RESUMO
CONTEXT: Targeted secretion inhibitors (TSIs), a new class of recombinant biotherapeutic proteins engineered from botulinum toxin, represent a novel approach for treating diseases with excess secretion. They inhibit hormone secretion from targeted cell types through cleavage of SNARE (soluble N-ethylmaleimide-sensitive factor-activating protein receptor) proteins. qGHRH-LH(N)/D is a TSI targeting pituitary somatotroph through binding to the GHRH-receptor and cleavage of the vesicle-associated membrane protein (VAMP) family of SNARE proteins. OBJECTIVE: Our objective was to study SNARE protein expression in pituitary adenomas and to inhibit GH secretion from somatotropinomas using qGHRH-LH(N)/D. DESIGN: We analyzed human pituitary adenoma analysis for SNARE expression and response to qGHRH-LH(N)/D treatment. SETTING: The study was conducted in University Hospitals. PATIENTS: We used pituitary adenoma samples from 25 acromegaly and 47 nonfunctioning pituitary adenoma patients. OUTCOME: Vesicle-SNARE (VAMP1-3), target-SNARE (syntaxin1, SNAP-23, and SNAP-25), and GHRH-receptor detection with RT-qPCR, immunocytochemistry, and immunoblotting. Assessment of TSI catalytic activity on VAMPs and release of GH from adenoma cells. RESULTS: SNARE proteins were variably expressed in pituitary samples. In vitro evidence using recombinant GFP-VAMP2&3 or pituitary adenoma lysates suggested sufficient catalytic activity of qGHRH-LH(N)/D to degrade VAMPs, but was unable to inhibit GH secretion in somatotropinoma cell cultures. CONCLUSIONS: SNARE proteins are present in human pituitary somatotroph adenomas that can be targeted by TSIs to inhibit GH secretion. qGHRH-LH(N)/D was unable to inhibit GH secretion from human somatotroph adenoma cells. Further studies are required to understand how the SNARE proteins drive GH secretion in human somatotrophs to allow the development of novel TSIs with a potential therapeutic benefit.
Assuntos
Adenoma/tratamento farmacológico , Antineoplásicos/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Adenoma Hipofisário Secretor de Hormônio do Crescimento/tratamento farmacológico , Proteínas de Neoplasias/antagonistas & inibidores , Hipófise/efeitos dos fármacos , Proteínas SNARE/antagonistas & inibidores , Via Secretória/efeitos dos fármacos , Acromegalia/etiologia , Acromegalia/prevenção & controle , Adenoma/metabolismo , Adenoma/patologia , Antineoplásicos/química , Toxinas Botulínicas/química , Toxinas Botulínicas/genética , Toxinas Botulínicas/farmacologia , Desenho de Fármacos , Hormônio Liberador de Hormônio do Crescimento/análogos & derivados , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Adenoma Hipofisário Secretor de Hormônio do Crescimento/metabolismo , Adenoma Hipofisário Secretor de Hormônio do Crescimento/patologia , Hormônio do Crescimento Humano/antagonistas & inibidores , Hormônio do Crescimento Humano/genética , Hormônio do Crescimento Humano/metabolismo , Humanos , Ligantes , Terapia de Alvo Molecular , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Hipófise/metabolismo , Hipófise/patologia , Neoplasias Hipofisárias/tratamento farmacológico , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/patologia , Engenharia de Proteínas , Estrutura Terciária de Proteína , Receptores LHRH/antagonistas & inibidores , Receptores LHRH/genética , Receptores LHRH/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/farmacologia , Proteínas SNARE/genética , Proteínas SNARE/metabolismo , Células Tumorais CultivadasRESUMO
Ghrelin is a 28 amino acid peptide hormone that is produced both centrally and peripherally. Regulated by the ghrelin O-acyl transferase enzyme, ghrelin exerts its action through the growth hormone secretagogue receptor, and is implicated in a diverse range of physiological processes. These implications have placed the ghrelin signaling pathway at the center of a large number of candidate gene and genome-wide studies which aim to identify the genetic basis of human heterogeneity. In this review we summarize the available data on the genetic variability of ghrelin, its receptor and its regulatory enzyme, and their association with obesity, stature, type 2 diabetes, cardiovascular disease, eating disorders, and reward seeking behavior.
Assuntos
Aciltransferases/genética , Mucosa Gástrica/metabolismo , Grelina/genética , Isoformas de Proteínas/genética , Precursores de Proteínas/genética , Receptores de Grelina/genética , Transdução de Sinais/genética , Acilação , Aciltransferases/metabolismo , Animais , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/fisiopatologia , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/fisiopatologia , Transtornos da Alimentação e da Ingestão de Alimentos/metabolismo , Transtornos da Alimentação e da Ingestão de Alimentos/fisiopatologia , Estudo de Associação Genômica Ampla , Grelina/química , Grelina/metabolismo , Humanos , Modelos Moleculares , Obesidade/metabolismo , Obesidade/fisiopatologia , Polimorfismo de Nucleotídeo Único , Postura/fisiologia , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Precursores de Proteínas/química , Precursores de Proteínas/metabolismo , Receptores de Grelina/metabolismoRESUMO
BACKGROUND: Ghrelin and its receptor play an important role in glucose metabolism and energy homeostasis, and therefore they are functional candidates for genes carrying susceptibility alleles for type 2 diabetes. METHODS: We assessed common genetic variation of the ghrelin (GHRL; five single nucleotide polymorphisms (SNP)) and the ghrelin-receptor (GHSR) genes (four SNPs) in 610 Caucasian patients with type 2 diabetes and 820 controls. In addition, promoter reporter assays were conducted to model the regulatory regions of both genes. RESULTS: Neither GHRL nor GHSR gene SNPs were associated with type 2 diabetes. One of the ghrelin haplotypes showed a marginal protective role in type 2 diabetes. We observed profound differences in the regulation of the GHRL gene according to promoter sequence variants. There are three different GHRL promoter haplotypes represented in the studied cohort causing up to 45% difference in the level of gene expression, while the promoter region of GHSR gene is primarily represented by a single haplotype. CONCLUSION: The GHRL and GHSR gene variants are not associated with type 2 diabetes, although GHRL promoter variants have significantly different activities.
Assuntos
Diabetes Mellitus Tipo 2/genética , Grelina/genética , Receptores de Grelina/genética , Animais , Linhagem Celular Tumoral , Estudos de Coortes , DNA/química , DNA/genética , Feminino , França , Predisposição Genética para Doença , Variação Genética , Haplótipos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Ratos , TransfecçãoRESUMO
BACKGROUND: The GH secretagogue receptor type 1a gene (GHSR) encodes the cognate receptor of ghrelin, a gut hormone that regulates food intake and pituitary GH secretion. Previous studies in U.S. families and a German population suggested GHSR to be a candidate quantitative locus for association with human obesity and growth. AIM: The aim of the study was to test common genetic variation in GHSR for association with body size in children and adults. METHODS: Sequencing was performed to systematically identify novel single nucleotide polymorphisms (SNPs) in GHSR. A set of three haplotype-tagging SNPs that captured all the genetic variation in GHSR was identified. These three haplotype-tagging SNPs were then genotyped in three large population-based U.K. cohort studies (two adult and one childhood cohort) comprising 5807 adults and 843 children. RESULTS: No significant genotype or haplotype associations were found with adult or childhood height, weight, or body mass index. CONCLUSION: Common variation in GHSR is not associated with body size in U.K. adults or children.
Assuntos
Tamanho Corporal/genética , Polimorfismo de Nucleotídeo Único , Receptores de Grelina/genética , Adulto , Idoso , Criança , Pré-Escolar , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Frequência do Gene , Genótipo , Humanos , Lactente , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Reino UnidoRESUMO
CONTEXT: Recently, a quantitative trait locus for stature was reported on chromosome 3p26 in patients with type 2 diabetes. OBJECTIVE: Given that ghrelin is a peptide involved in GH release and located on 3p26, we hypothesized that variation within its gene (GHRL) may be responsible for the quantitative trait locus on 3p26. DESIGN: The evidence for linkage around GHRL was refined with the genotyping of an additional four microsatellites (D3S4545, D3S1537, D3S1597, and D3S3611), giving a total of 27 markers, followed by multipoint variance components linkage analysis. Probands from the linkage families were typed for five common single nucleotide polymorphisms (SNPs) within GHRL and tested for association with adult stature using haplotype trend regression. RESULTS: The maximum multipoint evidence for linkage between adult stature and the 27 microsatellites yielded an LOD score of 2.58 (P = 0.0003) between D3S1297 and D3S1304. Five common (frequency of > or =5%) SNPs were typed in the probands [two promoter SNPs (rs27647 and rs26802), two exonic (rs696217 and rs4684677), and one intronic (rs35683)] capturing 80% of the total common variation in GHRL. No association was found between any SNP (or haplotypes thereof) and adult stature. CONCLUSION: Common genetic variation within GHRL is not responsible for variation in adult stature in this population.
Assuntos
Estatura/genética , Diabetes Mellitus Tipo 2/genética , Hormônios Peptídicos/genética , Adulto , Idoso , Feminino , Ligação Genética , Variação Genética , Grelina , Haplótipos , Humanos , Masculino , Repetições de Microssatélites , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Locos de Características QuantitativasRESUMO
Ghrelin and its receptor are widely distributed in cardiovascular tissues, and there is no doubt that the effects of ghrelin in the cardiovascular system are mediated not only via its growth-hormone-releasing effect but also by direct effects on the heart. Indeed, new pharmacological approaches with animal and cell models using elegant study designs have described new functions of ghrelin, providing new potential therapeutic opportunities for ghrelin in cardiovascular medicine.
Assuntos
Baixo Débito Cardíaco/tratamento farmacológico , Hipertensão/tratamento farmacológico , Isquemia Miocárdica/tratamento farmacológico , Hormônios Peptídicos/fisiologia , Animais , Grelina , Humanos , Hormônios Peptídicos/genética , Hormônios Peptídicos/uso terapêuticoRESUMO
Endocannabinoids and ghrelin are potent appetite stimulators and are known to interact at a hypothalamic level. However, both also have important peripheral actions, including beneficial effects on the ischemic heart and increasing adipose tissue deposition, while ghrelin has direct effects on carbohydrate metabolism. The AMP-activated protein kinase (AMPK) is a heterotrimeric enzyme that functions as a fuel sensor to regulate energy balance at both cellular and whole body levels, and it may mediate the action of anti-diabetic drugs such as metformin and peroxisome proliferator-activated receptor gamma agonists. Here we show that both cannabinoids and ghrelin stimulate AMPK activity in the hypothalamus and the heart, while inhibiting AMPK in liver and adipose tissue. These novel effects of cannabinoids on AMPK provide a mechanism for a number of their known actions, such as the reduction in infarct size in the myocardium, an increase in adipose tissue, and stimulation of appetite. The beneficial effects of ghrelin on heart function, including reduction of myocyte apoptosis, and its effects on lipogenesis and carbohydrate metabolism, can also be explained by its ability to activate AMPK. Our data demonstrate that AMPK not only links the orexigenic effects of endocannabinoids and ghrelin in the hypothalamus but also their effects on the metabolism of peripheral tissues.