Regulation of ghrelin receptor by microbial and inflammatory signals in human osteoblasts

Abstract: Recently, it has been suggested that the anti-inflammatory hormone ghrelin (GHRL) and its receptor GHS-R may play a pivotal role in periodontal health and diseases. However, their exact regulation and effects in periodontitis are not known. The aim of this in-vitro study was to investigate the effect of microbial and inflammatory insults on the GHS-R1a expression in human osteoblast-like cells. MG-63 cells were exposed to interleukin (IL)-1β and Fusobacterium nucleatum in the presence and absence of GHRL for up to 2 d. Subsequently, gene expressions of GHS-R1a, inflammatory mediators and matrix metalloproteinase were analyzed by real-time PCR. GHS-R protein synthesis and NF-κB p65 nuclear translocation were assessed by immunocytochemistry and immunofluorescence microscopy, respectively. IL-1β and F. nucleatum caused a significant upregulation of GHS-R1a expression and an increase in GHS-R1a protein. Pre-incubation with a MEK1/2 inhibitor diminished the IL-1β-induced GHS-R1a upregulation. IL-1β and F. nucleatum also enhanced the expressions of cyclooxygenase 2, CC-chemokine ligand 2, IL-6, IL-8, and matrix metalloproteinase 1, but these stimulatory effects were counteracted by GHRL. By contrast, the stimulatory actions of IL-1β and F. nucleatum on the GHS-R1a expression were further enhanced by GHRL. Our study provides original evidence that IL-1β and F. nucleatum regulate the GHS-R/GHRL system in osteoblast-like cells. Furthermore, we demonstrate for the first time that the proinflammatory and proteolytic actions of IL-1β and F. nucleatum on osteoblast-like cells are inhibited by GHRL. Our study suggests that microbial and inflammatory insults upregulate GHS-R1a, which may represent a protective negative feedback mechanism in human bone.

Future Treatment of Constipation-associated Disorders: Role of Relamorelin and Other Ghrelin Receptor Agonists

There is an unmet need for effective pharmacological therapies for constipation, a symptom that significantly deteriorates patients' quality of life and impacts health care. Ghrelin is an endogenous ligand for the growth hormone secretagogue receptor and has been shown to exert prokinetic effects on gastrointestinal (GI) motility via the vagus and pelvic nerves. The pharmacological potential of ghrelin is hampered by its short half-life. Ghrelin receptor (GRLN-R) agonists with enhanced pharmacokinetics were thus developed. Centrally penetrant GRLN-R agonists stimulate defecation and improve impaired lower GI transit in animals and humans. This review summarizes the current knowledge on relamorelin, a potent ghrelin mimetic, and other GRLN-R analogs which are in preclinical or clinical stages of development for the management of disorders with underlying GI hypomotility, like constipation.

Leu72Met and Other Intronic Polymorphisms in the GHRL and GHSR Genes Are Not Associated with Type 2 Diabetes Mellitus, Insulin Resistance, or Serum Ghrelin Levels in a Saudi Population

BACKGROUND: Ghrelin (GHRL), a gastric peptide encoded by the GHRL gene, is known to be involved in energy homeostasis via its G protein receptor, encoded by the growth hormone secretagogue receptor (GHSR) gene. Some studies have shown associations between plasma GHRL levels and GHRL single-nucleotide polymorphisms (SNPs), namely the Leu72Met polymorphism (rs696217 TG), with type 2 diabetes mellitus (T2DM) and insulin resistance (IR), while others have not. The controversies in these associations raise the issue of ‘which SNPs in which populations.’ The aim of this study was to investigate whether SNPs in GHRL and/or GHSR genes were associated with T2DM, IR, or plasma GHRL levels among Arab Saudis. METHODS: Blood was collected from 208 Saudi subjects with (n=107) and without (n=101) T2DM. DNA samples from these subjects were analyzed by real-time polymerase chain reaction to genotype five intronic SNPs in the GHRL (rs696217 TG, rs27647 CT, rs2075356 CT, and rs4684677 AT) and GHSR (rs509030 GC) genes. In addition, plasma GHRL levels were measured by a radioimmunoassay. RESULTS: None of the SNPs were associated with T2DM, IR, or plasma GHRL levels. The frequencies of the alleles, genotypes, and haplotypes of the five SNPs were comparable between the T2DM patients and the non-diabetic subjects. A large number of the GHRL haplotypes indicates the molecular heterogeneity of the preproghrelin gene in this region. CONCLUSION: Neither the Leu72Met polymorphism nor the other intronic GHRL and GHSR SNPs were associated with T2DM, IR, or GHRL levels. Further investigations should be carried out to explain the molecular basis of the association of the GHRL peptide with T2DM and IR.

Ghrelin attenuates the growth of HO-8910 ovarian cancer cells through the ERK pathway

Ovarian cancer is one of the most common causes of death from gynecologic tumors and is an important public health issue. Ghrelin is a recently discovered bioactive peptide that acts as a natural endogenous ligand of the growth hormone secretagogue receptor (GHSR). Several studies have identified the protective effects of ghrelin on the mammalian reproductive system. However, little research has been done on the effects of ghrelin on ovarian cancer cells, and the underlying mechanisms of these effects. We sought to understand the potential involvement of mitogen-activated protein kinases (MAPKs) in ghrelin-mediated inhibition of growth of the ovarian line HO-8910. We applied different concentrations of ghrelin and an inhibitor of the ghrelin receptor (D-Lys3-GHRP-6) to HO-8910 cells and observed the growth rate of cells and changes in phosphorylation of the MAPKs ERK1/2, JNK and p38. We discovered that ghrelin-induced apoptosis of HO-8910 cells was though phosphorylated ERK1/2, and that this phosphorylation (as well as p90rsk phosphorylation) was mediated by the GHSR. The ERK1/2 pathway is known to play an essential part in the ghrelin-mediated apoptosis of HO-8910 cells. Hence, our study suggests that ghrelin inhibits the growth of HO-8910 cells primarily through the GHSR/ERK pathway.

Time course study of growth hormone releasing peptide-6-induced c-fos expression in neurons of feeding-related nuclei of hypothalamus / 生理学报

The present study was aimed to explore the effects of intraperitoneal injection of growth hormone releasing peptide-6 (GHRP-6), a ghrelin receptor agonist, on food intake and neuronal activity of feeding-related nuclei in the hypothalamus of NMRI mice. Accumulated amount of food intake was measured, and total number of c-fos immunoreactive neurons in arcuate nucleus (ARC), paraventricular nucleus (PVN) and supraoptic nucleus (SON) was counted by immunohistochemistry at 1, 3 and 6 h after the GHRP-6 injection. The results showed that GHRP-6 significantly increased the amount of food intake with a peak at 3 h after the GHRP-6 injection. Meanwhile, GHRP-6 could promote c-fos expression in the ARC and PVN independent of food intake, and the total number of c-fos immunoreactive neurons was peaked at 1 h after injection and then decreased gradually. These results suggest that GHRP-6 may increase food intake in time-dependent manner, which is associated with up-regulations of c-fos protein expression in the ARC and PVN.

Effect of Zhizhu Pill on Gastric Smooth Muscle Contractile Response and Protein Expression of Growth Hormone Secretagogue Receptor in Functional Dyspepsia Rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the therapeutic mechanism of Zhizhu Pill (ZP) for treating functional dyspepsia (FD) rats.</p><p><b>METHODS</b>Totally 30 ten-day-old male rats were randomly divided into the normal control group (n =10) and the model group (n = 20). The FD rat model was induced using gastric administration of 0.1% iodoacetamide (IA) combined tail clamping. The model was evaluated when rats were 8-week old. Successfully modeled rats were randomly divided into the model group (n = 10) and the ZP group (n = 10). Rats in the normal group and the model group were administered with normal saline by gastrogavage, while those in the ZP group were administered with ZP Decoction (2 mL/100 g) by gastrogavage. All medication lasted for 7 successive days. The contractile activity in in vitro longitudinal gastric muscle was recorded using Power Lab biological signal collecting system. The expression of growth hormone secretagogue receptor (GHSR) in stomach of FD rats was detected using Western blot and immunohistochemistry (IHC).</p><p><b>RESULTS</b>Compared with the normal group, average frequencies of gastric contraction and changing rates of amplitude obviously decreased in the model group (P < 0.05). Results of Western blot and IHC showed that the expression of GHSR decreased in the model group (P < 0.01). Compared with the model group, average frequencies of gastric contraction and changing rates of amplitude obviously increased in the ZP group (P < 0.05). Results of Western blot and IHC showed that the expression of GHSR increased in the ZP group (P < 0.01).</p><p><b>CONCLUSION</b>ZP could promote the gastric motility in FD rats induced by gastric administration of IA combined tail clamping, and its mechanism might be related to up-regulating GHSR protein level.</p>

Effect of Electroacupuncture on Expression of Ghrelin and mRNA Expression of Its Receptor in Functional Dyspesia Rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To explore the effect of electroacupuncture (EA) on the expression of Ghrelin and mRNA expression of its receptor in functional dyspepsia (FD) rats.</p><p><b>METHODS</b>Totally 80 rats were divided into the normal group, the model group, the drug therapy group, and the EA group according to random digit table, 20 in each group. FD model was duplicated by clipping tail modeling. Drug containing cisapride [2 mL/100 g, 0.09 g/(kg x d)] was administered to rats in the drug therapy group from the 3rd day after successful modeling, once per day. EA at Zusanli (ST36) (0.3-0.5 cun) and Taichong (LR3) (0.1-0.2 cun) was performed in the EA group. The twirling of needle was performed to the subsidence of needle, and then the needle was connected to HANS-200A Acupoint Nerve Stimulating Device using disperse-dense wave at 2 Hz, 2 mA, 30 min each time, once per day. Six days consisted of one therapeutic course, two courses in total with an interval of one day. The intestinal propulsive rate of ink was observed. Ghrelin protein expression in gastric tissue was detected by Western blot. mRNA expression of growth hormone secretagogue receptor (GHS-R) in stomach, hypothalamus, and hippocampus was detected using Real-time PCR respectively.</p><p><b>RESULTS</b>Compared with the normal group, the intestinal propulsive rate of ink, Ghrelin protein expression in gastric tissue, mRNA expression of GHS-R in stomach, hypothalamus, and hippocampus decreased in the model group (P < 0.05, P < 0.01). Compared with the model group, the intestinal propulsive rate of ink, Ghrelin protein expression in gastric tissue, mRNA expression of GHS-R in stomach, hypothalamus, and hippocampus increased in the EA group (P < 0.01); mRNA expression of GHS-R in stomach, hypothalamus, and hippocampus increased in the drug therapy group (P < 0.01). Compared with the drug therapy group, Ghrelin protein expression in gastric tissue, mRNA expression of GHS-R in hypothalamus increased in the EA group (P < 0.05, P < 0.01).</p><p><b>CONCLUSION</b>EA could regulate Ghrelin content and GHS-R mRNA expression of FD rat hypothalamus, hippocampus, and gastric tissue, and promote the intestinal propulsive rate of ink.</p>

Effects of peptidic growth hormone secretagogue receptor (GHS-R) antagonist [D-Lys3] on some of serum hormonal and biochemical parameters in Wistar rat model / Efeitos do antagonista [D-Lys3] do receptor do peptídeo secretagogo do hormônio do crescimento (GHS-R) sobre alguns parâmetros bioquímicos e hormonais séricos em um modelo em ratos Wistar

Objective : The present study investigated the effects of different dosages of a GHS-R antagonist [D-Lys3] on some serum hormonal (cortisol, T3 and T4) and biochemical parameters in a rat.Materials and methods : Thirty-six 60-day-old male rats were assigned to four treatments. [D-Lys3]-GHRP-6 solutions were infused via intraperitoneal injections. Blood was collected and analyzed.Results : The large dosages of a GHS-R antagonist (200 ng/kg BW) caused increases in cortisol, whereas no significant changes occurred when low dosages were injected. There were no significant changes in T3 and T4 following the administration of the GHS-R antagonist, but a considerable increase was observed in blood glucose levels of the groups (G50, G100, and G200 ng/kg BW). There was a significant increase in total protein when the greatest dose was administrated (G200 ng/kg BW). However, total cholesterol, triglycerides, and albumin showed no significant changes.Conclusions : Exogenous GHS-R antagonist can cause an increase in glucose and moderate increases in cortisol and total protein, yet it has no significant effect on T3 and T4 levels or on the concentrations of serum lipids. The effect of GHS-R antagonist is not completely adverse to the effects of ghrelin. Further molecular studies are necessary to identify the physiological effects of the peptidic GHS-R antagonist. Arq Bras Endocrinol Metab. 2014;58(3):288-91.

Objetivo : O presente estudo investigou os efeitos de diferentes doses do antagonista do GHS-R [D-Lys3] sobre alguns parâmetros hormonais (cortisol, T3 e T4) e bioquímicos em ratos.Materiais e métodos : Trinta e seis ratos machos com 60 dias de idade foram alocados para quatro tratamentos. Soluções de [D-Lys3]-GHRP-6 foram administradas por meio de injeções intraperitoneais e foram coletadas e analisadas amostras.Resultados : Doses altas de antagonista de GHS-R (200 ng/kg PC) levaram a aumento do cortisol, enquanto não houve diferença significativa quando foram injetadas doses baixas. Não houve alterações significativas em T3 e T4 depois da administração do antagonista do GHS-R, mas foi observado aumento considerável nos níveis de glicose sanguínea dos grupos (G50, G100 e G200 ng/kg PC). Houve aumento significativo na proteína total quando foi administrada a maior dose (G200 ng/kg PC), entretanto, não foram observadas alterações no colesterol total, nos triglicérides e na albumina.Conclusões : O antagonista do GHS-R exógeno pode causar aumento da glicose e aumento moderado do cortisol e proteína total, embora não haja efeitos significativos nos níveis de T3 e T4 ou na concentração de lipídios séricos. O efeito do antagonista de GHS-R não é completamente adverso aos efeitos da grelina. Devem ser feitos outros estudos moleculares para se identificar os efeitos fisiológicos do peptídeo antagonista do GHS-R. Arq Bras Endocrinol Metab. 2014;58(3):288-91.

Sinalização da grelina no coração de camundongos obesos após hipernutrição durante a lactação / Ghrelin signaling in the heart of obese mice after overnutrition during lactation

A grelina é um ligante endógeno do receptor secretagogo do hormônio do crescimento (GHSR), potente estimulador da liberação do hormônio de crescimento (GH), ingestão alimentar, e adiposidade. Além disso, sua ação hormonal inclui regulação do metabolismo energético cardíaco. Entretanto, a hipernutrição no início da vida leva ao desenvolvimento da obesidade, induz hipertrofia cardíaca, compromete a função cardíaca, e gera insuficiência cardíaca na vida adulta. Avaliar proteínas chaves no processo de sinalização da grelina no remodelamento cardíaco no coração de camundongos obesos após a hipernutrição na lactação. A obesidade foi induzida por redução de ninhada e camundongos adultos (180 dias) foram divididos em: grupo hiperalimentado, GH com obesidade decorrente de hipernutrição na lactação e controle, GC. Cardiomiócitos (cmi) do ventrículo esquerdo foram analisados por microscopia de luz e estereologia, o conteúdo e fosforilação de proteínas cardíacas: receptor de grelina (hormônio do crescimento secretagogo receptor 1a, GHSR-1a), proteína quinase-B (AKT e pAKT), phosphatidil inositol 3-quinase (PI3K), proteína quinase ativada por AMP (AMPK e pAMPK), m-TOR, pmTOR, Bax, Bcl2 e actina foram analizados por western blotting. A expressão gênica do GHSR-1a foi analisada por PCR em tempo real. A respirometria de alta resolução dos cardiomiócitos foi analisada por oxígrafo OROBOROS®. Significância estatística (P< 0,05) determinada por teste t-Student não-pareado. Nossos dados demonstram que a hipernutrição na lactação induz aumento no peso corporal, iniciado aos 10 dias de idade, persistindo até os 180 dias de idade. A glicemia, peso do fígado, e da gordura visceral foram maiores no grupo GH. Além disso, o grupo GH também apresentou aumento no peso do coração e razão peso do coração/CT (comprimento da tíbia), indicando hipertrofia e remodelamento cardíaco, aumento na expressão e conteúdo de GHSR-1a no coração, associado ao maior conteúdo de PI3K e maior conteúdo...

Ghrelin, an endogenous ligand of the growth hormone secretagogue receptor (GHS-R), has been suggested to be associated to obesity, insulin secretion, cardiovascular growth and homeostasis. GHS-R has been found in most of the tissues, and among the hormone action it is included the regulation of heart energy metabolism. Therefore, hypernutrition during early life leads to obesity, induces cardiac hypertrophy, compromises myocardial function, inducing heart failure in adulthood. We examined ghrelin signaling process in cardiac remodeling in these obese adult mice. We examined key proteins of cardiomyocyte metabolism in heart left ventricle from overfed (OG) and control (CG) groups from adult mice (180 days) overfed during lactation. Obesity was induced by litter reduction. Therefore, the study was done in adult mice 180 days old (OG, obese group (n=10) and CG, control group (n=10). The cardiomyocytes (cmy) of left ventricle were analyzed by light microscopy and stereology. The content and phosphorylation of cardiac proteins: growth hormone secretagogue receptor 1a (GHSR-1a), protein kinase B (AKT and pAKT), phosphatidil inositol 3 kinase (PI3K), AMP-activated protein kinase (AMPK and pAMPK), mTOR and pmTOR, BAX, Bcl2 and actin was achieved by western blotting. GHSR-1a gene expression was analyzed to RT-PCR. We performed high-resolution respirometry of cardiomyocytes with OROBOROS® Oxygraph-2k. Statistical significance was determined by Student t-test for unpaired. P< 0.05 was considered statistical significant. Body weight, blood glucose, liver weight, and visceral fat weight were higher in OG than CG group. Obese mice had increased heart weight and heart weight/TL (tibia length) indicating cardiac remodeling and hypertrophy, increased GHSR-1a content and expression in the heart, associated to PI3K content, increased AKT content and phosphorylation (P< 0.05), decreased Bcl2 content. In contrast, AMPK and mTOR content and phosphorylation in heart were not...

Role of Ghrelin in the Pathophysiology of Gastrointestinal Disease

Ghrelin is a 28-amino-acid peptide that plays multiple roles in humans and other mammals. The functions of ghrelin include food intake regulation, gastrointestinal (GI) motility, and acid secretion by the GI tract. Many GI disorders involving infection, inflammation, and malignancy are also correlated with altered ghrelin production and secretion. Although suppressed ghrelin responses have already been observed in various GI disorders, such as chronic gastritis, Helicobacter pylori infection, irritable bowel syndrome, functional dyspepsia, and cachexia, elevated ghrelin responses have also been reported in celiac disease and inflammatory bowel disease. Moreover, we recently reported that decreased fasting and postprandial ghrelin levels were observed in female patients with functional dyspepsia compared with healthy subjects. These alterations of ghrelin responses were significantly correlated with meal-related symptoms (bloating and early satiation) in female functional dyspepsia patients. We therefore support the notion that abnormal ghrelin responses may play important roles in various GI disorders. Furthermore, human clinical trials and animal studies involving the administration of ghrelin or its receptor agonists have shown promising improvements in gastroparesis, anorexia, and cancer. This review summarizes the impact of ghrelin, its family of peptides, and its receptors on GI diseases and proposes ghrelin modulation as a potential therapy.

Diltiazem enhances food intake and gastrointestinal function in rats / 生理学报

The present study was to investigate the effects of diltiazem, a ghrelin receptor agonist, on food intake and gastrointestinal functions in rats. Rats were intragastrically administered with diltiazem solution (daily 16 mg/kg, 30 mg/kg or 80 mg/kg, 30 d), and the rats with saline as control. To detect the effects of diltiazem on food intake and body weight, the average daily food intake and body weight were recorded, and the serum metabolic hormones of plasma growth hormone (GH) and neuropeptide Y (NPY) were tested by radioimmunoassay. By means of the spectrophotometer and the modified Mett's method, the effects of diltiazem on rat's gastrointestinal function and pepsin activity were tested, respectively. In addition, the gastric juice's acidity of rats was detected by titration and the secretion amount was calculated. The results showed that the food intake and body weight were maximally promoted by diltiazem at the dose of 30 mg/kg daily (30 d). The average daily food intake and body weight were significantly increased, and the serum concentrations of GH and NPY were also remarkably increased in diltiazem-treated groups compared with those in control group. The results also showed that the gastric emptying rate, gastric acid secretion and the activity of pepsin were significantly increased in diltiazem-treated group compared with those in control group. These results suggest that diltiazem induces enhancement of eating, in the same time, it can also stimulate the gastrointestinal function and regulate growth of rat.

Ghrelin stimulates in vitro angiogenic capacity of rat cardiac microvascular endothelial cells / 中华心血管病杂志

<p><b>OBJECTIVE</b>To clarify whether ghrelin could promote in vitro rat cardiac microvascular endothelial cells (CMECs) angiogenesis and related mechanisms.</p><p><b>METHODS</b>CMECs were isolated from myocardial tissue of adult male SD rats and characterized by the immunocytochemistry staining with Factor VIII and the capacity of in vitro capillary tube-like formation. The mRNAs and protein expressions of ghrelin and its receptor (growth hormone secretagogue receptor, GHS-R) of CMECs were determined by RT-PCR, Immunofluorescence, ELISA and Western blot. Proliferation, migration and in vitro angiogenesis as well as ERK2 phosphorylation of CMECs were tested in the presence of ghrelin (10(-9) - 10(-7) mol/L) with or without pretreatment with specific MAPK/ERK2 inhibitor PD98059.</p><p><b>RESULTS</b>Purity of CMECs characterized by immunocytochemistry staining with Factor VIII was about 95%, and the cells showed a high ability to form the capillary tube-like structures on Matrigel. Ghrelin and GHS-R were constitutively expressed in CMECs. Proliferation, migration and in vitro angiogenesis capacities of CMECs (72.20 ± 5.72 vs. 28.60 ± 5.13, P < 0.001; 71.00 ± 7.78 vs. 28.60 ± 5.13, P < 0.001) as well as ERK2 phosphorylation (0.92 ± 0.13 vs. 0.29 ± 0.04, P < 0.001; 1.15 ± 0.16 vs. 0.29 ± 0.04, P < 0.001) were significantly enhanced by exogenous ghrelin (10(-8) - 10(-7) mol/L). PD98059 abolished ghrelin-induced ERK2 phosphorylation and in vitro angiogenesis.</p><p><b>CONCLUSIONS</b>Ghrelin and its receptor are expressed in CMECs and ghrelin could stimulate CMECs in vitro angiogenesis through activation of MAPK/ERK2 signaling pathway.</p>

Pesquisa de mutações no gene do receptor do secretagogo de hormônio de crescimento (GHSR) em crianças com baixa estatura idiopática e deficiência isolada de hormônio de crescimento / Growth hormone secretatogue receptor gene (GHSR) analysis in patients with idiopathic short stature (ISS) and patients with isolated growth hormone deficiency

A ghrelina, hormônio secretado principalmente por células gástricas, liga-se ao seu receptor, o receptor de secretagogo de GH (GHSR - Growth hormone secretagogue receptor), localizado no hipotálamo e na hipófise, estimulando a síntese e secreção do GH. Recentemente foram identificadas mutações no gene GHSR em crianças com baixa estatura idiopática (BEI) e com deficiência isolada de GH (DGH). No presente estudo investigamos a presença de mutações no gene GHSR em crianças com DGH isolada de causa não identificada e crianças com BEI, incluindo um subgrupo de crianças com atraso constitucional de crescimento e desenvolvimento (ACCD). Foram selecionados 14 pacientes com deficiência isolada de GH sem alterações anatômicas da região hipotálamo-hipofisária e 96 pacientes com BEI, destes 31 (32%) apresentavam ACCD. Também foram estudados 150 controles adultos e 197 crianças controle com crescimento e puberdade normais. A região codificadora do GHSR foi amplificada utilizando-se oligonucleotídeos iniciadores específicos, seguida de purificação enzimática e seqüenciamento automático. Encontramos 6 variantes alélicas em heterozigose no GHSR: nenhuma delas presente nos controles estudados, e quatro destas variantes estão localizadas em regiões conservadas do gene. Uma variante foi encontrada em uma paciente do grupo DGH (p.Val249Leu) e as outras cinco (c.-6 G>C, p.Ser84Ile, p.Val182Ala, p.Ala169Thr e p.Ala358Thr) foram encontradas em pacientes do subgrupo ACCD do grupo BEI. As variantes missense foram submetidas a estudo funcional que evidenciou que as mutações p.Ser84Ile e p.Val182Ala possuem diminuição na atividade basal associadas à diminuição da expressão do receptor na superfície celular. Adicionalmente, a mutação p.Ser84Ile também apresenta redução na atividade do GHSR induzida pelo ligante. A variante p.Val249Leu foi encontrada em uma paciente do sexo feminino com diagnóstico de DGH isolado...

Ghrelin, hormone secreted by gastric cells, stimulates growth hormone secretion by acting on its receptor GHSR, located in the hypothalamus and pituitary. Recently, mutations in the GHSR gene were described in patients with growth hormone deficiency (GHD) and idiopathic short stature (ISS). In the present study we analyzed the GHSR gene in patients with isolated GHD and patients with ISS, including a subgroup of patients with constitutional delay of growth and puberty (CDGP). We studied 14 GHD patients with normal pituitary magnetic resonance imaging and 96 patients with ISS, 31 of them with CDGP. We also studied 150 adults and in 197 children with normal stature. The entire coding region as well as the exon-intron boundaries of GHSR were PCR amplified in all patients and control group and PCR products were bidirectionally sequenced. Six different heterozygous variants in GHSR were identified: none of them were found in the control group and four of these amino acid substitutions occurred at a conserved position within the GHSR. One variant (p.Val249Leu) was found in a GHD patient and the other five (c.-6 G>C, p.Ser84Ile, p.Val182Ala, p.Ala169Thr e p.Ala358Thr) were found in patients with CDGP. The missense variants were submitted to functional studies. Two of these variants (p.Ser84Ile and p.Val182Ala) result in a decrease in basal activity that was in part explained by a reduction in cell surface expression. The p.Ser84Ile mutation was also associated with a defect in ghrelin potency. The p.Val249Leu variant, found in a female patient with isolated GHD, did not segregate with the phenotype in the family and had no functional impairment in vitro. This suggests that p.Val249Leu is not the cause of the GHD in the family and may be a rare allelic variant. The other variants (c.-6 G>C, p.Ser84Ile, p.Val182Ala, p.Ala169Thr e p.Ala358Thr) were identified only in patients with CDGP (3 male and 2 female)...

Effect of the expression of ghrelin receptors on the postoperative underpowered small intestinal motility in rats / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the effect of the expression of ghrelin receptors on the postoperative small intestine dysmotility in rat models.</p><p><b>METHODS</b>The effect of different concentrations of ghrelin (0, 0.01, 0.1, 0.5, 1.0 μmol/L) on the contraction of smooth muscle strips of rat small intestine in the presence or absence of carbachol was observed in vitro. End-to-side anastomosis was performed in the study group and sham controls were used. The expression of ghrelin receptors(GHS-R1a) in small intestine muscle layers was detected by immunohistochemistry and Western blot.</p><p><b>RESULTS</b>In vitro, ghrelin enhanced the contraction of smooth muscle strips in the presence of carbachol, and the differences in contraction induced by different concentrations of ghrelin(0.1, 0.5, 1.0 μmol/L) were statistically significant [(223±18)%, (245±22)%, (264±25)%, P<0.01]. Immunohistochemistry study showed that GHS-R1a mainly located in the muscular layer of the bowel wall. The expression of GHS-R1a in the circular and longitudinal muscle was significantly weaker than that in the control group. The expression of ghrelin receptors after surgery was down-regulated in the study group, which was lower than that in the control group(0.51±0.02 vs. 0.71±0.01, P<0.01).</p><p><b>CONCLUSION</b>Down regulation of ghrelin receptors in small intestine muscle layers may contribute to the occurrence of small intestine dysmotility after intestinal surgery.</p>

Expression of ghrelin and its receptor GHS-R in the hypothalamus and gastrointestinal tract in rats with chronic renal failure / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the expression of ghrelin and its receptor, growth hormone secretagogue receptor (GHS-R), in the hypothalamus and gastrointestinal tract in rats with chronic renal failure (CRF) and explore their relationship with the disorder of gastrointestinal tract motility.</p><p><b>METHODS</b>SD rats were randomly divided into sham-operated group (n=8) and CRF group (n=16), and in the latter group, the rats were subjected to 5/6 nephrectomy to induce CRF. Real-time PCR and immunohistochemical staining were used to detect the distribution of mRNA and protein of ghrelin and GHS-R in the gastric fundus, duodenum, and hypothalamus.</p><p><b>RESULTS</b>The rats in the CRF group showed a significantly higher expression of ghrelin mRNA and protein in the gastric fundus but a lower expression in the hypothalamus than those in the sham-operated group (P<0.01), but the expression in the duodenum was similar between the two groups (P>0.05). The expression of GHS-R mRNA and protein in the gastric fundus was significantly higher in the CRF group than in the sham-operated group (P<0.01), while in the hypothalamus and duodenum, the expression was significantly lower in the CRF group (P<0.01).</p><p><b>CONCLUSION</b>The different distribution patterns of ghrelin and GHS-R in the tissues may be an important pathological basis of gastrointestinal motility disorder in CRF.</p>

Relationship between Ghrelin and growth hormone secretagogue receptor expression and catch-up growth in rats with intrauterine growth restriction / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the relationship between Ghrelin and growth hormone secretagogue receptor (GHSR) expression and the catch-up growth in rats with intrauterine growth restriction (IUGR).</p><p><b>METHODS</b>The rat model of IUGR was established by food restriction during pregnancy. The small for gestational age (SGA) and appropriate for gestational age (AGA) rat pups from the pregnant rats were used as the experimental group. The AGA rat pups from the pregnant rats without food restriction served as the control group. The samples from the stomach fundus and hypothalamus were taken postnatal days 0, 20 and 40. Ghrelin mRNA and GHSR mRNA expression were determined by real-time fluorescence quantitative PCR (real-time FQ-PCR). Ghrelin protein and GHSR protein expression were examined by immunohistochemistry (IHC).</p><p><b>RESULTS</b>At postnatal day 0, both Gherlin mRNA and protein levels in the stomach fundus were significantly higher, while GHSR mRNA expression in the hypothalamus were significantly lower in SGA rats from food restriction group than those in AGA rats from restriction and control groups. At postnatal day 20, the ghrelin protein expression in the stomach of fundus, and GHSR mRNA and protein expression in the hypothalamus in SGA catch-up rats were significantly higher than those in SGA non-catch-up growth rats and AGA rats from the control group. At postnatal day 40, there were no significant differences among SGA catch-up growth rats, SGA non-catch-up growth rats and normal AGA rats.</p><p><b>CONCLUSIONS</b>Ghrelin-GHSR might be involved in the physiological regulation and pathological process in IUGR rats. It is also possibly involved in the regulation of catch-up growth in the early life of SGA rats.</p>

Ghrelin down-regulates ACAT-1 in THP-1 derived foam cells via growth hormone secretagogue receptor-dependent pathway / 中华心血管病杂志

<p><b>OBJECTIVE</b>To investigate the effects of Ghrelin on the expression of acyl coenzyme A:cholesterol acyltransferases-1 (ACAT-1) in THP-1 derived foam cells.</p><p><b>METHODS</b>The human monocytic leukemia cell line (THP-1) was chosen in our study. The differentiation of THP-1 cells into macrophages was induced by phorbol 12-myristate 13-acetate. Macrophages were then incubated with oxidized LDL (ox-LDL) to generate foam cells. Ghrelin and [D-Lys3]-GHRP-6, the special antagonist of growth hormone secretagogue receptor (GHS-R), were treated during foam cells formation. The ACAT-1 protein and mRNA levels were detected by Western blot and RT-PCR. The effect of variance of cholesterol content was measured by zymochemistry via-fluorospectrophotometer.</p><p><b>RESULTS</b>Ghrelin reduced the content of cholesterol ester in foam cells obviously. ACAT-1 protein and mRNA levels were also decreased. The antagonist of GHS-R inhibited the effects of Ghrelin on ACAT-1 expression in dose-dependent manner. The ACAT-1 mRNA levels of the GHS-R specific antagonist groups (10(-5), 5 x 10(-5), 10(-4) mol/L) were 1.14 +/- 0.04, 1.58 +/- 0.03, 2.40 +/- 0.16, significantly higher than that of the Ghrelin group (0.89 +/- 0.05). And the protein expressions were 1.25 +/- 0.09, 1.77 +/- 0.11, 2.30 +/- 0.09, also higher than that of the Ghrelin group (0.86 +/- 0.08).</p><p><b>CONCLUSIONS</b>Ghrelin might interfere atherosclerosis by down-regulating the expression of ACAT-1 via GHS-R pathway.</p>

Ghrelina e secretagogos do hormônio de crescimento (GHS): modulação da secreção do hormônio de crescimento e perspectivas terapêuticas: [revisão] / Ghrelin and growth hormone secretagogues (GHS): modulation of growth hormone secretion and therapeutic applications: [review]

A secreção do hormônio de crescimento (GH) é modulada pelo hormônio liberador de hormônio de crescimento (GHRH) e pela somatostatina. Na última década foi descoberto um terceiro mecanismo de controle, envolvendo os secretagogos de GH (GHS). A ghrelina, o ligante endógeno do receptor dos GHS, é um peptídeo acilado produzido no estômago, que também é sintetizado no hipotálamo. Este peptídeo é capaz de liberar GH, além de aumentar a ingesta alimentar. A ghrelina endógena parece amplificar o padrão básico de secreção de GH, ampliando a resposta do somatotrofo ao GHRH, estimulando múltiplas vias intracelulares interdependentes. Entretanto, seu local de atuação predominante é o hipotálamo. Neste trabalho, será apresentada revisão sobre a descoberta da ghrelina, os mecanismos de ação e o possível papel fisiológico dos GHS e da ghrelina na secreção de GH e, finalmente, as possíveis aplicações terapêuticas destes compostos.

Growth hormone-releasing hormone (GHRH) and somatostatin modulate growth hormone (GH) secretion. A third mechanism was discovered in the last decade, involving the action of growth hormone secretagogues (GHS). Ghrelin, the endogenous ligand of the GHS-receptor, is an acylated peptide mainly produced by the stomach, but also synthesized in the hypothalamus. This compound increases both GH release and food intake. Endogenous ghrelin might amplify the basic pattern of GH secretion, optimizing somatotroph responsiveness to GHRH, activating multiple interdependent intracellular pathways. However, its main site of action is the hypothalamus. In the current paper it is reviewed the available data on the discovery of this peptide, the mechanisms of action and possible physiological roles of the GHS and ghrelin on GH secretion, and finally, the possible therapeutic applications of these compounds.

Expression of growth hormone secretagogue receptor type 1a in visceral vagal and spinal afferent pathways / 生理学报

In this study, the expressions of growth hormone secretagogue receptor type 1a (GHS-R1a) in the rat dorsal root ganglion (DRG) and nodose ganglion (NG) were investigated by using immunohistochemistry and in situ hybridization. The results clearly showed the presence of GHS-R1a mRNA and GHS-R1a-positive neurons in the rat DRG and NG. GHS-R1a was also co-localized with calcitonin gene-related peptide (CGRP) in some DRG and NG neurons, indicating the existence of subpopulations of the visceral afferents. The extrinsic primary afferent visceroceptive DRG and NG neurons from the stomach were identified by retrograde tracing fluorogold and stained for GHS-R1a and CGRP. Some neurons both positive for CGRP and GHS-Rla were labled by fluorogold. Our results not only demonstrate the expression of GHS-R1a in the vagal afferents but also provide the first and direct morphological evidence for its presence in the spinal visceral afferents, and gherin might have a modulatory role in the visceral afferent signaling.

Ghrelin receptor gene polymorphisms are associated with female metabolic syndrome in Chinese population / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The ghrelin plays an important role in the regulation of food intake and energy homeostasis. Therefore, the ghrelin receptor gene (GHSR) is an excellent candidate for studying metabolic syndrome. This study aimed to investigate whether polymorphisms in ghrelin receptor gene are associated with metabolic syndrome in Chinese population.</p><p><b>METHODS</b>Subjects consisted of 698 patients aged 41 to 80 years, diagnosed as metabolic syndrome by International Diabetes Federation (IDF) 2005 criteria, and 762 age- and gender-matched controls. Three variants within the GHSR were selected and genotyped using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Odds ratios were estimated using a case-control study design by controlling confounding factors.</p><p><b>RESULTS</b>The A/A genotype (rs2922126) in the promoter was associated with metabolic syndrome (OR 1.41, 95% CI 1.03-1.94), increased waist circumference (OR 1.75, 95% CI 1.26-2.42), and increased fast blood glucose (OR 1.49, 95% CI 1.07-2.06) in women. The A/A genotype (rs509030) in the intron was associated with lower plasma high density lipoprotein in women (OR 1.37, 95% CI 1.02-1.84).</p><p><b>CONCLUSION</b>The polymorphisms within GHSR might be a genetic risk factor for metabolic syndrome in women.</p>