Effects of growth hormone and hypophysectomy of pregnant rats on serum concentrations of pregnancy-associated murine protein-1

Continuous infusion of bovine GH to hypophysectomized non-pregnant rats increased serum concentrations of pregnancy-associated murine protein-1 (PAMP-1) to the levels of adult female rats and pregnant rats. Serum concentrations of PAMP-1 were followed from Day 16 of gestation until 3 days after parturition in hypophysectomized (on Day 14 of gestation) and intact pregnant rats. In the intact pregnant rat there was a decrease in PAMP-1 values from Day 16 until delivery. The serum concentrations of PAMP-1 in hypophysectomized pregnant rats were similar to those in intact pregnant rats before parturition, but PAMP-1 concentrations decreased markedly after parturition in the hypophysectomized rats. We suggest that the serum concentrations of PAMP-1 can be maintained without pituitary GH in late pregnancy, while serum values of PAMP-1 in non-pregnant rats is dependent upon a continuous secretion of pituitary GH.     

Induction of pregnancy-associated murine protein-1 in dwarf mice by human growth hormone

Pregnancy-associated murine protein-1 (PAMP-1) could not be detected in peripheral blood of female dwarf mice (genotype dw/dw of the DW strain). By contrast the normal size females of the DW strain (genotypes +/+ and +/dw) had PAMP-1 serum levels of 18.9 AU +/- 15.7 AU/ml. Following administration of biosynthetic human growth hormone (hGH) every 2 h for 52 h PAMP-1 was detected in all dwarf females at concentrations of 16.0 AU +/- 3.3 AU/ml. The albumin levels in the circulation of DW females of normal size were significantly higher (P less than 0.05) than those of DW dwarfs, and the hGH administration did not change the serum albumin levels. The present experiment adds weight to the suggestion that the PAMP-1 serum level is regulated by GH.     

Secretory pattern of growth hormone regulates plasma concentration of pregnancy-associated murine protein-1 in the non-pregnant rat

Serum concentrations of pregnancy-associated murine protein-1 (PAMP-1) were followed in hypophysectomized adult female rats during treatment with oestradiol and continuous or intermittent human growth hormone (hGH). After hypophysectomy a rapid decrease in PAMP-1 values was recorded while concentrations of albumin and the acute phase alpha 2-macroglobulin were unaffected. PAMP-1 values were completely restored by continuous infusion of hGH (1.4 i.u./kg/day). Neither the same dose of hGH given as two daily injections nor oestrogen replacement treatment had any effect. It is concluded that the serum concentration of PAMP-1 in the non-pregnant rat is regulated by the sexually dimorphic secretory pattern of GH.     

Effects of growth hormone and insulin-like growth factor-1 on hepatocyte antioxidative enzymes

The physiological decline that occurs in aging is thought to result, in part, from accumulation of oxidative damage generated by reactive oxygen species during normal metabolic processes. Elevated levels of antioxidative enzymes in liver tissues are present in the Ames dwarf, a growth hormone (GH)-deficient mouse that lives more than 1 year longer than wild-type mice from the same line. In contrast, transgenic mice that overexpress GH exhibit depressed hepatic levels of catalase and have significantly shortened life spans. In this study, we evaluated the in vitro effects of GH and insulin-like growth factor 1 (IGF-1) on antioxidative enzymes in mouse hepatocytes. Hepatocytes were isolated from wild-type mice following perfusion of livers with a collagenase-based buffer. Dispersed cells were plated on Matrigel and treated with rat GH (0.1, 1.0, or 10 microg/ml) or IGF-1 (0.5, 5.0, or 50 nM) for 24 hr. Hepatocytes were recovered and protein was extracted for immunoblotting and enzyme activity assays of catalase (CAT), glutathione peroxidase (GPX), and manganese superoxide dismutase (MnSOD). A 41% and 27% decrease in catalase activity was detected in cells treated with GH, whereas IGF-1 reduced CAT activity levels to a greater extent than GH (P < 0.0001). The activity and protein levels of GPX were also significantly depressed in cells treated with GH, whereas activity alone was decreased in cells treated with IGF-1 (P < 0.04). GH significantly suppressed MnSOD levels by 40% and 66% in 1.0 and 0.1 microg/ml concentrations, respectively. Similarly, IGF-1 decreased MnSOD protein levels (5 nM; P < 0.05). These results suggest that GH and IGF-1 may decrease the ability of hepatocytes to counter oxidative stress. In addition, these experiments provide an explanation for the differing antioxidative defense capacity of GH-deficient versus GH-overexpressing mice, and they suggest that GH is directly involved in antioxidant regulation and the aging process.     

Effects of growth hormone on cognitive function

Whether growth hormone deficiency (GHD) and/or treatment in childhood and adolescence influences cognitive outcome in children with GHD or girls with Turner syndrome (TS) is controversial. Previous studies also suggest that quality of life (QoL) is reduced in adults with GHD, particularly in the areas of social isolation and fatigue. Baseline QoL scores were significantly lower in patients with GHD than in the general population of the same age, gender, and nationality. Unfortunately, few data are available describing QoL in children with GHD. TS is a genetic disorder characterized by short stature, gonadal dysgenesis, and a particular neurocognitive profile of normally developed language abilities (particularly verbal intelligence quotients) and impaired visual-spatial and/or visual-perceptual abilities. This study evaluated the effects of GH treatment on neurocognitive function in girls with TS who were enrolled in a long-term, double-blind, placebo-controlled trial of the effects of GH treatment on final adult height. Treatment duration ranged from 1 to 7 years. The major result of this study was the absence of GH treatment effects on cognitive function in girls with TS. GHD and/or treatment in childhood and adulthood influences cognitive and/or QoL outcomes in some but not all studies. This study did not support a role for GH in influencing the characteristic nonverbal neurocognitive deficits associated with TS. However, evaluation of QoL should be a part of the routine clinical management of patients with GHD or TS.     

Quantitative analysis of insulin-like growth factor-modulated proteolysis of insulin-like growth factor binding protein-4 and -5 by pregnancy-associated plasma protein-A

The metzincin metalloproteinase pregnancy-associated plasma protein-A (PAPP-A, pappalysin-1, EC 3.4.24.79) specifically cleaves insulin-like growth factor binding protein (IGFBP)-4 and -5. Regulation of insulin-like growth factor (IGF) bioavailability through cleavage of these inhibitory binding proteins is an important mechanism for the control of growth and development of vertebrate cells. Although proteolysis of IGFBP-4 and -5 by PAPP-A has been extensively studied in many systems, quantitative analyses have been lacking. We have characterized the cleavage of its natural substrates, IGFBP-4 and -5, in the absence and presence of IGF-I or -II and determined the kinetic parameters (Km and kcat) for the different combinations of IGFBP and IGF. The rate of IGFBP-4 proteolysis is dramatically increased upon addition of IGF-I or -II. Kinetic analysis revealed that IGF-II was a more potent activator of IGFBP-4 proteolysis than IGF-I. Proteolysis of IGFBP-5 is slightly inhibited by IGF, and we find that IGF-I and -II display a similar degree of inhibition of IGFBP-5 cleavage. We show that the mechanism of IGF-modulated proteolysis of IGFBP-4 and -5 involves changes in both the recognition of substrate (Km) and the turnover rate (kcat). In addition, we have devised a novel method of revealing potential consequences of substrate modification for kinetic analysis, and we have used this method to establish that there is no apparent difference in the behavior of radiolabeled IGFBP-4 and -5 compared to the behavior of the unmodified protein substrates. We also propose experimental conditions for the proper analysis of IGFBP proteolysis, and we demonstrate their usefulness by quantitatively evaluating the effect of inhibitory compounds on the rate of proteolysis. Finally, we have compared PAPP-A to other proteinases thought to have IGFBP-4 or -5 as a substrate. This emphasizes the potential of PAPP-A to specifically and efficiently function as a regulator in the IGF system.     

Effects of growth hormone on skeletal muscle

Human growth hormone (GH) is widely abused as a performance-enhancing anabolic drug by athletes and bodybuilders. However, the effects of GH on skeletal muscle mass, strength and fibre composition remain unclear. We therefore summarize in the following the current knowledge on the physiological role of GH in the regulation of skeletal muscle growth and function and evaluate its potential therapeutic potency as a muscle anabolic hormone. In states of GH deficiency, reduced muscle mass and strength are characteristic findings which can be reversed successfully by the supplementation of GH. In contrast, the currently available data suggest that GH administration alone or in combination with strength exercise has little, if any, effect on muscle volume, strength and fibre composition in non-GH-deficient healthy young individuals. This assumption is supported by the lack of evidence for a significant performance-enhancing effect of GH in athletes. However, further studies will be necessary to define patient populations which might benefit from GH treatment like frail elderly individuals in whom a GH-induced change into a more youthful muscle fibre composition has been reported.     

Effects of inhibitors of signal transduction pathways on transforming growth factor B1 and osteogenic protein-1-induced insulinlike growth factor binding protein-3 expression in human bone cells

Signal transduction initiated by TGFB1 and OP-1 was studied in MG63 human osteosarcoma cells and in normal human bone cells (HBCs) in the presence of inhibitors of signal transduction events, using insulinlike growth factor binding protein-3 (IGFBP-3) production as an end point. Treatment of serum-free MG63 cells and normal HBCs with TGFB1 increased IGFBP-3 protein level several fold in the conditioned medium. This effect of TGFB1 was mediated by increased de novo synthesis because mRNA level increased to the same extent as protein level and TGFB1 treatment had very little effect on IGFBP-3 protease activity. The stimulatory effect of TGFB1 on IGFBP-3 production was inhibited in a dose-dependent manner by pretreatment with staurosporine, a protein kinase C inhibitor, or with vanadate, a phosphotyrosyl protein phosphatase inhibitor in both MG63 cells and normal HBCs. In addition, pretreatment with okadoic acid, an inhibitor of serine/threonine protein phosphatase, counteracted TGFB1 induction of IGFBP-3 production. Interestingly, pretreatment of MG63 cells or HBCs with staurosporine, vanadate, or okadoic acid augmented OP-1 stimulation of IGFBP-3 production. Staurosporine- or vanadate-induced changes in IGFBP-3 protein levels in the presence of TGFB1 and OP-1 were associated with corresponding changes in IGFBP-3 mRNA levels in MG63 cells. These findings are consistent with the hypothesis that TGFB1 and OP-1 increase IGFBP-3 expression via distinct intracellular signal transduction pathways. J. Cell. Physiol. 173:28–35, 1997. © 1997 Wiley-Liss, Inc.     

Effects of human growth hormone on erythrocyte insulin binding in growth hormone deficient children

In this paper, the effect of acute human growth hormone (GH) administration on erythrocyte insulin binding in GH deficient children (N = 6) was studied. Following GH (0.25 U/kg) administration, the blood levels of GH peaked within 4 to 8 h and returned to basal levels 24 h later. However, the changes in somatomedin activity, free fatty acid (FFA), urea, blood glucose and 125I-insulin binding to erythrocyte were observed around 24 h following the injection, and there was a converse relationship between maximum percent 125I-insulin binding (IBmax) and FFA (P less than 0.02). By Scatchard analysis it was found that the decrease in IBmax is mainly due to the change in the number of insulin receptors. These results suggest that GH may possibly affect the insulin binding to erythrocyte indirectly through metabolic changes as a result of hormonal changes in GH deficient children.     

Effects of insulin-like growth factor-1/binding protein-3 complex on muscle atrophy in rats

Muscle atrophy and wasting is a serious problem that occurs in patients with prolonged debilitating illness, burn injury, spinal injury, as well as with space flight. Current treatment for such atrophy, which often relies on nutritional supplementation and physical therapy, is of limited value in preventing the muscle wasting that occurs. Considerable recent attention has focused on the use of anabolic growth factors such as insulin-like growth factor (IGF-1) in preventing muscle atrophy during limb disuse or with various catabolic conditions. However, potential side effects such as hypoglycemia appear to be limiting factors in the usefulness of IGF-1 for clinical treatment of muscle wasting conditions. The formulation of IGF-1 used in this study (IGF-1/BP3) is already bound to its endogenous-binding protein (BP3) and, as a result, has a greater specificity of action and significantly less hypoglycemic effect. Using a rat model of hind limb suspension (HLS) for 10 days, we induced marked muscle atrophy that was accompanied by enhanced muscle proteolysis and reduced muscle protein content. When HLS rats were treated with IGF-1/BP3 (50 mg/kg, b.i.d.), they retained greater body and muscle mass. Muscle protein degradation was significantly reduced and muscle protein content was preserved. The rate of protein synthesis, although somewhat reduced in HLS muscle, was not significantly elevated by IGF-1/BP3 treatment. Volume density of HLS-treated muscles were increased compared to untreated HLS rats and the actual number of fibers per area of muscle was likewise increased. The results of the current study suggest that IGF-1/BP3 might be useful for inhibiting muscle proteolysis in catabolic conditions and thus preserving muscle protein content and mass.     

Interaction of insulin-like growth factor binding protein-3 with latent transforming growth factor-beta binding protein-1

Insulin-like growth factor binding protein-3 (IGFBP-3) inhibits the replication and promotes apoptosis in various cell lines in an IGF-independent manner. We utilized a yeast two-hybrid system to identify binding partners for IGFBP-3 in a mouse embryo cDNA library. A partial cDNA encoding mouse latent transforming growth factor beta (TGF-) binding protein-1 (LTBP-1) was identified. This cDNA encoded a mouse LTBP-1 mRNA fragment corresponding to amino acid residues 1160–1712. Analysis of C-terminal deleted mutants indicated that the IGFBP-3 interacting domain resides in the 552 residue C-terminal fragment encoding amino acids 831–1383. The interaction of IGFBP-3 with recombinant human LTBP-1 immobilized on nitrocellulose was also demonstrated. Neither binding of IGF-I to IGFBP-3 nor binding of latency associated protein (LAP) with LTBP-1 inhibited the interaction of IGFBP-3 with LTBP-1. Furthermore the large latent complex, ¹²⁵I-TGF-/LAP/LTBP-1 was able to bind to immobilized IGFBP-3. These data demonstrate that IGFBP-3 can bind to LTBP-1 and provide a potential mechanism whereby IGFBP-3 can interact with the TGF- system.     

Effects of cholinergic drugs on growth hormone release

Acetylcholine and the cholinergic agonists, pilocarpine and physostigmine, increased GH release $\text{in}$$\text{vivo}$. The increase in GH release by pilocarpine was reversed by concurrent administration of the cholinergic receptor blocker, atropine, whereas atropine alone did not alter serum GH concentrations. Cholinergic stimulation of GH release appears to be partially mediated through a catecholaminergic system since the response was partially inhibited by pimozide, a dopamine receptor blocker, or phentolamine, an α-adrenergic receptor blocker. The cholinergic system may function physiologically to help regulate GH release.     

Pregnancy-associated murine protein-1 (PAMP-1) in wild rodents.

The presence of pregnancy-associated murine protein-1 (PAMP-1) in a number of wild rodent species was examined using tandem crossed immunoelectrophoresis and anti PAMP-1 antisera raised in rabbits. PAMP-1 is a protein present in female laboratory mice and rats; analogues were readily detected in the following species: Musculus domesticus, M. musculus, Apodemus sylvaticus, A. agrarius, A. flavicollis, and Microtus arvalus. Immunologically, cross-reacting proteins could not be detected in the laboratory hamster and laboratory guinea pig. Maternal PAMP-1 serum levels were recorded throughout pregnancy in the two species of house mouse: M. domesticus and M. musculus. Both proteins exhibited maximum concentration at day 11 of gestation. During early pregnancy, M. domesticus females had significantly higher serum levels than those of M. musculus females.     

Effects of growth hormone administration on wool growth in merino sheep

The effects of daily administration of 10 mg of highly purified ovine growth hormone (GH) for a period of 4 weeks on wool growth have been measured in 12 Merino ewes fed either a calculated maintenance energy intake or 1.6 times this amount (six on each ration). Concentrations of hormones, glucose, urea, alpha-amino N and amino acids in the blood were monitored and faeces and urine collected for measurement of nitrogen balance. Wool growth rate decreased by 20% during the 4 weeks of GH treatment in sheep fed the high energy diet, largely because of reduced wool fibre diameter. This was followed by restoration of normal growth and then an increase of up to 20% above control levels, a response which persisted for 12 weeks following cessation of GH administration, and which was due to increases in both fibre length and diameter. GH administration caused marked increases in plasma concentrations of GH, insulin and somatomedin C, glucose and free fatty acids, all of which returned to basal levels following cessation of GH administration. No consistent changes in plasma concentration of T3, T4, cortisol, prolactin or alpha amino N were detected. Plasma urea and methionine levels decreased during GH treatment and returned to, or were raised above, basal levels after the GH treatment period. GH injection also resulted in a net retention of N during treatment, followed by a transient period of net N loss. The GH-induced changes in wool growth may be caused by a change in the partitioning of amino acids between the muscle mass and the skin. No other contributing factor(s) were identified.