Role Of GH Research Articles

Bovine growth hormone transgenic mice display alterations in locomotor activity and brain monoamine neurochemistry.

Recent clinical and experimental data indicate a role for GH in mechanisms related to anhedonia/hedonia, psychic energy, and reward. In the present study we have investigated whether bovine GH (bGH) transgenic mice and nontransgenic controls differ in spontaneous locomotor activity, a behavioral response related to brain dopamine (DA) and reward mechanisms, as well as in locomotor activity response to drugs of abuse known to interfere with brain DA systems. The animals were tested for locomotor activity once a week for 4 weeks. When first exposed to the test apparatus, bGH transgenic animals displayed significantly more locomotor activity than controls during the entire registration period (1 h). One week later, after acute pretreatment with saline, the two groups did not differ in locomotor activity, whereas at the third test occasion, bGH mice were significantly more stimulated by d-amphetamine (1 mg/kg, ip) than controls. At the fourth test, a tendency for a larger locomotor stimulatory effect of ethanol (2.5 g/kg, ip) was observed in bGH transgenic mice. bGH mice displayed increased tissue levels of serotonin and 5-hydroxyindoleacetic acid in several brain regions, decreased DA levels in the brain stem, and decreased levels of the DA metabolite 3,4-dihydroxyphenylacetic acid in the mesencephalon and diencephalon, compared with controls. In conclusion, bGH mice display more spontaneous locomotor activity than nontransgenic controls in a novel environment and possibly also a disturbed habituation process. The finding that bGH mice were also more sensitive to d-amphetamine-induced locomotor activity may suggest that the behavioral differences observed are related to differences in brain DA systems, indicating a hyperresponsiveness of these systems in bGH transgenic mice. These findings may constitute a neurochemical basis for the reported psychic effects of GH in humans.

Lactogenic hormone-inducible phosphorylation and gamma-activated site-binding activities of Stat5b in primary rat Leydig cells and MA-10 mouse Leydig tumor cells.

The signal transducer and activator of transcription Stat5b has been implicated in signal transduction pathways for a number of cytokines and growth factors, including GH and PRL. Although these lactogenic hormones have the potential to enhance gonadotropin-induced steroidogenesis, the role of GH and PRL in the testis has long been and remains the subject of controversy. In this report we provide, to our knowledge, the first evidence of Stat5b protein expression in the testis and characterize the activation of Stat5b by these lactogenic hormones in primary rat progenitor, immature and adult Leydig cells, and mouse MA-10 Leydig tumor cells. In MA-10 cells, both GH and PRL mediate tyrosine phosphorylation of Janus kinase (JAK) 2 and Stat5b and induce DNA-binding activity of Stat5b. GH enhances both PIE (PRL-inducible element) and Fc gammaRI gamma-activated sites (GAS), but PRL modulates only PIE GAS. In primary Leydig cells isolated from 18-day-old rats, GH, but not PRL, activates cytoplasmic Stat5b and induces the binding of translocated nuclear Stat5b to GAS elements. Although Stat5b protein is expressed in both Percoll- and elutriator-purified adult rat Leydig cells, neither GH nor PRL treatment results in Stat5b-DNA binding. Our studies indicate that the MA-10 cell has the capacity to bind both GH and PRL and provides a useful model system with which to study the distinct testicular roles of these hormones. Moreover, our findings suggest that progenitor and immature Leydig cells are functional targets for GH in the immature rat, suggestive of a role for GH-Stat5b in testicular development. Our data indicate that lactogenic hormone-inducible transcriptional activation may target distinct gene expression in a signaling cascade(s) involving Stat5b but also imply coordinate control by multiple Leydig cell factors.

Effects of GH and IGF-I administration on GHRH and somatostatin mRNA levels: I. A study on ad libitum fed and starved adult male rats.

The individual role played by GH and IGF-I in the regulation of hypothalamic GHRH and SRIF gene expression is still object of debate. We have investigated the effect of exogenously administered recombinant hGH (rhGH) and recombinant hIGF-I (rhIGF-I) in ad libitum fed control and starved rats, the latter an animal model which is characterized by low circulating levels of endogenous GH and IGF-I. Adult male rats were fed ad libitum (C) or food-deprived (S) for 72 hours; rats in either C or S groups were treated with systemic administration of rhGH and rhIGF-I for 3 days. GHRH, SRIF and GH mRNA levels were evaluated by Northern and slot blot hybridization. Administration of rhGH (250 micrograms/kg/twice daily, sc) induced a significant inhibition of GHRH and a significant stimulation of SRIF mRNA levels in C rats; GH treatment was, however, ineffective on both neuropeptide mRNA levels in the S group. Continuous infusion of rhIGF-I (300 micrograms/kg/day, sc) induced a significant increase of SRIF levels in both C and S rats but did not modify GHRH mRNA levels in either group. In the pituitary, GH mRNA levels followed a pattern very similar to that of GHRH. These results provide evidence for a direct role of GH in the inhibition of GHRH mRNA levels; IGF-I appears more involved in the direct stimulation of SRIF mRNA levels.

Non-growth hormone (GH) deficient short children--what role for GH?

Non-growth hormone (GH) deficient short children--what role for GH?

The role of growth hormone in fetal mouse reproductive tract differentiation.

Although GH plays a key role in postnatal growth and differentiation, its role in fetal differentiation is not clear at the present. The aim of the present study was to investigate whether GH plays a role in fetal sexual differentiation, and we used in vitro organ culture assay of sexual differentiation to determine this. The results showed that anti-rGH antibody blocked Wolffian duct differentiation specifically in the presence of fetal testes. Exogenous GH supplemented in the above experiment reversed the blocking effect of anti-GH. Among the other related products, insulin-like growth factor I was highly effective in reversing the anti-GH effect, insulin-like growth factor II was partially effective, but PRL was unable to reverse the anti-GH effect. GH itself was found to produce some masculinizing effect, as demonstrated by its ability to stabilize the Wolffian duct in female fetuses. The role of GH was further demonstrated by the observation that GH-immunoreactive material of the size of authentic GH was detected in the 18-day fetal reproductive tract, and the concentration of this material increased in response to progression of sexual differentiation. Determination of androgen-binding activity using Scatchard analysis on the cells isolated from the 18-day fetal reproductive tract indicated that androgen-binding activity increased after GH treatment of the cells. Thus, it may be concluded that GH influences male sexual differentiation and alters the androgen-binding activity of the fetal reproductive tract.

Evidence for a direct action of GH on haemopoietic cells of a marine fish, the gilthead sea bream (Sparus aurata)

Receptors for GH were characterized in the head kidney of gilthead sea bream (Sparus aurata), using radioiodinated and biotinylated ligands. The specific binding of radiolabelled recombinant gilthead sea bream GH (rsbGH) to head kidney membrane preparations was dependent on membrane concentration. Salmon prolactin, salmon gonadotrophin and carp gonadotrophin did not compete for 125I-labelled rsbGH-binding sites. Unlabelled rsbGH competitively displaced 125I-labelled rsbGH bound to head kidney membranes. Scatchard plots were always linear, denoting the presence of a single class of binding sites. The binding affinity (Ka = 2.7 x 10(9) M-1) was equivalent to that found in liver membrane preparations, but the binding capacity (2.5 +/- 0.30 fmol/mg protein) was 50- to 75-fold lower. To identify the cells which express the GH receptor, head kidney smears were incubated with biotinylated rsbGH, followed by incubation with an avidin-biotin complex conjugated to alkaline phosphatase. The reaction with the new-fuchsin substrate gave a red precipitate, showing a specific and intense labelling in erythroblasts, polychromatophilic erythroblasts and myeloblasts. Noticeable binding was observed in myelocytes and immature granulocytes, tending to disappear at the latter stages of granulocyte maturation. Light but appreciable binding was also observed in monocytes, lymphocytes and acidophilic erythroblasts, whereas it was completely absent in proerythrocytes and erythrocytes. The proliferative action of rsbGH and recombinant human IGF-I on in vitro cultures of head kidney cells was demonstrated by a 5-bromo-2'-deoxy-uridine immunoassay. To our knowledge, this is the first report that provides suitable evidence for a role of GH as a haemopoietic growth and differentiation factor in lower vertebrate species.

Growth hormone and longitudinal bone growth in vivo: short-term effect of a growth hormone antiserum.

The control of longitudinal growth is poorly understood but GH is considered to be one of the major hormones regulating postnatal growth. However, there is dispute as to whether it has a direct or indirect action. To study the role of GH we used a polyclonal antiserum to rat GH and investigated changes in cell proliferation and enzyme activities associated with bone formation and resorption during longitudinal growth. IGF-I levels were measured by two independent RIAs, DNA synthesis by bromodeoxyuridine incorporation followed by immunocytochemistry and enzyme activities were quantified in situ by microdensitometry. After 1 day the percentage of chondrocytes undergoing DNA synthesis within the proliferative zone was reduced but no other parameters were affected. By day 4 the labelling index was the same as in pair-fed animals but the number of chondrocytes synthesising DNA was reduced as was the total width of the growth plate and that of the proliferative zone. Alkaline phosphatase (associated with mineralisation) was unchanged but glucose 6-phosphate dehydrogenase activity (associated with cell proliferation) was decreased. Osteoclastic tartrate-resistant acid phosphatase activity (associated with bone resorption) was also significantly reduced. Similar changes were apparent after 10 days. At no time was the circulating level of IGF-I decreased. These data suggest that, during longitudinal growth, GH affects the number of proliferating chondrocytes but not the percentage of cells undergoing DNA synthesis, indicating that its primary role may be on the commitment of prechondrocytes to a proliferative state. Furthermore, while GH does not seem to have any effect on skeletal mineralisation it may stimulate osteoclastic resorption of the primary spongiosa.

Control of the thymic microenvironment by growth hormone/insulin-like growth factor-I-mediated circuits.

The thymus gland is a central lymphoid organ in which bone marrow-derived T cell precursors undergo maturation, eventually leading to the migration of positively selected thymocytes to the T-dependent areas of peripheral lymphoid organs. This process occurs under the influence of the thymic microenvironment, by means of secretory polypeptides and cell-cell contacts. The thymic microenvironment is a tridimensional cellular network composed of epithelial cells (its major component), macrophages, dendritic cells, fibroblasts and extracellular matrix elements. The epithelial reticulum is a heterogeneous tissue, in which a particular lymphoepithelial structure has been isolated in vitro: the thymic nurse cell complex, which possibly creates particular microenvironmental conditions for thymocyte differentiation. Additionally, thymic nurse cells are useful tools to study mechanisms involved in intrathymic T cell migration, including neuroendocrine influences. Previous data showed that thymic hormonal function can be modulated by hormones and neuropeptides, including growth hormone. Interestingly, GH acts pleiotropically on the thymic epithelium increasing cell growth and expression of extracellular matrix ligands and receptors, the latter resulting in an enhancement of thymocyte adhesion to the epithelial cells and thymocyte release from thymic nurse cells. The role of GH on thymus development is further stressed by the findings obtained with GH-deficient dwarf mice. Besides the precocious decline in serum thymulin found in these animals, a progressive thymic hypoplasia occurs, with decreased numbers of CD4+CD8+thymocytes, both defects being largely restored by long-term GH treatment. The effects of GH in the thymus are apparently mediated by IGF-1. Enhancement of thymulin secretion induced by GH, as well as the stimulation of thymocyte adhesion to thymic epithelial cells can be prevented in vitro by treatment with antibodies for IGF-I or IGF-I receptor. Moreover, in both systems IGF-I alone can yield similar effects. Also, the enhanced concanavalin-A mitogenic response and IL-6 production by thymocytes observed in GH-treated mice can be detected in animals treated with IGF-I. Lastly, mouse substrains selected for high or low IGF-I circulating levels exhibited differential thymus developmental patterns correlating with IGF-I levels. A further conceptual aspect concerning the GH-IGF-I-mediated control of thymus physiology is the recent demonstration of an intrathymic production of these molecules, leading to the hypothesis that, in addition to the classical endocrine pathway, GH-IGF-I-mediated paracrine and autocrine pathways may also be implicated in the control of thymus physiology. In any case, such control is exerted pleiotropically, with modulation in the expression of several genes in different cell types of the organ. In this respect, it is exciting to imagine a role of GH-IGF-I loops in shaping the intrathymically generated T cell repertoire.

Serum growth hormone-binding protein is unchanged in adult panhypopituitarism.

The role of GH in regulating GH-binding protein (GHBP) and GH receptor concentrations in humans is not clear. Studies performed mostly in children and on a minor scale in adults are somewhat controversial. The key question as to whether GHBP levels are altered in hypopituitarism before GH treatment is instituted remains unanswered. In this study, we have selected a severely GH-deficient group of adult patients with panhypopituitarism, acquired as a result of surgery and irradiation of hypothalamic-pituitary tumors, to evaluate the GHBP/receptor status by measuring GHBP activity in plasma. Twenty panhypopituitary patients (8 males and 12 females; age range, 20-74 yr) and 20 age (22-68 yr)- and sex-matched normal subjects were studied. GH deficiency was confirmed by insulin-induced hypoglycemia and arginine infusion tests; the peak GH response was less than 2 micrograms/L. Plasma insulin-like growth factor-I levels were below or in the low normal range (mean +/- SD, 88.3 +/- 53.6 micrograms/L) and were significantly different from insulin-like growth factor-I (IGF-I) levels in the normal group (mean +/- SD, 189 +/- 49.8 micrograms/L; P < 0.01). Plasma GHBP activity, measured using a GH-binding/gel chromatography assay, showed similar values in the GH-deficient group (mean +/- SD, 14.1 +/- 3.83%) and the control group (mean +/- SD, 13.7 +/- 3.79%), with no statistically significant difference. Neither the intra- nor intergroup comparison of GHBP levels according to age and sex showed statistically significant differences or age trends. In the light of these data and considering that GHBP activity in plasma probably reflects the GH receptor status in tissues, we may assume that the GH receptor was unaffected by chronic GH deficiency. These findings also support the previously reported concept that the GHBP/receptor level is a relatively fixed determinant of growth, established individually and independently of GH secretory status in early life, perhaps on a genetic basis.

Role of GH and IGF-I in the regulation of IGF-I, IGF-I receptor and IGF binding protein gene expression in the rat spleen

To characterize the expression of the IGF-I system in the spleen and its role in spleen growth, we have studied the effect of hypophysectomy and the action of either GH or IGF-I treatment on the expression of several components of the IGF system in the rat. Female Sprague-Dawley rats were hypophysectomized (Hx) on postnatal day 50, and five animals each received twice-daily sc injections of saline, bovine GH (bGH; 84 μg/animal/day), or recombinant human IGF-I (rhIGF-I; 125 μg/animal/day) for 11 days. Compared to sham-operated controls, Hx animals exhibited a reduction in both body ( 192.6 ± 5.6 g (mean ± S.E.M.) vs. 268.6 ± 6.0 g ; P<0.001) and spleen weights ( 0.42 ± 0.03 g vs. 0.84 ± 0.06 g ; P<0.001). The reduction in body and spleen weights in Hx animals was partially prevented by both bGH and rhIGF-I. Body weights were 234.2 ± 5.3 g ( P<0.001) after bGH and 213.8 ± 6.3 g ( P<0.05) after rhIGF-I. Spleen weights were 0.56 ± 0.048 after bGH P<0.01 and 0.53 ± 0.05 g after rhIGF-I ( P<0.05). Serum GH and IGF-I levels were markedly reduced in Hx animals and bGH partially maintained IGF-I levels. Hypophysectomy reduced spleen IGF-I mRNA levels ( 30.6 ± 7.5% of control values; P<0.05) and this reduction was prevented by bGH ( 96.6 ± 24.2%; NS) but not by rhIGF-I ( 39.9 ± 5.0% NS vs. Hx). There were no changes in GH receptor or IGF-I receptor mRNA levels in Hx or bGH or rhIGF-I-treated animals. When IGF-I binding protein (IGFBP) mRNA levels were studied under these conditions, we found that IGFBP-1 mRNA was not detected in spleen; IGFBP-2 mRNA levels were reduced in Hx rats ( 67.9 ± 7.4% of control values, P<0.05) and bGH treatment prevented this reduction ( 95.5 ± 12.2%, NS). IGFBP-3 mRNA levels were not affected by hypophysectomy or by bGH treatment, but were reduced in rhIGF-treated rats ( 69.6 ± 3.0%, P<0.05). On the other hand, IGFBP-4 mRNA levels were increased in Hx rats ( 136.4 ± 15.9% of control values, P<0.05) and bGH treatment prevented this increase. These data demonstrate that several components of the IGF system are expressed in rat spleen and that following hypophysectomy, there is a reduction in spleen weight that correlates with a reduction in local IGF-I gene expression and, presumably, a reduction in IGF-I bioavailability due to a decrease in IGFBP-2 and an increase in IGFBP-4 gene expression.

Effects of a polyclonal antiserum to rat growth hormone on circulating insulin-like growth factor (IGF)-I and IGF-binding protein concentrations and the growth of muscle and bone.

A polyclonal antiserum to rat GH (anti-rGH) injected into rats for 3 or 8 weeks markedly reduced the weight, total protein and RNA content of muscles of the hind limb. These effects were prevented when bovine GH (bGH) was administered simultaneously. In a second experiment, the effects of 8 weeks of treatment with anti-rGH on the growth of the whole body, muscle and bone were investigated. Body weights of rats were decreased by 58% by treatment with anti-rGH; muscle weights were reduced by slightly more than the decrease in body weight (by 64%, 65% and 61% respectively for plantaris, soleus and gastrocnemius). The weight of the tibia was decreased by 54%, its length was decreased by 23%, cortical width and overall width were reduced by 26% and 18% respectively, suggesting a possible role for GH in osteoclastic activity. Serum total insulin-like growth factor-I (IGF-I) concentrations were decreased by 80-90% in both experiments by anti-rGH; these changes were prevented in the first experiment by concurrent treatment with anti-rGH and bGH. The serum IGF-binding protein-3 (IGFBP-3) concentration was also decreased by anti-rGH in experiment 1 (by 86%); the response of the 28-32 kDa IGFBPs was smaller (-35%), and was restored to control values by simultaneous injection of bGH. Western immunoblotting using an antiserum to IGFBP-2 showed that there was a marked decrease from neonatal to adult stages which was independent of anti-rGH treatment. This clearly demonstrated a dissociation of the reciprocal relationship supposed to exist between IGFBPs-2 and -3. The 24 kDa IGFBP-4 was unaffected by anti-rGH but replacement therapy with bGH doubled its concentration. Although the effects on body and muscle weight were prevented when rats were given anti-rGH and bGH simultaneously, the possibility of mediation by other hormones cannot be precluded.

Insulin-like growth factor-I regulation of renal 25-hydroxyvitamin D-1-hydroxylase activity.

Controversy exists regarding the role of GH and insulin-like growth factor-I (IGF-I) in the modulation of calcitriol production. While their administration increases serum levels of 1,25-dihydroxyvitamin D, the mechanism remains unknown. Investigations have also implicated GH as a causal factor underlying renal 25-hydroxyvitamin D-1 alpha-hydroxylase activity [1(OH)ase] secondary to phosphate depletion. Thus, we investigated the effects of IGF-I on 1(OH)ase and the relationships between these actions and those of phosphate depletion. Our studies indicate that IGF-I administration to normal mice results in a dose-dependent (0-10 micrograms/h) increase in 1(OH)ase with maximum effects evident after 24 h, independent of changes in serum calcium, phosphorus, and glucose levels. Similarly, hormone administration to phosphate-depleted mice increases enzyme activity (6.06 +/- 0.96 vs. 13.97 +/- 1.67 fmol/mg.min) but to a level significantly greater than that achieved in normals (2.72 +/- 0.4 vs. 5.01 +/- 0.56 fmol/mg.min). Furthermore, the response represents an additive increment of the effects elicited by maximum doses of IGF-I and phosphate depletion, suggesting that the hormone- and phosphate-dependent enzyme stimulation occur by different mechanisms. Thus, our data establish that IGF-I stimulates renal 1(OH)ase activity in a time- and dose-dependent fashion. However, they do not support the hypothesis that IGF-I modulates the effects of phosphate-depletion on 1(OH)ase activity. Regardless, the documentation that IGF-I stimulates enzyme function provides an explanation for many observed physiological states associated with concomitant alterations of hormone levels and calcitriol production.

Characterization of transgenic mice lineages. I. Overexpression of hGH causes the formation of liver intranuclear pseudoinclusion bodies and renal and hepatic injury

AbstractThe expression of the human Growth Hormone (hGH) in transgenic mice is accompanied by health afflictions and structural changes in several organs and tissues. A macroscopic and light microscopic study was conducted to examine the morphological alterations related to the exposure of endogenously expressed hGH. Although many transgenic mice were under cachectic status, their body and organ weights were significantly higher than those of the controls matched. Kidney, liver and female genital organs were massively damaged.The most common findings in these organs were glomeruli hyalinosis and glomerulosclerosis in the kidneys accompanied by a varying degree of segmental sclerosis and necrosis. Tubuli dilatation was observed with hyaline casts and atrophied epithelium as well as interstitial fibrosis with mononuclear infiltrate. In the liver the hepatocytes were pleomorphic and exhibited vacuolar degeneration. Intranuclear lipid droplets were also found. Individual necrosis, mononuclear infiltrate and nuclear pseudoinclusions were also detected. The uterus exhibited cystic glandular hyperplasia and ovaries were observed to have several, large foamy cells as well as stromal infiltration of adipose cells. Other changes were found in the heart, the spleen and the salivary glands. A possible role of GH and/or the Insulin‐like Growth Factor‐I is suggested according to the morphological changes found. The similarity to kidney changes with human diabetic glomerulosclerosis was noticed.

Investigation of the mechanism of action of growth hormone in stimulating lactation in the rat.

The role of GH was examined using an antiserum to rat GH (anti-rGH). When administered to lactating rats on day 2 of lactation it was without effect, whereas bromocriptine markedly suppressed milk production, with no additional effect of combined treatment. On day 6 of lactation, treatment with anti-rGH was also without effect, whilst bromocriptine again suppressed milk production. Combined treatment, however, suppressed milk synthesis completely, suggesting that GH was capable of maintaining about 50% of normal milk yield in the absence of prolactin at day 6 of lactation. By day 14 of lactation, anti-rGH treatment alone was capable of decreasing milk yield by about 20%, and again milk secretion only stopped completely when GH and prolactin were suppressed. These data suggest that the role of GH in supporting lactation increases as lactation progresses. The effects of GH in stimulating growth and in increasing milk yield in ruminants have been proposed to be mediated via insulin-like growth factor-I (IGF-I). In rats treated with anti-rGH, both IGF-I and IGF-II were decreased in serum. The concentration of the major IGF-binding protein (IGFBP-3) was not, however, affected by inhibition of GH or prolactin individually, but was decreased in animals treated with bromocriptine and anti-rGH. In animals given both bromocriptine and anti-rGH, concurrent treatment with recombinant bovine GH maintained milk yield at 50% of control values and normalized serum IGF-I, IGF-II and IGFBP-3 concentrations. By contrast, concurrent treatment with IGF-I or IGF-II, despite normalizing their respective concentrations in serum, failed to affect milk yield.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of the growth hormone receptor, identified by immunocytochemistry, in second trimester human fetal tissues and in placenta throughout gestation.

Pituitary GH secretion appears largely unnecessary for the attainment of normal birth size in many species, including man. This is believed to be due to an immaturity and/or an absence of GH receptors in many fetal tissues. However, in vitro studies using late first trimester human fetal tissues have demonstrated mitogenic actions of GH on liver and stimulation of insulin biosynthesis in pancreas. To resolve this discrepancy, we have employed immunocytochemistry to identify the presence and distribution of GH receptors in various human fetal tissues. Fetuses of 14-16 weeks gestation were obtained after therapeutic abortion, tissues were fixed, and immunocytochemistry was performed using monoclonal antibodies against purified rat or rabbit GH receptor. The specificity of staining was confirmed by preabsorption of the antibodies with 1) adult rat liver membranes or 2) human fetal liver membranes, both of which possess specific GH-binding sites, or 3) human fetal skeletal muscle membranes, which do not specifically bind GH. Positive staining was seen in a subpopulation of liver parenchymal cells, in the ductal and endocrine tissue of pancreas, in the germinal layer of the epidermis and the deeper dermal layers of skin, and in the tubular epithelium of kidney. No immunopositive staining was seen in skeletal or cardiac muscle, epiphyseal growth plate, lung, intestine, or adrenal. Positive staining was present in the neuronal cell bodies of the cerebral cortex. GH receptor was also detectable as early as 8 weeks gestation in syncytial layers of the placenta and was maintained until term. Results demonstrate the presence of immunoreactive GH receptor/binding protein in some human fetal tissues early in development. In particular, these results would support a role for GH in the growth and function of liver and pancreas.

Growth hormone and insulin-like growth factor-I stimulate hormonal function and proliferation of thymic epithelial cells.

We have investigated the role of GH and insulin-like growth factor-I (IGF-I) in controlling the secretion of thymulin, a hormone produced by thymic epithelial cells (TEC). Thymulin plasma concentrations (mean +/- SD) were increased in 21 patients with acromegaly compared to those in 30 controls, as assessed by bioassay (4.24 +/- 0.97 vs. 2.67 +/- 0.87; P less than 0.001) and RIA (561 +/- 241 vs. 315 +/- 113 pg/L; P less than 0.01). Good correlations were observed between plasma levels of thymulin and IGF-I (P less than 0.001). In vitro experiments demonstrated that both recombinant human GH and IGF-I significantly increased thymulin production in culture supernatants of normal human TEC and a rat TEC line. In parallel, IGF-I also significantly stimulated the proliferation of human TEC, as measured by bromodeoxyuridine incorporation. Additionally, the stimulatory effect of GH on thymulin production was abrogated by both an anti-IGF-I antibody and an anti-IGF-I receptor antibody. These results support a role for GH and IGF-I in the control of thymic hormonal function in man and suggest that the effect of GH may be mediated by local secretion of IGF-I within the thymus.

Regulation of testicular insulin-like growth factor-I in pubertal growth hormone-deficient male rats

GH plays a major role in pubertal growth, effects mainly mediated by stimulation of insulin-like growth factor-I (IGF-I) production by the liver. However, the role of GH in the regulation of pubertal onset, spermatogenesis and fertility is still under debate. GH and FSH have, in addition, been implicated in the regulation of IGF-I production by Sertoli cells in a number of studies, although conflicting results have been reported. The interpretation of studies using GH-deficient mutant mice has been complicated by the presence of additional defects in the hypothalamic-pituitary-gonadal axis of these animals. We have therefore used GH-deficient mutant male rats with no other documented hormonal deficiencies to study the effect of GH administration on somatic and testicular development, circulating and testicular IGF-I concentrations and testicular histology. Body weights in GH-deficient rats substituted with GH were not significantly different from untreated or GH-treated normal rats and were significantly higher than body weights in untreated dwarf rats. Similarly, circulating IGF-I concentrations in GH-treated GH-deficient rats were not significantly different from those in untreated or GH-treated normal rats but were significantly higher than circulating IGF-I concentrations in untreated dwarf rats. No differences in testicular IGF-I concentrations were observed in any of the groups studied. Testicular weights remained low in both untreated and GH-treated GH-deficient animals compared with control animals but spermatogenesis was qualitatively and quantitatively normal in all groups at the end of the observation period.(ABSTRACT TRUNCATED AT 250 WORDS)

The Effects of Thyroid Hormone on Insulin-Like Growth Factor (IGF) and IGF-Binding Protein (IGFBP) Expression in the Neonatal Rat: Prolonged High Expression of IGFBP-2 in Methimazole- Induced Congenital Hypothyroidism*

In the rat a developmental switch in the serum insulin-like growth factor (IGF) and IGF-binding protein (IGFBP) profile takes place during the first 3 postnatal weeks. The fetal expression pattern of high IGF-II and IGFBP-2 is replaced by the adult pattern of low levels of IGF-II and IGFBP-2 and high levels of IGF-I and IGFBP-3. The regulatory mechanisms mediating these changes are unknown, but may include perinatal changes in endocrine function. To study the effects of thyroid function and the perinatal thyroid secretory burst on IGF and IGFBP expression, we established a rat model of congenital hypothyroidism, leading to marked postnatal growth retardation during the perinatal period. The hypothyroid animals lacked the steep rise in serum IGF-I levels normally occurring during the third week of life, showing only a modest rise to approximately 50% of control levels. The pattern of serum IGF-II decline in hypothyroid animals was slightly different from that in controls, with lower IGF-II levels during the second week of life and a slower decline down to the very low final levels. The hypothyroid pups continued to express high levels of IGFBP-2 up to the age of 19 days, while the control animals, after a slow initial decline, showed an abrupt fall of IGFBP-2 serum levels during the third week of life. Liver IGFBP-2 mRNA levels reflected the serum changes, with elevated IGFBP-2 mRNA in hypothyroid animals. The expression of other IGFBPs did not differ from that in the control group. At the age of 18 days, serum GH levels in the hypothyroid animals were approximately one third of control GH levels, which suggests a role for GH as a possible mediator of thyroid hormone actions on the IGF system. The changes in growth parameters and in the IGF and IGFBP profile of hypothyroid pups could be abolished by thyroid hormone replacement from birth. We conclude that thyroid hormone is, directly or indirectly, essential for some of the neonatal changes in IGF and IGFBP profiles.

An Antisense Oligodeoxynucleotide to Growth Hormone Messenger Ribonucleic Acid Inhibits Lymphocyte Proliferation*

The role of GH in lymphocyte proliferation was studied by examining the effect of an antisense oligodeoxynucleotide (ODN) complementary to GH mRNA. The results of these studies showed that antisense GH ODN treatment inhibits lymphocyte production of immunoreactive GH (irGH). Lymphocytes treated with the GH antisense ODN produced less irGH than did lymphocytes treated with control sense GH ODN. Antisense GH ODN-mediated inhibition of irGH production resulted in a decrease in lymphocyte proliferation. Cells with the antisense GH ODN had less (87%) incorporation of [3H]thymidine [( 3H]TdR) in both resting and Concanavalin-A-stimulated lymphocytes, whereas the incorporation of [3H]TdR in cells treated with a control ODN was not significantly affected. The effect of the antisense ODN on [3H]TdR incorporation was specific, since it could be reversed by hybridization competition with a complementary GH sense ODN or by the addition of exogenous rat GH. Collectively, the data indicate that lymphocytes synthesize and secrete irGH and that irGH produced by these cells can stimulate proliferation, suggesting that GH may play an autocrine/paracrine role in lymphocyte replication.

Cellular Localization of the Growth Hormone Receptor/ Binding Protein in the Male and Female Reproductive Systems*

We have used immunohistochemistry to localize GH receptor/binding protein (BP) in the male and female reproductive systems of adult rats. Testes and ovaries from neonatal animals were also examined to determine if GH receptor/BP expression in these tissues is developmentally regulated. Two monoclonal antibodies (MAb 43 and 263) were immunoreactive in identical locations whereas no immunoreactivity was evident when control monoclonal antibodies 7 and 50.8 were used. Localization of the receptor/BP was observed in both the nucleus and cytoplasm of immunopositive cells confirming our recent report of a nuclear GH receptor. Intense GH receptor/BP immunoreactivity in the male reproductive system was evident in the epithelium of the vas deferens and coagulating gland, the prostatic epithelium during the secretory phase, and the ductular epithelium of the coagulating and bulbourethral glands, respectively. Strong immunoreactivity was detectable in the Leydig and Sertoli cells, the epithelium of the ductus epididymis and seminal vesicles and smooth muscle of the tunica muscularis of the vas deferens, septae of the seminal vesicles, and in prostatic fibromuscular stroma. Cells of the seminiferous tubules (spermatogonia, primary and secondary spermatocytes, and spermatids) were moderately immunoreactive. No immunoreactivity was detectable in spermatozoa in the ductus epididymis or vas deferens, in scattered epithelial cells of the ductus epididymis, the prostatic epithelium in the nonsecretory phase, and mucous secreting cells of the bulbourethral glands. Leydig cells of 10-day postnatal rat testis were intensely immunoreactive whereas seminiferous tubular cells displayed homogenous immunoreactivity from moderate to strong. Intense GH receptor/BP immunoreactivity in the female reproductive system was evident in the germinal epithelium, the vascular endothelium of the myometrium, the epithelial lining of the fimbriae and oviduct, the endometrial epithelium and scattered endometrial glands, the mesothelium of the perimetrium, and the vascular endothelium of the endometrium. Strong immunoreactivity was exhibited by scattered oocytes, lutein cells of the corpus luteum, scattered endometrial glands, and the vascular endothelium of the endometrium. Moderate immunoreactivity was evident in scattered oocytes, granulosa cells, theca interna and externa, smooth muscle of the oviduct and myometrium, scattered endometrial glands, and luminally placed endometrial stroma cells. Ovarian granulosa cells from 10-day postnatal rats displayed strong immunoreactivity in contrast to moderate immunoreactivity in adult granulosa cells. In conclusion, we report a widespread distribution of the GH receptor/BP in the reproductive system of the rat by which GH may exert a direct action on reproductive function. The distribution is concordant with a role for GH in epithelial function and/or maintenance and also with a possible role for GH in the integrity of the endometrial vasculature.