Human Growth Hormone-releasing Research Articles

Intranasal delivery of a ghrelin mimetic engages the brain ghrelin signalling system in mice

Abstract Ghrelin, the endogenous ligand of the growth hormone secretagogue receptor (GHSR), promotes food intake, other feeding behaviours and stimulates growth hormone (GH) release from the pituitary. Growth hormone secretagogues (GHS), such as GHRP-6 and MK-0677, are synthetic GHSR ligands that activate orexigenic Neuropeptide Y neurons that co-express Agouti-Related Peptide (AgRP) in the arcuate nucleus of the hypothalamus when administered systemically. Systemic GHRP-6 also stimulates GH release in humans and rats. Thus, GHS and ghrelin have therapeutic relevance in patients who could benefit from its orexigenic and/or GH-releasing effects. This study examined whether intranasal delivery of ghrelin, GHRP-6, or MK-0677 engages the brain ghrelin signalling system. Effective compounds and doses were selected based on increased food intake after intranasal application in mice. Only GHRP-6 (5 mg/kg) increased food intake without adverse effects, prompting detailed analysis of meal patterns, neuronal activation in the arcuate nucleus (via Fos mapping) and neurochemical identification of c-fos mRNA-expressing neurons using RNAscope. We also assessed the impact of intranasal GHRP-6 on serum GH levels. Intranasal GHRP-6 increased food intake by increasing meal frequency and size. Fos expression in the arcuate nucleus was higher in GHRP-6-treated mice than saline controls. When examining the neurochemical identity of c-fos-mRNA-expressing neurons, we found co-expression with 63.5±1.9% Ghsr-mRNA, 79±6.8% Agrp-mRNA and 11.4±2.5% Ghrh-mRNA, demonstrating GHRP-6’s ability to engage arcuate nucleus neurons involved in food intake and GH release. Additionally, intranasal GHRP-6 elevated GH serum levels. These findings suggest that intranasal GHRP-6, but not ghrelin or MK-0677, can engage the brain ghrelin signalling system.

Preparation and characterization of the injectable pH- and temperature-sensitive pentablock hydrogel containing human growth hormone-loaded chitosan nanoparticles via electrospraying

This research investigated the in vivo gelation, biodegradation, and drug release efficiency of a novel injectable sensitive drug delivery system for human growth hormone (HGh). This composite system comprises pH- and temperature-sensitive hydrogel, designated as oligomer serine-b-poly(lactide)-b-poly(ethylene glycol)-b-poly(lactide)-b-oligomer serine (OS-PLA-PEG-PLA-OS) pentablock copolymer, as matrix and electrosprayed HGh-loaded chitosan (HGh@CS) nanoparticles (NPs) as principal material. The proton nuclear magnetic resonance spectrum of the pH- and temperature-sensitive OS-PLA-PEG-PLA-OS pentablock copolymer hydrogel proved that this copolymer was successfully synthesized. The HGh was encapsulated in chitosan (CS) NPs by an electrospraying system in acetic acid with appropriate granulation parameters. The scanning electron microscopy images and size distribution showed that the HGh@CS NPs formed had an average diameter of 366.1 ± 214.5 nm with a discrete spherical shape and dispersed morphology. The sol–gel transition of complex gel based on HGh@CS NPs and OS-PLA-PEG-PLA-OS pentablock hydrogel was investigated at 15 °C and pH 7.8 in the sol state and gelled at 37 °C and pH 7.4, which is suitable for the physiological conditions of the human body. The HGh release experiment of the composite system was performed in an in vivo environment, which demonstrated the ability to release HGh, and underwent biodegradation within 32 days. The findings of the investigation revealed that the distribution of HGh@CS NPs into the hydrogel matrix not only improved the mechanical properties of the gel matrix but also controlled the drug release kinetics into the systematic bloodstream, which ultimately promotes the desired therapeutic body growth depending on the distinct concentration used.

Silica-collagen nanoformulations with extended human growth hormone release

Growth hormone deficiency has been treated by the daily administration of recombinant human growth hormone (hGH) for decades. Patient compliance to this treatment is generally incomplete due to challenges including dose frequency and lack of perceived benefits. This stimulates the research on new formulations to reduce the number of periodic administrations. In this study silica nanoparticles and silica-collagen nanocomposites were evaluated for hGH loading and release. Bare nanoparticles showed higher hGH adsorption capacity than thiol- and isobutyl-bearing particles of similar diameters. Monitoring of bound protein conformation changes indicated hGH structure retention when adsorbed on bare silica nanoparticles and suggested no alterations on protein activity. Protein-loaded particles incorporated into collagen matrices (silica-collagen nanocomposites) showed a progressive protein release profile different from the observed for hGH-loaded silica nanoparticles and hGH-loaded collagen matrices. While both the collagen and the silica nanoparticle systems reached a 100 % release after 4 and 7 days respectively, silica-collagen nanocomposites showed a bi-phasic prolonged hGH release reaching approximately an 80 % after 15 days. These findings suggest that biocompatible silica-collagen nanocomposites could be used as vehicles for the prolonged delivery of hGH which could lead to a potential reduction in the number of periodic administrations.

Actively separated microneedle patch for sustained-release of growth hormone to treat growth hormone deficiency

Growth hormone deficiency (GHD) has become a serious healthcare burden, and presents a huge impact on the physical and mental health of patients. Here, we developed an actively separated microneedle patch (PAA/NaHCO3-Silk MN) based on silk protein for sustained release of recombinant human growth hormone (rhGH). Silk protein, as a friendly carrier material for proteins, could be constructed in mild full-water conditions and ensure the activity of rhGH. After manually pressing PAA/NaHCO3-Silk MN patch to skin for 1min, active separation is achieved by absorbing the interstitial fluid (ISF) to trigger HCO3 ‒ in the active backing layer to produce carbon dioxide gas (CO2). In rats, the MN patch could maintain the sustained release of rhGH for more than 7 days, and produce similar effects as daily subcutaneous (S.C.) injections of rhGH in promoting height and weight with well tolerated. Moreover, the PAA/NaHCO3-Silk MN patch with the potential of painless self-administration, does not require cold chain transportation and storage possess great economic benefits. Overall, the PAA/NaHCO3-Silk MN patch can significantly improve patient compliance and increase the availability of drugs, meet current unmet clinical needs, improve clinical treatment effects of GHD patients.

562 Neuroendocrinology of human scalp hair follicles: Growth hormone-releasing hormone (GHRH) stimulates hair growth and controls a fully functional intrafollicular hypothalamic-pituitary-somatotropic (HPS) signaling axis

Human scalp hair follicles (HFs) are a major non-classical site of neurohormone production. While we had previously shown that growth hormone (GH), unexpectedly, inhibits female human scalp HFs growth ex vivo, it is unknown whether human HFs also produce key HPS axis neurohormones themselves, i.e. GH, its upstream regulator, GHRH, and/or the negative feedback control, somatostatin (SST), and whether GHRH modulates hair growth. To answer these questions, human scalp HFs were organ-cultured and stimulated with human GH (100 or 300ng/mL), GHRH (100 or 300ng/mL), or vehicle for mRNA or protein analysis of key HPS axis elements.

Constitutive signal bias mediated by the human GHRHR splice variant 1

Alternative splicing of G protein-coupled receptors has been observed, but their functions are largely unknown. Here, we report that a splice variant (SV1) of the human growth hormone-releasing hormone receptor (GHRHR) is capable of transducing biased signal. Differing only at the receptor N terminus, GHRHR predominantly activates Gs while SV1 selectively couples to β-arrestins. Based on the cryogenic electron microscopy structures of SV1 in the apo state or GHRH-bound state in complex with the Gs protein, molecular dynamics simulations reveal that the N termini of GHRHR and SV1 differentiate the downstream signaling pathways, Gs versus β-arrestins. As suggested by mutagenesis and functional studies, it appears that GHRH-elicited signal bias toward β-arrestin recruitment is constitutively mediated by SV1. The level of SV1 expression in prostate cancer cells is also positively correlated with ERK1/2 phosphorylation but negatively correlated with cAMP response. Our findings imply that constitutive signal bias may be a mechanism that ensures cancer cell proliferation.

Antagonists of Growth Hormone-Releasing Hormone Inhibit the Growth of Pituitary Adenoma Cells by Hampering Oncogenic Pathways and Promoting Apoptotic Signaling.

Simple SummaryMany studies have demonstrated that the antagonists of growth hormone-releasing hormone (GHRH) exert inhibitory activities in a variety of experimental cancers; however, their potential antitumor role in pituitary adenomas (PAs) remains largely unknown. Here, we show that GHRH antagonists of Miami (MIA) class, MIA-602 and MIA-690, are able to reduce the growth and promote cell death in hormone-secreting PA cell lines, through the inhibition of mechanisms implicated in tumorigenesis and cancer progression. MIA-602 and MIA-690 also decreased the viability of tumor cells derived from human pituitary tumors. Overall, these findings suggest that GHRH antagonists may represent new therapeutic tools for the treatment of PAs, both alone or in combination with standard pharmacological treatments.Pituitary adenomas (PAs) are intracranial tumors, often associated with excessive hormonal secretion and severe comorbidities. Some patients are resistant to medical therapies; therefore, novel treatment options are needed. Antagonists of growth hormone-releasing hormone (GHRH) exert potent anticancer effects, and early GHRH antagonists were found to inhibit GHRH-induced secretion of pituitary GH in vitro and in vivo. However, the antitumor role of GHRH antagonists in PAs is largely unknown. Here, we show that the GHRH antagonists of MIAMI class, MIA-602 and MIA-690, inhibited cell viability and growth and promoted apoptosis in GH/prolactin-secreting GH3 PA cells transfected with human GHRH receptor (GH3-GHRHR), and in adrenocorticotropic hormone ACTH-secreting AtT20 PA cells. GHRH antagonists also reduced the expression of proteins involved in tumorigenesis and cancer progression, upregulated proapoptotic molecules, and lowered GHRH receptor levels. The combination of MIA-690 with temozolomide synergistically blunted the viability of GH3-GHRHR and AtT20 cells. Moreover, MIA-690 reduced both basal and GHRH-induced secretion of GH and intracellular cAMP levels. Finally, GHRH antagonists inhibited cell viability in human primary GH- and ACTH-PA cell cultures. Overall, our results suggest that GHRH antagonists, either alone or in combination with pharmacological treatments, may be considered for further development as therapy for PAs.

Release of human growth hormone from an in-situ implant modulated by poly(ethylene glycol) dimethyl ether and tris(hydroxymethyl) aminomethane

A patient-friendly delivery system to release human growth hormone (hGH) is very desirable. In situ forming implant systems (ISIs) can provide a long acting and effective protein delivery. In these systems, solvents and additives play major roles in drug release. In this study, four groups of PLGA-based ISIs containing hGH were prepared in N-methyl-2-pyrrolidone (NMP) and poly(ethylene glycol) dimethyl ether (PEG-DME) as solvents with and without tris(hydroxymethyl) aminomethane (Tris) as stabilizer. Several analyses were used to investigate the implants, which include release profile, viscosity, contact angle, gel permeation chromatography (GPC), scanning electron microscopy (SEM), hGH and IGF-1 serum measurements and histopathology. In in vitro release experiments, the hGH cumulative release from PEG-DME system was twice that from NMP system during 14 days, and hGH release was tripled in the presence of Tris. With the addition of Tris to the ISIs containing PEG-DME, the water penetration, interconnectivity of pores and inner channels, surface pores and hydrophilicity were increased. Moreover, the effect of Tris on the hGH stabilization synergized its positive effects and increased the hGH final cumulative release. Results of the ISIs containing PEG-DME and Tris injection in rabbits demonstrated a reduced tissue inflammation. Moreover, the 14-days serum levels of the hGH and IGF-1 of this system in recipient rabbits were comparable to those of the commercial daily injection samples.

MON-LB003 Clinical Case Series of Augmented Fertility in Females After Administration of an Amino Acid Blend That Enhances Release of Human Growth Hormone

Supplementation with adjuvant therapies, including growth hormone (hGH), is commonly used to improve fertility treatment outcomes. hGH is important for normal female fertility; low hGH has been associated with causes of infertility and impaired fertility, including polycystic ovarian syndrome (PCOS), endometriosis, diminished ovarian reserve (DOR), and advanced maternal age. A novel, low dose, orally administered amino acid blend has been previously shown in a double-blind, randomized, placebo-controlled, crossover clinical trial to produce a statistically significant increase in endogenous hGH secretion. In this clinical case series, we report outcomes in 7 women with infertility or impaired fertility and conditions associated with low hGH who administered the amino acid blend during fertility treatment (n=5) or timed intercourse/spontaneous pregnancy (n=2). Medical history included conditions associated with impaired fertility and low hGH: endometriosis (n=3), PCOS (n=2), and poor response to ovarian stimulation/history of failed in vitro fertilization (IVF) (n=3). The amino acid blend (containing 2.9 g of L-lysine, L-arginine, oxo-proline, N-acetyl-L-cysteine, L-glutamine, and schizonepeta) was administered daily on an empty stomach. Outcomes included embryo quality and success of embryo transfer (for IVF) and successful pregnancy/live births. Mean±SD age was 33±5 years (range 27-38) and BMI was 27±7 kg/m2 (range 21-37). Time to pregnancy ranged from 1 week to 9 months (median 3 months) prior to egg retrieval for IVF (n=4), intrauterine insemination (n=1), or timed intercourse/spontaneous pregnancy (n=2). For women with a history of failed IVF (n=3), there was an improvement in oocyte retrieval, a higher fertilization rate, and a greater number of high-quality embryos compared to previous IVF attempts. There were 2 twin pregnancies (both following IVF). All 7 pregnancies resulted in live births. Consistent with previous studies, the amino acid blend was well tolerated; no adverse events were observed. We report a case series of successful pregnancy in 7 women with conditions associated with low hGH including PCOS, endometriosis, and poor response to ovarian stimulation/history of failed IVF who administered the amino acid blend concomitant with fertility treatment or who reported spontaneous pregnancy. This may represent a potential low-risk and cost-effective treatment to improve IVF success and increase pregnancy rates in individuals with infertility or impaired fertility.

Orphan designation: Recombinant protein consisting of modified human growth hormone releasing hormone and the translocation and endopeptidase domains of botulinum toxin serotype D, Treatment of acromegaly

Orphan designation: Recombinant protein consisting of modified human growth hormone releasing hormone and the translocation and endopeptidase domains of botulinum toxin serotype D, Treatment of acromegaly

Abacavir, nevirapine, and ritonavir modulate intracellular calcium levels without affecting GHRH-mediated growth hormone secretion in somatotropic cells in vitro

Growth Hormone (GH) deficiency is frequent in HIV-infected patients treated with antiretroviral therapy. We treated GH3 cells with antiretrovirals (nevirapine, ritonavir or abacavir sulfate; 100 pM-1 mM range), after transfection with human growth hormone releasing hormone (GHRH) receptor cDNA. Cells viability, intracellular cAMP, phosphorylation of CREB and calcium increase, GH production and secretion were evaluated both in basal condition and after GHRH, using MTT, bioluminescence resonance energy transfer, western blotting and ELISA.Antiretroviral treatment did not affect GHRH 50% effective dose (EC50) calculated for 30-min intracellular cAMP increase (Mann-Whitney's U test; p ≥ 0.05; n = 4) nor 15-min CREB phosphorylation. The kinetics of GHRH-mediated, rapid intracellular calcium increase was perturbed by pre-incubation with drugs, while GHRH failed to induce the ion increase in ritonavir pre-treated cells (ANOVA; p < 0.05; n = 3). Antiretrovirals did not impact 24-h intracellular and extracellular GH levels (ANOVA; p ≥ 0.05; n = 3).We demonstrated the association between antiretrovirals and intracellular calcium increase, without consequences on somatotrope cells viability and GH synthesis. Overall, these results suggest that antiretrovirals may not directly impact on GH axis in HIV-infected patients.

Growth Hormone Deficiency Is Frequent After Recent Stroke.

Introduction: The incidence of pituitary dysfunction after severe ischemic stroke is unknown, however given the increasing attention to pituitary dysfunction after neurological injuries such as traumatic brain injury, this may represent a novel area of research in stroke.Methods: We perform an arginine and human growth hormone releasing hormone challenge on ischemic stroke patients within a week of symptom onset.Results: Over the study period, 13 patients were successfully tested within a week of stroke (baseline NIHSS 10, range 7–16). Overall, 9(69%) patients had a poor response, with 7(54%) of these patients meeting the criteria for had human growth hormone deficiency. Other measures of pituitary function were within normal ranges.Conclusion: After major ischemic stroke, low GH levels are common and may play a role in stroke recovery.

Analysis of new growth promoting black market products

Detecting agents allegedly or evidently promoting growth such as human growth hormone (GH) or growth hormone releasing peptides (GHRP) in doping controls has represented a pressing issue for sports drug testing laboratories. While GH is a recombinant protein with a molecular weight of 22 kDa, the GHRPs are short (3–6 amino acids long) peptides with GH releasing properties. The endogenously produced GH (22 kDa isoform) consists of 191 amino acids and has a monoisotopic molecular mass of 22,124 Da. Within this study, a slightly modified form of GH was discovered consisting of 192 amino acids carrying an additional alanine at the N-terminus, leading to a monoisotopic mass of 22,195 Da. This was confirmed by top-down and bottom-up experiments using liquid chromatography coupled to high resolution/high accuracy mass spectrometry.Additionally, three analogues of GHRPs were identified as Gly-GHRP-6, Gly-GHRP-2 and Gly-Ipamorelin, representing the corresponding GHRP extended by a N-terminal glycine residue. The structure of these peptides was characterised by means of high resolution (tandem) mass spectrometry, and for Gly-Ipamorelin and Gly-GHRP-2 their identity was additionally confirmed by custom synthesis. Further, established in-vitro experiments provided preliminary information considering the potential metabolism after administration.

Sustained Release of Recombinant Human Growth Hormone from Bioresorbable Poly(ester urea) Nanofibers

Recombinant human growth hormone (rhGH) has been used clinically for several decades. However, rhGH has a short half-life in vivo and therefore for efficacy requires frequent subcutaneous injection, leading to pain and poor patient compliance. An effective rhGH delivery system able to continuously release rhGH over a comparatively long period (generally over a month) would be a significant clinical advance. In this study, a novel long-duration release strategy of rhGH was developed by encapsulating sugar glass-stabilized rhGH in electrospun, bioresorbable poly(ester urea) (PEU) nanofibers. The rhGH was found to be randomly dispersed among the fibers, and sustained rhGH release with only a modest burst was observed for at least 6 weeks as studied by a bicinchoninic acid (BCA) protein assay. Significantly, the released rhGH remained bioavailable and bioactive as determined by an Nb2 cell bioassay specific for rhGH. Our results suggest the feasibility of this system as an effective long-term sustained releas...

Assessment of Adhesion and Proliferation of Bone Marrow Mesenchymal Stem Cells in Polymer Matrices with rhGH

Biomaterials, as an alternative to autogenous bone and other biologic tissues, have been widely used in oral and maxillofacial surgery. In this context, a biomaterial that functions as a scaffold (osteoconductor), combined with a growth factor (osteoinductor), would be of great interest for clinical application. Biodegradable polymers used for slow drug release have been investigated, demonstrating good results and interesting potential. Growth hormone (GH) may be released by incorporating it into these polymers. This study aimed to evaluate cell adhesion and proliferation of a polymeric biomaterial for slow release of recombinant human GH (rhGH). Poly(lactic-co-glycolic acid) (PLGA) and PLGA/polycaprolactone (PCL) (at a 70/30 ratio of PLGA to PCL) matrices were prepared by the solvent evaporation method, combined or not with GH. Biomaterials were tested for cell adhesion and proliferation by culture in mesenchymal stem cells derived from Wistar rat bone marrow, 4',6-diamidino-2-phenylindole (DAPI) staining, and subsequent cell counting, in addition to scanning electron microscopy. Cell adhesion and proliferation was assessed at 24 and 72 hours of biomaterial exposure to culture medium. All tested polymers exhibited cell adhesion and proliferation. However, PLGA-based biomaterials, especially when combined with GH, showed greater cell proliferation when the difference in growth from 24 to 72 hours was evaluated. GH appeared to modify the polymer surface, with increased roughness and microporosity. This feature was more evident in the PLGA + GH combination. The biomaterials tested showed pronounced cell adhesion in all test groups, and GH appeared to contribute to the increase in cell proliferation, especially when combined with PLGA as compared with pure PLGA. Further studies are required to clarify this potential for development of new biomaterials.

Synthesis and structure-activity studies on novel analogs of human growth hormone releasing hormone (GHRH) with enhanced inhibitory activities on tumor growth

The syntheses and biological evaluations of new GHRH analogs of Miami (MIA) series with greatly increased anticancer activity are described. In the design and synthesis of these analogs, the following previous substitutions were conserved: D-Arg2, Har9, Abu15, and Nle27. Most new analogs had Ala at position 8. Since replacements of both Lys12 and Lys21 with Orn increased resistance against enzymatic degradation, these modifications were kept. The substitutions of Arg at both positions 11 and 20 by His were also conserved. We kept D-Arg28, Har29 –NH2 at the C-terminus or inserted Agm or 12-amino dodecanoic acid amide at position 30. We incorporated pentafluoro-Phe (Fpa5), instead of Cpa, at position 6 and Tyr(Me) at position 10 and ω-amino acids at N-terminus of some analogs. These GHRH analogs were prepared by solid-phase methodology and purified by HPLC. The evaluation of the activity of the analogs on GH release was carried out in vitro on rat pituitaries and in vivo in male rats. Receptor binding affinities were measured in vitro by the competitive binding analysis. The inhibitory activity of the analogs on tumor proliferation in vitro was tested in several human cancer cell lines such as HEC-1A endometrial adenocarcinoma, HCT-15 colorectal adenocarcinoma, and LNCaP prostatic carcinoma. For in vivo tests, various cell lines including PC-3 prostate cancer, HEC-1A endometrial adenocarcinoma, HT diffuse mixed β cell lymphoma, and ACHN renal cell carcinoma cell lines were xenografted into nude mice and treated subcutaneously with GHRH antagonists at doses of 1–5μg/day. Analogs MIA-602, MIA-604, MIA-610, and MIA-640 showed the highest binding affinities, 30, 58, 48, and 73 times higher respectively, than GHRH (1-29) NH2. Treatment of LNCaP and HCT-15 cells with 5μM MIA-602 or MIA-690 decreased proliferation by 40%–80%. In accord with previous tests in various human cancer lines, analog MIA-602 showed high inhibitory activity in vivo on growth of PC-3 prostate cancer, HT-mixed β cell lymphoma, HEC-1A endometrial adenocarcinoma and ACHN renal cell carcinoma. Thus, GHRH analogs of the Miami series powerfully suppress tumor growth, but have only a weak endocrine GH inhibitory activity. The suppression of tumor growth could be induced in part by the downregulation of GHRH receptors levels.

Protection of neonatal rat cardiac myocytes against radiation-induced damage with agonists of growth hormone-releasing hormone

Despite the great clinical significance of radiation-induced cardiac damage, experimental investigation of its mechanisms is an unmet need in medicine. Beneficial effects of growth hormone-releasing hormone (GHRH) agonists in regeneration of the heart have been demonstrated. The aim of this study was the evaluation of the potential of modern GHRH agonistic analogs in prevention of radiation damage in an in vitro cardiac myocyte-based model.Cultures of cardiac myocytes isolated from newborn rats (NRVM) were exposed to a radiation dose of 10Gy. The effects of the agonistic analogs, JI-34 and MR-356, of human GHRH on cell viability, proliferation, their mechanism of action and the protein expression of the GHRH/SV1 receptors were studied.JI-34 and MR-356, had no effect on cell viability or proliferation in unirradiated cultures. However, in irradiated cells JI-34 showed protective effects on cell viability at concentrations of 10 and 100nM, and MR-356 at 500nM; but no such protective effect was detected on cell proliferation. Both agonistic analogs decreased radiation-induced ROS level and JI-34 interfered with the activation of SAFE/RISK pathways. Using Western blot analysis, a 52kDa protein isoform of GHRHR was detected in the samples in both irradiated and unirradiated cells.Since GHRH agonistic analogs, JI-34 and MR-356 alleviated radiation-induced damage of cardiac myocytes, they should be tested in vivo as potential protective agents against radiogenic heart damage.

Assessment of dose–effect and therapeutic time window in preclinical studies of rhEGF and GHRP-6 coadministration for stroke therapy

Background: Stroke continues to be a leading cause of mortality and morbidity worldwide, and novel therapeutic options for ischaemic stroke are urgently needed. In this context, drug combination therapies seem to be a viable approach, which has not been fully explored in preclinical studies.Objectives: In this work, we assessed the dose–response relationship and therapeutic time window, in global brain ischaemia, of a combined therapeutic approach of recombinant human epidermal growth factor (EGF) and growth hormone-releasing peptide-6 (GHRP-6).Methods: Mongolian gerbils underwent 15 minutes occlusion of both common carotid arteries. Four different doses of rhEGF, GHRP-6 and these combined agents were intraperitoneally administered immediately after the onset of reperfusion. Having identified a better response with both agents, rhEGF+GHRP-6 were administered at 2, 4, 6, 8 or 24 hours after the onset of reperfusion to assess the time window of effectiveness. Animals were evaluated daily for neurological deficits. Three days post-occlusion, the animals were sacrificed and 2,3,5-triphenyltetrazolium chloride was used to quantify infarcted tissues.Results: The coadministration of rhEGF and GHRP-6 at doses of 100 and 600 μg/kg, respectively, administered up to 4 hours following the ischaemic insult, significantly improved survival and neurological outcome, and reduced infarct volume compared with vehicle treatment. These results are considered as an additional proof of concept as supporting a combined therapeutic approach and justify the further development of this preclinical research.

Central and peripheral metabolic changes induced by gamma-hydroxybutyrate.

Gamma-hydroxybutyrate (GHB) was originally introduced as an anesthetic but was first abused by bodybuilders and then became a recreational or club drug.1 Sodium salt of GHB is currently used for the treatment of cataplexy in patients with narcolepsy. The mode of action and metabolism of GHB is not well understood. GHB stimulates growth hormone release in humans and induces weight loss in treated patients, suggesting an unexplored metabolic effect. In different experiments the effect of GHB administration on central (cerebral cortex) and peripheral (liver) biochemical processes involved in the metabolism of the drug, as well as the effects of the drug on metabolism, were evaluated in mice. C57BL/6J, gamma-aminobutyric acid B (GABAB) knockout and obese (ob/ob) mice were acutely or chronically treated with GHB at 300 mg/kg. Respiratory ratio decreased under GHB treatment, independent of food intake, suggesting a shift in energy substrate from carbohydrates to lipids. GHB-treated C57BL/6J and GABAB null mice but not ob/ob mice gained less weight than matched controls. GHB dramatically increased the corticosterone level but did not affect growth hormone or prolactin. Metabolome profiling showed that an acute high dose of GHB did not increase the brain GABA level. In the brain and the liver, GHB was metabolized into succinic semialdehyde by hydroxyacid-oxoacid transhydrogenase. Chronic administration decreased glutamate, s-adenosylhomocysteine, and oxidized gluthathione, and increased omega-3 fatty acids. Our findings indicate large central and peripheral metabolic changes induced by GHB with important relevance to its therapeutic use.

Butyrate increases intracellular calcium levels and enhances growth hormone release from rat anterior pituitary cells via the G-protein-coupled receptors GPR41 and 43.

Butyrate is a short-chain fatty acid (SCFA) closely related to the ketone body ß-hydroxybutyrate (BHB), which is considered to be the major energy substrate during prolonged exercise or starvation. During fasting, serum growth hormone (GH) rises concomitantly with the accumulation of BHB and butyrate. Interactions between GH, ketone bodies and SCFA during the metabolic adaptation to fasting have been poorly investigated to date. In this study, we examined the effect of butyrate, an endogenous agonist for the two G-protein-coupled receptors (GPCR), GPR41 and 43, on non-stimulated and GH-releasing hormone (GHRH)-stimulated hGH secretion. Furthermore, we investigated the potential role of GPR41 and 43 on the generation of butyrate-induced intracellular Ca2+ signal and its ultimate impact on hGH secretion. To study this, wt-hGH was transfected into a rat pituitary tumour cell line stably expressing the human GHRH receptor. Treatment with butyrate promoted hGH synthesis and improved basal and GHRH-induced hGH-secretion. By acting through GPR41 and 43, butyrate enhanced intracellular free cytosolic Ca2+. Gene-specific silencing of these receptors led to a partial inhibition of the butyrate-induced intracellular Ca2+ rise resulting in a decrease of hGH secretion. This study suggests that butyrate is a metabolic intermediary, which contributes to the secretion and, therefore, to the metabolic actions of GH during fasting.