ and lit/lit Mice

The present study investigated the diurnal variation in GH receptor (GHR) mRNA in liver and skeletal muscle of 3-month-old GH-deficient and -sufficient mice using quantitative real-time RT-PCR. lit/lit (GH deficient) or lit/+ (GH sufficient) mice were fed ad libitum and lights were on between 0600 and 2000. Tissues were collected at 0800-1000, 1200-1400 and 2000-2200. Hepatic GHR mRNA levels of lit/+ mice at 0800-1000 were significantly lower than those at 1200-1400 and 2000-2200. There was no significant variation in hepatic GHR mRNA of lit/lit mice. In skeletal muscle, GHR mRNA levels of both lit/+ and lit/lit mice at 1200-1400 were significantly higher than those at 0800-1000 and 2000-2200. There was also a diurnal change in hepatic IGF-I mRNA levels of lit/+ but not of lit/lit mice; the levels were lowest at 0800-1000 in lit/+ mice. On the other hand, there was no variation in IGF-I mRNA levels in skeletal muscle. These results suggest that 1) there is a diurnal variation in GHR expression in liver and skeletal muscle and the pattern of the variation is tissue specific; 2) GH deficiency blunted the diurnal variation in GHR mRNA in liver but not that in skeletal muscle; 3) IGF-I mRNA expression in liver is more closely related to GHR mRNA expression than that in skeletal muscle.

Endotoxin at low doses stimulates pituitary GH whereas it decreases IGF-I and IGF-binding protein-3 in rats.

While it is well known that sepsis inhibits serum IGF-I and its gene expression in the liver, the effect on pituitary GH and IGF-binding protein-3 (IGFBP-3) is poorly understood. The GH-IGF-I-IGFBP-3 response to different doses of lipopolysaccharide (LPS) administration has been investigated in adult male rats. Two experiments were performed, administration of low doses of LPS (5, 10, 50 and 100 microg/kg) and high doses of LPS (100, 250, 500 and 1000 microg/kg). Rats received two i.p. injections of LPS (at 1730 h and 0830 h the following day) and were killed 4 h after the second injection. LPS administration induced a biphasic response in serum concentrations of GH, with an increase at the 10 microg/kg dose, followed by a decrease at higher doses (100 microg/kg on up). Pituitary GH mRNA was also increased by the administration of 10 and 50 microg/kg LPS, whereas at higher doses LPS did not modify pituitary GH mRNA. We also analyzed the GH response to LPS in primary pituitary cell cultures. When exposed to LPS, in the culture medium, there was an increase in GH release at the concentration of 0.1 and 10 ng/ml, whereas more concentrated LPS did not modify GH release. Serum concentrations of IGF-I declined in a dose-dependent fashion after LPS administration in the rats injected with 10 microg/kg LPS on up. This decrease is secondary to modifications in its synthesis in the liver, since endotoxin injection decreased both IGF-I and its mRNA in the liver. The liver GH receptor mRNA was also decreased by LPS administration, but only in the animals injected with high LPS doses. There was a decrease in both the IGFBP-3 serum levels and its gene expression in the liver with all LPS doses studied. These data suggest a biphasic LPS effect on pituitary GH, a stimulatory effect at low doses and an inhibitory effect at higher doses, whereas it has a clear inhibitory effect on IGF-I and IGFBP-3 synthesis in the liver. The decrease in liver IGFBP-3 mRNA and in serum concentrations of IGFBP-3 in the rats injected with LPS may contribute to the decrease in serum concentrations of IGF-I.

Reduced Serum Insulin-Like Growth Factor (IGF) I Is Associated with Reduced Liver IGF-I mRNA and Liver Growth Hormone Receptor mRNA in Food-Deprived Cattle

Nutritional deprivation decreases blood insulin-like growth factor I (IGF-I) concentrations in a variety of species. In this study, we explored the underlying mechanism by determining the effects of food deprivation on the levels of total IGF-I mRNA and total growth hormone receptor (GHR) mRNA, as well as the levels of individual IGF-I mRNA variants and GHR mRNA variants in the liver of steers. Food deprivation for nearly 3 d decreased the levels of serum IGF-I by 63% (P < 0.01), and this decrease was associated with a 75% decrease (P < 0.01) in total IGF-I mRNA in the liver. The food deprivation-induced decrease in liver total IGF-I mRNA was associated with an equivalent decrease in the levels of both class 1 and class 2 IGF-I mRNA. In addition to IGF-I mRNA, food deprivation also decreased the levels of total GHR mRNA in the liver (P < 0.05), and this decrease was associated with a decrease in the liver expression of GHR mRNA variants 1C3 (P < 0.05) and 1A (P = 0.08). Food deprivation did not affect the levels of two other major GHR mRNA variants, 1B and 1C2, in the liver. These results demonstrate that the food deprivation-induced decrease in circulating IGF-I in steers is associated with a coordinate decrease in the expression of different IGF-I mRNA variants and a specific decrease in the expression of GHR mRNA variants 1C3 and 1A in the liver.

The GH/IGF-I axis in the brushtail possum (Trichosurus vulpecula) pouch young.

Plasma and pituitary GH concentrations and liver GH receptor (GHR), IGF-I and IGF-binding protein-3 (IGFBP-3) mRNA expression were determined in brushtail possum (Trichosurus vulpecula) pouch young aged 12-150 days post-partum and in adults. Mean plasma GH concentrations were highest, measuring around 150 ng/ml, from 12 to 100 days post-partum, and thereafter declined so that by 150 days post-partum levels were not significantly different from those in adults (10.8+/-1.8 ng/ml (S.E.M.)). In contrast to plasma levels, pituitary GH content increased markedly throughout pouch life, with an 87-fold increase between 12 and 150 days post-partum. However, when expressed per gram body weight, pituitary content was relatively constant between 25 and 150 days post-partum, indicating that the decline in plasma GH after 100 days post-partum was not due to decreased synthesis and/or storage of GH in the pituitary gland. Expression of GHR, IGF-I and IGFBP-3 mRNAs was determined by semi-quantitative RT-PCR. Liver GHR and IGF-I mRNA expression were low at 12 and 25 days post-partum and did not show sustained and significant increases (P<0.05) until 125 and 150 days post-partum. IGFBP-3 expression was also low at 12 days post-partum but then increased rapidly to a maximum at 50 days post-partum and thereafter declined. For all three mRNAs, liver expression at day 150 was not significantly different from that in adults. These patterns of gene expression for GHR and IGF-I suggest that the possum liver is resistant to the high plasma GH concentrations during early pouch life and in this way is similar to the fetal liver of some eutherian mammals.

Effect of recombinant growth hormone on expression of growth hormone receptor, insulin-like growth factor mRNA and serum level of leptin in growing pigs

Sixteen Large White x Landrace castrated male pigs were allotted into treatment and control group. The treatment group was injected intramuscularly with recombinant porcine growth hormone (rpGH, 4 mg d(-1)) and the control group with vehicle for 28 days. Animals were slaughtered 4 h after final injection for liver, longissimus dorsi (LD) muscle and blood sampling. Serum concentration of insulin-like growth factor 1 (IGF-I) and leptin were determined by RIA. The total RNA was extracted from tissues to measure the abundance of growth hormone receptor (GHR), IGF-I mRNA by RT-PCR with 18S rRNA internal standard. Results showed that rpGH enhanced the average daily weight gain by 26.1% (P < 0.05), the serum IGF-I concentration by 70.94% (P < 0.01), decreased serum leptin by 34.8% (P < 0.01). The relative abundance of GHR and IGF-I mRNA in liver were increased by 24.45% (P < 0.05) and 45.30% (P < 0.01), respectively, but no difference of GHR (P > 0.05) and IGF-I mRNA (P > 0.05) in LD between GH treated and control group was found. These results suggest that rpGH can up-regulate hepatic GHR and IGF-I gene expression and improve animal growth. However the effect of rpGH on GHR and IGF-I gene expression are tissue-specific.

Regulation of the growth hormone receptor/binding protein, insulin-like growth factor ternary complex system in human cirrhosis

Background/Aims: The liver is the central organ of the endocrine growth hormone/insulin-like growth factor I (GH/IGF-I) axis and cirrhosis effects a state of acquired GH resistance. Low IGF-I levels are associated with adverse clinical outcomes in cirrhotic patients and may be pathogenic to the complications of cirrhosis. We examined the impact of cirrhosis on hepatic mRNA and serum protein levels for the GH receptor (GHR)/binding protein (GHBP), IGF-I, IGF binding protein (IGFBP)-3 and the acid-labile subunit (ALS). Methods: Fifty patients with cirrhosis were studied and liver tissue was obtained from 18. Gene expression was assessed by Northern analysis and serum protein levels by immunoassay. Results: In cirrhotic liver GHR mRNA and GH binding to microsomal membranes were decreased by 61 and 56%, respectively. Serum GHBP levels were decreased only in severe disease, not correlating with GHR mRNA or GH binding. Hepatic IGF-I and ALS mRNA were significantly decreased by 84 and 68%, respectively, in parallel with serum protein, suggesting transcriptional regulation. Hepatic IGFBP-3 mRNA was unchanged but low serum IGFBP-3 suggested post-transcriptional regulation. Conclusions: The decreased mRNA and serum levels for the GH-dependent, hepatocyte produced proteins IGF-I and ALS confirm the importance of GH receptor loss to the GH resistance of cirrhosis.

Expression of two variants of growth hormone receptor messenger ribonucleic acid in porcine liver.

Transcription of GH receptor (GHR) mRNA is initiated from multiple promoters. Most GHR mRNA arise from GHR Promoter 1 (GHR P1) and GHR P2, which transcribe GHR 1A and GHR 1B mRNA, respectively. Our objective was to characterize the expression of GHR 1A and GHR 1B mRNA in liver of neonatal intact (1 d of age) and castrated (14, 28, and 42 d of age) male pigs (Exp. 1; n = 6 per age group), intact male pigs treated with recombinant porcine ST (rpST) or control (Exp. 2; 21, 42, and 77 d of age; n = 4 pigs per treatment per age), and pregnant gilts treated with rpST (n = 6) or control (n = 7) (Exp. 3). Tissue samples were collected at slaughter for mRNA analyses. The porcine GHR 1A and GHR 1B cDNA were cloned and were homologous to GHR cDNA isolated from other species. Ribonuclease protection assays were used to measure GHR 1A and GHR 1B mRNA. Liver expressed GHR 1A and GHR 1B mRNA, whereas muscle, uterus, and ovary expressed GHR 1B mRNA. The GHR 1A mRNA in the liver of neonatal intact and castrated male pigs (Exp. 1) was expressed at very low levels on d 1, 14, and 28, and two of six pigs expressed a high level of GHR 1A on d 42. The GHR 1B mRNA, however, was detected at all ages (d 1 through 42), and the amount of GHR 1B increased (P<.05) on d 42. The liver of intact male pigs (Exp. 2) expressed GHR 1B mRNA by 21 d, whereas a high level of GHR 1A mRNA was not detected until d 42 (P<.10). Administration of rpST had no effect on expression of GHR 1A or GHR 1B mRNA in pigs younger than 77 d (Exp. 2), but it tended to decrease (P<.10) GHR 1A mRNA but not GHR 1B mRNA in pregnant gilts (Exp. 3). In conclusion, GHR mRNA in porcine liver was composed of at least two variants (GHR 1A and GHR 1B). The GHR 1B mRNA was the major GHR mRNA in pig liver before 77 d of age. The GHR 1A mRNA increased after 42 d of age and tended to undergo specific down-regulation in response to rpST in pregnant gilts.

Expression of growth hormone receptor 1A messenger ribonucleic acid in liver of dairy cows during lactation and after administration of recombinant bovine somatotropin.

The mRNA for growth hormone receptor is transcribed from at least three different promoters in cattle. The first promoter (P1) is liver-specific and transcribes growth hormone receptor mRNA containing exon 1A (growth hormone receptor 1A). The second and third promoters (P2 and P3) are active in a variety of tissues and transcribe growth hormone receptor mRNA containing exon 1B and 1C. The objective was to characterize P1 activity by measuring the amount of growth hormone receptor 1A mRNA in liver of dairy cows at different stages of lactation as well as after administration of recombinant bovine somatotropin (rbST). In study 1, liver RNA was isolated from Holstein cows during the dry period (nonlactating, n = 6) and during early (n = 6), mid (n = 6), and late (n = 11) stages of lactation. Six of the late-lactation cows received injections of rbST (25 mg/d) for 7 d prior to collection of liver tissue. In study 2, lactating Holstein cows received either no infusion (control, n = 10) or continuous infusion of rbST (29 mg/d, n = 10) for 63 d. The amount of growth hormone receptor 1A mRNA was decreased in early- and mid-lactation cows compared with late-lactation cows or nonlactating cows (study 1). Administration of rbST increased growth hormone receptor 1A mRNA (studies 1 and 2). The total amount of growth hormone receptor transcribed from alternative promoters (growth hormone receptor P2 and P3) remained unchanged during different stages of lactation or in response to rbST. We conclude that changes in liver growth hormone receptor mRNA in lactating dairy cattle primarily depend on growth hormone receptor P1 activity.

Interactions between environmental temperature and porcine growth hormone (pGH) treatment in neonatal pigs

Age-dependent interactions between environmental temperature and porcine growth hormone (pGH) treatment on the function of the somatotrophic axis were evaluated in the neonatal pig. At 3 d of age, 40 Landrace x Yorkshire x Duroc piglets received intraperitoneal implants containing either recombinant pGH (0.5 mg/d; n = 20) or vehicle (control; n = 20). Piglets were maintained at either a low (21°C, 50% relative humidity; n = 20) or high (32°C, 50% relative humidity; n = 20) temperature. At 4 and 6 wk of age, 5 pGH-treated and 5 control piglets from each thermal group were sacrificed for tissue collection. Blood samples were collected at the time of sacrifice and analyzed for serum concentrations of GH, insulin-like growth factor 1 (IGF-1), and IGF-2. Liver RNA was analyzed for mRNAs specific for the GH receptor, IGF-1, IGF-2, and IGF binding protein 3. There was no effect of pGH treatment (P = 0.4) on average daily gain; however, both age (P = 0.002) and temperature (P = 0.001) had an effect on average daily gain such that those animals maintained at a low temperature and those sacrificed at 6 wk had greater average daily gains. Serum concentration of GH was elevated (P = 0.003) by pGH treatment and was lowest in the 6-wk-old group (P = 0.008). Serum concentration of IGF-1 was elevated (P = 0.007) by pGH treatment and increased with age (P = 0.01). Liver GH receptor mRNA was unaffected (P > 0.5) by pGH treatment, but was greater in the 6-wk-old group (P < 0.0001) and in piglets maintained at the high temperature (P = 0.04). IGF-1 mRNA was enhanced by pGH treatment (P = 0.0003) and by exposure to the high temperature (P = 0.04), but did not differ (P > 0.5) between age groups. IGF-2 mRNA was greater (P = 0.0009) in the 4-wk-old group and in piglets maintained at the high temperature (P = 0.007), but was unaffected (P = 0.5) by pGH treatment. IGF binding protein 3 mRNA increased with age (P = 0.0004) and was stimulated by pGH treatment in the 6-wk-old group (P = 0.034). The relatively lower level of GH receptor and IGF mRNAs in conjunction with greater growth in the cold environment suggests that somatotrophic gene expression in the liver is not rate limiting for growth in the neonatal pig.

Regulation of growth hormone (GH) gene expression and secretion during pregnancy and lactation in the rat: role of insulin-like growth factor-I, somatostatin, and GH-releasing hormone.

GH appears to play an important metabolic role during late pregnancy and in lactation maintenance. In this study, pregnant (days 8, 15, and 20 of gestation) and postpartum (days 3 and 8 postpartum, including lactating and nonlactating dams) Wistar rats were used to investigate pituitary GH gene expression and hormone secretion, and the potential alterations of the major signals regulating GH secretion and action [somatostatin (SS) and GH-releasing hormone (GHRH), GH receptor (GH-R), and insulin-like growth factor-I (IGF-I)]. GH and SS messenger RNA (mRNA) were quantitated by Northern blot, and both IGF-I and GH-R mRNA were analyzed by the ribonuclease protection assay technique. Pituitary IR-GH content and GH mRNA increased at midpregnancy. IR-GH content was decreased in lactating rats. Plasma GH levels progressively increased during pregnancy, whereas no significant alterations were shown during lactation. Elevated GH levels persisted during lactation. Levels at this time were higher in nonsuckling compared with suckling dams. Liver GH-R mRNA progressively decreased during pregnancy, but it remained unchanged during lactation. Plasma IGF-I and liver IR-IGF-I constantly decreased during pregnancy, and no significant modifications were seen either in suckling or in nonsuckling animals. IGF-I mRNA accumulation in the liver decreased during pregnancy. After delivery, a progressive decrease of liver IGF-I mRNA occurred. At the hypothalamic level, a progressive increase in the IR-SS content was found during pregnancy, with no SS mRNA modification. After delivery, a higher hypothalamic IR-SS content was found in lactating than in nonlactating rats, with no changes in SS mRNA levels. Hypothalamic IR-IGF-I also showed a progressive increase during pregnancy with no significant alterations during lactation. Hypothalamic IR-GHRH presented a nonsignificant mild increase during pregnancy with no modifications during lactation. In the pituitary, IR-IGF-I content progressively increased during gestation, reaching its highest concentration at day 20. During lactation, pituitary IGF-I did not change. In summary, our data show that the mechanisms of the increase in plasma GH levels occurring during pregnancy include an increase in GH gene expression in the pituitary, a decrease in SS secretion from the hypothalamus, an increase in IR-IGF-I content in the hypothalamus and in the pituitary, and a significant decrease in circulating IGF-I. Plasma and liver IR-IGF-I and IGF-I mRNA in the liver decreased throughout gestation due to a lower GH-R gene expression in the liver. This state of GH resistance with a higher GH/IGF-I ratio could be important in providing supplementary nutrients to the fetus. During lactation, GH and its regulatory machinery did not show important modifications.

Developmental changes and differential regulation by testosterone and estradiol of growth hormone receptor expression in the rabbit.

To investigate the effects of testosterone and estradiol (E2) on growth hormone receptor (GH-R) gene expression, we measured GH-R mRNA levels in relation to the changes of sex steroid concentrations in the normal male rabbits aged 1-12 months and after administration of testosterone or E2 to castrated male rabbits. In the normal animals, E2 levels were below the detection limit in all age groups, and testosterone levels were below the detection limit at 1 month, increased at 2 months and reached the plateau of the adult levels after 4 months. Liver GH-R mRNA levels were low at 1 month, reached a peak at 2 months and then decreased slightly thereafter. In the castrated animals, liver and growth plate GH-R mRNA levels were increased in the testosterone-treated group (162.0 +/- 12.0%, p < 0.025; 128.4 +/- 7.6%; p < 0.025) and reduced in the E2-treated group (29.6 +/- 6.2%, p < 0.005; 53.6 +/- 11.3%, p < 0.025). Sex steroid administration did not result in any significant change in GH-R mRNA levels in striated muscle, kidney and heart. Serum GH concentrations were increased in E2 (15.3 +/- 7.7 microg/l vs 4.8 +/- 2.2 microg/l, p < 0.025) but the increase was not significant in testosterone-treated animals (8.4 +/- 7.7 microg/l vs 4.8 +/- 2.2 microg/l). Both testosterone and E2 treatment resulted in a reduction of mean serum growth hormone-binding protein (GHBP) levels compared to control animals (1077 +/- 422 pmol/l, p < 0.01; 1137 +/- 443 pmol/l, p < 0.01; 2308 +/- 565 pmol/l). We conclude that in addition to their stimulatory effect on GH secretion, testosterone and E2 have opposite effects on GH-R gene expression in liver and growth plate in the rabbit. The modulation of GH-R expression by sex steroids may be important for growth during sexual maturation in mammals.

Ontogeny of GH receptor and GH-binding protein in the pig.

The present study was undertaken to examine the developmental pattern of GH receptor (GHR) and GHR gene expression in skeletal muscle (longissimus dorsi and trapezius (TR)) and liver from the last third of gestation until 1 year of age in male Large White (LW) and Meishan (MS) pigs. Plasma GH-binding protein (GHBP) levels were also measured. 125I-Labelled bovine GH (bGH) specific binding (not determined in foetal TR) and GHR mRNA were detected in skeletal muscle from 75 days of gestation until the adult stage with no clear age-related changes. By contrast, 125I-labelled bGH specific binding and GHR mRNA were undetectable or barely detectable in foetal liver. After birth, 125I-labelled bGH specific binding (P < 0.001) and GHR mRNA in liver increased with age. The level of bGH binding to liver membranes was higher in MS than in LW pigs at 1, 45, 80 and 120 days of age and did not differ between breeds at the other ages. Specific binding of 125I-labelled human GH (hGH) to plasma GHBP was easily detected as early as 75 days of gestation and increased with age (P < 0.001). The level of hGH binding to plasma GHBP was higher in MS than in LW pigs at 1, 80 and 120 days of age. It can be concluded that (1) the developmental expression of the GHR is tissue-specific, (2) the presence of GHBP in foetuses despite the absence of GHR in liver suggests that other tissues such as skeletal muscle could contribute to the generation of GHBP and (3) the presence of GHR in skeletal muscle as early as 75 days of gestation suggests that GH may play a role in foetal muscle growth.

Effect of fasting and growth hormone (GH) administration on gh receptor (GHR) messenger ribonucleic acid (mRNA) and GH-binding protein (GHBP) mRNA levels in male rats

To elucidate whether GHR and GHBP are coordinately regulated or not, we studied the effect of fasting with or without GH administration on the GHR and GHBP mRNAs in the liver as well as in extrahepatic tissues in rats. Tissues were collected from 7-week-old male rats by decapitation 1, 3, and 7 days after the start of fasting. Liver GHR mRNA levels were not affected 1 day after the start of fasting but progressively decreased for the subsequent 3 and 7 days of fasting as compared with those in control rats fed ad libitum. In contrast, liver GHBP mRNA levels significantly rose after 1 day fasting, returned to the control level after 3 days and further reduced after 7 days of fasting. Changes in GHBP mRNA level after fasting were different among the tissues. A transient increase in GHBP mRNA levels was observed in muscle and heart as well as liver, while the GHBP mRNA levels in fat tissues did not change throughout 7 days of fasting. Next, bovine GH(bGH) was administered ip to the fasted rats and control fed rats for either 1 day(100mg, tid) or 5 days(150mg, daily). In fed rats, liver GHR mRNA level was significantly increased by 1 day bGH treatment, but after 5 days treatment with bGH it was not different from the level in saline-injected control. Accordingly, net increment of plasma IGF-I was 296.0 ng/ml with 1 day bGH treatment and 234.2 rig/ml with bGH administration for 5 days. In fasted rats, liver GHR mRNA level did not changed after 1 day treatment with bGH, but markedly decreased 5 days after bGH administration. Net increment of plasma IGF-I was slightly reduced to 284 ng/ml with 1 day treatment with bGH, and markedly decreased to 37.0 with bGH administration for 5 days. The effect of GH administration on liver GHBP mRNA level was virtually absent in either fasting or fed state. These findings suggest that GHR and GHBP mRNAs in the liver are expressed in different ways and that expression of GHBP mRNA is differently regulated among tissues.