Diurnal rhythms of leptin and ghrelin in the systemic circulation and in the gastric mucosa are related to food intake in rats

We investigated diurnal changes in leptin and ghrelin levels in the stomach and in the systemic circulation and their relation to food intake rhythms in Wistar rats housed at 22 °C with a 12-h light/dark cycle and free access to food and water. Animals were sacrificed every 3 h over a 24-h period. Leptin and ghrelin levels in serum and in the gastric mucosa were analysed by immunoassay. Leptin mRNA levels were determined in the gastric mucosa by RT-PCR and in different adipose tissue depots (epididymal, retroperitoneal and mesenteric) by Northern blot. Ghrelin mRNA levels were determined by Northern blot. Gastric and serum leptin levels displayed similar diurnal rhythms, rising during the dark phase and decreasing gradually during the light phase. Leptin expression in the different adipose tissue depots correlated positively with circulating leptin levels (P<0.05), although there were some depot-associated differences. Leptin mRNA levels in the mesenteric depot correlated positively with food intake (P<0.05). In blood, ghrelin levels rose sharply just before the onset of the dark phase and dropped suddenly just after. In the stomach, ghrelin levels were high during the fasting period of light and low during the night, and correlated inversely with food intake, gastric contents and serum leptin levels (P<0.05). Leptin and ghrelin in the stomach and in the systemic circulation thus show diurnal variations that are influenced by food intake rhythms. The results agree with a role for ghrelin as a stimulant of meal initiation.     

Gastric capacity, test meal intake, and appetitive hormones in binge eating disorder

Binge eating disorder (BED), characterized by ingestion of very large meals without purging afterwards, is found in a subset of obese individuals. We showed previously that stomach capacity is greater in obese than in lean subjects, and in this study, we investigated capacity in obese individuals with BED. We also determined ad-libitum intake of a test meal until extremely full. Furthermore, we measured various appetitive hormones (insulin, leptin, glucagon, CCK, ghrelin) and glucose before a fixed meal and for 120 min afterwards. An acetaminophen tracer was used to assess gastric emptying rate. We compared three groups of overweight women: 11 BED, 13 BE (subthreshold BED), and 13 non-binge-eating normals. The BED individuals had the largest stomach capacity as assessed by either maximum volume tolerated (P=.05) or by gastric compliance to pressure (P=.02) using an intragastric balloon. Although test meal intake did not differ between groups, it correlated (P=.03) with gastric capacity. The BED group showed a tendency (P=.06) to have greater area under the curve (AUC) and had higher values at 5 and 60 min (P<.05) for insulin compared to normals. Moreover, the BED subjects had lower ghrelin baselines premeal, and lower AUC for ghrelin, which then declined less postmeal than for the normals (P<.05). None of the other blood values differed, including glucose, leptin glucagon, and CCK, as well as acetaminophen, reflecting gastric emptying. The lower ghrelin in BED, although contrary to what was expected, is consistent with lower ghrelin in obesity, and suggests down-regulation of ghrelin by overeating. The lack of differences in CCK is consistent with the lack of differences in gastric emptying rate, given that CCK is released when nutrients reach the intestine. The results show that BED subjects have a large gastric capacity as well as abnormalities in meal-related ghrelin and insulin patterns that may be factors in binge eating.     

脑肠肽Ghrelin对血管紧张素Ⅱ诱导的大鼠脑主动脉血管平滑肌细胞胞内钙浓度上升的影响

目的研究脑肠肽Ghrelin对血管紧张素Ⅱ（AngⅡ）诱导的大鼠胸主动脉血管平滑肌细胞（VSMCs）胞内钙离子浓度上升的作用,并初步探讨其机制。方法采用细胞培养的方法获得大鼠胸主动脉VSMCs,经Fluo-4-AM染色后,采用激光共聚焦显微镜技术分别检测加入AngⅡ后,1×10^-7mol·L^-1和1×10^-5mol·L^-1的Ghrelin对胞内钙荧光强度（FI）的影响,以及腺苷酸环化酶抑制剂Sq22536对Ghrelin作用的影响。结果①2×10^-8mol·L^-1的AngⅡ可以使得VSMCs胞内钙FI上升到静息状态的（165±76）％;②随后加入1×10^-7mol·L^-1的Ghrelin可以使FI下降到（117±34）％（P〈0．01vs①）;③加入1×10^-5mol·L^-1的Ghrelin可以使FI下降到（87±22）％（P〈0．01vs①,P〈0．05vs②）;④预先以1×10^-5mol·L^-1的Sq22536孵育,再加入2×10^-8mol·L^-1AngⅡ和1×10^-7mol·L^-1的Ghrelin,胞内FI为原来的（143±24）％（P〈0．01vs②）。结论Ghrelin能够降低AngⅡ引起的大鼠胸主动脉VSMCs胞内钙离子升高作用,其机制可能与激活胞内cAMP／PKA信号通路有关。     

急性胰腺炎患者外周血NGAL和胃饥饿素水平变化及其意义

目的:探讨中性粒细胞明胶酶相关脂质运载蛋白(NGAL)及胃饥饿素(ghrelin)在急性胰腺炎(AP)患者外周血中水平高低与患者病情的关系。方法:收集既往收治的199例AP患者资料,其中急性轻症胰腺炎(MAP)103例,急性重症胰腺炎(SAP)96例;治疗期间17例(8.54%)死亡,分别比较不同病情与不同治疗转归及患者间入院第1天NGAL、ghrelin水平和其他指标的差异。结果:SAP患者的NGAL、ghrelin、C反应蛋白(CRP)、白细胞(WBC)、血淀粉酶、降钙素原(PCT)、APACHE II评分、Balthazar CT评分、BISAP指数均明显高于MAP组患者(均P0.05);治疗期间死亡患者的以上指标也均明显高于存活患者(均P0.05);199例患者外周血中NGAL、ghrelin水平与BISAP指数呈明显正相关关系(r=0.579、0.482,均P0.05),且两者与BISAP指数的相关性优于血淀粉酶、WBC、PCT。结论:AP患者外周血中NGAL、ghrelin水平与病情密切相关,两者水平增高预示着患者预后的不良的风险增大。     

Characterization of turkey and chicken ghrelin genes, and regulation of ghrelin and ghrelin receptor mRNA levels in broiler chickens

Ghrelin, a peptide hormone produced by the stomach in mammals, stimulates growth hormone release and food intake. Recently, ghrelin was identified and characterized in chicken proventriculus and shown to stimulate growth hormone release but inhibit feed intake. The purpose of this work was to identify and further characterize the ghrelin gene in chickens and in turkeys. Using molecular cloning techniques we have sequenced cDNAs corresponding to chicken (White Leghorn) and turkey ghrelin mRNAs. A total of 844 (chicken) or 869 (turkey) bases including the complete coding regions (CDS), and the 5'- and 3'-untranslated regions (UTRs) were determined. Nucleotide sequence (CDS) predicted a 116 amino acid precursor protein (preproghrelin) for both the chicken and the turkey that demonstrated complete conservation of an N-terminal 'active core' (GSSF) including a serine (position 3 of the mature hormone) known to be a modification (acylation) site important for ghrelin bioactivity. Additional nucleotide sequence was found in the 5'-UTRs of both Leghorn and turkey cDNAs that was not present in broilers or the red jungle fowl. The turkey ghrelin gene, sequenced from genomic DNA templates, contained five exons and four introns, a structure similar to mammalian and chicken ghrelin genes. Ghrelin was highly expressed in proventriculus with much lower levels of expression in other tissues such as pancreas, brain, and intestine. RT-PCR was used to quantify ghrelin mRNA levels relative to 18S rRNA in 3-week-old male broiler chickens. The level of ghrelin mRNA increased in proventriculus in response to fasting but did not decline with subsequent refeeding. Plasma ghrelin levels did not change significantly in response to fasting or refeeding and did not appear to reflect changes in proventriculus ghrelin mRNA levels. Ghrelin mRNA levels declined in broiler pancreas after a 48 h fast and increased upon refeeding. Expression of the gene encoding the receptor for ghrelin (growth hormone secretagogue receptor, GHS-R) and a variant form was detected in a variety of tissues collected from 3-week-old male broiler chickens possibly suggesting autocrine/paracrine effects. These results offer new information about the avian ghrelin and ghrelin receptor genes and the potential role that this system might play in regulating feed intake and energy balance in poultry.     

Presence and distribution of ghrelin-immunopositive cells in the chicken gastrointestinal tract

The presence and distribution patterns of ghrelin, a gastric acylated peptide, were studied in the entire gastrointestinal tract of the chicken (Gallus domesticus) using the peroxidase-antiperoxidase immunohistochemical method, western blot analysis and a specific antibody against the C-terminal region of rat ghrelin. Ghrelin-immunopositive cells were observed in the mucosal layer of all segments examined. The largest numbers of ghrelin-positive cells were located at the base of lobuli of the proventriculus gland, along villi of the intestines and in crypts of the duodenum. Lower numbers of ghrelin-immunostained cells were located in crypts of jejunum and ileum and only few ghrelin-immunostained cells were detected at the base of crypts of the large intestine. Closed and open types of cells were observed in all segments. Western blot analysis confirmed the presence of ghrelin-like protein in the entire chicken gastrointestinal tract. The anatomical distribution patterns and the morphological characteristics of chicken ghrelin-positive cells suggest that they are endocrine cells. Furthermore, it is concluded that ghrelin shows a high degree of preservation during evolution from non-mammalian vertebrates to mammals.     

Ghrelin对糖尿病大鼠海马神经元凋亡及其认知功能的影响

目的观察Ghrelin对糖尿病大鼠海马神经元凋亡及其认知功能的影响。方法 40只雄性SD大鼠随机均分为对照组、糖尿病组、糖尿病+Ghrelin组和糖尿病+Ghrelin+D-lys3-GHRP-6组,每组10只。建立STZ糖尿病模型,open-field实验排除合并抑郁症的糖尿病大鼠,Morris水迷宫测试学习与记忆能力,RT-PCR检测海马caspase-3mRNA的表达,免疫组化检测海马神经元caspase-3和BCL-xl蛋白表达,原位末端标记法检测海马神经元的凋亡。结果与对照组大鼠相比,糖尿病组和糖尿病+Ghrelin+D-lys3-GHRP-6组学习与记忆能力显著受损(P<0.05),海马部位的caspase-3 mRNA及蛋白表达明显升高(P<0.05),BCL-xl蛋白表达显著下降(P<0.05),海马神经元的凋亡增加了49%(P<0.05)。糖尿病+Ghrelin组学习和记忆成绩明显优于糖尿病组(P<0.05),其海马部位的caspase-3 mRNA及蛋白表达显著下降(P<0.05),BCL-xl蛋白表达明显升高(P<0.05),海马神经元的凋亡减少了42%。结论 Ghrelin对糖尿病大鼠...     

Ghrelin levels are associated with hunger as measured by the Three-Factor Eating Questionnaire in healthy young adults

Weight gain and appetite regulation are complex interplays between internal and external cues. Our aim was to investigate the association of eating behaviors with ghrelin taking into account lifestyle. We conducted a cross-sectional analysis in a sample of first-year university students at the Université de Sherbrooke. We collected medical history, anthropometric measurements, vital signs, fitness index, and fasting blood samples. Questionnaires included a lifestyle questionnaire and the Three-Factor Eating Questionnaire (TFEQ) estimating dietary restraint, disinhibition and hunger. We recruited 308 participants aged 20.7 ± 3.2 years and a mean BMI of 23.3 ± 3.4 kg/m². Hunger score was significantly associated with ghrelin levels (r = 0.11, P < 0.05). In women, this association was independent of age, BMI, dietary and lifestyle factors (P = 0.02). The association between ghrelin level and hunger score was observable in leaner individuals (r = 0.28, p < 0.0001) but not in heavier individuals (r = − 0.08, p = 0.34; stratified by BMI < vs > 22.6 kg/m²). Restraint (R) and disinhibition (D) were not associated with ghrelin levels. The three eating behaviors demonstrated expected correlations with lifestyle supporting the validity of the TFEQ in this cohort. In conclusion, we demonstrated that ghrelin, a biological marker, is associated with self-reported perception of hunger, independently of anthropometric measures and lifestyle.     

枳术丸对功能性消化不良大鼠胃排空功能及Ghrelin、5-HT、CGRP的影响

目的研究枳术丸对碘乙酰胺（iodoacetamide,IA）诱导的功能性消化不良（FD）大鼠胃排空功能及胃促生长素（Ghrelin）、5-羟色胺（5-HT）、降钙素基因相关肽（CGRP）的影响,初步探讨枳术丸治疗FD的机制。方法雄性10日龄SD幼鼠80只,随机分为正常组30只、造模组50只。采用0.1%IA灌胃诱导大鼠FD模型。大鼠8周龄时进行模型评定,将造模成功的50只大鼠随机分为模型组30只、枳术丸组10只和阳性药组10只。正常组与模型组予0.9%氯化钠溶液1m L/100 g灌胃,枳术丸组予生药含量1.575 g/mL的水煎剂1 mL/100 g灌胃,阳性药组给予0.3 mg/mL多潘立酮溶液1 mL/100 g灌胃。采用苏木素-伊红（Hematoxin eosin,HE）染色观察大鼠胃组织病理变化;采用放射免疫法检测大鼠胃组织髓过氧化物酶（MPO）活性以及血清中Ghrelin、5-HT、CGRP的含量。结果枳术丸组和阳性药组大鼠3 h摄食量和胃排空率显著大于模型组（P〈0.05）,而3 h后胃内食物残余量小于模型组（P〈0.05）。枳术丸组和阳性药组均可不程度地升高FD大鼠血清中Ghrelin和5-HT的含量（P〈0.05,P〈0.01）,但2组间差异无统计学意义;枳术丸组和阳性药组均可不程度地降低FD大鼠血清中CGRP的含量（P〈0.05）,但2组间差异无统计学意义。结论枳术丸对IA诱导的FD大鼠具有加快胃排空、降低胃肠感觉过敏的作用,其机制可能与外周血Ghrelin、5-HT含量提高,CGRP含量降低有关。