GHRP-6 Research Articles

Growth hormone releasing peptide-6 (GHRP-6) and other related secretagogue synthetic peptides: A mine of medical potentialities for unmet medical needs

Growth hormone releasing peptide-6 (GHRP-6) and other related secretagogue synthetic peptides: A mine of medical potentialities for unmet medical needs

Human Ghrelin: A Gastric Hormone with Cardiovascular Properties.

Ghrelin is a growth hormone-releasing peptide, isolated from the stomach. Researches in progress documented that ghrelin participates in the stimulation of the hypothalamus-pituitary-adrenal axis at the hypothalamic level and in the regulation of energy balance. Growth hormone-independent functions have been ascribed to ghrelin. Among others, a large body of literature demonstrated the presence of specific receptors for ghrelin, distributed at the level of cardiomyocytes and endothelial cells. Therefore, a link between ghrelin and cardiovascular system has been hypothesized and, then, demonstrated in both experimental and clinical studies. Ghrelin has largely documented cardiac beneficial effects, including protection from ischemia/reperfusion injury, attenuation of left ventricular remodeling following myocardial infarction, and improvement of left ventricular function. Exercise level in patients with chronic heart failure had also been seen. Ghrelin exerts these effects through several mechanisms, including the inhibition of apoptosis. At the level of blood vessels, ghrelin exerts a significant impact on vascular function. In particular, acutely infused, ghrelin reverses endothelial dysfunction by increasing NO availability and restores the endothelin-1/nitric oxide imbalance in the peripheral microcirculation of patients with metabolic syndrome. Antioxidant/anti-inflammatory effects, and-or an ameliorated insulin sensitivity are proposed mechanisms whereby ghrelin exerts its vascular protective actions. At higher doses, ghrelin also decreases blood pressure, by mechanisms that involve the modulation of sympathetic nervous system. This finding highlights the ghrelin system as a promising candidate for cardiovascular drug discovery.

Long-term follow-up results of growth hormone therapy for patients with adult growth hormone deficiency.

We evaluated the long-term effects of growth hormone (GH) on markers of quality of life, glucose metabolism, and lipid metabolism to validate the adequacy of long-term GH replacement therapy for adult GH deficiency (AGHD). Eighty-three of 100 sequentially followed patients who received GH therapy were selected for this study. Forty-nine were men aged 26 to 78 years (mean, 52 years) and 34 were women aged 20 to 78 years (mean, 56 years). The GH-releasing peptide-2 stimulation test and arginine stimulation test were used to diagnose AGHD. The adult hypopituitarism questionnaire (AHQ) and biochemical parameters such as cholesterol (TC), triglyceride (TG), high-density lipoprotein (HDL) cholesterol and low-density lipoprotein (LDL) cholesterol, and gyrated hemoglobin (HbA1c) were determined before treatment, at 6 months of treatment, and at 1, 2, 3, 4, 5, 6, 7, and 8 years of treatment. Considering age and sex as factors potentially influencing the effect of GH therapy, the patients were divided into age groups of <60 and ≥60 years and sex groups of men and women. Repeated measured analysis of variance (ANOVA) was employed. ANOVA demonstrated significant changes in mean AHQ scores during follow-up. Comparison of individual AHQ scores with baseline values revealed sequential improvements, stabilization, and decline in QOL. A significant elevation in HbA1c level was demonstrated. LDL-C and HDL-C levels changed significantly upon GH treatment regardless of sex or age. Levels of glucose, TC or TG did not change significantly. The effect of GH therapy on QOL showed sequential improvements and stabilization until 6-year follow-up.

Oral administration of the growth hormone secretagogue-6 (GHRP-6) enhances growth and non-specific immune responses in tilapia (Oreochromis sp.)

Aquaculture is part of the solution for meeting the growing world demand for food, mainly as an animal protein source. However, current yields are insufficient for aquaculture to play this crucial role. Growth-rate enhancement is one of the approaches that have been exploited in this regard. Also, losses caused by bacteria, viruses and parasites remain a significant problem. It has been demonstrated that growth stimulants contribute not only to growth enhancement but also to fish health improvement. Growth Hormone Releasing Peptide-6 (GHRP-6) is one of the earliest-developed, synthetic, peptidyl growth hormone secretagogue receptor agonists. These compounds mimic the effect of the endogenous ligand, ghrelin. This peptide has shown its benefits for both growth enhancement and immune system stimulation in fish. In the present study, we demonstrate that intraperitoneal administration of GHRP-6 induces liver insulin growth factor-I messenger RNA, and increases growth hormone levels in juvenile tilapia (Oreochromis sp.), in a time-course experiment. GHRP-6 also stimulates growth using non-encapsulated and encapsulated peptide administered by a plastic tube to the pharyngeal cavity to juveniles. In addition, administration of GHRP-6 in formulated feed to tilapia larvae was assessed. Growth and immune parameters such as lectin titers and intestinal intraepithelial lymphocyte numbers were increased in treated larvae. Based on these results, GHRP-6 could be an interesting candidate for development into a new biotechnology product for improving fish farming. Statement of relevanceThe results presented in the paper are a pioneer experimentation to look for molecules to improve the aquaculture mainly in the intensive culture in order to protect the fish from the pathogens and at the same time to improve the growth rate.

Solid‐phase extraction of small biologically active peptides on cartridges and microelution 96‐well plates from human urine

Currently liquid chromatography - mass spectrometry (LC-MS) analysis after solid-phase extraction (SPE) on weak cation-exchange cartridges is a method of choice for anti-doping analysis of small bioactive peptides such as growth hormone releasing peptides (GHRPs), desmoporessin, LHRH, and TB-500 short fragment. Dilution of urine samples with phosphate buffer for pH adjustment and SPE on weak cation exchange microelution plates was tested as a means to increase throughput of this analysis. Dilution using 200 mM phosphate buffer provides good buffering capacity without affecting the peptides recoveries. SPE on microelution plates was performed on Waters Positive Pressure-96 Processor with subsequent evaporation of eluates in nitrogen flow. Though the use of smaller sample volume decreases the pre-concentration factor and increases the limits of detection of 5 out of 17 detected peptides, the recovery, linearity, and reproducibility of the microelution extraction were comparable with cartridge SPE. The effectiveness of protocols was confirmed by analysis of urine samples containing ipamorelin, and GHRP-6 and its metabolites. SPE after urine sample dilution with buffer can be used for faster sample preparation. The use of microelution plates decreases consumption of solvents and allows processing of up to 96 samples simultaneously. Cartridge SPE with manual рН adjustment remains the best option for confirmation. Copyright © 2015 John Wiley & Sons, Ltd.

Growth hormone-releasing peptide-biotin conjugate stimulates myocytes differentiation through insulin-like growth factor-1 and collagen type I.

Based on the potential beneficial effects of growth hormone releasing peptide (GHRP)-6 on muscle functions, a newly synthesized GHRP-6-biotin conjugate was tested on cultured myoblast cells. Increased expression of myogenic marker proteins was observed in GHRP-6-biotin conjugate-treated cells. Additionally, increased expression levels of insulin-like growth factor-1 and collagen type I were observed. Furthermore, GHRP-6-biotin conjugate-treated cells showed increased metabolic activity, as indicated by increased concentrations of energy metabolites, such as ATP and lactate, and increased enzymatic activity of lactate dehydrogenase and creatine kinase. Finally, binding protein analysis suggested few candidate proteins, including desmin, actin, and zinc finger protein 691 as potential targets for GHRP6-biotin conjugate action. These results suggest that the newly synthesized GHRP-6-biotin conjugate has myogenic stimulating activity through, at least in part, by stimulating collagen type I synthesis and several key proteins. Practical applications of the GHRP-6-biotin conjugate could include improving muscle condition. [BMB Reports 2015; 48(9): 501-506]

Randomized, dose-finding trial of ghrelin treatment for chronic respiratory failure.

Ghrelin, a growth hormone-releasing peptide, has shown efficacy in chronic obstructive pulmonary disease (COPD) patients in previous trials. This study was designed to evaluate the effective dose of ghrelin in chronic respiratory failure patients. In this randomized, double-blind, dose-finding, single-center study, 18 patients, including 16 with COPD, were randomly assigned to receive pulmonary rehabilitation (PR) with intravenous ghrelin at 1 μg/kg or 2 μg/kg, twice daily for 3 weeks. The primary outcome was the change in peak oxygen uptake ( V˙o2). Secondary outcomes included changes in plasma vascular endothelial growth factor (VEGF)-A levels, and exertional cardio-respiratory functions with blood gas analysis. With incremental exercise, there was no significant differences in the mean difference (high-dose ghrelin minus low-dose ghrelin) of peak V˙o2 (1.0 mL/kg/min, 95% CI: -0.6 to 2.6 mL/kg/min, between-group, P = 0.193). However, there were significant differences in the mean difference of (i) O2 -pulse (0.6 mL/beats, 95% CI: 0.0 to 1.1 mL/beats, between-group, P = 0.035) at iso-time; and ii) PaO2 (4.2 mmHg, 95% CI: 0.2 to 8.2 mmHg, between-group, P = 0.041) and PaCO2 (-3.1 mmHg, 95% CI: -6.0 to -0.3 mmHg, between-group, P = 0.034) at peak exercise. The mean difference in the plasma VEGF-A level was significantly inhibited by high dose-ghrelin with PR (-125.4 pg/mL, 95% CI: -235.2 to -15.5 pg/mL, between-group, P = 0.028). Although the primary outcome of the study was not met, high-dose ghrelin with PR improved exertional cardiac function and blood gas values, and inhibited circulating VEGF-A levels.

Ghrelin Inhibits Interleukin-6 Production Induced by Cigarette Smoke Extract in the Bronchial Epithelial Cell Via NF-κB Pathway.

Cigarette smoke (CS)-induced airway inflammation is the main pathogenesis of COPD. The present study was designed to evaluate whether ghrelin, a novel growth hormone-releasing peptide, can affect the pro-inflammatory cytokine interleukin-6 (IL-6) production induced by cigarette smoke extract (CSE) in the human bronchial epithelial cell line (16-HBE) and its possible mechanism. 16-HBE cells were pre-incubated with vehicle or ghrelin (0.1 to 1000ng/mL) in a concentration-dependent manner, and then CSE (0 to 16%) was added. The protein levels of IL-6 in the medium were determined by ELISA, and the mRNA expressions of IL-6 was detected by RT-PCR. We also detected the phosphorylation of IKKα/β/p65 protein and the degradation of inhibitory protein-κB (I-κB) by Western blot analysis. And the generation of reactive oxygen species (ROS) in 16-HBE was evaluated by labeling specific fluorescence probes DCFH-DA. 16-HBE Cells treated with CSE (8%) exhibited significantly higher IL-6 production compared with cells treated with vehicle alone (P < 0.05). Ghrelin suppressed CSE-induced IL-6 production at both mRNA and protein levels in a concentration-dependent manner (P < 0.05). Moreover, ghrelin attenuated CSE-triggered NF-κB activation in 16-HBE, but the intracellular ROS level after application of CSE was not affected by ghrelin (0.1 to 1000ng/mL). Together, these results suggest that ghrelin inhibits CSE-induced IL-6 production in 16-HBE cells by targeting on NF-κB pathway, but not by scavenging intracellular ROS.

The effect of obestatin on anxiety-like behaviour in mice

Obestatin is a 23 amino acid-peptide, derived from the same preproghrelin-gene as ghrelin. Obestatin was originally reported as a ghrelin antagonist with anorexigenic activity, but later it was proven to be involved in multiple processes including sleep, memory retention, anxiety, morphine-induced analgesia and withdrawal. In the present study, in male CFLP mice, by using computerised open field (OF) and elevated plus maze (EPM) tests we have investigated the behavioural effects of the acute intracerebroventricular (icv) administration of obestatin alone, and following ghrelin receptor blockage with [d-Lys3]-Growth Hormone Releasing Peptide-6 ([d-Lys3]- GHRP6) or corticotropin-releasing hormone (CRH) receptor 1 antagonism with antalarmin. Plasma corticosterone levels were measured for each treatment group by using chemofluorescent assay. Our results in the EPM test showed that obestatin reduced the percent of time spent in the open arms. The basal locomotor activity (ambulation distance and time, rearing and jumping) was not influenced significantly neither in the obestatin-treated groups, nor in those receiving pre-treatment with antalarmin or [d-Lys3]-GHRP6. The percentage of central ambulation distance however was decreased by obestatin, while the percentage of time spent in the central zone showed a decreasing tendency. The administration of antalarmin or [d-Lys3]-GHRP6 have both reversed the effect of obestatin on central ambulation. Plasma corticosterone levels were elevated by obestatin, which effect was antagonised by the injection of antalarmin. These are the first results to indicate that obestatin exerts anxiogenic-like effect in mice, which might be mediated through ghrelin receptor and CRH activation.

Levels of hormones and cytokines associated with growth in Honamlı and native hair goats.

This study was designed to assess alterations of hormone and cytokine levels associated with growth period during puberty in Honamlı goats which were identified as a new goat breed and had one of the highest meat production potential among the other goat breeds in Turkey. Honamlı goats are originated from native hair goats, so parallel studies of sampling and analyzing were conducted also in native hair goats which have moderate meat production. Blood serum samples of Honamlı (n=90) and native hair goats (n=90) were obtained from the pure herds in Korkuteli and Ka districts of Anatolia. Concentrations of growth hormone (GH), myostatin (MSTN), insulin-like growth factor (IGF), growth hormone releasing hormone (GHRH), growth hormone releasing peptide (GHRP), leptin, transforming growth factor-betal (TGF-β1) and vascular endothelial cell growth factor (VEGF) levels were measured by ELISA in each breed in the age groups of 4, 8 and 12 months. The present results indicate interesting correlations among the age groups and all the examined hormone and cytokine parameters exhibited significant (P<0.05 and P<0.001) differences. The parameters investigated were usually begun to increase after 4 months of age in the both breeds and sexes. Therefore, this paper supported the view that the beginning of hormonal alterations of goats could occur at 4th month of age. The results reported here emphasize the primary role played by GH, MSTN, IGF-1, leptin, GHRH, GHRP, TGF-βi and VEGF in the first year growth period of goats.

6D.02

Objective:Essential hypertensive patients (EH) are characterized by endothelial dysfunction caused by a reduced nitric oxide (NO) availability due to reactive oxygen species excess and low-grade inflammatory condition. Ghrelin is a recently identified growth hormone-releasing peptide, with recognize

One-year intranasal application of growth hormone releasing peptide-2 improves body weight and hypoglycemia in a severely emaciated anorexia nervosa patient.

BackgroundIn Japan, growth hormone releasing peptide-2 (GHRP-2) is clinically used as a diagnostic agent for growth hormone secretion deficiency, but the therapeutic application of GHRP-2 has not been studied in anorexia nervosa. GHRP-2 reportedly exhibits agonistic action for ghrelin receptor and increases food intake.MethodsWe administered GHRP-2 to a patient with a 20-year history of anorexia nervosa to determine whether GHRP-2 treatment increases food intake and body weight. GHRP-2 was administered before every meal by an intranasal approach for 1 year.ResultsAlthough the patient reported a decreased fear of eating and decreased desire to be thin by our previous treatment, she was unable to increase food intake or body weight because of digestive tract dysfunction. Vomiting after meals caused by delayed gastric emptying and incurable constipation were prolonged, and sub-ileus and hypoglycemia were observed. GHRP-2 increased the feeling of hunger and food intake, decreased early satiety and improved hypoglycemia. The patient's body weight gradually increased by 6.7 kg (from 21.1 kg to 27.8 kg) in 14 months after starting GHRP-2 administration. The fatigability and muscle strength improved, and the physical and mental activities were also increased. No obvious side effects were observed after long-term intranasal administration of GHRP-2.ConclusionsPatients with a long-term history of eating disorder occasionally recover from the psychological problems such as fear for obesity but remain emaciated. We believe that ghrelin agonists such as GHRP-2 may be promising agents for the effective treatments of severe anorexia nervosa in a chronic condition.

Determination of growth hormone releasing peptides metabolites in human urine after nasal administration of GHRP-1, GHRP-2, GHRP-6, Hexarelin, and Ipamorelin.

Growth hormone releasing peptides (GHRPs) stimulate secretion of endogenous growth hormone and are listed on the World Anti-Doping Agency (WADA) Prohibited List. To develop an effective method for GHRPs anti-doping control we have investigated metabolites of GHRP-1, GHRP-2, GHRP-6, Hexarelin, and Ipamorelin in urine after nasal administration. Each compound was administrated to one volunteer. Samples were collected for 2 days after administration, processed by solid-phase extraction on weak cation exchange cartridges and analyzed by means of nano-liquid chromatography - high resolution mass spectrometry. Six metabolites of GHRP-1 were identified. GHRP-1 in the parent form was not detected. GHRP-1 (2-4) free acid was detected in urine up to 27 h. GHRP-2, GHRP-2 free acid and GHRP-2 (1-3) free acid were detected in urine up to 47 h after administration. GHRP-6 was mostly excreted unchanged and detected in urine 23 h after administration, its metabolites were detectable for 12 h only. Hexarelin and Ipamorelin metabolized intensively and were excreted as a set of parent compounds with metabolites. Hexarelin (1-3) free acid and Ipamorelin (1-4) free acid were detected in urine samples after complete withdrawal of parent substances. GHRPs and their most prominent metabolites were included into routine ultra-pressure liquid chromatography-tandem mass spectrometry procedure. The method was fully validated, calibration curves of targeted analytes were obtained and excretion curves of GHRPs and their metabolites were plotted. Our results confirm that the detection window after GHRPs administration depends on individual metabolism, drug preparation form and the way of administration.

Synthesis of Mono-PEGylated Growth Hormone Releasing Peptide-2 and Investigation of its Biological Activity.

The purpose of this study was to investigate an efficient synthetic route to the mono-PEGylated growth hormone releasing peptide-2 (GHRP-2) and its biological activity in vivo. The commercially available key PEGylating reagent, mPEG-NHS ester, was successfully utilized to the synthesis of mono-PEGylated GHRP-2, during which the PEGylation profiles of GHRP-2 were monitored by high-performance liquid chromatography (HPLC). The product was purified by cation exchange chromatography, and its biological activity was conducted in rats. The desired mono-PEGylated GHRP-2 as the major product was readily obtained in anhydrous aprotic solvent, such as dimethyl formamide (DMF) and dimethylsulfoxide (DMSO), when the molar ratio of mPEG-NHS ester to GHRP-2 was fixed to be 0.8:1. The products were characterized by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. The evaluation of the biological activity for the products showed that the mono-PEGylated GHRP-2 gave a more stable activity than GHRP-2, suggesting that PEGylation led to the increase in the half-life of GHRP-2 in plasma without greatly impairing the biological activity. PEGylation of the GHRP-2 is a good choice for the development of the GHRP-2 applications.

Inhibitory Effect of Oleic Acid on Octanoylated Ghrelin Production.

Ghrelin is a growth hormone-releasing peptide that also displays orexigenic activity. Since serine-3 acylation with octanoylate (octanoylation) is essential for the orexigenic activity of ghrelin, suppression of octanoylation could lead to amelioration or prevention of obesity. To enable the exploration of inhibitors of octanoylated ghrelin production, we developed a cell-based assay system using AGS-GHRL8 cells, in which octanoylated ghrelin concentration increases in the presence of octanoic acid. Using this assay system, we investigated whether fatty acids contained in foods or oils, such as acetic acid, stearic acid, oleic acid, linoleic acid, and α-linolenic acid, have inhibitory effects on octanoylated ghrelin production. Acetic acid did not suppress the increase in octanoylated ghrelin production in AGS-GHRL8 cells, which was induced by the addition of octanoic acid. However, stearic acid, oleic acid, linoleic acid, and α-linolenic acid significantly suppressed octanoylated ghrelin production, with the effect of oleic acid being the strongest. Additionally, oleic acid decreased the serum concentration of octanoylated ghrelin in mice. The serum concentration of des-acyl ghrelin (without acyl modification) was also decreased, but the decrease was smaller than that of octanoylated ghrelin. Decreased octanoylated ghrelin production likely resulted from post-translational ghrelin processing, as there were no significant differences in gene expression in the stomach between oleic acid-treated mice and controls. These results suggest that oleic acid is a potential inhibitor of octanoylated ghrelin production and that our assay system is a valuable tool for screening compounds with suppressive effects on octanoylated ghrelin production.

Ghrelin protects alveolar macrophages against lipopolysaccharide-induced apoptosis through growth hormone secretagogue receptor 1a-dependent c-Jun N-terminal kinase and Wnt/β-catenin signaling and suppresses lung inflammation.

Alveolar macrophages (AMs) undergo increased apoptosis during sepsis-induced acute respiratory distress syndrome (ARDS). Ghrelin exhibits an antiapoptotic effect in several cell types and protects against sepsis-induced ARDS in rats; however, the molecular mechanisms underlying this antiapoptotic effect remain poorly understood. In this study, we first examined the antiapoptotic effect of ghrelin on lipopolysaccharide (LPS)-stimulated AMs in vitro. In AMs, GH secretagogue receptor-1a (GHSR-1a), the ghrelin receptor, was expressed, and treatment of AMs with ghrelin markedly reduced LPS-induced apoptosis, mitochondrial transmembrane potential decrease, and cytochrome c release. These effects of ghrelin were mediated by GHSR-1a because a GHSR-1a-targeting small interfering RNA abolished the antiapoptotic action of ghrelin. LPS treatment activated the c-Jun N-terminal kinase (JNK) signaling pathway but inhibited the Wnt/β-catenin pathway. Interestingly, combined LPS-ghrelin treatment reduced JNK activation and increased Wnt/β-catenin activation. Furthermore, like ghrelin treatment, the addition of the JNK inhibitor SP600125 or the glycogen synthase kinase-3β inhibitor SB216763 rescued AMs from apoptosis. We also demonstrated that ghrelin altered the balance of Bcl-2-family proteins and inhibited caspase-3 activity. Next, we investigated whether ghrelin protected against septic ARDS in vivo. Sepsis was induced in male rats by performing cecal ligation and puncture; administration of ghrelin reduced sepsis-induced AMs apoptosis, pulmonary injury, protein concentrations in the bronchoalveolar lavage fluid, the lung neutrophil infiltration, and wet to dry weight ratio. However, administration of a specific ghrelin-receptor antagonist, [D-Lys-3]-GH-releasing peptide-6, abolished the beneficial effects of ghrelin. Collectively our results suggest that ghrelin exerts an antiapoptotic effect on AMs at least partly by inhibiting JNK and activating the Wnt/β-catenin pathway and thereby helps alleviate septic ARDS in rats.

Detecting peptidic drugs, drug candidates and analogs in sports doping: current status and future directions

With the growing availability of mature systems and strategies in biotechnology and the continuously expanding knowledge of cellular processes and involved biomolecules, human sports drug testing has become a considerably complex field in the arena of analytical chemistry. Proving the exogenous origin of peptidic drugs and respective analogs at lowest concentration levels in biological specimens (commonly blood, serum and urine) of rather limited volume is required to pursue an action against cheating athletes. Therefore, approaches employing chromatographic–mass spectrometric, electrophoretic, immunological and combined test methods have been required and developed. These allow detecting the misuse of peptidic compounds of lower (such as growth hormone-releasing peptides, ARA-290, TB-500, AOD-9604, CJC-1295, desmopressin, luteinizing hormone-releasing hormones, synacthen, etc.), intermediate (e.g., insulins, IGF-1 and analogs, ‘full-length’ mechano growth factor, growth hormone, chorionic gonadotropin, erythropoietin, etc.) and higher (e.g., stamulumab) molecular mass with desired specificity and sensitivity. A gap between the technically possible detection and the day-to-day analytical practice, however, still needs to be closed.

The cardiovascular action of hexarelin.

Hexarelin, a synthetic growth hormone-releasing peptide, can bind to and activate the growth hormone secretagogue receptor (GHSR) in the brain similar to its natural analog ghrelin. However, the peripheral distribution of GHSR in the heart and blood vessels suggests that hexarelin might have direct cardiovascular actions beyond growth hormone release and neuroendocrine effects. Furthermore, the non-GHSR CD36 had been demonstrated to be a specific cardiac receptor for hexarelin and to mediate its cardioprotective effects. When compared with ghrelin, hexarelin is chemically more stable and functionally more potent. Therefore, it may be a promising therapeutic agent for some cardiovascular conditions. In this concise review, we discuss the current evidence for the cardiovascular action of hexarelin.

Abstract 068: Ghrelin Restores Nitric Oxide Availability In The Forearm Microcirculation Of Essential Hypertensive Patients

Objective: Essential hypertensive patients (EH) are characterized by endothelial dysfunction caused by a reduced nitric oxide (NO) availability due to reactive oxygen species excess and low-grade inflammatory condition. Ghrelin is a recently identified growth hormone-releasing peptide, with recognized cardiovascular actions. Possible effects on endothelial dysfunction have been never investigated in EH. In this study we evaluated whether exogenous ghrelin can improve endothelial dysfunction in the forearm microcirculation of untreated mild-moderate EH. Methods: In 9 EH (51.8±8.1 yrs) and 9 normotensive subjects (NS, 50.5±3.5 yrs), we studied the forearm blood flow (FBF, strain-gauge plethysmography) response to intrabrachial acetylcholine (ACh, 0.15-15 mg/100 ml/min) with and without NO synthase blockade by L-NMMA (100 μg/100 ml/min), or the antioxidant vitamin (Vit) C (8 mg/100 ml/min). The protocol was repeated under exogenous ghrelin intra-arterial infusion (200 ng/min, 30’ pre-infusion). Results: In NS, maximal vasodilation (VD) to ACh (480±20%) was inhibited by L-NMMA (292±22, -39±7%; P&lt;0.001) and unchanged by Vit C (482±34%). Ghrelin failed to modify these vascular responses. In EH, VD to ACh was blunted vs NS (337±45%; P&lt;0.001) and resistant to L-NMMA (313±32, -7±3%). Vit C increased the response to ACh (509±57%; P&lt;0.01 vs ACh alone) and restored the inhibiting effect of L-NMMA (332±42, -34±8%; P&lt;0.001). Ghrelin, while not modifying the basal FBF, it increased (P&lt;0.001) the VD to ACh (448±55%) and restored the inhibitory effect of L-NMMA on ACh (355±43, -20±6%; P&lt;0.001). Vit C only slightly improved VD to ACh under ghrelin infusion (486±45%). In EH ghrelin significantly (P&lt;0.05) decreased plasma venous malodialdehyde (from 6.9±1.5 to 5.2±1.0 μmol/L), lipoperoxides (from 9.1±1.9 to 6.6±2.3 μmol/L) and IL-6 (from 11.1±0.6 to 9.3±1.0 pg/mL) and increased plasma antioxidant capacity (from 407±109 to 630±97 mmol/L). Response to sodium nitroprusside was similar between EH and NS and not affected by ghrelin. Conclusions: Exogenous ghrelin is able to ameliorate endothelial dysfunction by restoring NO availability in the forearm microcirculation of EH, an effect probably determined by antioxidant and/or anti-inflammatory activities.

Identification of black market products and potential doping agents in Germany 2010-2013.

The desire to increase the athletic performance, to 'optimize' an individual's appearance, and to complement but also to arguably substitute exercise by means of drugs and drug candidates has generated a considerable (illicit) market for compounds such as anabolic-androgenic steroids, stimulants, growth promoting peptide hormones, and so on. Genuinely developed for therapeutic use, their abuse/misuse generates enormous health risks, which has necessitated comprehensive controls of compound trafficking by customs and anti-doping authorities. From 2012 to 2013, the Bureau of Customs Investigation confiscated products containing anabolic-androgenic steroids (AAS; 259 kg), stimulants (13 kg), selective estrogen receptor modulators (SERMs; 24 kg), and human growth hormone (hGH; 3500 ampules). In cooperation with the Bureau and under the umbrella of the European Monitoring Center for Emerging Doping Agents (EuMoCEDA), the Cologne Anti-Doping Laboratory analyzed an additional 337 (black market) products between 2010 and 2013, allowing to monitor developments in drug use and, hence, the anticipation of new challenges in sports drug testing. Main tools utilized in characterizing confiscated materials were liquid chromatography-high resolution mass spectrometry (LC-HRMS), gas chromatography-high resolution mass spectrometry (GC-HRMS), and polyacrylamide gel electrophoresis (PAGE) with subsequent bottom-up identification of peptidic compounds using nano liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS). Among the 337 substances analyzed in the doping control laboratory in Cologne, 67 active ingredients were found, 49 of which being categorized as doping agents by the World Anti-Doping Agency (WADA). A total of 83.7 % accounted for steroidal substances (predominantly testosterone, trenbolone, and nandrolone and corresponding esters), 12.8 % accounted for peptide hormones and growth factors (predominantly hGH and growth hormone releasing peptides (GHRPs)), 3.2 % of the products contained hormones and metabolic modulators, and 0.3 % accounted for diuretic agents. Outstanding findings were the detection of the selective androgen receptor modulator (SARM) LGD-4033, the thymic hormone thymosin β4, and a fusion protein of unknown biological activity. Trafficking of considerable amounts of arguably performance and/or body-enhancing compounds has been observed during the past 4 years, the majority of which is categorized as relevant to sports drug testing. Several substances are of fake/non-approved nature and represent enormous health risks to the 'customer'.