Elemental iron does repress transferrin, haemopexin and haemoglobin receptor expression in Haemophilus influenzae

The present study was undertaken to determine the effect of GH administration on GH and IGF-I receptors in skeletal muscle compared with liver in growing pigs. Plasma IGF-I and GH-binding protein (GHBP) levels were also determined. Twelve Large White pigs (castrated males) were treated daily with 100 micrograms pituitary porcine GH (pGH) per kg body weight or vehicle for 41 days intramuscularly. Relative to controls, pGH administration increased plasma IGF-I concentrations by 3.3-fold. Administration of pGH had no effect on plasma GHBP levels. In liver, 125I-labelled bovine GH (bGH)-specific binding (P < 0.05) and GH receptor (GHR) mRNA levels (P < 0.05) were higher in pGH-treated than in control pigs. In longissimus dorsi (LD), 125I-labelled bGH specific binding did not differ significantly between the two groups while GHR mRNA levels (P < 0.05) were lower in pGH-treated than in control pigs. Administration of pGH had no effect on 125I-labelled bGH-specific binding and GHR mRNA levels in trapezius (TR). 125I-Labelled IGF-I-specific binding in liver was unaffected by pGH administration. Similarly, in liver, LD and TR, IGF-I receptor mRNA levels were not different between pGH-treated and control animals. It can be concluded that (1) GH binding and IGF-I receptor mRNA are not affected by GH in skeletal muscle, (2) GH influences GHR in a tissue-specific manner and (3) hepatic GHR and GHBP levels are not co-regulated.     

Monoclonal antibodies to bovine growth hormone potentiate hormonal activity in vivo by enhancing growth hormone binding to hepatic somatogenic receptors

Administration of GH complexed with monoclonal antibodies (MABs) potentiates the in vivo actions of the hormone. In particular, growth and serum IGF-I concentrations of GH-treated hypophysectomized rats are increased by concomitant injection of anti-GH MABs. Among 37 anti-bovine GH (bGH) MABs, we selected one MAB with the most potentiating effects to investigate the mechanisms responsible for this phenomenon. Hypophysectomized rats were killed 18 h after a single s.c. injection of bGH (100 micrograms/rat), alone or complexed with increasing doses of MAB (4, 40, 400 micrograms/rat; MAB:bGH molar ratio: 0.005, 0.05, 0.5). IGF-I was measured by radioimmunoassay in acid-extracted sera and livers, whereas liver IGF-I mRNA was quantified by Northern blot hybridization. The in vivo occupancy of liver somatogenic (GH) receptors was derived from the determinations of total and free 125I-labelled bGH binding to liver homogenates treated with 4 mol MgCl2/l or water. Injection of MAB-bGH complexes enhanced body weight gain and raised serum IGF-I, liver IGF-I and liver IGF-I mRNA more than bGH alone (1.6-, 6-, 10- and 7-fold increases at the highest dose of MAB, compared with bGH alone; P < 0.001). These potentiating effects of the MAB were dose-dependent and significant potentiation of the growth response was already observed with the lowest dose of MAB. In vivo occupancy of liver GH receptors was markedly higher 18 h after injection of MAB-bGH complexes than after bGH alone, and this effect was also dose-dependent (receptor occupancy of 28%, 37% and 83% after 4, 40 and 400 micrograms of MAB respectively compared with 6% after bGH alone; P < 0.05, 0.05 and 0.001 respectively). In contrast, the in vitro binding of 125I-labelled bGH to liver homogenates was decreased in the presence of high doses of MAB. We conclude that low amounts of MABs complexed with bGH potentiate the stimulation by the hormone of liver IGF-I synthesis and secretion in a dose-dependent manner. These effects are mediated, at least in part, through changes in hormone-receptor interaction in vivo, leading to enhanced and/or prolonged binding of bGH to its somatogenic receptors.     

Cross-linked growth hormone dimers have enhanced biological activity

In this study we have investigated the effect on the bioactivity of pituitary-derived human growth hormone (hGH) and recombinant bovine (b) GH after the addition of various concentrations of the water soluble cross-linking agent 1-ethyl-3(3-dimethylaminopropyl) carbodiimide (EDC; 6.25-100 mg/ml). The biological activity of resulting cross-linked reactions were determined by its ability to promote incorporation of 35SO4(2-) into costal cartilage of hypopituitary Snell dwarf mice in vivo. Administration of EDC-treated hGH solutions resulted in a significant enhancement of hormone activity in vivo compared with non-cross-linked samples. A similar significant enhancement of bGH activity in vivo was also observed when solutions containing recombinant bGH were cross-linked using EDC. For both hGH and bGH the degree of enhancement appears to be dose-dependent for the concentration of EDC (6.25-100 mg/ml for hGH; 6.25-50 mg/ml for bGH) present in the cross-linking reactions. SDS-PAGE analysis of EDC cross-linked solutions containing hGH and bGH spiked with 125I-hGH and 125I-bGH respectively revealed that dimeric GH was the primary cross-linked component. Increasing the concentration of EDC in cross-linking reactions resulted in increased formation of dimeric hGH and bGH. There was a significant correlation between the amount of GH dimer present and the increase in biological activity, suggesting that GH dimers were responsible for the enhanced biological activity. This was confirmed by the enhanced biological activity of a purified preparation of EDC cross-linked dimeric hGH. In conclusion, covalently cross-linked GH dimers reported here have enhanced bioactivity in vivo. However, since naturally occurring GH dimers are known to have reduced biological activity, this work suggests that the structure of EDC cross-linked GH dimers differs fundamentally from that of native dimeric hGH.     

Suppression of anti-hapten antibody response in vitro by hapten-carrier conjugates

Production of antibodies was stimulated or suppressed arbitrarily by antigen treatment in vitro of spleens cultured at various time intervals after in vivo immunization. Spleens of mice immunized to the 2,4-dinitrophenyl or (4-hydroxy-3-iodo-5-nitrophenyl)acetyl haptenic determinants produced antibodies in culture when no antigen was applied in vitro. When a conjugate of the hapten to the same carrier employed for priming was given in vitro, an initial reduction of the response was observed, the level of which was dependent on antigen dose. Subsequently, increased amounts of antibodies were measured. In contrast, in vitro exposure to the hapten conjugated to an unrelated carrier resulted in significant reduction of the response for the entire period of the test. This suppressive effect manifested with various carrier proteins (ovalbumin, bovin IgG, bovine and rabbit serum albumin and keyhole limpet hemocyanin), when when applied to cultures in doses which were potentially immunogenic.     

Hypophysectomy or adrenalectomy of rats with insulin-dependent diabetes mellitus partially restores their responsiveness to growth hormone

The studies reported herein were conducted to confirm that the pituitary gland is involved in maintaining growth hormone (GH) resistance in rats with insulin-dependent diabetes mellitus (IDDM) and to determine whether the adrenocorticotropic hormone (ACTH)-adrenal cortical axis is responsible. The rats were made diabetic by injecting streptozotocin (85 mg/kg body wt) IP once daily on two consecutive days. They were then injected with 15 IU insulin SC twice daily on two consecutive days to enable them to survive hypophysectomy or adrenalectomy. Intact nondiabetic (NonDb), diabetic (Db), hypophysectomized diabetic (HxDb), and adrenalectomized diabetic (AxDb) rats were injected twice daily with 50 micrograms porcine (p) GH or with 0.9% saline for 2 weeks following the surgeries. Serum glucose levels of the saline-injected Db, HxDb, and AxDb rats were significantly greater than those of the NonDb rats by 106%, 65% and 49%, respectively. However, the levels in the HxDb and AxDb animals were significantly lower than those of the Db group by 20% and 28%, respectively. Injections of pGH into NonDb rats increased serum glucose concentrations by 38%, over their saline-treated controls, and by 29% in AxDb rats. This diabetogenic effect of GH was not seen in any other group. Administration of pGH to Db rats failed to increase body weight gain, tall growth, tibial epiphysial plate width, or serum IGF-I concentration over saline-injected controls. By contrast, HxDb and AxDb rats injected with pGH showed significant increases in all four growth parameters. Total serum IGF-I concentrations in AxDb rats injected with pGH equaled those in NonDb controls. To determine whether the lack of corticosterone (B) in the AxDb rats was responsible for the reduced hyperglycemia and restored responsiveness to pGH, AxDb rats were given B in their drinking water at 5 or 25 micrograms/ml. Administration of B reduced the beneficial effects of adrenalectomy by restoring hyperglycemia and growth impairment, and partially restored resistance to the pGH injections. These studies confirm that the pituitary contributes to diabetic growth impairment and show that the ACTH-adrenal cortical axis is primarily responsible for the GH-resistant state that develops in rats with IDDM.     

Calculated magneto-optical Kerr effect in Fe, Co, and Ni

To examine whether GH and IGF-I participate in the regulation of obese (ob) mRNA expression we determined ob mRNA levels in epididydimal fat pads of hypophysectomised (hypox) rats, hypox rats treated with recombinant human (rh) GH or rhIGF-I and normal, weight-matched controls. We found that 1. ob mRNA was markedly suppressed after hypophysectomy (37 +/- 25% of controls), 2. GH infusion had no effect on ob mRNA, but stimulated IGF-I mRNA in fat pads, 3. IGF-I treatment further suppressed ob mRNA (3.5% +/- 0.6% of controls) and 4. serum insulin levels were decreased in all hypox groups (11.2 to 15.9% of controls). In conclusion, exogenous and GH-induced IGF-I differ in their effects on ob mRNA expression and GH is unable to restore ob mRNA towards normal at low insulin levels.     

Structure and function of bovine growth hormone. Bovine growth hormone as an experimental model for studies of protein-protein interactions

Growth hormone (GH) is a polipeptide that controls the differentiation, growth and metabolism of many cell types, and is secreted from the hypophysis of all vertebrate species tested so far. Despite the overlapping evolutionary, structural, immunological and biological properties, it is well-known that GHs from distinct mammalian species have significant species-specific characteristics. The main purpose of this review is to highlight bovine GH (bGH) structural features related to its species-specific properties. Novel interest in bGH is also aroused by the advent of biotechnological methods for production of recombinant proteins. In fact recombinant bGH will have a great importance in veterinary medicine research and as a 'high tech' drug that needs to be monitored in zootechnical productions.     

Multiple eruptive dermatofibromas in three men with HIV infection

Several recent studies have investigated the effects of growth hormone (GH) on the healing of fractures and bone ingrowth, but with conflicting results. The negative results may be due to antibody formation against injected GH or because some experimental models are able to prove only positive GH effects. In this study, we wanted to investigate the effect of GH on implant integration in bone. To avoid potential formation of antibodies against injected GH, we used a model with transgenic mice overexpressing bovine GH (bGH). Titanium implants were inserted in the forehead of the mice. 4 months after insertion, the implants were cut out en bloc with the surrounding bone. The calcified specimens were cut and ground to a thickness of approximately 10 microns. Histomorphometry demonstrated significantly more direct bone-to-metal contact in the transgenic mice than in the nontransgenic littermates. Our findings indicate that systemic administration of GH in humans may improve implant integration in bone.     

Application of enzyme-linked immunosorbent assay for epidemiological studies of diseases of livestock in the tropics of Mexico

This study was conducted at the Centre for Research, Teaching and Extension in Tropical Livestock (Centro de Investigación, Ense?anza y Extensión en Ganadería Tropical) of the Faculty of Veterinary Medicine of the National Autonomous University of Mexico. During the latter part of 1986 and throughout 1988 and 1989, the herd of Holstein x zebu cattle at the University was tested for IgG antibodies to twenty-one viral, bacterial, rickettsial and parasitic agents. Antigens prepared from twenty infectious disease agents were used as the solid phase in an enzyme-linked immunosorbent assay, and the agar gel immunodiffusion procedure was used to test for antibodies against bovine leukaemia virus. The prevalence of IgG antibodies was high (> 50%) for bluetongue virus, Anaplasma marginale and Mycoplasma bovis. Antibodies to Brucella abortus were absent and antibodies against bovine virus diarrhoea virus and infectious bovine rhinotracheitis virus showed a very low prevalence (< 5%). Antibodies to fifteen other antigens showed intermediate prevalence (15-46%). Antibodies to Campylobacter fetus, A. marginale, bluetongue virus, bovine leukaemia virus and Haemophilus somnus displayed seasonal variations. Levels of antibody to bovine leukaemia virus, M. bovis and Listeria monocytogenes exhibited increasing secular trends while antibodies to bovine virus diarrhoea virus and C. fetus showed declining trends. Prevalence of antibodies increased with the age of animals tested. No consistent difference in antibody prevalence was found between three genotypic groups examined.     

Competitive enzyme immunoassay for bovine growth hormone

We developed an enzyme immunoassay (EIA) for bovine GH (bGH) which is based on indirect competitive immunoassay in culture medium from a bovine pituitary cell culture. 40 microliters cell culture samples (or bGH standard) and bGH antibody (rabbit anti-bGH) were added to the 96 well microplate coated with secondary antibody (Goat anti-rabbit IgG), and incubated for 24 h at 37 degrees C. Biotin-label bGH was added and incubated further for 24 h at 37 degrees C, and biotinylated bGH was linked with streptoavidin-peroxidase. Substrates for peroxidase were added to the plate and incubated for 1 h at 4 degrees C. The enzyme reaction was stopped with 4N H2SO4, and the absorbency at 450 nm was measured with an ELISA Reader. The coefficients of intra-assay and inter-assay variations were 4.13 approximately 7.59% and 3.71 approximately 8.27%, respectively. The regression equation and correlation coefficients with the radioimmunoassay (RIA) were y(RIA) = 1.9986 x (EIA) - 1.3921 and 0.9701 (n = 27), respectively. Collectively, the present assay provides a reliable alternative to RIA and offers the major advantage of eliminating radioactive reagents and counting equipment.     

Effects of iodinated contrast agents in MR imaging

At least two classes of mRNA for the GH receptor (GHR) and GH binding protein (GH BP) with different 5' untranslated first exons exist in the rat. One such class, the GHR1 is predominantly expressed in the liver of female rats. The hepatic expression of the GHR1 mRNA in normal and hypophsectomized rats of both sexes was studied by employing an RNase protection/solution hybridization assay. Normal females expressed 10-fold more GHR1 mRNA than males, hypophysectomy of female rats decreased the GHR1 level to that observed in male rats. Continuous GH treatment of hypophysectomized male and female rats for 6 days increased the expression of GHR1 mRNA to levels found in normal females, whereas intermittent GH treatment without effect. Bovine GH(bGH) induced the GHR1 expression in a time- and dose-dependent manner in primary cultures of adult rat hepatocytes as determined by solution hybridization. Maximal induction was achieved after 72 h of treatment with 50 ng bGH/ml medium. Female enriched expression of receptor and binding protein mRNAs raises the possibility that they participate in determining the ability of the liver to respond differently to the male and female GH secretory patterns. Our in vitro model utilizing cultures of primary adult rat hepatocytes could be used to address this issue as well as explore a hormonal interplay in regulation of GHR1 expression.     

Cross-reactivity of monoclonal antibodies to defined human leucocyte differentiation antigens with bovine cells

Thirty-seven subpanels of monoclonal antibodies (mAbs) included within the Vth International Workshop on Human Leucocyte Differentiation Antigens (Vth Workshop) were assayed for reactivity with bovine peripheral blood leucocytes. Sixty-five of the 772 mAbs (8.4%) stained bovine cells. mAbs from each of the 27 different CD groups that contained a mAb reacting with cattle were further investigated to compare the cellular expression of the antigen in cattle with that reported for the different CD antigens in humans. Two-colour immunofluorescence staining of the Vth Workshop mAbs against characterized bovine leucocyte subpopulation markers that identified monocytes, B cells, CD4, CD8 and WC1 +T cells were used for these analyses. Eighteen of the mAbs to different human CD antigens (CD11a, CD14, CD18, CD21, CD27, CD29, CD49a, CD49b, CD49d, CD49e, CD51, CD61, CD62L, CD62P, CD63, CDw78, CD98, CD100) stained bovine antigens with an almost identical cellular distribution to that reported in humans. This implies that these mAb react with the homologous cattle molecules. Nine mAbs (CD35, CD37, CD49c, CD50, CD54, CD66, CD81, CD88, CD102) stained bovine cells but the cellular distribution of the bovine antigen was different to that reported in humans implying either a different cellular distribution for these antigens in cattle or a reaction with a different molecule. The investigation has allowed the identification of several bovine homologues of human CD antigens that have not been previously defined in cattle and the cross-reacting mAbs will be valuable reagents for future investigations of bovine immunology.     

Experimental Goodpasture's syndrome in Wistar-Kyoto rats immunized with alpha3 chain of type IV collagen

BACKGROUND: Glomerulonephritis and lung hemorrhage of autoimmune Goodpasture syndrome develop due to immune reactions against epitope(s) of the non-collagenous (NC1) domain of alpha3-chain of type IV collagen [alpha3(IV) NC1]. Whether thymic mechanisms have a role in the loss of tolerance to the Goodpasture epitope has not been established. We studied the renal and pulmonary effects of immunization with different forms (monomer, dimer, or hexamer) of alpha3(IV) NC1 collagen in Wistar-Kyoto (WKY) rats, and assessed whether the intrathymic inoculation of the antigen may protect against anti-GBM disease. METHODS: WKY rats were immunized with bovine alpha3(IV) monomer, dimer, or hexamer, or with alpha3(IV) NC1 synthetic peptide. Renal function, kidney and lung immunohistology, and circulating and tissue bound antibodies to type IV collagen chains were analyzed. Effects of intrathymic inoculation of antigen on subsequent disease induction were analyzed in WKY rats given alpha3(IV) NC1 dimer or GBM preparation intrathymically 48 hours before immunization. RESULTS: Proteinuria, linear IgG deposition in GBM, and crescentic glomerulonephritis developed in WKY rats immunized with alpha3(IV) NC1 dimer or hexamer. Lesions were dose-dependent upon injections of 10 to 100 microgram dimer. The alpha3(IV) NC1 monomer induced less severe proteinuria and no crescents. Pulmonary hemorrhage was detectable in 35% of rats immunized with 25 to 100 microgram alpha3(IV) NC1 dimer; alpha3(IV) synthetic peptide (36 carboxyl terminal) did not induce disease. Rats injected intrathymically with up to 100 microgram alpha3(IV) NC1 dimer or with GBM 48 hours before immunization were not protected against subsequent development of proteinuria and glomerulonephritis. CONCLUSIONS: These findings document that glomerulonephritis and lung hemorrhage can be elicited in WKY rats by immunization with alpha3(IV) NC1. Failure of the intrathymic inoculation of antigen to prevent disease suggests that immunological tolerance cannot be achieved by this intervention, in contrast to other autoimmune conditions, and may imply independent roles for cellular and humoral nephritogenic pathways in anti-GBM nephritis.     

Hypothalamic growth hormone secretagogue-receptor (GHS-R) expression is regulated by growth hormone in the rat

Synthetic GH secretagogues (GHSs) act via a receptor (GHS-R) distinct from that for GH-releasing hormone (GHRH). We have studied the hypothalamic expression and regulation of this receptor by in situ hybridization using a homologous riboprobe for rat GHS-R. GHS-R mRNA is prominently expressed in arcuate (ARC) and ventromedial nuclei (VMN) and in hippocampus, but not in the periventricular nucleus. Little or no specific hybridization could be observed in the pituitary under the conditions that gave strong signals in the hypothalamus. No sex difference in GHS-R expression was found in ARC or hippocampus, though expression in VMN was lower in males than in females. Compared with GHRH and neuropeptide Y (NPY), GHS-R was expressed in a distinct region of ventral ARC, and in regions of VMN not expressing GHRH or NPY. GHS-R expression was highly sensitive to GH, being markedly increased in GH-deficient dw/dw dwarf rats, and decreased in dw/dw rats treated with bovine GH (200 microg/day) for 6 days. Similar changes were observed in GHRH expression, whereas NPY expression was reduced in dw/dw rats and increased by bGH treatment. Continuous sc infusion of GHRP-6 in normal female rats did not alter ARC or VMN GHS-R expression. Our data implicate ARC and VMN cells as major hypothalamic targets for direct GHS action. The sensitivity of ARC GHS-R expression to modulation by GH suggests that GHS-Rs may be involved in feedback regulation of GH.     

Occurrence of ateliosis in Japanese Black calves and their secretory function of growth hormone

Unthrifty calves occurred sporadically in Japanese Black (beef cattle) in an area in northeastern Japan. The states of unthrifty development, pedigree, clinico-biochemistry and the secretory function of bovine growth hormone (bGH) in pituitary were investigated. The total cholesterol concentration and CK, AST and LDH activities in the serum showed higher values than those of control calves. Basal bGH concentrations in the serum and bGH secretory reactivity in the insulin tolerance test (ITT) were showed to be significantly lower than those of the control calves. Furthermore, sperm donated from a specific bull had been used for these unthrifty calves. This study suggested that the present occurrence of unthrifty calves represented ateliosis possibly caused by congenital hypopituitarism which decreased of bGH secretory function.     

Transgenic rabbits overexpressing growth hormone develop acromegaly and diabetes mellitus

Transgenic rabbits expressing the bovine growth hormone (bGH) gene in liver and kidney were obtained to study the long-term effects of chronic exposure to GH in nonrodent animals. These rabbits presented high levels of bGH and insulin-like growth factor I in serum. In spite of chronic exposure to bGH, transgenic rabbits had similar body weight to controls. However, enlargement of the head and limbs and reduction of visceral fat were observed in these animals. They also showed marked hyperinsulinemia, hyperglycemia, and hypertriglyceridemia, indicating that they developed insulin resistance. Furthermore, serious histopathological alterations, including marked fibrosis, were observed in liver, kidney, and skeletal muscle. These anatomical, metabolic, and histological alterations closely resemble those found in patients with acromegaly. Thus, transgenic rabbits overexpressing GH may be a good model of the human disease.     

Viral antibodies in bovine fetuses in Argentina

In order to establish the prevalence of viral infections of the bovine fetus in Argentina, a serological survey for antibodies against viral agents currently affecting cattle in this country was conducted. Antibodies against foot-and-mouth disease virus (FMDV), bovine herpesvirus-1 (BHV-1), bovine leukaemia virus (BLV), bovine rotavirus (BRV), bovine coronavirus (BCV), bovine viral diarrhoea virus (BVDV) and parainfluenza-3 (PI-3) were investigated in a total of 315 fetal serum samples. Conventional techniques were used: indirect immunofluorescence (FMDV, BHV-1, BVDV and BCV), radial immunodiffusion (BLV), ELISA (BRV) and haemagglutination inhibition (PI-3). Antibodies against BHV-1, BVDV and PI-3 were detected in samples from fetuses in the second and third trimester of gestation, with a prevalence of 1.21 per cent (two of 165), 2.03 per cent (four of 197) and 5.08 per cent (nine of 177), respectively. Either antibodies or non-antibody factors able to bind to BRV and BCV antigens were detected with a prevalence of 2.44 per cent (five of 205) and 4.54 per cent (five of 110), respectively. In addition, 14.68 per cent of non-specific inhibitors of PI-3 mediated haemagglutination were found. No seropositives against FMDV and BLV were detected.     

Induction of antigen-specific isotype switching by in vitro immunization of human naive B lymphocytes

The use of in vitro immunization technology for the generation of human antigen-specific antibodies has essentially resulted in low affinity IgM antibodies, resembling an in vivo primary immune response. We now describe a detailed reproducible protocol for a two-step in vitro immunization, which yields isotype switched, antigen-specific human antibodies. The immunizing antigen was a 30aa synthetic peptide, containing both a B (15aa V3 peptide of the HIV-1) and a T helper cell epitope (15aa peptide from tetanus toxin). The immunization protocol includes: (i) a selection procedure of donors with a memory T cell response against tetanus toxoid; (ii) immunization of mature naive peripheral B lymphocytes in two distinct phases, involving a primary and a secondary step. None of the donors which were examined after primary immunization showed at any time an IgG anti-V3 specific antibody response, while all the donors showed an IgM response. After the secondary immunization step, anti-V3 antibodies of both IgM and IgG isotypes were detected. The switch frequency event was high among the tested donors (5/8).     

Stimulatory effect of human, but not bovine, growth hormone expression on numbers of tuberoinfundibular dopaminergic neurons in transgenic mice

Mice transgenic for heterologous and ectopic GH expression serve as models for studying the feedback effects of elevated nonregulated GH on hypothalamic hypophysiotropic neurons as well as on peripheral function. For example, hypothalamic somatostatin expression has been shown to be increased markedly in mice bearing either bovine (b) or human (h) GH transgenes. Human, but not bovine, GH has lactogenic properties in mice, and appears to stimulate PRL-inhibiting tuberoinfundibular dopaminergic (TIDA) neurons. The present study was designed to determine the effect of a lifelong excess of hGH on dopamine (DA) expression in and numbers of TIDA neurons. Male mice of four transgenic lines were examined. The transgenic animals bore constructs of either bGH or hGH fused to either metallothionein (MT) or phosphoenolpyruvate carboxykinase (PEPCK) promoters; brains of transgenic mice were compared morphologically with those of nontransgenic littermates. Formaldehyde-induced catecholamine histofluorescence and tyrosine hydroxylase (TH) immunocytochemistry were examined in alternate brain sections; cell number was quantified for TIDA neurons (area A12) and a nonhypophysiotropic diencephalic DA area, the medial zona incerta (A13). Body weights were higher (P < 0.01) in PEPCK-GH than in MT-GH transgenic mice, as were serum levels of heterologous GH in those lines. In MT-hGH, but not MT-bGH or PEPCK-bGH, transgenic mice, A12 perikaryal fluorescence was enhanced, and ME fluorescence was reduced compared with those in control animals. The reduced ME DA is likely to reflect stimulation of TIDA neurons, because A12 TH-immunoreactive neuron number was increased by 34% in MT-hGH mice compared with that in controls (P < 0.05). In mice bearing the PEPCK-hGH construct, A12 TH neuron number was increased 47% (P < 0.001) compared with that in littermate controls. There were no differences in A13 cell number among animals, and A12 cell numbers in mice expressing bGH did not differ from control values. These results suggest that although extremely high levels of circulating bGH do not stimulate TIDA neurons, lifelong high levels of hGH have a stimulatory and graded effect on developmental differentiation of these cells for TH and DA production, supporting the concept of PRL as a trophic factor for TIDA neurons.     

The serum half-life of somatomedin activity: evidence for growth hormone dependence

The serum half-life of somatomedin (SM) activity has been measured following the intravenous injection of SM activity into hypophysectomized rats. A [3H]thymidine incorporation assay in chick embryo fibroblasts (CEFs) has been utilized to measure SM activity. Cell cycle analysis data obtained with the flow microfluorometer shows that the [3H]thymidine incorporation data reflects actual DNA synthesis. When normal rat serum was injected, a half-life for SM activity of approximately 3 h was determined. In marked contrast, when serum from hypophysectomized (hypox) rats acutely treated with growth hormone (GH) was used as the source of SM actively, the half-life of the SM actively was short, approximately 8 min. However, when serum from hypox rats chronically treated with GH was injected, the half-life was again long, approximately 4 h. SM activity has been separated from its binding proteins by boiling under acid conditions or chromatography on Sephadex G-50 in 1 N acetic acid. The half-life of this partially purified SM activity was short, in the range of 10-30 min finally, recombination of the partially purified sm activity with the large proteins in normal serum extended in half-life of the sm from 10-03 min to about 2 h. These data indicate that the serum half-life of SM activity is GH dependent and suggest that the serum binding protein, like SM itself, is under GH control.