The effects of ovine placental lactogen and bovine growth hormone on hepatic and mammary gene expression in lactating sheep

The ability of ovine placental lactogen (oPL) to bind to the growth hormone receptor (GHR) raises the possibility that oPL may exert a growth hormone (GH)-like action on galactopoiesis. We have compared the effects of treating lactating ewes for 5 days with an equimolar dose (0.1 mg/kg/day, administered as two equal doses 12 hourly) of either bovine growth hormone (bGH) (n = 10), oPL (n = 10) or saline (n = 9) on hepatic and mammary GHR, insulin-like growth factor-I (IGF-I) and IGF-binding protein-3 (IGFBP-3) gene expression and hepatic GHR number. Hepatic GHR and IGFBP-3 mRNA were unaltered by bGH or oPL treatment. Hepatic IGF-I mRNAs increased following bGH (P < 0.05) but not oPL treatment. GHR gene expression was greater in liver compared to mammary gland extracts. There was no effect of either bGH or oPL treatment on mammary GHR, IGF-I or IGFBP-3 mRNA or hepatic GHR number. These studies confirm the galactopoietic effects of bGH in lactating ruminants and suggest that the mechanism of this action is not via increased hepatic GHR number or gene expression. In addition, the increase in hepatic but not mammary IGF-I mRNA with bGH treatment suggests an endocrine action of IGF-I on milk synthesis. These studies also demonstrate that an equimolar dose of oPL is not galactopoietic or somatogenic in the lactating ewe.     

Insulin-like growth factors, insulin-like growth factor-binding proteins, insulin-like growth factor-binding protein-3 protease, and growth hormone-binding protein in lipodystrophic human immunodeficiency virus-infected patients

Human immunodeficiency virus (HIV)-lipodystrophy is associated with impaired growth hormone (GH) secretion. It remains to be elucidated whether insulin-like growth factors (IGFs), IGF-binding proteins (IGFBPs), IGFBP-3 protease, and GH-binding protein (GHBP) are abnormal in HIV-lipodystrophy. These parameters were measured in overnight fasting serum samples from 16 Caucasian males with HIV-lipodystrophy (LIPO) and 15 Caucasian HIV-infected males without lipodystrophy (NONLIPO) matched for age, weight, duration of HIV infection, and antiretroviral therapy. In LIPO, abdominal fat mass and insulin concentration were increased (>90%, P < .01) and insulin sensitivity (Log10ISI(composite)) was decreased (-50%, P < .001). Total and free IGF-I, IGF-II, IGFBP-3, and IGFBP-3 protease were similar between groups (all P > .5), whereas, in LIPO, IGFBP-1 and IGFBP-2 were reduced (-36%, P < .05 and -50%, P < .01). In pooled groups, total IGF-I, free IGF-I, total IGF-II, and IGFBP-3, respectively, correlated inversely with age (all P < .01). In pooled groups, IGFBP-1 and IGFBP-2 correlated positively with insulin sensitivity (age-adjusted all P < .05). IGFBP-3 protease correlated with free IGF-I in pooled groups (r(p) = 0.47, P < .02), and in LIPO (r(p) = 0.71, P < .007) controlling for age, total IGF-I, and IGFBP-3. GHBP was increased, whereas GH was decreased in LIPO (all P < .05). GH correlated inversely with GHBP in pooled groups (P < .05). Taken together the similar IGFs and IGFBP-3 concentrations between study groups, including suppressed GH, and increased GHBP in LIPO, argue against GH resistance of GH-sensitive tissues in LIPO compared with NONLIPO; however, this notion awaits examination in dose-response studies. Furthermore, our data suggest that IGFBP-3 protease is a significant regulator of bioactive IGF-I in HIV-lipodystrophy.     

Changes in the growth hormone-IGF-I axis in non-obese diabetic mice

We investigated the changes in GH-IGF-I axis in non-obese diabetic (NOD)-mice, a model of insulin-dependent diabetes mellitus. Diabetic female NOD mice and their age- and sex-matched controls were sacrificed at 4, 14, 21 and 30 days (30d DM) after the onset of glycosuria. Serum GH levels increased and serum IGF-I levels decreased in the 30d DM group (182 +/- 32% and 45 +/- 24% of age-matched controls respectively, p < 0.05). Another group (30d DM + I) was given SC insulin, and its serum IGF-I levels remained decreased. Liver GH receptor (GHR) and GH binding protein (GHBP) mRNA levels, as well as liver membrane GH binding assays were deeply decreased in the 30d DM group in comparison to controls. GHR message and binding capacity remained decreased in the 30d DM + I group. Renal GHR mRNA was decreased at 21d DM but not at 14d DM, whereas GHBP mRNA remained unchanged throughout the experiment. In conclusion, increased serum GH levels are documented in NOD diabetic mice, similarly to the changes described in humans. The decrease in GHR levels and decreased serum IGF-I in spite of increased circulating GH suggest a state of GH resistance.     

Ontogeny of messenger RNA for the rat growth hormone receptor and serum binding protein.

Little is known of the regulation of gene expression for the family of growth hormone (GH) and prolactin (PRL) receptors (PRL-R). Furthermore, the relationship between expression of the GH receptor (GHR) and its soluble truncated form (GH-binding protein, GHBP) is unclear. The actions of both GH and PRL are developmentally regulated and several studies have examined the ontogeny of these receptors by classical hormone-binding techniques. In the current study we have examined the expression of GHR/GHBP and PRL-R mRNA in the male rat over a broad developmental range--fetal through to 110 days of age. The GHR mRNA (4.5 kb) was barely detectable in fetal and early (less than 20 days) postnatal livers, but was followed by a gradual increase up to 40 days of age by which time adult plateau levels were reached. In contrast, hepatic GHBP mRNA (1.2 kb) was clearly identifiable in the fetus and subsequently followed a similar pattern to the 4.5 kb GHR mRNA although there was a somewhat earlier rise. Hepatic membrane binding studies using 125I-bovine GH as ligand revealed no measurable binding activity at less than 20 days of age. Binding remained low thereafter. In contrast, the serum GHBP binding activity was detectable at 10 days of age and rose to adult levels by 50 days of age. These results indicate that mRNA species for GHR, GHBP, PRL-R and insulin-like growth factor I (IGF-I) are all developmentally regulated with the pattern for IGF-I correlating more closely with that of GHBP than GHR.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of zinc deficiency on serum somatomedin levels and skeletal growth in young rats

We have studied potential mechanisms by which zinc deficiency (ZD) may result in growth impairment in young animals. Dietary-induced ZD in young rats resulted in diminished skeletal growth as measured by tibial epiphyseal width. Treatment with bovine GH (bGH) did not increase skeletal growth suggesting GH resistance rather than GH deficiency in zinc-deficient rats. Serum levels of basic somatomedin (SM) were lower in zinc-deficient rats than in control rats receiving a zinc adequate diet, either ad libitum or in pair matched amounts, and were restored to normal by zinc repletion but not by bGH treatment, suggesting that SM production is impaired by ZD. There was a high correlation between tibial epiphyseal widths and serum or femur zinc concentrations. These findings, along with observations that despite similar levels of serum basic SM the bGH-treated zinc-deficient rats had smaller tibial epiphyseal widths than pair fed control rats, additionally suggest that the action of SM on skeletal growth is impaired by ZD.     

Upregulation of GH receptor and GH binding protein during pregnancy in the GH deficient rat

During pregnancy there are dramatic changes in the endocrine and metabolic status of the mother: growth hormone (GH) is an important regulator of growth and development. A proportion of GH is bound by specific GH binding proteins (GHBP) that closely resemble the GH receptor (GHR). In the rodent both GHBP and the GHR are considered to be GH dependent, and consequently during pregnancy the increase in serum GH is associated with an increase in GHBP. To examine whether an increase in maternal GH is obligatory for elevation of maternal GHBP or GHR during pregnancy, we used a unique GH-deficient (GHD) strain of rats, to avoid the methodological complications of hypophysectomy and assessed serum GH, GHBP and hepatic GHR binding during the course of pregnancy. In GH normal rats, serum GH concentrations increased twofold and GHBP levels increased threefold; there was no change in hepatic GHR binding. In GHD rats, serum GH concentrations were low and did not increase during pregnancy. Nonetheless, levels of both serum GHBP and hepatic GHR binding increased to that measured in normal rats. Thus, an increase in maternal GH concentration is not required for the gestational upregulation of maternal GHBP or hepatic GHR binding, suggesting that other hormones may be essential in modulating the GH axis during pregnancy.     

Short-term growth hormone or IGF-I administration improves the IGF-IGFBP system in arthritic rats

ObjectiveAdjuvant-induced arthritis is an experimental model of rheumatoid arthritis that inhibits the GH-IGF-I axis and decreases body weight gain and muscle mass. Although chronic GH or IGF-I treatment increases body weight gain in arthritic rats, muscle resistance to GH and IGF-I is a very common complication in inflammatory diseases. In this study we examine the effect of short-term administration of rhGH and rhIGF-I on liver and muscle IGF-I, IGFBP-3 and ? 5 as well as on the ubiquitin-ligases MuRF1 and atrogin-1 in the muscle of arthritic rats.DesignArthritis was induced in adult male Wistar rats by an intradermal injection of 4 mg of Freund's adjuvant. Fifteen days after adjuvant injection, 300 μg/kg of rhGH or 200 μg/kg of rhIGF or saline was administrated 18 and 3 h before decapitation. A pair-fed group injected with saline was included in order to discard a possible effect of decreased food intake. Gene expression of IGF-I, GHR, IGFBP-3, IGFBP-5, atrogin-1 and MuRF1 were quantified using RT-PCR. In serum, IGF-I was measured by radioimmunoassay (RIA) and IGFBP-3 by ligand blot.ResultsArthritis decreased serum IGF-I and IGF mRNA in liver (P < 0.05), but not in skeletal muscle. In arthritic rats, rhGH increased serum IGF-I and liver IGF-I mRNA similar to the levels of pair-fed rats. Arthritis increased atrogin-1, MuRF1, IGFBP-3 and IGFBP-5 mRNA in muscle (P < 0.01). IGFBP-3 mRNA was downregulated by rhIGF-I, but not by rhGH, administration in control and arthritic rats (P < 0.05). Administration of rhGH and rhIGF-I increased IGFBP-5 in the gastrocnemius of arthritic rats.ConclusionsShort-term rhGH and rhIGF-I administration was found to increase muscle IGFBP-5 mRNA, whereas only rhIGF-I administration decreased muscle IGFBP-3 mRNA in control and arthritic rats. These data suggest that arthritis does not induce GH or IGF-I resistance in skeletal muscle.     

Effects of growth hormone treatment on serum levels of insulin-like growth factors (IGFs) and IGF binding proteins 1-4 in postmenopausal women

BACKGROUND AND OBJECTIVE: Insulin-like growth factor binding proteins (IGFBPs) modulate the actions and bioavailability of insulin-like growth factors (IGFs), however, their regulation in vivo is incompletely understood. In this study we investigated the effects of different doses of growth hormone (GH) on circulating levels of IGFs and IGFBPs. DESIGN: The study was double-blind and placebo-controlled. Patients were treated with either GH in doses of 0.05, 0.10, or 0.20 lU/kg/day of placebo for one week. PATIENTS: Forty post-menopausal women aged 52-73 years with low bone mass. MEASUREMENTS: Serum IGF-I and IGF-II were measured by RIA while IGFBP-1-3 were measured by Western ligand blot (WLB) and compared with determinations by specific immunoassays. IGFBP-4 was measured by WLB alone. RESULTS: Both IGF-I (P < 0.001) and IGF-II (P < 0.01) increased significantly during GH treatment. Additionally, IGFBP-1 (P < 0.001) and IGFBP-2 (P < 0.001) decreased significantly while IGFBP-3 (P < 0.001) and IGFBP-4 (P < 0.05) increased all in a dose-dependent manner. Stepwise (backwards) multiple regression analyses showed that the changes in IGF-I and IGF-II, and age correlated with the change in serum IGFBP-1. Both GH-dosage, the increase in IGF-II, and body mass index correlated with the decrease in IGFBP-2. Furthermore, the increase in serum IGF-I, IGF-II, and triiodothyronine correlated with the increase in IGFBP-3. Moreover, GH-dosage correlated with the increase in serum IGFBP-4. CONCLUSION: GH significantly increased serum IGF-I, IGF-II, IGFBP-3, and IGFBP-4 and decreased serum IGFBP-1 and IGFBP-2 in post-menopausal women.     

Growth and endocrine effects of recombinant bovine growth hormone treatment in non-transgenic and growth hormone transgenic coho salmon

To examine the relative growth, endocrine, and gene expression effects of growth hormone (GH) transgenesis vs. GH protein treatment, wild-type non-transgenic and GH transgenic coho salmon were treated with a sustained-release formulation of recombinant bovine GH (bGH; Posilac). Fish size, specific growth rate (SGR), and condition factor (CF) were monitored for 14 weeks, after which endocrine parameters were measured. Transgenic fish had much higher growth, SGR and CF than non-transgenic fish, and bGH injection significantly increased weight and SGR in non-transgenic but not transgenic fish. Plasma salmon GH concentrations decreased with bGH treatment in non-transgenic but not in transgenic fish where levels were similar to controls. Higher GH mRNA levels were detected in transgenic muscle and liver but no differences were observed in GH receptor (GHR) mRNA levels. In non-transgenic pituitary, GH and GHR mRNA levels per mg pituitary decreased with bGH dose to levels seen in transgenic salmon. Plasma IGF-I was elevated with bGH dose only in non-transgenic fish, while transgenic fish maintained an elevated level of IGF-I with or without bGH treatment. A similar trend was seen for liver IGF-I mRNA levels. Thus, bGH treatment increased fish growth and influenced feedback on endocrine parameters in non-transgenic but not in transgenic fish. A lack of further growth stimulation of GH transgenic fish suggests that these fish are experiencing maximal growth stimulation via GH pathways.     

The decrease in hepatic IGF-I gene expression in arthritic rats is not associated with modifications in hepatic GH receptor mRNA

OBJECTIVE: Adjuvant-induced arthritis induces a catabolic response, and a decrease in circulating IGF-I. Hypermetabolism and GH insensitivity have been described in acute inflammation. The aim of this study was to analyze whether impaired IGF-I secretion in arthritic rats can be attributed to hepatic GH resistance. DESIGN AND METHODS: Male Wistar rats were injected with complete Freund's adjuvant, and 14 days afterwards arthritic and control rats were injected daily with recombinant human GH (rhGH) (3 IU/kg) or saline for 8 days. GH receptor (GHR) gene expression in the liver and the effect of rhGH on hepatic IGF-I synthesis in arthritic rats were examined. RESULTS: There was a significant decrease in hepatic concentrations of IGF-I (P < 0.01) as well as in the IGF-I gene expression in arthritic but not in pair-fed rats. In contrast, arthritis did not modify GHR mRNA levels in the liver. The 8 day administration of rhGH resulted in an increase in body weight gain in arthritic but not in control rats. There was an increase in hepatic IGF-I synthesis and in GHR mRNA levels after rhGH treatment, both in control and in arthritic rats. Two endotoxin lipopolysaccharide (LPS) (1 mg/kg) injections decreased hepatic concentrations of IGF-I and IGF-I mRNA (P < 0.01). Contrary to the results obtained in arthritic rats, mRNA expression of GHR in the liver was lower in LPS- than in saline-treated rats (P < 0.01). CONCLUSION: These data suggest that the decrease in IGF-I synthesis induced by chronic arthritis is not secondary to GH resistance.     

Growth hormone-binding protein is directly and IGFBP-3 is inversely associated with risk of female breast cancer

OBJECTIVE: Several components of the GH and IGF systems have been implicated in the development of malignancies. All components of these hormonal systems have never been jointly evaluated in female breast cancer, and previous studies have not examined the role of IGF-binding proteins (IGFBP-4, IGFBP-6) or GH-binding protein (GHBP). DESIGN: Hospital-based case-control study. METHODS: In this sample of primarily postmenopausal women, we obtained serum measures of IGF-I, IGF-II, and binding proteins IGFBP-1, IGFBP-3, IGFBP-4, IGFBP-6, as well as GHBP, insulin, and leptin from 74 breast cancer cases and 76 control subjects. RESULTS: In crude analyses, we found lower age-standardized mean IGF-I, IGFBP-3, IGFBP-4, IGFBP-6, and higher IGFBP-1 and GHBP in breast cancer cases when compared with controls. Multivariate models mutually adjusted for other GH-IGF system components and classical breast cancer risk factors demonstrated an inverse association between IGFBP-3 and risk of breast cancer (odds ratio (OR) = 0.2, P < 0.01) and a direct association between GHBP and disease risk (OR = 3.3, P < 0.01). No significant associations were detected in multivariate analyses among IGF-I, IGF-II or IGFBP-1, IGFBP-4, IGFBP-6 with risk of breast cancer, indicating that these factors may not have effects independent of and/or comparable with IGFBP-3 and GHBP. CONCLUSIONS: These results support a protective role of IGFBP-3 and demonstrate for the first time an increased risk of breast cancer with higher GHBP, after accounting for variation in IGFs, IGFBPs, and classical breast cancer risk factors.     

Coordinated regulation of the GH/IGF system genes during refeeding in rainbow trout (Oncorhynchus mykiss)

The GH/IGF system is a complex regulation network strongly dependent on nutrient availability. While the effect of starvation on the GH/IGF system has been extensively studied, the time course of events leading to the restoration of GH/IGF system activity after starvation is largely unknown. We, therefore, measured the plasma levels of GH, IGF-I and IGF-II and the expression of the GH/IGF system in liver and muscle. Starvation increased the plasma GH level and 1 day of refeeding completely restored it (1.10 +/- 0.27 vs 1.12 +/- 0.28 ng/ml). Thereafter, plasma GH continued to decrease until day 7 and returned to control values from day 15. Starvation decreased plasma IGF-I and IGF-II and refeeding raised plasma IGF-I only from day 4. In contrast, the plasma IGF-II level doubled after 1 day's refeeding (26.5 +/- 1.9 vs 44.0 +/- 3.4 ng/ml; P < 0.01). Starved fish exhibited higher GH receptor (GHR)1 mRNA abundance in liver and muscle than in controls, whereas GHR2 mRNA abundance was increased only in muscle. In liver, 1 day of refeeding, decreased GHR1 (twofold), but increased GHR2 mRNA abundance (twofold). Thereafter, a progressive return to normal values was observed. Liver IGFBP-4 mRNA abundance was lowered in starved fish followed by a progressive restoration during refeeding. Starvation had no effect on liver IGFBP-2 and IGFBP-6 mRNA abundance, whereas refeeding provoked a peak of IGFBP-2 and IGFBP-6 expression at day 7. In muscle, starvation led to a decrease of the IGFBP-2 mRNA level, which was restored only from day 7. IGFBP-4 mRNA abundance in starved fish was lower than in the controls and refeeding led to a transient upregulation (sevenfold) of IGFBP-4 gene at day 1. IGF-I, IGFBP-5, and IGFBP-related protein 1 (rP1) expression profiles were similar, showing a decrease of expression after starvation, a first peak of expression at day 2, a second peak at day 7, and a return to normal value from day 15. Moreover, IGF-I, IGFBP-5, and IGFBP-rP1 mRNA abundance were positively correlated (r = 0.6-0.8; P < 0.0001). In conclusion, plasma IGF-I was restored later than plasma GH level, which suggests that plasma IGF-I levels cannot account for plasma GH changes. The coordinated regulation of IGF-I, IGFBP-5, and IGFBP-rP1 expression would be a signature for the resumption of myogenic activity.     

Differential expression of the insulin-like growth factor binding proteins in spontaneously diabetic rats.

Diabetes-induced growth retardation in the rodent is associated with both reduced circulating insulin-like growth factor-I (IGF-I) and enhanced levels of inhibitors of somatomedin activity. IGF-binding proteins (IGFBPs) are present in the circulation and tissue fluids and are believed to modulate the actions of IGF-I. Since elevated concentrations of the IGFBPs may contribute to the enhanced somatomedin-inhibitor activity observed in serum from diabetic animals, we have examined the amounts of hepatic IGFBP-1, -2, -3 and -4 mRNA in the spontaneously diabetic BioBreeding/Worcester rat. The study used two types of diabetic animal: mildly diabetic animals, which received suboptimal insulin treatment (0.5-1 U/day) and diabetic animals, which received intensive insulin treatment (3-6 U/day). A significant increase in the amount of IGFBP-1 and IGFBP-2 mRNA was seen 1 month and 3 months after the onset of diabetes. Intensive insulin treatment for 3 weeks normalized the amount of IGFBP-1 mRNA in diabetic rats and resulted in a decrease in IGFBP-2 mRNA. In contrast to the increase in IGFBP-1 and IGFBP-2 mRNA, a significant decrease in IGFBP-3 mRNA was seen in diabetic rats (54.6% of control, P less than 0.0005 and 64.6% of control, P less than 0.005 for 1 and 3 months respectively) and intensive insulin treatment for 3 weeks did not restore the IGFBP-3 mRNA level in diabetic rats. No significant difference in IGFBP-4 mRNA levels was seen in diabetic compared with non-diabetic rats. When serum was analysed by ligand blotting the major finding was a reduction in the 39-42 kDa binding protein. No increase in 29-30 kDa IGFBP in the serum was detected in the diabetic rats. From these studies we conclude that the major change in IGFBPs in mildly hyperglycaemic spontaneously diabetic rats is a decrease in IGFBP-3. The changes in hepatic IGFBP-1 and -2 mRNA do not appear to be of sufficient magnitude to result in an increase in serum concentrations of these binding proteins.     

A single arginine residue determines species specificity of the human growth hormone receptor.

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.