Acute changes in growth hormone-releasing hormone secretion after injection of BIM 23014, a long acting somatostatin analog, in rams.

BIM 23014 is a somatostatin analog displaying an increased biological half life due to resistance to enzymatic degradation. This peptide inhibits GH release directly at the level of pituitary somatotrophs. In addition, an action of BIM 23014 at the level of the hypothalamus is possible since somatostatinergic fibers and receptors have been identified on GH-RH neurons. To evaluate the effect of BIM 23014 on GH-RH secretion, hypophysial portal blood (HPB) was continuously collected in conscious sheep. Twelve rams (40-45 kg, 9-month-old) with chronically implanted perihypophysial cannulae were i.v. injected with BIM 23014 (1 mg) or saline. HPB and jugular blood were collected for 3-5 hours before and after the injection for the determinations of GH-RH and GH concentrations respectively. The acute injection of BIM 23014 induced a rapid decrease of plasma GH within the first two hours. Simultaneously, GH-RH in HPB decreased significantly. After reaching a nadir, GH concentrations increased to values greater than baseline. A similar rebound in GH-RH levels in HPB was also observed. These data indicate that BIM 23014 acts at the level of GH-RH hypothalamic neurons, in addition to its well-know effect on the pituitary gland.     

Design and characterization of a fluorescent ghrelin analog for imaging the growth hormone secretagogue receptor 1a

Ghrelin is a 28-amino acid peptide hormone produced in the stomach. It binds to the growth hormone secretagogue receptor 1a (GHS-R1a), a class A G-protein-coupled receptor. In the present study, we describe the design, synthesis and characterization of a truncated, 18-amino acid analog of ghrelin conjugated to a fluorescent molecule, fluorocein isothiocyanate (FITC), through the addition of a lysine at its C terminus ([Dpr(octanoyl)(3), Lys(fluorescein)(19)]ghrelin(1-19)). Receptor binding affinity of this novel fluorescein-ghrelin(1-18) was similar to that of wild-type ghrelin and a synthetic GHS-R1a ligand, hexarelin. Live cell imaging in CHO/GHS-R1a cells demonstrated cell surface receptor labeling and internalization, and agonist activity of fluorescein-ghrelin(1-18) was confirmed by increased phosphorylation of ERK1/2. We also show that GHS-R1a protein is expressed primarily in the heart when compared to all other organs, suggesting high receptor density in the left ventricle. Finally, we demonstrate that fluorescein-ghrelin(1-18) binds specifically to heart tissue in situ, and its binding is displaced by both wt ghrelin and hexarelin. We have therefore developed a novel imaging probe, fluorescein-ghrelin(1-18), that can be used to image GHS-R1a in situ, for the purposes of investigating mechanisms of receptor trafficking or pharmacological agents that target GHS-R1a.     

The benefits of regular kinesiotherapy once a week for postmenopausal women: an aged-matched study

Regular exercise training improves overall physical fitness and quality of life in postmenopausal women. The exigent training frequency depends on a user-specified training aim. The aim of this study was to confirm the benefits of regular once a week exercise training for the maintenance of fitness in postmenopausal women. The test group included 20 postmenopausal women (65 +/- 3.1 years) who have been attending the exercise training program conducted by the physiotherapist once a week for three years. The age-matched control group included 20 healthy women (65.5 +/- 2.4 years) who did not regularly attend the training program. The outcomes were: right and left lateral trunk flexion, right and left shoulder flexion, right and left grip strength, endurance capacity of the trunk extensors, lower limb muscle strength (1' chair stand test), and balance (one-leg standing duration time with eyes open and closed). Women from the test group achieved statistically significant better results in the following outcomes: right lateral trunk flexion (15.4 cm: 12.6 cm, p < 0.001), left lateral trunk flexion (15.4 cm: 12.6 cm, p = 0.001), trunk extension muscle endurance (53.4 s: 40.5 s, p < 0.001), lower limb muscle strength (28.4 x: 25 x, p < 0.001), and one-leg standing duration time with open eyes (33.5 s: 19.7 s, p < 0.001). The results suggest that a regular once a week exercise training program designed and conducted by the physiotherapist, may be helpful in the improvement or maintenance of flexibility, muscle strength and capacity, and balance in postmenopausal women. The better fitness proved by our study could be a result of other causes and not solely that of the designed training program.     

Plerocercoid growth factor: a homologue of human growth hormone

Plerocercoids of the tapeworm, Spirometra mansonoides, produce a factor with characteristics similar to those of mammalian growth hormone (GH). Plerocercoid growth factor (PGF) stimulates growth and mimics other actions of GH but does not possess the anti-insulin/diabetogenic activities intrinsic to mammalian growth hormones. Duplication of activities unique to human GH, chemical and physical similarities, plus crossreactivity with strictly specific anti-hGH monoclonal antibodies, underlie the hypothesis that S. mansonoides has obtained and expresses a human gene for GH. In this article, Kirk Phares discusses the similarities between the two hormones.     

A bicyclo-somatostatin analog,highly specific for the inhibition of growth hormone release

The tetradecapeptide somatostatin was cyclized by a combination of conventional and solid phase peptide synthesis methods, to a homodetic cyclic disulfide tetradecapeptide, Wy-40,391:

The analog inhibits the release of growth hormone (GH) $\text{in vivo}$ without affecting either insulin or glucagon secretion. A correlation between binding affinity to the receptors and specificity is suggested.     

Purification of a pyrogen-free human growth hormone antagonist

Human growth hormone (hGH) is a polypeptide with 191 amino acids and a molecular mass of 22 kilodaltons. With the aid of computer molecular simulation, an hGH analog was created by altering an hGH gene to reflect the change of one amino acid (glycine [G] 120 to arginine [R]) within the third alpha-helix of the hGH molecule. This hGH analog, named hGHG120R, was found to be an hGH antagonist. It may have important implications in treating human conditions in which hGH levels are abnormally high, as found in type I diabetics. Several hundred milligrams of purified hGHG120R were needed to determine the biological activity of the antagonist in animal models. A multistep downstream process was developed to purify hGHG120R from cultured mouse L cells transfected with the hGHG120R gene. The process consisted of cell clarification, salt precipitation, membrane ultrafiltration, reversed phase high performance liquid chromatography, phase separation, and lyophiliation. This work discusses the rationale for the design of the process and experimental results on the purification of hGHG120R using the process. (c) 1995 John Wiley & Sons, Inc.     

Effects of a leucine analog on growth hormone processing and secretion by cultured cells

Bovine and rat growth hormones (bGH and rGH, respectively) possess signal peptides that direct the hormone to the secretory pathway and are proteolytically cleaved prior to secretion. Previous in vitro translation studies indicated that incorporation of the polar leucine analog beta-hydroxyleucine into de novo synthesized polypeptides inhibits signal peptide function. To test the effects of this analog on GH secretion by cultured animal cells, transfections of mouse L-cells with a bGH expression plasmid or metabolic labeling of endogenous rGH in anterior pituitary cells was performed in the absence or presence of beta-hydroxyleucine. Transient expression of bGH in mouse L-cells or endogenous expression of rGH in anterior pituitary cells resulted in an accumulation of GH in the culture medium. Treatment with beta-hydroxyleucine resulted in a block in secretion as evidenced by an accumulation of GHs within these cells. Amino-terminal sequencing of the intracellular form of the analog-substituted GHs demonstrated accurate signal peptide cleavage. In contrast, in vitro translations of bGH RNA performed in the presence of beta-hydroxyleucine and microsomal membranes resulted in the inhibition of signal peptide cleavage. The results suggest that beta-hydroxyleucine can uncouple signal peptide processing and protein secretion in cultured cells.     

The treadmill exhausting test is not suitable for screening of growth hormone deficiency!

BACKGROUND/METHOD: We compared the growth hormone response to a modified exercise test--the treadmill exhausting test--to pharmacological stimulation tests in 77 children with short stature. Each child underwent the treadmill test to individual exhaustion and at least one pharmacological test for GH stimulation. To determine the point of individual exhaustion, the heart rate, workload and oxygen consumption were measured. RESULTS: The mean +/- SEM peak GH concentration (ng/ml) in 47 small, normally growing children (group 1) was 16.1 +/- 1.3 in the pharmacological tests vs. 5.0 +/- 0.6 after a treadmill exhausting test. Thirty children with GH deficiency (group 2) had mean +/- SEM peak GH concentrations (ng/ml) of 5.5 +/- 0.5 in the pharmacological tests and 4.1 +/- 0.7 after physical exercise. The groups differed significantly in the pharmacological tests (p < 0.001) but not in the exhausting test. We found a 90% sensitivity but only a 11% specificity for the treadmill exhausting test compared to the diagnosis obtained by pharmacological testing. CONCLUSION: We do not recommend the treadmill exhausting test in clinical practice of pediatric endocrinology at all.     

Adipocyte studies: systems for investigating effects of growth hormone and other chronically acting hormones

Adipose tissue is very amenable to study in vitro. Collagenase digestion yields free adipocytes which usually respond well to acute stimulation/inhibition by hormones and other factors. Chronic effects of hormones are best studied using explants of adipose tissue which, from some species (e.g. sheep), can be maintained in culture for up to a week without loss of function. Alternatively, preadipocytes can be readily isolated from adipose tissue and induced to proliferate and differentiate in culture, while various adipocyte-like cell-lines have been established, which can be used for chronic studies. Use of these various systems for investigating the mechanisms of action of growth hormone are described.     

Synthesis of a nonreducible cyclic analog of somatostatin having only growth hormone release inhibiting activity

A nonreducible cyclic analog of somatostatin (SRIF) was prepared by a combination of solid phase and solution peptide synthesis. The compound, gamma-Abu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-Asp-OH, was tested for its effect on the release of growth hormone, glucagon and insulin in rats. It significantly suppressed pentobarbital-stimulated growth hormone release but showed no effect on arginine-stimulated glucagon or insulin release. The linear form, NH2-gamma-Abu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-Asp-OH, was also prepared and tested in vivo. It was shown to have only slight activity.