Growth Hormone Control of Hepatic Lipid Metabolism

The anabolic actions of GH are mediated by the production of insulin-like growth factor I (IGF-I) from the liver and by local production of IGF-I in extrahepatic tissues. Insulin facilitates the hepatic production of IGF-I by up-regulating GH receptors (GHRs) in the liver and augmenting the IGF-I response to GH. Although GHRs have also been identified in extrahepatic tissues that produce IGF-I, the possibility that IGF-I and insulin might partake in GHR regulation, thereby modulating the effects of GH locally has not received detailed study. The aim of this study was to investigate whether IGF-I and insulin are involved in the local regulation of GHRs, using osteoblasts as a model of GH-responsive extrahepatic tissues. We have used UMR106.06, a well differentiated rat osteoblast-like cell line that expresses GHRs and exhibits a mitogenic response to GH. IGF-I and insulin (0-10 nM) increased cell number and reduced [125I]GH binding in a concentration-dependent manner, with ED50 values of 0.8 and 0.3 nM, respectively. Although IGF-I increased cell number maximally by 36.9 +/- 1.2% (mean +/- SE) above the control value and insulin by 104.8 +/- 5.7% (P < 0.001), they decreased GH binding to 47.0 +/- 9.3% (P < 0.01) and 29.8 +/- 8.7% of the control value (P < 0.001), respectively. Scatchard analysis revealed that the down-regulation of GH binding was attributed to reduced receptor numbers and not binding affinity. The effects of IGF-I and insulin at submaximal concentrations were additive, although the combined effects did not exceed the maximal effect of either growth factor alone. Addition of an anti-IGF-I receptor antibody (alpha IR3) reversed the inhibition of GH binding induced by IGF-I, but not that caused by insulin; similarly, an antiinsulin receptor antibody (29B4) attenuated the inhibitory effect of insulin only. Addition of alpha IR3 alone or an ant-IGF-I antibody (Sm1.2) decreased cell number and increased GH binding in a concentration-dependent mode. GH at 1.5 nM significant increased cell number by 19.3 +/- 2.4% above the control level (P < 0.01), an increase that was reversed by alpha IR3. GH increased GH binding by 32.4 +/- 7.2% (P < 0.05) in cells treated with alpha IR3 to remove the secondary effect of IGF-I. In summary, IGF-I and insulin acted via specific receptors to stimulate cell proliferation and down-regulate GHRs in osteoblasts. GH stimulated cell proliferation, an action mediated by local production of IGF-I, and GH enhanced its own binding. The collective data suggest the presence of a peripheral negative feedback loop that allows IGF-I to limit locally the response of extrahepatic tissues to circulating GH.

A genomic deletion of exon 3 (d3-GHR) of the growth hormone (GH) receptor (GHR) has been linked to the effectiveness of GH therapy in children with GH deficiency. Carriers of the d3-GHR genotype had higher GH-induced growth rates than children homozygous for the full-length (fl)-GHR. The aim of this study was to test whether the relationship between GH and insulin-like growth factor-1 (IGF-1) concentrations is influenced by the GHR genotype in patients with acromegaly. Study participants were 44 adult patients with established diagnosis of acromegaly. The genotype of the GHR was determined in leukocyte DNA from peripheral blood. Clinical and biochemical findings at the time of diagnosis of acromegaly were obtained from the medical records of the patients. fl-GHR homozygosity was found in 22 (50%) of patients, and 22 (50%) of patients had at least 1 d3 allele (d3-GHR). Demographic and clinical characteristics (age, height, weight, estimated duration of disease, and mean tumor size) of the 2 groups were comparable. Median (range) serum IGF-1 concentrations at the time of diagnosis were 670 (447-1443) microg/L in the fl-GHR group and 840 (342-1494) microg/L in the d3-GHR group (P = not significant). Basal GH concentrations were higher in the fl-GHR group [29.7 (3.8-159) microg/L] than in the d3-GHR group [8.4 (2.6-74 microg/L), P = 0.002], and so were mean (30.4 vs 6.1 microg/L, P = 0.005) and nadir (20.5 vs 5.1 microg/L, P = 0.003) GH concentrations during an oral glucose tolerance test. The GHR fl/d3 genotype modulates the relationship between GH and IGF-1 concentrations in patients presenting with acromegaly.

The growth hormone (GH) receptor (GHR) binds GH in its extracellular domain and transduces activating signals via its cytoplasmic domain. Both GH-induced GHR dimerization and JAK2 tyrosine kinase activation are critical in initiation of GH signaling. We previously described a rapid GH-induced disulfide linkage of GHRs in human IM-9 cells. In this study, three GH-induced phenomena (GHR dimerization, GHR disulfide linkage, and enhanced GHR-JAK2 association) were examined biochemically and immunologically. By using the GH antagonist, G120K, and an antibody recognizing a dimerization-sensitive GHR epitope, we demonstrated that GH-induced GHR disulfide linkage reflects GH-induced GHR dimerization. GH, not G120K, promoted both GHR disulfide linkage and enhanced association with JAK2. Measures that diminished GH-dependent JAK2 and GHR tyrosine phosphorylation diminished neither GH-induced GHR disulfide linkage nor GH-enhanced GHR-JAK2 association. By using both transient and stable expression systems, we determined that cysteine 241 (an unpaired extracellular cysteine) was critical for GH-induced GHR disulfide linkage; however, GH-induced GHR dimerization, GHR-JAK2 interaction, and GHR, JAK2, and STAT5 tyrosine phosphorylation still proceeded when this cysteine residue was mutated. We conclude GH-induced GHR disulfide linkage is not required for GHR dimerization, and activation and GH-enhanced GHR-JAK2 association depends more on GHR dimerization than on GHR and/or JAK2 tyrosine phosphorylation.

The cytokine-inducible SH2 domain-containing protein CIS inhibits signaling from the growth hormone (GH) receptor (GHR) to STAT5b by a proteasome-dependent mechanism. Here, we used the GH-responsive rat liver cell line CWSV-1 to investigate the role of CIS and the proteasome in GH-induced GHR internalization. Cell-surface GHR localization and internalization were monitored in GH-stimulated cells by confocal immunofluorescence microscopy using an antibody directed against the GHR extracellular domain. In GH naïve cells, GHR was detected in small, randomly distributed granules on the cell surface and in the cytoplasm, with accumulation in the perinuclear area. GH treatment induced a rapid (within 5 min) internalization of GH.GHR complexes, which coincided with the onset of GHR tyrosine phosphorylation and the appearance in the cytosol of distinct granular structures containing internalized GH. GHR signaling to STAT5b continued for approximately 30-40 min, however, indicating that GHR signaling and deactivation of the GH.GHR complex both proceed from an intracellular compartment. The internalization of GH and GHR was inhibited by CIS-R107K, a dominant-negative SH2 domain mutant of CIS, and by the proteasome inhibitors MG132 and epoxomicin, which prolong GHR signaling to STAT5b. GH pulse-chase studies established that the internalized GH.GHR complexes did not recycle back to the cell surface in significant amounts under these conditions. Given the established specificity of CIS-R107K for blocking the GHR signaling inhibitory actions of CIS, but not those of other SOCS/CIS family members, these findings implicate CIS and the proteasome in the control of GHR internalization following receptor activation and suggest that CIS-dependent receptor internalization is a prerequisite for efficient termination of GHR signaling.

GH stimulates tyrosyl phosphorylation of GH receptors in 3T3-F442A fibroblasts, and highly purified GH receptor preparations exhibit tyrosine kinase activity. Paradoxically, however, the GH receptor cloned from liver exhibits no sequence similarity to receptors with known signal transduction mechanisms, including those exhibiting ligand-activated tyrosine kinase activity. These observations raise the possibility that there are two kinds of receptors for GH: the first represented by the cloned liver GH receptor, and the second by a tyrosine kinase-containing GH receptor. To inquire into the possibility of two distinct GH receptors, we determined whether the cloned liver GH receptor shares structural similarities with the tyrosine kinase-associated GH receptor. When the cloned rabbit liver GH receptor is expressed in human kidney 293 cells, it migrates with a mol wt appropriate for the tyrosine kinase-associated GH receptor, despite the calculated mol wt of the cloned GH receptor being 60,000 smaller than that of the tyrosine kinase-associated GH receptor. The recognition of tyrosine kinase-associated GH receptor by antipeptide antibodies to three different epitopes on the cytoplasmic domain of the cloned liver GH receptor was also tested. Tyrosyl phosphorylated [125I]human GH-receptor complexes were prepared by immunoprecipitation with phosphotyrosyl-binding antibody; this subpopulation of GH-receptor complexes was recognized by all three antipeptide antibodies. The antibodies also recognized similarly isolated tyrosyl phosphorylated GH-receptor complexes, which had been further phosphorylated in solution on tyrosyl residues upon addition of [gamma 32P] ATP. Furthermore, highly purified GH receptors prepared by sequential immunoprecipitation using phosphotyrosyl-binding antibody and any one of the three antipeptide antibodies incorporated 32P into tyrosyl residues upon the addition of [gamma 32P] ATP. These results provide evidence that tyrosine kinase-associated GH receptors share sequence similarity in the cytoplasmic domain with the cloned liver GH receptor. The cloned GH receptor and the tyrosine kinase-associated GH receptor, therefore, are likely to be the same receptor or related receptor isoforms.

The presence of growth hormone (GH) and GH receptors (GHRs) in the lung suggests it is an autocrine/paracrine target site for pulmonary GH action and/or an endocrine site of pituitary GH action. Roles for GH in lung growth or pulmonary function are, however, uncertain. The possibility that pituitary and/or pulmonary GH have physiological roles in lung development has therefore been investigated in GHR knockout (KO or -/-) mice, using a proteomics approach to determine if an absence of GH-signaling affects the proteome of the developing lung. More than 600 proteins were detected by 2-DE in the lungs of control [GHR (+/+)] and GHR (-/-) mice at the end of the alveolarization period (at day 14 postnatally). Of these, 39 differed significantly in protein content at the p>0.05 level [6 were of higher abundance in the GHR (-/-) group, 33 were of lower abundance] and 17 differed at the p>0.02 level [5 of higher abundance in the GHR (-/-) group, 12 of lower abundance] and 7 were definitively identified by MS. Vimentin, a protein involved in cellular proliferation, was reduced in content by approximately 75% in the lungs of the GHR (-/-) mice. Three proteins involved in oxidative protection [SH3 domain-binding glutamic acid-rich-like protein, peroxiredoxin 6 (Prdx6), and isocitrate dehydrogenase 1] were also of lower content in the GHR (-/-) lungs (by approximately 88%, 81% and 70%, respectively). Prdx6 is also involved in lipid and surfactant metabolism, as is apolipoprotein A-IV, the lung content of which was reduced by approximately 73% in these mice. Proteasome 26S ATPase subunit 4, a protein involved in the non-lysosomal degradation of intracellular proteins, and electron flavoprotein alpha subunit , involved in intracellular metabolism, were also reduced in content in the lungs of the GHR (-/-) mice (by approximately 70% and 49%, respectively). These results therefore suggest that these proteins are normally dependent upon GH signaling, and that GH is normally involved in early lung growth, oxidative protection, lipid and energy metabolism and in proteasomal activity. These roles may reflect endocrine actions of pituitary GH and/or local autocrine/paracrine actions of GH produced within the lung.

Mutations in the growth hormone receptor (GHR) gene can cause growth hormone (GH) resistance. Given the sequence homology between the extracellular domain of the GHR and a soluble GH-binding protein (GH-BP), it is remarkable that GH-BP binding activity is absent from the serum of patients with Laron-type GH insensitivity, a hereditary form of severe dwarfism. We have previously identified a mutation within the extracellular domain of this receptor, replacing phenylalanine by serine at position 96 of the mature protein, in a patient with Laron syndrome. We have now investigated the effect of this Phe----Ser substitution on hormone binding activity by expressing the total human GHR cDNA and mutant form in eukaryotic cells. The wild-type protein expressed was able to bind GH but no plasma membrane binding was detectable on cells transfected with the mutant cDNA; this was also the case of cells transfected with a Phe96----Ala mutant cDNA, suggesting that the lack of binding activity is not due to a posttranslational modification of serine. Examination of the variant proteins in subcellular fractions revealed the presence of specific GH binding activity in the lysosomal fraction, whereas immunofluorescence studies located mutant proteins in the cytosol. Our findings suggest that these mutant GHRs fail to follow the correct intracellular transport pathway and underline the potential importance of this phenylalanine residue, which is conserved among the GH, prolactin, and erythropoietin receptors that belong to the same cytokine receptor superfamily.

Growth hormone receptor (GHR) is a cytokine receptor superfamily member that binds growth hormone (GH) via its extracellular domain and signals via interaction of its cytoplasmic domain with JAK2 and other signaling molecules. GHR is a target for inducible metalloprotease-mediated cleavage in its perimembranous extracellular domain, a process that liberates the extracellular domain as the soluble GH-binding protein and leaves behind a cell-associated GHR remnant protein containing the transmembrane and cytoplasmic domains. GHR metalloproteolysis can be catalyzed by tumor necrosis factor-alpha-converting enzyme (ADAM-17) and is associated with down-modulation of GH signaling. We now study the fate of the GHR remnant protein. By anti-GHR cytoplasmic domain immunoblotting, we observed that the remnant induced in response to phorbol ester or platelet-derived growth factor has a reliable pattern of appearance and disappearance in both mouse preadipocytes endogenously expressing GHR and transfected fibroblasts expressing rabbit GHR. Lactacystin, a specific proteasome inhibitor, did not appreciably change the time course of remnant appearance or clearance but allowed detection of the GHR stub, a receptor fragment slightly smaller than the remnant but containing the C terminus of the remnant (receptor cytoplasmic domain). In contrast, MG132, another (less specific) proteasome inhibitor, strongly inhibited remnant clearance and prevented stub appearance. Inhibitors of gamma-secretase, an aspartyl protease, also prevented the appearance of the stub, even in the presence of lactacystin, and concomitantly inhibited remnant clearance in the same fashion as MG132. In addition, mouse embryonic fibroblasts derived from presenilin 1 and 2 (PS1/2) knockouts recapitulated the gamma-secretase inhibitor studies, as compared with their littermate controls (PS1/2 wild type). Confocal microscopy indicated that the GHR cytoplasmic domain became localized to the nucleus in a fashion dependent on PS1/2 activity. These data indicate that the GHR is subject to sequential proteolysis by metalloprotease and gamma-secretase activities and may suggest GH-independent roles for the GHR.

A 2 x 2 factorial experimental design was used to evaluate effects of ovariectomy and implantation (200 mg trenbolone acetate and 28 mg estradiol benzoate; Synovex-Plus) on the performance, serum urea nitrogen, serum insulin like growth factor -1 (IGF-1), and mRNA expression of hepatic IGF-1, growth hormone (GH) receptor and estrogen receptor-α as well as pituitary GH, estrogen receptor-α and GH releasing hormone receptor in feedlot heifers. Thirty-two British x Continental heifers were randomly assigned to either an ovariectomized (OVX) or intact group and then to either a control group or a Synovex-Plus implant group. Heifers were fed a 900 g/kg concentrate steam-flaked maize based diet for 42 d. Liver biopsies were taken before OVX for baseline expression of IGF-1, GH receptor and hepatic estrogen receptor-α, which did not differ between treatment groups. Blood and body weight (BW) were taken on d 0, 28, and 42 and one heifer/pen was slaughtered on d 42 for liver and pituitary tissue samples. Initial and final BW did not differ due to OVX or implant. No gender x treatment interaction was observed for average daily gain (ADG) or final BW. Neither OVX nor implant affected ADG for the final 14 d of the feeding period, but 28 and 42 d ADG were greater in implanted than in control heifers. No 2- or 3-way interactions with day were observed for serum urea nitrogen. Serum urea nitrogen was greater in control heifers than in implanted heifers, but gender had no effect. Both implant and OVX increased serum IGF-1 and a gender x treatment interaction was observed. Neither gender nor implant treatment affected hepatic estrogen receptor-α, hepatic IGF-1, hepatic GH receptor, GH releasing hormone receptor, GH, or pituitary estrogen receptor-α. Data indicate that implanting heifers with Synovex-plus increases serum IGF-1, but the reason for this increase cannot be explained by mRNA expression of key somatotropic genes in the present study.A 2 x 2 factorial experimental design was used to evaluate effects of ovariectomy and implantation (200 mg trenbolone acetate and 28 mg estradiol benzoate; Synovex-Plus) on the performance, serum urea nitrogen, serum insulin like growth factor -1 (IGF-1), and mRNA expression of hepatic IGF-1, growth hormone (GH) receptor and estrogen receptor-α as well as pituitary GH, estrogen receptor-α and GH releasing hormone receptor in feedlot heifers. Thirty-two British x Continental heifers were randomly assigned to either an ovariectomized (OVX) or intact group and then to either a control group or a Synovex-Plus implant group. Heifers were fed a 900 g/kg concentrate steam-flaked maize based diet for 42 d. Liver biopsies were taken before OVX for baseline expression of IGF-1, GH receptor and hepatic estrogen receptor-α, which did not differ between treatment groups. Blood and body weight (BW) were taken on d 0, 28, and 42 and one heifer/pen was slaughtered on d 42 for liver and pituitary tissue samples. Initial and final BW did not differ due to OVX or implant. No gender x treatment interaction was observed for average daily gain (ADG) or final BW. Neither OVX nor implant affected ADG for the final 14 d of the feeding period, but 28 and 42 d ADG were greater in implanted than in control heifers. No 2- or 3-way interactions with day were observed for serum urea nitrogen. Serum urea nitrogen was greater in control heifers than in implanted heifers, but gender had no effect. Both implant and OVX increased serum IGF-1 and a gender x treatment interaction was observed. Neither gender nor implant treatment affected hepatic estrogen receptor-α, hepatic IGF-1, hepatic GH receptor, GH releasing hormone receptor, GH, or pituitary estrogen receptor-α. Data indicate that implanting heifers with Synovex-plus increases serum IGF-1, but the reason for this increase cannot be explained by mRNA expression of key somatotropic genes in the present study. Keywords : Synovex-plus, mRNA, gender South African Journal of Animal Science Vol. 38 (3) 2008: pp. 207-216

Although growth hormone (GH) receptors (GHRs) in many species bind human (h) GH as well as their own GH, the hGHR only binds primate GH. Arg43 in hGHR interacts with Asp171 of hGH. Nonprimates have a His in the position equivalent to residue 171 of primate GH and a Leu in position 43 of primate GHR. To determine whether Arg43 accounts for the species specificity of the hGHR, point mutations that changed Leu43 to Arg were introduced into the cDNAs encoding the bovine (b) GHR or the rat GH binding protein (GHBP) and these mutants or their wild-type (WT) counterparts were expressed in mouse L cells. Binding of hGH or bGH to transfected cells or to GHBP secreted into the incubation medium was assessed by displacement of 125I-labeled hGH. WT and mutant bGHR bound hGH with similar affinity, but the affinity of the mutant receptors for bGH was reduced 200-fold. Likewise, WT and mutant GHBP bound hGH with equal affinity, but only WT GHBP bound bGH. Cross-linking of 125I-labeled hGH to WT or mutant GHR produced a 141-kDa labeled complex whose appearance was blocked by unlabeled hGH, but bGH blocked cross-linking only to WT receptors. Both hGH and bGH stimulated tyrosine phosphorylation of a 95-kDa protein in cells transfected with WT GHR, but bGH was less effective in cells expressing mutant GHR. We conclude that incompatibility of Arg43 in the hGHR with His171 in nonprimate GH is the major determinant of species specificity.

Sex-linked dwarfism (SLD) in chickens is characterized by impaired growth despite normal or supranormal plasma growth hormone (GH) levels. This resistance to GH action is thought to be due to mutations of the GH receptor (GHR) gene that reduce or prevent GH binding to target sites. The genetic lesion causing GH resistance in Cornell SLD chickens is, however, not known. Previous studies have shown that hepatic GH-binding activity is abnormally low in these birds, yet the GHR gene is transcribed into a transcript of appropriate size and abundance. Point mutations or defects in translation could therefore account for the impaired GHR activity in this strain. These possibilities were addressed in the present study. A missense mutation resulting in the substitution of serine for the conserved phenylalanine was identified in the region of the GHR cDNA encoding the extracellular domain. Translation of this mutant transcript was indicated by the presence of GHR/GH-binding protein (GHBP)-immunoreactive proteins in liver (55, 70 and 100 kDa) and serum (70 kDa) of normal (K) and SLD birds. Radiolabelled GH did not, however, bind to the hepatic membranes of most SLD chickens. Serum GH-binding activity, in contrast, was readily detectable, although at significantly lower levels than in normal birds. The missense mutation in the SLD GHR gene may thus affect targeting of GHRs to hepatic plasma membranes.

Background/Aim: The exon 3-deleted/full-length (d3/fl) growth hormone (GH) receptor (GHR) polymorphism has been associated with responsiveness to GH therapy in some diagnostic groups. However, there are still controversies on this issue. To evaluate the effect of the GHR exon 3 polymorphism on growth after 1 and 2 years of GH therapy in Turkish patients with GH deficiency (GHD) and Turner’s syndrome (TS) and the distribution of GHR exon 3 isoforms. Materials and Methods: 218 patients with GHD (125 males/93 females) and 43 patients with TS were included in the study. The control group included 477 healthy adults aged from 18 to 57 years (54 females/423 males). Anthropometric parameters and insulin-like growth factor (IGF)-1 and IGF binding protein (IGFBP)-3 were evaluated annually. GHR isoforms were studied using simple multiplex PCR. Height and body mass index were expressed as standard deviation score (SDS). Results: There were no differences among TS, GHD and healthy adults regarding the distribution of GHR exon 3 isoforms (fl/fl, fl/d3 and d3/d3). There was a significant increase in height SDS in both diagnostic groups on GH therapy; however, there were neither differences in height SDS and Δheight velocity between fl/fl, fl/d3 and d3/d3 groups nor a correlation between the distribution of GHR exon 3 isoforms and change in IGF-1 SDS and IGFBP-3 SDS levels on GH therapy in either of the diagnostic groups. There was also no gender difference in GHR isoforms in healthy adults. Conclusion: The results suggest that responsiveness to GH therapy does not depend on the exon 3 GHR genotypes in GHD and TS patients.

Growth hormone (GH) receptor (GHR) is abundantly expressed in neurons that co-release the agouti-related protein (AgRP) and neuropeptide Y (NPY) in the arcuate nucleus of the hypothalamus (ARH). Since ARHAgRP/NPY neurons regulate several hypothalamic-pituitary-endocrine axes, this neuronal population possibly modulates GH secretion via a negative feedback loop, particularly during food restriction, when ARHAgRP/NPY neurons are highly active. The present study aims to determine the importance of GHR signaling in ARHAgRP/NPY neurons on the pattern of GH secretion in fed and food-deprived male mice. Additionally, we compared the effect of two distinct situations of food deprivation: 16 h of fasting or four days of food restriction (40% of usual food intake). Overnight fasting strongly suppressed both basal and pulsatile GH secretion. Animals lacking GHR in ARHAgRP/NPY neurons (AgRP∆GHR mice) did not exhibit differences in GH secretion either in the fed or fasted state, compared to control mice. In contrast, four days of food restriction increased GH pulse frequency, basal GH secretion, and pulse irregularity/complexity (measured by sample entropy), whereas pulsatile GH secretion was not affected in both control and AgRP∆GHR mice. Hypothalamic Ghrh mRNA levels were unaffected by fasting or food restriction, but Sst expression increased in acutely fasted mice, but decreased after prolonged food restriction in both control and AgRP∆GHR mice. Our findings indicate that short-term fasting and prolonged food restriction differentially affect the pattern of GH secretion, independently of GHR signaling in ARHAgRP/NPY neurons.

Introduction: The combined effects of circulating cytokines and reduced caloric intake may cause growth hormone (GH) resistance in children with Crohn’s disease. Anabolic effects of GH are mediated via phosphorylation of the STAT5b transcription factor and subsequent up regulation of IGF-1 gene expression. The molecular basis for GH resistance in colitis was not known. We hypothesized that liver GH dependent phosphorylation of STAT5b would be impaired in colitis, and that this would precede growth failure. Methods: Interleukin 10 (IL-10) null mice on the C3H/HejBir background develop chronic colitis beginning at 4–6 weeks of age. Growth, body composition, and GH signaling in liver and skeletal muscle were characterized in IL-10 null mice and wild type (WT) littermates at 8 and 20 weeks of age. Results: Colitis was present in both sexes by 8 weeks of age. Maximal reductions in growth and body composition were observed in both sexes by 20 weeks of age; these were first apparent in male mice at 8 weeks. As expected, liver IGF-1 expression was reduced in colitic mice. This was associated with a significant reduction in nuclear phospho-STAT5 abundance at both 8 and 20 weeks, and down regulation of GH receptor (GHR) abundance. Down regulation of liver GHR expression was associated with reduced nuclear abundance and DNA binding of the GHR gene promoter transactivator, Sp3. By comparison, skeletal muscle STAT5 phosphorylation was relatively preserved in colitic mice, despite reductions in lean body mass in males. Tumor Necrosis Factor (TNF) pre-treatment of rat H4IIE hepatoma cells and C2C12 mouse myoblast cells reduced GHR abundance and impaired GH dependent STAT5 activation in the liver, but not muscle, cell line. Conclusion: Growth failure and reduced IGF-1 expression in experimental colitis were associated with down regulation of liver GHR abundance and impaired STAT5 phosphorylation. The defect in liver STAT5 phosphorylation occurred early in the course of colitis, coincident with the onset of reduced weight gain, and was reproduced by TNF treatment in a liver cell line. Therapies which block TNF may improve growth in children with Crohn’s disease in part by restoring hepatic GH signaling and IGF-1 expression.

In the rat, the GH receptor (GHR) and the GH-binding protein (GHBP), which arise from alternative splicing of the same gene, show a sexually dimorphic and GH-dependent expression pattern. Multiple alternative 5'-untranslated regions (UTRs) are present in GHR and GHBP transcripts in the rat, one of which, GHR1, has recently been shown to be liver specific and found at higher levels in females. We have measured the hepatic GHR1, GHR, and GHBP transcript levels, by RNase protection and solution hybridization assay, in animals with differing hormonal status, in which hepatic GHR binding and plasma GHBP have been previously assayed. Estradiol (E2) induced GHR1 in males, whereas ovariectomy or the antiestrogen tamoxifen reduced GHR1 expression in females. The induction of GHR1 by E2 was GH dependent, being lower in GH-deficient dwarf rats and absent in hypophysectomized rats, paralleling previous measurements of plasma GHBP and hepatic GHR binding in these animals. Significant changes in GHR1 could explain the trends seen in the same extracts when coding region probes were used. Short-term adrenalectomy had no effect on GHR and GHBP expression, but dexamethasone markedly reduced both protein and messenger RNA (mRNA) levels. Corticosterone treatment had no effect alone but reduced the E2-induced increase in GHR1 levels, whereas methylprednisolone administered orally reduced hepatic GH binding, plasma GHBP, and GHR1 mRNA levels. Thus, 5'-UTRs, encoded by different first exons, are involved in the regulation of hepatic GHR and GHBP expression and need to be considered when comparing effects of hormonal manipulation on the mRNA transcripts and protein products of the GHR gene. Previous studies have found discrepancies between levels of protein expression and mRNA transcripts measured only with coding region probes. Our results suggest that posttranscriptional differences related to 5'-UTR heterogeneity in the GHR gene explain some of these discrepancies.