The living record of
peptide science.

Role of serum myostatin during the lactation period.

J Reprod Dev

Tohru Hosoyama, Keitaro Yamanouchi, Masugi Nishihara

Myostatin, also known as GDF-8 (Growth/Differentiation Factor-8), is a member of the TGF-beta superfamily that negatively regulates skeletal muscle mass in mammals. Mutation of the myostatin gene in mice, cattle, and humans causes a massively developed skeletal muscle, characterized by muscle hypertrophy and hyperplasia. Although myostatin is predominantly expressed in skeletal muscle tissue, several recent studies have shown the presence of myostatin protein in blood and suggested a possible role for circulating myostatin in the regulation of skeletal muscle mass. In the present study, we examined changes in the levels of active form myostatin (13 kDa) in serum after birth by Western blot analysis to predict the role of serum myostatin in early postnatal muscle growth in the rat. Interestingly, the amount of active form myostatin in serum increased after birth and then decreased along with ageing after weaning. To clarify the role of increased serum myostatin during the postnatal period, we administrated follistatin, an inhibitor of myostatin activity, into postnatal rats intraperitoneally just after birth. Follistatin-administration during the postnatal period caused selective hypertrophy of type II muscle fibers in the soleus muscle. These results demonstrate that myostatin in serum acts on skeletal muscle and negatively regulates early postnatal muscle growth.

Influence of long-term treatment with tuftsin analogue TP-7 on the anxiety-phobic states and body weight.

Pharmacol Rep

Róza Czabak-Garbacz, Beata Cygan, Lukasz Wolański +1 more

TP-7 is a synthetic analogue of tuftsin. It has a structure of tuftsin, to which three natural L-amino-acids Pro-Gly-Pro are attached. This heptapeptide improves learning and memorization and causes antidepressant and anxiolytic effect. It is possible to use TP-7 in the future to optimize cognitive functions and as a potential new anxioselective, fast-acting and easy-dosed drug. Therefore, it was purposeful to study such properties of the heptapeptide as its influence on anxiety-fear and body weight under a long-term treatment regimen. The experiment was performed on 24 preselected Wistar rats with the use of Rodina's method. There were three experimental groups of animals with high initial emotional reactivity: passive control group (P), active control group (A, receiving distilled water) and group treated with TP-7 at the dose of 0.3 mg/kg (T). The rats of A and T groups received intraperitoneal injections every day. The experiments were conducted 15 min after the administration of the drug, one and two days after initial testing day, then 1, 2, 3 and 4 weeks after that. The heptapeptide reduced the anxiety-phobic states significantly starting from the second day of drug application. The observed effects persisted throughout four weeks of the experiment, which confirmed effective long-term anxiolytic properties of the heptapeptide. TP-7 did not cause any changes in the body mass by itself.

Pulsatile secretion of growth hormone (GH) persists during continuous stimulation by CJC-1295, a long-acting GH-releasing hormone analog.

J Clin Endocrinol Metab

Madalina Ionescu, Lawrence A Frohman

Pulsatile GH secretion is considered important for many of the hormone's physiological effects. Short-term GHRH infusions enhance GH pulsatility and increase IGF-I, but the short GHRH half-life limits its therapeutic use. A synthetic GHRH analog (CJC-1295) that binds permanently to endogenous albumin after injection (half-life = 8 d) stimulates GH and IGF-I secretion in several animal species and in normal human subjects and enhances growth in rats.

The effects of adulthood olanzapine treatment on cognitive performance and neurotrophic factor content in male and female rats neonatally treated with quinpirole.

Eur J Neurosci

Stephanie K Thacker, Marla K Perna, Jeffery J Ward +4 more

Male and female Sprague-Dawley rats were administered quinpirole (1 mg/kg, i.p.) or saline once daily from postnatal day (P)1 to P21. This drug treatment has been shown to produce long-term priming of the D2 receptor. Beginning on P62, rats were administered the atypical antipsychotic olanzapine (2.5 mg/kg) or saline twice daily (i.p.) for 28 days. One day after olanzapine treatment ceased, rats were tested on the place and match-to-place versions of the Morris water maze (MWM) for seven consecutive days. Dopamine D2 receptor priming was verified through a yawning behavioural test, a D2 receptor-mediated event, before olanzapine was administered as well as after olanzapine treatment and behavioural testing were complete. Results showed that neonatal quinpirole treatment induced D2 priming that was eliminated by olanzapine treatment. On the MWM place version, D2-primed rats demonstrated a significant impairment that was eliminated by olanzapine treatment, but olanzapine treatment to animals neonatally treated with saline produced a significant deficit on the place version of the MWM. There were no significant deficits on the match-to-place version. Brain tissue analyses revealed that neonatal quinpirole treatment produced a significant decrease in hippocampal NGF, BDNF and ChAT that was eliminated by olanzapine treatment. Neonatal quinpirole treatment produced a significant decrease in BDNF and ChAT in the frontal cortex that was unaffected by olanzapine treatment. These results show that olanzapine eliminates D2 receptor priming and cognitive impairment and also alleviates decreases in neurotrophins and acetylcholinergic markers produced by D2 priming in the hippocampus.

Once-daily administration of CJC-1295, a long-acting growth hormone-releasing hormone (GHRH) analog, normalizes growth in the GHRH knockout mouse.

Am J Physiol Endocrinol Metab

Maria Alba, Danilo Fintini, Alessia Sagazio +4 more

Although the majority of children with isolated growth hormone (GH) deficiency have a good growth response to GH-releasing hormone (GHRH), the use of this therapeutic agent is limited by its very short half-life. Indeed, we have shown that, in mice with GHRH gene ablation (GHRH knockout; GHRHKO), even twice-daily injections of a GHRH analog are unable to normalize growth. CJC-1295 is a synthetic GHRH analog that selectively and covalently binds to endogenous albumin after injection, thereby extending its half-life and duration of action. We report the effects of CJC-1295 administration in GHRHKO animals. Three groups of 1-wk-old GHRHKO mice were treated for 5 wk with 2 microg of CJC-1295 at intervals of 24, 48, and 72 h. Placebo-treated GHRHKO mice and mice heterozygous for the GHRHKO allele served as controls. GHRHKO animals receiving daily doses of CJC-1295 exhibited normal body weight and length. Mice treated every 48 and 72 h reached higher body weight and length than placebo-treated animals, without full growth normalization. Femur and tibia length remained normal in animals treated every 24 and 48 h. Relative lean mass and subcutaneous fat mass were normal in all treated groups. CJC-1295 caused an increase in total pituitary RNA and GH mRNA, suggesting that proliferation of somatotroph cells had occurred, as confirmed by immunohistochemistry images. These findings demonstrate that treatment with once-daily administration of CJC-1295 is able to maintain normal body composition and growth in GHRHKO mice. The same dose is less effective when administered every 48 or 72 h.

Melanocortin peptide therapeutics: historical milestones, clinical studies and commercialization.

Peptides

Mac E Hadley, Robert T Dorr

The melanocortins (MCs) are a family of multifunctional peptidergic hormones. Several superpotent, prolonged acting, enzymatically resistant, MC analogs have been designed and synthesized to help clarify the nature and role of MCs and their receptors (MCRs) in physiological functions. Two of these analogs, a linear peptide, melanotan I, and a cyclic truncated peptide, melanotan II (MTI and MTII, respectively) have been patented and tested clinically for studies on tanning of the skin (MTI) and for diagnosis and treatment of male erectile dysfunction (MTII). A new MTII analog, PT-141 (Palatin Technologies) has initial phase I/II trials and is scheduled to enter pivotal stage III clinical trials leading to commercialization. MTI may provide a therapeutic tan with the potential to lower the risk of skin cancer. PT-141 may improve sexual functionality in both males and females.

Seabream ghrelin: cDNA cloning, genomic organization and promoter studies.

J Endocrinol

Chung-Man Yeung, Chi-Bun Chan, Norman Y S Woo +1 more

Recent studies have indicated that ghrelin stimulates growth hormone release from the pituitary via the growth hormone secretagogue receptor (GHSR). We have previously isolated two GHSR subtypes from the pituitary of the black seabream Acanthopagrus schlegeli. In the present study, we have cloned and characterized ghrelin from the same fish species at both the cDNA and gene levels. The full-length seabream ghrelin cDNA, isolated from sea-bream stomach using a novel approach by exploiting a single conserved region in the coding region, was found to encode a prepropeptide of 107 amino acids, with the predicted mature ghrelin peptide consisting of 20 amino acids (GSSFLSPSQKPQNRGKSSRV). Embedded in this full-length cDNA is a putative fish orthologue of the recently reported mammalian obestatin peptide. The ghrelin gene in black seabream, obtained by genomic PCR, was found to encompass four exons and three introns, possessing the same structural organization as in tilapia and goldfish, but different from that in rainbow trout. In addition, a 2230-bp 5'-flanking region of the seabream ghrelin gene was obtained by genome walking. Sequence analysis revealed that, as in the case of the human ghrelin gene, there is neither a GC box nor a CAAT box present in the isolated 5'-flanking region. However, a number of putative transcription factor-binding sites different from the human counterpart were found in the 5'-flanking region of the seabream ghrelin gene, suggesting that different cis- and trans-acting elements are involved in controlling their gene expression. Functional activity of this 5'-flanking region was examined by cloning it into the pGL3-Basic vector upstream of the luciferase reporter gene and transfected into various cell lines. Positive promoter activity could only be recorded in the colon-derived Caco-2 cells, suggesting that the cloned 5'-flanking region represents the functional promoter of the seabream ghrelin gene, which exhibits tissue-specific promoter activity. Using reverse transcriptase PCR analysis, expression of ghrelin was detected only in the seabream stomach, but not in the other tissues examined, including the brain, gill, intestine, kidney, liver and spleen. This stomach-specific expression of ghrelin in seabream is subject to regulation, as administration of growth hormone or ipamorelin to the fish in vivo was demonstrated to enhance its expression. Reminiscent of the homologous upregulation found in the transcriptional control of the seabream GHSR gene, a similar homologous regulatory mechanism might also exist in controlling the expression of seabream ghrelin. The identification of both GHSR and ghrelin from a single fish species would facilitate our subsequent studies on the elucidation of the physiological functions of the ghrelin/GHSR system in teleost. The possible existence of obestatin in teleost opens up new research avenues on the somatotropic axis in fish.

Prolonged stimulation of growth hormone (GH) and insulin-like growth factor I secretion by CJC-1295, a long-acting analog of GH-releasing hormone, in healthy adults.

J Clin Endocrinol Metab

Sam L Teichman, Ann Neale, Betty Lawrence +3 more

Therapeutic use of GHRH to enhance GH secretion is limited by its short duration of action.

[Evenly tritium-labeled peptides and their in vivo and in vitro biodegradation].

Bioorg Khim

Iu A Zolotarev, A K Dadaian, O V Dolotov +19 more

Biologically active peptides evenly labeled with tritium were used for studying the in vitro and in vivo biodegradation of the peptides. Tritium-labeled peptides with a specific radioactivity of 50-150 Ci/mmol were obtained by high temperature solid phase catalytic isotope exchange (HSCIE) with spillover tritium. The distribution of the isotope label among all amino acid residues of these peptides allows the simultaneous determination of practically all possible products of their enzymatic hydrolysis. The developed analytical method includes extraction of tritium-labeled peptides from organism tissues and chromatographic isolation of individual labeled peptides from the mixture of degradation products. The concentrations of a peptide under study and the products of its biodegradation were calculated from the results of liquid scintillation counting. This approach was used for studying the pathways of biodegradation of the heptapeptide TKPRPGP (Selank) and the tripeptide PGP in blood plasma. The pharmacokinetics of Selank, an anxiolytic peptide, was also studied in brain tissues using the intranasal in vivo administration of this peptide. The concentrations of labeled peptides were determined, and the pentapeptide TKPRP, tripeptide TKP, and dipeptides RP and GP were shown to be the major products of Selank biodegradation. The study of the biodegradation of the heptapeptide MEHFPGP (Semax) in the presence of nerve cells showed that the major products of its biodegradation are the pentapeptide HFPGP and tripeptide PGP. The enkephalinase activity of blood plasma was studied with the use of evenly tritium-labeled [Leu]enkephalin. A high inhibitory effect of Semax on blood plasma enkephalinases was shown to arise from its action on aminopeptidases. The method, based on the use of evenly tritium-labeled peptides, allows the determination of peptide concentrations and the activity of enzymes involved in their degradation on a tg scale of biological samples both in vitro and in vivo.

Impairment of IGF-I gene splicing and MGF expression associated with muscle wasting.

Int J Biochem Cell Biol

Geoffrey Goldspink

The characterisation of a local tissue repair factor (mechano growth factor, MGF) that is produced by exercised and/or damaged muscle by differential splicing of the IGF-I gene provides understanding of how muscle is maintained in the young normal individual. Mechano growth factor, or MGF, is different to the systemic IGF-I as it has an insert of 49 base pairs in exon 5 that introduces a reading frame shift resulting in a C terminal peptide with unique properties. Muscle is a post-mitotic tissue and as cell replacement is not a means of tissue repair there has to be an efficient local repair mechanism otherwise the damaged cells undergo cell death. The extra nuclei for muscle repair and hypertrophy are provided by the muscle satellite (stem) cells. The pool of these stem cells is apparently replenished by the action of MGF, which is produced as a pulse following a mechanical challenge. Unfortunately, the production of MGF is deficient in certain diseases such as in the muscular dystrophies in which the mechanotransduction mechanism, which may involve the dystrophin complex, is defective. In elderly muscles, decreased levels of growth hormone apparently mean that there is less primary RNA transcript of the IGF-I gene to be spliced towards MGF. Consequently, there is an increasing inability to maintain muscle mass during ageing. Delivery of MGF and cDNA or peptide produces marked increases in the strength of normal as well as diseased muscle and, therefore, MGF has considerable potential as a generic means of treating muscle cachexia.

Effects of ghrelin and synthetic GH secretagogues on the cardiovascular system.

Trends Endocrinol Metab

Ji-Min Cao, Huy Ong, Chen Chen

Ghrelin, a newly discovered endogenous hormone that is produced by the stomach, and synthetic peptides have been identified recently as potent growth-hormone secretagogues. This effect is exerted through interaction with a specific G-protein-coupled receptor, GHS-R1a, which is expressed mainly in the hypothalamus-pituitary complex. A study of the peripheral distribution of GHS receptors has shown that it is also present in cardiovascular tissue, which has led to the exploration of the cardiovascular functions of ghrelin and synthetic, growth-hormone-releasing peptides. These ligands have several cardiovascular activities, including a cardioprotective effect against myocardial ischemia, and vasoactive and cardiotropic effects in both experimental models and humans. These effects are mediated by the interaction of these ligands with binding sites, including GHS-1Ra, for which the signalling pathways are not documented fully. Identification of the cardiac and vascular binding sites for ghrelin and synthetic, growth-hormone-releasing peptides will provide new perspectives for treating cardiovascular diseases with these ligands.

[Effects of the new peptide anxiolytic drug selank on the cardiovascular system functioning and respiration in cats].

Eksp Klin Farmakol

T S Gan'shina, I I Kozlovskiĭ

The effects of the new heptapeptide anxiolytic drug selank (300 mg/kg, i.v.) on the arterial pressure (AP), cerebral blood flow, heart rhythm, respiration, and the AP depression induced by acetylcholine injections was recorded in narcotized male cats. Selank produced a 32 +/- 4.3% decrease in the AP for the first 1-3 min after injection. The drug also induced a 24 +/- 2.8% increase in the cerebral blood flow for the first 5 - 10 min upon injection and then the blood flow was gradually restored on the initial level. The heart rate and the respiratory rate were not affected by the drug injections. Nor did selank influence the AP depression in response to the acetylcholine introduction.

[Reduced thymulin production during occupational exposure to lead].

G Ital Med Lav Ergon

L Santarelli, L Di Lorenzo, M Valentino +8 more

Thymulin is a thymic hormone that being activated by binding a zinc ion promotes differentiation and several functions of T lymphocytes. It has been demonstrated only in experimental animals that metallic lead (Pb) is able to cause adverse effects on thymocyte number and function. The objective of this study is to evaluate the plasmatic level of active thymulin of 58 male workers being exposed for more than one year to low lead doses with respect to 59 male never exposed workers. All these were subjected to anamnesis collection, medical examination and determination of blood lead (PbB), plasmatic lead (PbPl), plasmatic thymulin, urinary lead (PbU) and urinary zinc (ZnU) levels. The mean plasma concentration of active thymulin was significantly lower in lead exposed than in non exposed workers. Active thymulin was also significantly and negatively correlated to PbB, PbPl and PbU level and resulted to be significantly and negatively influenced by PbB. Lead exposed workers had slightly higher zinc concentration in urine than non exposed workers, increasing ZnU levels by class of PbB. It is the first time that a toxic effect of lead on plasmatic active thymulin levels is demonstrated in humans, particularly in occupationally exposed workers. This study opens perspectives for further research that would both confirm the results and verify the mechanisms of action of lead on thymulin either direct or indirect and the possible role of zinc.

Muscle regeneration through myostatin inhibition.

Curr Opin Rheumatol

Kathryn R Wagner

Myostatin is an endogenous, negative regulator of muscle growth. Selective inhibition of myostatin may have broad clinical utility by improving regeneration in diverse and burdensome muscle disorders. An understanding of this potential is relevant because inhibitors of myostatin have recently entered clinical trials.

[Comparative study of the long-term behavioral effects of noopept and piracetam in adult male rats and female rats in postnatal period].

Eksp Klin Farmakol

T A Voronina, L S Guzevatykh, S S Trofimov

Adult male and female rats were treated with the peptide nootrope drug noopept (daily dose, 0.1 mg/kg) and piracetam (200 mg/kg). In the period from 8th to 20th day, both drugs (cognitive enhancers) suppressed the horizontal and vertical activity and the anxiety in test animals as compared to the control group treated with 0.9 % aqueous NaCl solution. Early postnatal injections of the nootropes influenced neither the morphology development nor the behavior of adult female rats in the plus maze, extrapolational escape, passive avoidance, and pain sensitivity threshold tests. Animals in the "intact" group (having received neither drugs not physiological solution, that is, developing in a poor sensor environment), showed less pronounced habituation in the open field test as compared to the control and drug treated groups.

[Effect of new synthetic anxiolytic selank on gastric wall blood flow and mesenteryc lymphatic vessels contractility in anesthetized rats].

Ross Fiziol Zh Im I M Sechenova

T S Pavlov, L Ts Sanzhieva, G E Samonina +2 more

The influence of a new heptapeptide Selank on microcirculation in anesthetized white rats was investigated. Selank (Thr-Lys-Pro-Arg-Pro-Gly-Pro) is a new synthetic anxiolytic which exerts obvious antiulcerogenic action and promotes healing of experimental ulcers. Action of the peptide on gastric blood flow in the stomach by using the method ofhydrogenic clearance and Selank action on mesenteric lymphatic contractility by microscopical observation in situ, were studied. Selank fail to influence basal gastric blood flow but it can normalize blood flow reduced by indomethacin. The study of dose-depended of Selank effect on lymphatic vessels contractility showed that its low concentration (10(-12)--10(-14) M) enhanced amplitude and increase frequency of lymphatic vessel contractions which indicates an enhancement of lymphatic flow. The high doses of peptide (10(-6)--10(-10) M) also augmented the contraction amplitude but decreased its frequency. The maintenance of adequate blood flow and lymphatic vessel contractility can be one of the mechanisms of the Selank antiulcerogenic properties.

Chromatographic separation and mass spectrometric identification of positional isomers of polyethylene glycol-modified growth hormone-releasing factor (1-29).

J Chromatogr A

Yu Seok Youn, Dong Hee Na, Sun Dong Yoo +2 more

A one-step chromatographic method capable of separating all isomers of polyethylene glycol (PEG)-growth hormone-releasing factor (GRF) (1-29) conjugates was developed. The unmodified GRF (1-29) and seven different isomers of PEG-GRF (1-29) conjugates were separated by using a simple reversed-phase HPLC method depending on the differences of hydrophobicity due to the number and site of PEG attachment. The PEGylation sites of all isomers of PEG-GRF (1-29) conjugates were identified by determining the molecular masses of the Lys-C digested fragments with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This study is a first report for the separation of all PEG-conjugate isomers and would be useful for further studies to find the promising conjugate by evaluating biological activity and stability of each isomer.

A new property of the synthetic anxiolytic Selank and its derivatives.

Dokl Biol Sci

T S Pavlov, G E Samonina, L A Andreeva +2 more

Growth factors and muscle ageing.

Exp Gerontol

Geoffrey Goldspink, Stephen D R Harridge

Loss of muscle mass (sarcopenia) is one of the main problems associated with ageing as it has major health care as well as socioeconomic implications. The growth hormone (GH)/IGF-I axis is regarded as an important regulator of muscle mass. However, it is now appreciated that other tissues in addition to the liver express IGF-I and that there are local as well as systemic forms of IGF-I which have different functions. At least two different kinds of IGF-I that are expressed by skeletal muscle are derived from the IGF-I gene by alternative splicing, one of which is expressed in response to physical activity which has now been called 'mechano growth factor' (MGF). The other is similar to the systemic or liver type (IGF-IEa) and is important as the provider of mature IGF-I required for upregulating protein synthesis. MGF differs from systemic IGF-IEa in that it has a different peptide sequence which is responsible for replenishing the satellite (stem) cells in skeletal muscle. The ability to produce MGF declines with age, and this is commensurate with the decline in circulating GH levels. GH treatment up regulates the level of IGF-I gene expression in older people and when combined with resistance exercise more is spliced towards MGF and hence should improve the ability of muscle to respond to physical activity. The possibility of ameliorating sarcopenia using MGF is discussed.

Mechanical signals and IGF-I gene splicing in vitro in relation to development of skeletal muscle.

J Cell Physiol

Umber Cheema, Robert Brown, Vivek Mudera +3 more

It has been shown that the insulin-like growth factor (IGF-I) gene is spliced in response to mechanical signals producing forms of IGF-I which have different actions. In order to study how mechanical signals influence this gene splicing in developing muscle, C2C12 cells were grown in three-dimensional (3D) culture and subjected to different regimens of mechanical strain. IGF-IEa which initiates the fusion of myoblasts to form myotubes was found to be constitutively expressed in myoblasts and myotubes (held under endogenous tension) and its expression upregulated by a single ramp stretch of 1-h duration but reduced by repeated cyclical stretch. In contrast, mechano growth factor (MGF), which is involved in the proliferation of mononucleated myoblasts that are required for secondary myotube formation and to establish the muscle satellite (stem) cell pool, showed no significant constitutive expression in static cultures, but was upregulated by a single ramp stretch and by cycling loading. The latter types of force simulate those generated in myoblasts by the first contractions of myotubes. These data indicate the importance of seeking to understand the physiological signals that determine the ratios of splice variants of some growth factor/tissue factor genes in the early stages of development of skeletal muscle.