Growth hormone regulates the level of insulin-like growth factor-I mRNA in rat skeletal muscle

Levels of mRNA for the insulin-like growth factor-I (IGF-I) in rat heart and skeletal muscle and its dependence on GH were investigated using a solution hybridization assay. Levels of IGF-I mRNA decreased following hypophysectomy, and replacement therapy with human GH (hGH) normalized heart and skeletal muscle levels. The stimulatory effect of hGH was dose-dependent, the lowest effective dose being 100 micrograms. A significant increase of IGF-I mRNA was observed 60 min after s.c. administration of 100 micrograms hGH and the maximum increase was apparent 6-12 h after hGH injection. Administration of 200 micrograms IGF-I or 11 micrograms insulin did not significantly change levels of IGF-I mRNA. The results show that GH regulates the level of IGF-I mRNA in rat heart and skeletal muscle and give further support to the hypothesis that locally produced IGF-I might be a local mediator for the direct stimulatory effect of GH on the growth and development of heart and skeletal muscle.     

Thyroid hormone and growth hormone interact to regulate insulin-like growth factor-I messenger ribonucleic acid and circulating levels in the rat

Thyroid hormones influence growth in part by altering the secretion and effects of GH. GH, in turn, mediates its effects by regulating the synthesis and secretion of insulin-like growth factor-I (IGF-I). IGF-I is a pleiotropic growth factor that is synthesized by many tissues and acts on many tissues to regulate both cellular replication and differentiated function. We have studied the direct effects of thyroid hormones and the combined effects of thyroid hormones and GH on the regulation of IGF-I synthesis and secretion in hypophysectomized (hypox) rats in vivo. All rats, except normal littermates and a hypox control group, received 100 micrograms hydrocortisone/100 g BW for 10 days. Circulating IGF-I was measured by specific RIA (normal rats, 1 U/ml), and hepatic IGF-I mRNA was measured by Northern blot hybridization with an antisense cRNA probe. 1) Hypox rats treated with hGH (75 micrograms, ip, twice daily) for 10 days gained 17 g BW vs. 70 g for normal littermates. GH markedly increased hepatic IGF-I mRNA and circulating IGF-I (0.52 +/- 0.14 U/ml 12 h after the last GH injection vs. 0.03 +/- 0.02 for hypox controls). 2) T4 (1 micrograms/100 g BW, ip) for 10 days increased neither weight, hepatic IGF-I mRNA, nor circulating IGF-I. 3) Rats treated with T4 for 10 days followed by a single injection of 1 mg GH, ip, increased hepatic IGF-I mRNA and circulating IGF-I levels comparably as in rats receiving acute GH alone (IGF-I, 12 h, 0.31 +/- 0.09 vs. 0.36 +/- 0.06 U/ml). 4) Hypox rats treated with a single injection of T3 (1.5 micrograms/100 g BW, ip) had slightly increased hepatic IGF-I mRNA, but showed no significant change in circulating IGF-I levels. 5) A single injection of T3 plus GH to hypox rats increased IGF-I mRNA levels above those in rats injected with GH alone and increased serum IGF-I levels to 0.48 +/- 0.12 U/ml compared to 0.36 +/- 0.06 U/ml for GH alone. 6) After 10 days of GH treatment, a single injection of T3 lowered both hepatic IGF-I mRNA and circulating IGF-I (0.52 +/- 0.14 to 0.16 +/- 0.06 U/ml, 6 h after T3). These studies demonstrate that thyroid hormones have relatively little direct effect on IGF-I synthesis but can have major effects on GH-stimulated IGF-I synthesis and secretion. The pattern of these effects depends on the integrity of the pituitary gland, prior exposure of the liver to GH and/or thyroid hormones, and the temporal relationship between GH and thyroid hormone administration.     

Presence of insulin-like growth factor-I receptors and absence of growth hormone receptors in the antler tip.

Red deer antler tips in the growing phase were removed 60 days after the recommencement of growth for autoradiographical studies and RRAs. Sections were incubated with radiolabeled GH or insulin-like growth factor-I (IGF-I), with or without excess competing unlabeled hormones, and were analyzed autoradiographically. There was negligible binding of [125I]GH in any histological zone of antler sections. [125I]IGF-I showed highest specific binding in the chondroblast zone to a receptor demonstrating binding characteristics of the type 1 IGF receptor. The lowest specific binding of [125I]IGF-I was to prechondroblasts. RRAs on antler microsomal membrane preparations RRAs on antler microsomal membrane preparations confirmed the absence of GH receptors and the presence of type 1 IGF receptors found by autoradiography. These findings suggest that IGF-I may act in an endocrine manner in antler growth through a receptor resembling the type 1 IGF receptor. The presence of type 1 receptors in the chondroblast zone implicates IGF-I involvement in cartilage formation through matrixogenesis. There is no support for IGF-I having a major role in mitosis in the antler.     

Identification of insulin-like growth factor-I and its receptors in the rat testis

As part of a study of the testicular production and action of insulin-like growth factor-I (IGF-I), adult rat testes were extracted with acidified methanol, yielding an immunoreactive IGF-I fraction corresponding in size to human IGF-I. The mean IGF-I content (+/- SEM) of testes weighing approximately 1.1 g was 51.5 +/- 5.6 ng/testis, and was not due to serum contamination. After a 3-day fast testicular IGF-I decreased by 80%, whereas serum IGF-I levels declined by 90%. Testicular homogenates and isolated Leydig cells were shown to contain specific IGF-I receptors, Ka = 2 X 10(9) M-1, with 10% IGF-II cross-reactivity. The concentration of these receptors was 2 pmol binding sites per testis, or 3.3 fmol per 10(6) Leydig cells. However IGF-I at 250 ng/ml had no effect on basal or hCG-stimulated testosterone production by isolated Leydig cells, measured over 3 h. Although an effect of IGF-I over longer incubation periods cannot be excluded, it is also possible that testicular IGF-I has a mitogenic role, rather than acting on differentiated testicular functions.     

Thyroid hormone status regulates preprotachykinin-A gene expression in male rat anterior pituitary

Substance P (SP) and substance K (SK) are mammalian tachykinin peptides derived from a single preprotachykinin-A (PPT-A) gene and are widely but selectively distributed in neural and endocrine tissues. SP is present in the rat anterior pituitary, and its content there varies with the thyroid status of the animal. The present studies were undertaken to determine whether the PPT-A gene is expressed in the anterior pituitary and if so, whether PPT-A messenger RNA (mRNA) abundance is regulated by thyroid hormone status. Male rats were surgically or chemically thyroidectomized or made hyperthyroid by thyroid hormone (T3) injection. Total RNA was isolated from individual anterior pituitaries, and PPT-A mRNA abundance was determined by dot blot procedures. In parallel groups of rats, anterior pituitaries were extracted for measurement of SP and SK by specific RIAs. Surgical or chemical thyroidectomy increased PPT-A mRNA abundance 4 to 6-fold and increased both SP and SK content in the anterior pituitary. Administration of T3 to thyroidectomized rats reversed the increase in both PPT-A mRNA abundance and SP and SK content in the adenohypophysis. T3 administration to euthyroid rats also decreased PPT-A mRNA abundance and SP and SK content in the anterior pituitary. The coordinate presence of PPT-A mRNA with SP and SK in the anterior pituitary strongly suggests that these peptides are synthesized within this gland.     

Inhibition of neonatal rat growth and circulating concentrations of insulin-like growth factor-I using an antiserum to rat growth hormone

Treatment of rats for 24 h on day 2, 10 or 20 of age with a specific antiserum to rGH (anti-(rGH], GH, bromocriptine (CB-154) or prolactin failed to influence body weight gain or serum concentrations of insulin-like growth factor-I (IGF-I). On day 28 of age, however, anti-(rGH) completely inhibited body weight gain and markedly reduced circulating IGF-I concentrations, effects which were completely prevented by exogenous ovine GH (oGH). When administered to control rats on day 28 oGH caused supranormal weight gain and serum IGF-I concentrations. These results suggested that GH does not play a significant role in growth or regulation of serum IGF-I until after day 20 of age. By contrast, when anti-(rGH) was given for 4 consecutive days beginning on day 2 of life, body weight gain was reduced within 48 h and remained so until at least 28 days of age. Tail length was also significantly reduced. The effect was due to inhibition of GH effects since serum GH concentrations were low and exogenous GH prevented the effect. Inhibition of growth during the first 14 days of life occurred in the absence of any effect on serum IGF-I although by 21 days of age serum IGF-I was considerably lower than in control rats. The prolonged reduction in growth after treatment has stopped appeared to be due to a cytotoxic effect on the pituitary gland since pituitary weight and GH but not prolactin content were significantly decreased.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conventional pituitary irradiation is effective in lowering serum growth hormone and insulin-like growth factor-I in patients with acromegaly

BACKGROUND: There has been recent controversy as to the effectiveness of conventional pituitary irradiation in reducing circulating GH levels to less than 2.5 ng/ml and/or normalization of serum IGF-I. OBJECTIVES: Our objectives were to determine the effects of conventional pituitary irradiation on 1) lowering of serum GH and IGF-I levels, 2) the proportion of patients who achieve a GH level less than 2.5 ng/ml and a normal age-corrected IGF-I and the time taken to achieve this, and 3) the incidence of hypopituitarism and other adverse effects. DESIGN: We conducted retrospective data collection from 14 centers throughout the United Kingdom. PATIENTS: We studied 1840 patients with acromegaly, of whom 884 had received conventional pituitary irradiation. MEASUREMENTS: We assessed circulating GH and IGF-I levels and pituitary function at intervals after irradiation. RESULTS: Mean GH levels declined from 13.5 to 5.3 ng/ml at 2 yr after irradiation, to 2.0 ng/ml by 10 yr, and to 1.1 ng/ml at 20 yr. Twenty-two percent of patients achieved a level less than 2.5 ng/ml by 2 yr, 60% by 10 yr, and 77% by 20 yr. The interval to achieve this depended on the preirradiation GH level. IGF-I levels fell in parallel to those of GH with 63% of patients having a normal level by 10 yr. The proportions of patients with new pituitary hormone deficiencies 10 yr after irradiation were 18% for LH/FSH, 15% for ACTH, and 27% for TSH. No other side effects were noted. CONCLUSIONS: In this, the largest series reported, conventional pituitary irradiation is shown to be an effective and safe means of reducing both serum GH and IGF-I concentrations in patients with acromegaly.     

Treatment with growth hormone and insulin-like growth factor-I in critical illness

The wider availability of recombinant human growth hormone and insulin-like growth factor-I has resulted in an investigation into the potential benefits of the pharmacological administration of these anabolic peptides in a variety of clinical conditions, characterized by an increase in catabolic rate. The initial studies were small, often uncontrolled open investigations, but investigators have more recently concentrated on larger, controlled multi-centre trials. Studies to date have included patients with cardiac failure, sepsis, burns, cancer cachexia, end-stage renal failure, trauma and AIDS, and those prior to or following major surgery. The authors have in general cautiously interpreted positive effects of treatment with growth hormone and insulin-like growth factor-I, either alone or in combination, on net protein balance, body composition, well-being and performance. Two large, randomized, placebo-controlled European multi-centre studies have recently detailed the effects of growth hormone treatment in critically ill intensive care patients. Major increases in mortality and morbidity were associated with growth hormone treatment. The mechanism(s) accounting for the increased mortality remain poorly understood. These negative findings have led to a decrease in the clinical use of growth hormone and in research activity in the area of anabolic treatment in human illness.     

Corticotrophin-releasing hormone inhibits insulin-like growth factor-I release from primary cultures of rat granulosa cells

Corticotrophin-releasing hormone (CRH), a neuropeptide which modulates gonadal function during stress, is expressed by several cell types of the rat ovary and is able to suppress oestrogen release from rat granulosa cells. The mechanism of this effect is, however, not known. Since insulin-like growth factor (IGF)-I is produced by rat granulosa cells and exerts a synergistic role with FSH on granulosa cell steroidogenesis, we hypothesised that CRH may suppress oestrogen release from granulosa cells by inhibiting IGF-I release and/or stimulating the release of its binding protein (IGFBP-3). To test this hypothesis, granulosa cells were obtained from immature female Sprague-Dawley rats primed with diethylstilboestrol, and hormone concentrations were measured in the conditioned medium by radioimmunoassay. CRH suppressed oestrogen and IGF-I release stimulated by FSH used at a concentration of 1 IU/l, whereas it did not have any statistically significant effect on oestrogen and IGF-I release in basal conditions or in response to 5 IU/l FSH. The suppressive effects of CRH on oestrogen and IGF-I release were antagonised by a selective CRH receptor antagonist. CRH had no effects on IGFBP-3 release. CRH did not have any effect on oestrogen release stimulated by increasing concentrations of IGF-I and its suppressive effect on FSH-stimulated oestrogen release was overcome by the addition of low doses of exogenous IGF-I. In conclusion, CRH suppressed the release of oestrogen and IGF-I, but not of IGFBP-3. Thus, the inhibitory effects of CRH on oestrogen release could be mediated, partly, by a suppression of the autocrine/paracrine action of IGF-I.     

Non-lactogenic effects of growth hormone on growth and insulin-like growth factor-I messenger ribonucleic acid of rat mammary gland

In contrast to established dogma that PRL is central in mammary development, and GH mimics PRL in affecting growth because of structural similarities, we found that both hGH, which is lactogenic, and rGH, which is non-lactogenic, were significantly more potent than hPRL and rPRL in stimulating mammary growth in rats. Additionally, hGH was more potent than hPRL in increasing mammary IGF-I mRNA content. These data indicate that GH has separate effects on parameters of mammary gland growth, suggesting an independent role for GH in mammary growth.     

Growth hormone regulation of insulin-like growth factor-I mRNA in rat adipose tissue and isolated rat adipocytes.

The effects of hypophysectomy and hormonal replacement therapy on insulin-like growth factor-I (IGF-I) mRNA in rat adipose tissue and adipocytes were studied. The effects of GH and IGF-I in vitro on IGF-I mRNA and IGF-I production were also studied in cultured rat adipocytes. Male rats were hypophysectomized at about 50 days of age and given replacement therapy with cortisol (400 micrograms/kg per day) and thyroxine (10 micrograms/kg per day). GH was given as a single i.v. or s.c. injection and also as a continuous s.c. infusion for 6 days. Epididymal fat pads were excised and used either for isolation of adipocytes or for determination of IGF-I mRNA in adipose tissue. A solution hybridization assay was used. The IGF-I mRNA content of adipocytes was analysed either immediately after isolation or after short-term (2-3 days) culture with or without GH or IGF-I. Hypophysectomy resulted in a marked decrease in IGF-I mRNA in both tissue and cells. Replacement therapy (in vivo) with cortisol and thyroxine alone had no effect, whereas additional treatment with GH caused a dose-dependent increase in IGF-I mRNA. IGF-I mRNA was also increased after a continuous s.c. infusion of GH. A single i.v. injection of GH (100 micrograms) resulted in an increase in IGF-I mRNA after approximately 2 h, with maximal levels around 6 h after the injection. In cultured adipocytes, addition of GH to the culture medium increased IGF-I mRNA in a dose-dependent manner and a marked increase was observed with a concentration of GH of 1 ng/ml. Addition of IGF-I (100 ng/ml) had no effect. The increase in IGF-I mRNA after addition of GH (100 ng/ml) was detectable after 3 h. The concentration of IGF-I in the culture medium was increased 24 h after the addition of GH. These results demonstrate that GH induces IGF-I mRNA in both adipose tissue and isolated fully differentiated adipocytes and that this increase in IGF-I mRNA results in increased IGF-I production.     

The growth hormone/insulin-like growth factor-I axis in exercise and sport

The syndrome of adult GH deficiency and the effects of GH replacement therapy provide a useful model with which to study the effects of the GH/IGF-I axis on exercise physiology. Measures of exercise performance including maximal oxygen uptake and ventilatory threshold are impaired in adult GH deficiency and improved by GH replacement, probably through some combination of increased oxygen delivery to exercising muscle, increased fatty acid availability with glycogen sparing, increased muscle strength, improved body composition, and improved thermoregulation. In normal subjects, in addition to the long-term effects of GH/IGF-I status, there is evidence that the acute GH response to exercise is important in regulating substrate metabolism after exercise. Administration of supraphysiological doses of GH to athletes increases fatty acid availability and reduces oxidative protein loss, particularly during exercise, and increases lean body mass. Despite a lack of evidence that these metabolic effects translate to improved performance, GH abuse by athletes is widespread. Tests to detect GH abuse have been developed based on measurement in serum of 1) indirect markers of GH action, and 2) the relative proportions of the two major naturally occurring isoforms (20 and 22kDa) of GH. There is evidence that exercise performance and strength are improved by administration of GH and testosterone in combination to elderly subjects. The potential benefits of GH in these situations must be weighed against potential adverse effects.     

Growth regulation in the gobiid teleost, Gillichthys mirabilis: roles of growth hormone, hepatic growth hormone receptors and insulin-like growth factor-I.

Studies of the teleost Gillichthys mirabilis were undertaken to assess the role of GH in regulating hepatic GH receptors, insulin-like growth factor-I (IGF-I) activity and cartilage growth. Hypophysectomized G. mirabilis were injected with saline vehicle or with 10, 100 or 1000 ng tilapia GH (tGH)/g every other day for 2-3 weeks. Growth, judged by increased body weight and length, was inhibited by hypophysectomy and stimulated by increasing tGH doses. Hepatic GH receptors, measured by 125I-labelled tGH binding, were decreased 50% by hypophysectomy. An additional dose-dependent reduction in binding was observed 24 h after tGH injection, but MgCl2 stripping of membranes suggested receptor occupation by exogenous tGH. IGF-I activity, measured by 35SO4 incorporation into oral cartilage explants in vitro, was decreased 50% by hypophysectomy and increased to 200% of intact control levels after injection of 1000 ng tGH/g. In a second experiment, 35SO4 incorporation by oral cartilage from hypophysectomized fish injected with 10, 100 or 1000 ng tGH/g was stimulated to intact control levels. Effects of feeding and tGH injection on in-vitro responsiveness of oral cartilage to recombinant bovine IGF-I (rbIGF-I; 10-1000 ng/ml) were also assessed. Oral cartilage from fed fish showed a parabolic dose-response curve, whereas oral cartilage from starved fish had a lower basal rate of 35SO4 incorporation and a linear dose-response relationship. Oral cartilage from hypophysectomized G. mirabilis showed a significantly attenuated response to rbIGF-I which was restored by tGH injection, suggesting that the GH status of the animal is important for sensitivity of target tissue to IGF-I. Because of its similarity to other vertebrate systems, G. mirabilis presents a good teleost model of growth regulation and of the functions and interactions of GH and IGF-I.     

Developmental regulation of insulin-like growth factor-I and growth hormone receptor gene expression.

During development, the insulin-like growth factor I (IGF-I) gene is expressed in a tissue specific manner; however, the molecular mechanisms governing its developmental regulation remain poorly defined. To examine the hypothesis that expression of the growth hormone (GH) receptor accounts, in part, for the tissue specific expression of the IGF-I gene during development, the developmental regulation of IGF-I and GH receptor gene expression in rat tissues was examined. The level of IGF-I and GH receptor mRNA was quantified in RNA prepared from rats between day 17 of gestation (E17) and 17 months of age (17M) using an RNase protection assay. Developmental regulation of IGF-I gene expression was tissue specific with four different patterns of expression seen. In liver, IGF-I mRNA levels increased markedly between E17 and postnatal day 45 (P45) and declined thereafter. In contrast, in brain, skeletal muscle and testis, IGF-I mRNA levels decreased between P5 and 4M but were relatively unchanged thereafter. In heart and kidney, a small increase in IGF-I mRNA levels was observed between the early postnatal period and 4 months, whereas in lung, minimal changes were observed during development. The changes in GH receptor mRNA levels were, in general, coordinate with the changes in IGF-I mRNA levels, except in skeletal muscle. Interestingly, quantification of GH receptor levels by Western blot analysis in skeletal muscle demonstrated changes coordinate with IGF-I mRNA levels. The levels of the proteins which mediate GH receptor signaling (STAT1, -3, and -5, and JAK2) were quantified by Western blot analysis. These proteins also are expressed in a tissue specific manner during development. In some cases, the pattern of expression was coordinate with IGF-I gene expression, whereas in others it was discordant. To further define molecular mechanisms for the developmental regulation of IGF-I gene expression, protein binding to IGFI-FP1, a protein binding site that is in the major promoter of the rat IGF-I gene and is important for basal promoter activity in vitro, was examined. Gel shift analyses using a 34-base pair oligonucleotide that contained IGFI-FP1 did not demonstrate changes in protein binding that paralleled those in IGF-I gene expression, suggesting that protein binding to IGFI-FP1 does not contribute to the developmental regulation of IGF-I gene expression, at least in brain and liver. In summary, the present studies demonstrate coordinate expression of the IGF-I gene and GH receptor during development and suggest that GH receptor expression contributes to the tissue specific expression of the IGF-I gene during development.     

New concepts: growth hormone, insulin-like growth factor-I and the kidney.

Both growth hormone (GH) and IGF-1 have major effects on normal kidney growth, structure and function and participate in the pathogenesis of certain kidney diseases. Furthermore when the kidneys fail there are profound changes in the circulating GH-IGF-1 system and the renal and systemic responses to these hormones. In this brief review we address the advances that have been made in our understanding of the relationship between growth hormone GH and IGF-1 and the kidney in health and the systemic and local perturbations that occur in kidney disease and identify key unanswered questions.     

Regulation of insulin-like growth factor-I and growth hormone receptor gene expression by diabetes and nutritional state in rat tissues

Insulin-like growth factor-I (IGF-I) mRNA and GH receptor mRNA levels were analysed in different tissues from rats made diabetic with streptozotocin, fasted rats and rats fed with a protein-reduced diet. Diabetes decreased IGF-I mRNA levels in liver, heart, diaphragm, kidney and aorta, but not in brain. GH receptor mRNA levels were decreased in heart and diaphragm, but not in liver and kidney. Fasting decreased IGF-I mRNA in all tissues studied except brain, and decreased GH receptor mRNA in liver, heart and diaphragm, but not in kidney. A protein-reduced diet decreased hepatic IGF-I mRNA levels but did not significantly affect other tissues, while GH receptor mRNA levels were reduced in liver and diaphragm. In conclusion, both diabetes and limited nutrition affected IGF-I and GH receptor mRNA in different tissues, but the two mRNAs were not co-ordinately regulated in all tissues studied. While reduced GH receptor gene expression may thus be responsible for decreased IGF-I gene expression in some states and tissues, additional regulatory mechanisms may be of importance.     

Insulin and insulin-like growth factor-I acutely inhibit surface translocation of growth hormone receptors in osteoblasts: A novel mechanism of growth hormone receptor regulation

We previously have demonstrated that insulin and insulin-like growth factor-I (IGF-I) down-regulate growth hormone (GH) binding in osteoblasts by reducing the number of surface GH receptors (GHRs). The present study was undertaken to investigate the mechanism of GHR down-regulation. Treatment with 5 nM insulin or IGF-I for 18 hr significantly decreased surface GH binding to 26.4 ± 2.9% and 23.0 ± 2.7% of control (mean ± SE; P < 0.05), respectively. No corresponding reductions in the mRNA level and total cellular content of GHR were found, nor was the rate of receptor internalization affected. The effects on GHR translocation were assessed by measuring the reappearance of GH binding of whole cells after trypsinization to remove the surface receptors. GH binding of control cultures significantly increased (P < 0.05) over 2 hr after trypsinization, whereas no recovery of binding activity was detected in insulin and IGF-I-treated cultures, indicating that GHR translocation was impaired. Studies on the time course of GHR down-regulation revealed that surface GH binding was reduced significantly by 3-hr treatment (P ≤ 0.0005), whereas GHR translocation was completely abolished by 75–90 min with insulin and IGF-I. The inhibition of receptor translocation by insulin, but not IGF-I, was attenuated by wortmannin. In conclusion, insulin and IGF-I down-regulated GH binding in osteoblasts by acutely impairing GHR translocation, with their effects exerted through distinct postreceptor signaling pathways.