Ghrelin mRNA Expression Research Articles

Glucagon-Like Peptide-1 Receptor Agonist, Exendin-4, Regulates Feeding-Associated Neuropeptides in Hypothalamic Neurons in Vivo and in Vitro

Exendin-4, a long-acting glucagon-like peptide-1 receptor (GLP-1R) agonist, is a potential regulator of feeding behavior through its ability to inhibit gastric emptying, reduce food intake, and induce satiety. GLP-1R activation by exendin-4 induces anorexia; however, the specific populations of neuropeptidergic neurons activated by exendin-4 within the hypothalamus, the central regulator of energy homeostasis, remain unclear. This study determines whether exendin-4 regulates hypothalamic neuropeptide expression and explores the signaling mechanisms involved. The distribution and quantity of exendin-4-induced c-Fos immunoreactivity were evaluated to determine activation of α-melanocyte-stimulating hormone/proopiomelanocortin, neuropeptide Y, neurotensin (NT), and ghrelin neurons in hypothalamic nuclei during exendin-4-induced anorexia in mice. Additionally, exendin-4 action on NT and ghrelin transcript regulation was examined in immortalized hypothalamic neurons. With anorexia induced by intracerebroventricular exendin-4, α-melanocyte-stimulating hormone/proopiomelanocortin and neuropeptide Y neurons were activated in the arcuate nucleus, with simultaneous activation of NT-expressing neurons in the paraventricular nucleus, and ghrelin-expressing neurons in the arcuate nucleus, paraventricular nucleus, and periventricular hypothalamus, suggesting that neurons in one or more of these areas mediate the anorexic action of exendin-4. In the hypothalamic neuronal cell models, exendin-4 increased cAMP, cAMP response element-binding protein/activating transcription factor-1 and c-Fos activation, and via a protein kinase A-dependent mechanism regulated NT and ghrelin mRNA expression, indicating that these neuropeptides may serve as downstream mediators of exendin-4 action. These findings provide a previously unrecognized link between central GLP-1R activation by exendin-4 and the regulation of hypothalamic NT and ghrelin. Further understanding of this central GLP-1R activation may lead to safe and effective therapeutics for the treatment of metabolic disorders.

Expression of ghrelin and the ghrelin receptor in different stages of porcine corpus luteum development and the inhibitory effects of ghrelin on progesterone secretion, 3β-hydroxysteroid dehydrogenase (3β-honestly significant difference (HSD)) activity and protein expression

Recent studies have suggested that ghrelin plays a direct role in controlling female reproduction. The aim of the present study was to investigate the mRNA and protein expression of ghrelin and its receptor (via real time PCR, Western blot and immunohistochemistry analysis, respectively) in porcine corpora lutea (CL) collected during early (CL1: 1–2 days after ovulation), middle (CL2: 7–10 after ovulation), and late luteal phase (CL3: 13–15 after ovulation). Ghrelin expression and concentration of both acylated and unacylated forms of ghrelin significantly increased during CL development. Immunohistochemistry analysis shown localization of ghrelin protein in the cytoplasm of large luteal cells. No changes in the expression of the ghrelin receptor were observed. Direct in vitro effects of ghrelin on progesterone (P4) secretion and 3-beta-hydroxysteroid dehydrogenase (3β-honestly significant difference (HSD)) activity, which were measured by the conversion of pregnenolone (P5) to P4, and 3β-HSD protein expression were then analyzed. To assess 3β-HSD activities, mature luteal cells were first cultured for 24 h with ghrelin at 100, 250, 500 and 1000 pg/mL with P5, or with aminoglutethimide (AMG). AMG is an inhibitor of CYP11A1-mediated hydroxylation; an addition of AMG and P5 enabled P4 production to serve as an index of 3β-HSD activity. Inhibitory effects of ghrelin on P4 secretion, 3β-HSD activity and protein expression were observed. In conclusion, the presence of ghrelin and its receptor in porcine corpora lutea and the direct inhibitory effects of ghrelin on luteal P4 secretion and 3β-HSD suggest potential auto/paracrine regulation by ghrelin in the luteal phase of ovary function.

Mapping of Ghrelin Gene Expression and Cell Distribution in the Stomach of Morbidly Obese Patients—a Possible Guide for Efficient Sleeve Gastrectomy Construction

Ghrelin is secreted mainly in the stomach and plays a role in food intake regulation. Morbidly obese (MO) individuals report a decline in appetite after sleeve gastrectomy (SG), presumably due, in part, to ghrelin cell removal. Ghrelin cell distribution and expression were determined in three areas of resected stomach specimens from MO patients subjected to SG. Resected stomach specimens from 20 MO patients undergoing SG were analyzed. Real-time polymerase chain reaction of ghrelin mRNA and immunohistostaining for ghrelin cells in three stomach regions (fundus, body, and pre-antral areas) were performed. Body mass index (BMI) and total plasma ghrelin levels were obtained before and 3 months postoperatively. Ghrelin mRNA was detected throughout the stomach, its expression decreasing from the fundus towards the antrum. The relative quantification for ghrelin mRNA expression was 0.043, 0.026, and 0.015 at the fundus, body, and pre-antral region, respectively (P = 0.05, fundus vs. pre-antral region). Average ghrelin cell counts declined from 60 ± 40 to 45 ± 20 and 39 ± 13 cells/high power fields in the fundus, body, and pre-antral region, respectively. Three months after surgery, total plasma ghrelin levels decreased from 1,676 ± 470 to 1,179 ± 188 pg/ml (P < 0.00001) and BMI dropped from 46 ± 6 to 38 ± 5 kg/m2 (P < 0.00001). Distribution and expression of ghrelin-secreting cells throughout the stomach were defined, emphasizing the importance of meticulous resection of the fundus during SG for maximal ghrelin cell removal.

Underlying Mechanism of Aconitum Lizhong Acting on Experimental Hypothermia with Indigestion in Rats: Role of Ghrelin

This study is aimed to investigate the Aconitum Lizhong pill (ALZ) pharmacological actions on hypothermia with indigestion, especially the ghrelin roles. The littermate-matched rats were randomly divided into four groups. Control did sham operation or standard diet, Model carried out interscapular brown adipose (IBA) removal with standard diet, Fat-diet did IBA removal with fat-diet, and ALZ did IBA removal and fat-diet with 4.536 g/kg/d ALZ. The potency of adaptive thermogenesis, ghrelin levels in plasma or gastric mucosa, thyroid hormones and metabolite in sera, expression of ghrelin mRNA, and protein in gastric mucous membrane were determined. ALZ relieved the hypothermia processes with indigestion, via inhibiting ghrelin expression and increasing ghrelin secretion; the dynamics from the therapy is supported with the energy changes as less body weight loss, less plasma lipid decrease, more plasma T 3 or T 4 increase with TSH decrease, and more compensation of thermogenic AUC decrease. Ghrelin played key roles in the actions of ALZ on the hypothermia with indigestion. The pharmacological mechanisms of ALZ involved the homeostasis of ghrelin expression and secretion.

Influence ofHelicobacter pyloriinfection on ghrelin levels in children

To compare ghrelin levels in plasma and gastric mucosa before and after Helicobacter pylori (H. pylori) treatment in children with H. pylori-associated functional dyspepsia. Children with H. pylori-associated functional dyspepsia were enrolled in this study. H. pylori infection was confirmed by positive bacterial culture results. All of the children received triple H. pylori eradication therapy (a 2 wk course of omeprazole, amoxicillin, and clarithromycin). The children were divided into two groups based on the success of the H. pylori treatment: group 1 (eradicated) - patients who had a negative 13C-urea breath test 2 mo after the end of therapy; and group 2 (non-eradicated) - patients who had a positive 13C-urea breath test. Plasma ghrelin, gastric ghrelin mRNA, and the body mass index were evaluated in both groups before and after the H. pylori treatment. The plasma ghrelin levels were measured by a radioimmunoassay. The expression of gastric ghrelin mRNA was determined by real-time reverse transcription polymerase chain reaction. A total of 50 children with H. pylori-associated functional dyspepsia were treated with triple H. pylori eradication therapy. The mean age of the children was 5.52 ± 0.83 years, and there were 28 males and 22 females. Among the 50 H. pylori-positive children, 30 successfully achieved eradication, and 20 did not. The mean plasma ghrelin levels of group 1 were 22.17 ± 1.73 ng/L and 26.59 ± 2.05 ng/L before and after the treatment, respectively, which was a significant increase (P = 0.001). However, the mean plasma ghrelin level of group 2 before and after the H. pylori treatment was 21.34 ± 2.40 ng/L and 22.24 ± 2.10 ng/L (P = 0.785). The plasma ghrelin levels increased substantially after treatment in group 1 but showed only minor changes in group 2. Similarly, the gastric ghrelin mRNA expression in group 1 before treatment was 2.84 ± 0.08. After treatment, the level was 3.11 ± 0.65, which was significantly different (P = 0.023). The gastric ghrelin mRNA expression in group 2 did not change significantly during the treatment (2.82 ± 0.44 vs 2.79 ± 0.31, P = 0.875). The plasma ghrelin and gastric ghrelin mRNA levels in group 1 increased substantially after the treatment but did not do so in group 2. In addition, the body mass index the two groups did not differ significantly 2 mo before and after the H. pylori treatment. H. pylori eradication increases the plasma and tissue ghrelin levels in children with H. pylori-associated functional dyspepsia.

Cysteamine improves growth performance and gastric ghrelin expression in preweaning piglets

The aim of the present study was to investigate the effect of cysteamine on growth performance of preweaning piglets and gastric expression of ghrelin mRNA in vivo and in vitro. Twelve litters of newborn piglets were allocated randomly to control and treatment groups. From 15 d of age, piglets in the control group were fed basal creep diet, whereas the treatment group received basal diet supplemented with 120 mg cysteamine per kg of diet until weaning on 35 d of age. Body weight gain, creep feed consumption, and diarrhea rates were recorded, and gastric mucosal tissues were collected for quantifying mRNA expression. To evaluate the direct effect of cysteamine on gastric ghrelin expression, primary cultures of gastric mucosal cells isolated from 35-d-old piglets were exposed to cysteamine for 20 h at 0, 1, 10, and 100 μg/mL, respectively. Dietary cysteamine increased (P < 0.05) average daily creep feed consumption and BW gain in preweaning pigs, which was accompanied by reduction in diarrhea rates. At 35 d of age, piglets treated with cysteamine showed increased (P < 0.05) ghrelin and gastrin and decreased (P < 0.05) somatostatin mRNA expression in gastric mucosa. Moreover, dietary cysteamine treatment increased serum concentration of gastrin (P < 0.05). In vitro, cysteamine significantly increased ghrelin mRNA expression in gastric mucosal cells at the concentration of 10 μg/mL. In conclusion, dietary cysteamine is effective in improving the growth performance and health condition of preweaning piglets, which is associated with its stimulatory effects on gastric ghrelin mRNA expression both in vivo and in vitro.

Interferon-γ inhibits ghrelin expression and secretion via a somatostatin-mediated mechanism.

To investigate if and how the proinflammatory cytokine interferon γ (IFNγ) affects ghrelin expression in mice. The plasma concentration of ghrelin, and gastric ghrelin and somatostatin expression, were examined in wild-type mice and mice infected with Helicobacter pylori (H. pylori). Furthermore, ghrelin expression was examined in two achlorhydric mouse models with varying degrees of gastritis due to bacterial overgrowth. To study the effect of IFNγ alone, mice were given a subcutaneous infusion of IFNγ for 7 d. Finally, the influence of IFNγ and somatostatin on the ghrelin promoter was characterized. H. pylori infection was associated with a 50% reduction in ghrelin expression and plasma concentration. Suppression of ghrelin expression was inversely correlated with gastric inflammation in achlorhdyric mouse models. Subcutaneous infusion of IFNγ suppressed fundic ghrelin mRNA expression and plasma ghrelin concentrations. Finally, we showed that the ghrelin promoter operates under the control of somatostatin but not under that of IFNγ. Gastric infection and inflammation is associated with increased IFNγ expression and reduced ghrelin expression. IFNγ does not directly control ghrelin expression but inhibits it indirectly via somatostatin.

English

The aim of this study was to investigate the validity of exogenous ghrelin for early recovery in partial gastrectomied rats and its long-term significance. The subtotal gastrectomy (B-ltype) rats were randomly divided into two groups, and then they were treated with intraperitoneal injection of saline or ghrelin intervention respectively. Body weight and daily food intake were measured before and after the operation. After the seventh post-operation, rats were sacrificed, the fundus of stomach and anastomosis were taken to determine ghrelin mRNA expression by real-time fluorescent quantitive PCR and to evaluate the bursting pressure and hydroxyproline content. There was no difference in body weight between the two groups (P&gt;0.05); Cumulative food intake (P&lt;0.05) was 52.50&plusmn;6.77 g in ghrelin group, which was higher than that in the saline group (45.67&plusmn;7.47 g) (P&lt;0.05). However, the ghrelin mRNA expression in ghrelin treatment group was lower than the control group (P&lt;0.01). Compared to the saline group, ghrelin-treated rats displayed a higher bursting pressure (P&lt;0.05) and anastomotic hydroxyproline content (P&lt;0.01). Ghrelin treatment in the postoperative rats can promote energy metabolism and body weight gain. Ghrelin could effectively promote the healing of anastomosis. However, exogenous ghrelin intervention may inhibit the early compensatory mechanisms of residual stomach. &nbsp; Key words:&nbsp;Ghrelin, subtotal gastrectomy, rats, anastomosis, early recovery.

Ghrelin reduces rat myocardial calcification induced by nicotine and vitamin D3 in vivo

Ghrelin, a newly discovered bioactive peptide, initially was identified as a strong stimulant for the release of growth hormone (GH) and that has improved cardiac function in patients suffering from end-stage chronic heart failure. Increasing evidence has demonstrated that ghrelin may have myocardial protective effects. However, the role of ghrelin in the pathogenesis of cardiovascular diseases remains unclear. In this study, an in vivo model of rat myocardial calcification induced by vitamin D3 and nicotine was used to study the possible mechanism in the regulatory action of ghrelin on the calcified myocardium. Calcification increased total Ca2+ content and 45Ca2+ deposition in the myocardium and alkaline phosphatase (ALP) activation in the plasma. Compared with the control group, ghrelin mRNA expression was up-regulated and the myocardium calcium content was significantly increased in vitamin D3 and nicotine-treated rats. Rats were subcutaneously injected with 1 or 10 nmol/kg ghrelin. Rats treated with both low- and high-dose ghrelin decreased total Ca2+ content and 45Ca2+ deposition in cardiac muscle and inhibited ALP activation in the myocardium and plasma, in a concentration-dependent manner. In addition, osteopontin (OPN) mRNA expression significantly decreased and that of endothelin (ET-1) significantly increased with myocardial calcification. Ghrelin treatment increased OPN expression at the mRNA level and reduced ET-1 mRNA expression in a dose-dependent manner. These results indicate that exogenous administration with ghrelin attenuates myocardial calcification induced by nicotine and vitamin D3, and that the possible mechanism is via the ghrelin-induced increase in the OPN mRNA levels and decrease in the ET-1 mRNA expression in the myocardium.

CDNA cloning of ghrelin and ontogeny of ghrelin mRNA expression in the gastrointestinal tract of African ostrich chicks

Ghrelin is the endogenous ligand for the growth hormone secretagogue receptor (GHS-R). The sequence of ghrelin has been determined in many species ranging from fish to mammals. The ostrich is the largest herbivorous bird in the world. Although the distribution, morphological characteristics, and developmental changes of ghrelin-producing cells in the gastrointestinal tract of African ostrich chicks have recently been determined, the sequence and structure of ghrelin and its expression in the gastrointestinal tract of African ostrich chicks have not been studied. In the present study, the sequence and structure of ghrelin and its expression in the gastrointestinal tract of African ostrich chicks were investigated by reverse-transcriptase polymerase chain reaction (RT-PCR). Results of cDNA cloning revealed that African ostrich ghrelin is composed of 28 amino acid residues and the sequence of the 7 amino acids of the N-terminal region of African ostrich ghrelin was identical with that of other birds. Ninety-day-old female African ostriches were used to investigate the expression of ghrelin in the gastrointestinal tract. The results showed that ghrelin mRNA existed in the proventriculus, gizzard, duodenum, ileum, cecum, and rectum; there was no expression in the jejunum and colon. We observed developmental changes in the ghrelin mRNA expression in the stomach and small intestine of African ostriches. The results of the present study showed that ghrelin mRNA existed on day 1 in the proventriculus, but there was no expression in other tissues. On day 45, ghrelin mRNA existed in the proventriculus, gizzard, and ileum; however, there was no expression in the duodenum and jejunum. On day 90 and 334, we detected ghrelin mRNA in the proventriculus, gizzard, duodenum, and ileum, but there was no expression in the jejunum. The results of the present study clearly demonstrate that ghrelin mRNA exists and the distribution of ghrelin mRNA in the gastrointestinal tract of African ostriches changes with age (from postnatal day 1 to day 334).

1048 Ghrelin Mrna and Protein Expression in Fundus of Stomach in Postnatal Rats Born Small for Gestational Age

Objectives: To explore the changes of Ghrelin mRNA and protein expression in fundus of stomach of rats born small for gestational age (SGA) and to analyze the relationship with the catch-up growth of postnatal SGA rats. Methods: The SGA and appropriate for gestational age (AGA) rat models were established by food restriction during pregnancy. The newborn rats were acquired after delivery, among which 10 rats were selected randomized from SGA group and control group at postnatal day 0, day 20 and day 40. The fundus of stomach were taken to determine Ghrelin mRNA and protein expression. Results: At postnatal day 0, stomach fundus Gherlin mRNA expression is higher in SGA rats than those in AGA rats. At postnatal day 20 and day 40, there is no significant difference in Gherlin mRNA expression among SGA non-catching-up rats, SGA catching-up rats and normal AGA rats. At postnatal day 0, Gherlin IOD in SGA rats is higher than that in AGA rats. Ghrelin IOD in SGA catching-up rats is higher than that in SGA non-catching-up rats and normal AGA rats (p< 0.05) at postnatal day 20, and there is no significant difference among them at postnatal day 40. Conclusion: Ghrelin might be involved in the physiological regulation or pathological process of IUGR, and intrauterine nutritional status or endocrine circumstances may have effects on the cells of gastric fundus of the fetuses, which results in the up-regulation of Ghrelin expression. Ghrelin might also be involved in the regulation of catch-up growth in their early life.

Effects of Orexin A on mRNA Expression of Various Neuropeptides in the Hypothalamus and Pituitary, and on Serum LH Levels in Ovariectomized Gilts

Orexin has several biological functions, including the regulation of reproductive endocrine signaling, which has received much attention. However, little is known about the mechanism through which orexin regulates the levels of neuroendocrine hormones and peptides. We injected orexin A or physiological saline into the lateral ventricle of 10 ovariectomized (OVX) gilts, and determined the subsequent changes in serum luteinizing hormone (LH) concentration by using radioimmunoassay (RIA). We also examined the expression of GnRH, NPY, and POMC mRNAs in the hypothalamus and that of LH, follicle-stimulating hormone (FSH), POMC, and ghrelin mRNAs in the pituitary by using semi-quantitative reverse transcription polymerase chain reaction. We found the following results: (1) Orexin A transiently promoted LH secretion; serum LH concentration started to increase at 10 min after the orexin injection, peaked at 30 min, and returned to its initial level at 1.5 h; (2) orexin A upregulated GnRH mRNA expression and downregulated NPY and POMC mRNAs expression in the hypothalamus; (3) orexin A upregulated LH and FSH mRNAs expression (FSH, P > 0.05), but downregulated ghrelin mRNA expression in the pituitary. No significant effects were observed on the pituitary expression of FSH and POMC mRNAs. Our data suggest that orexin A regulates reproductive function by stimulating GnRH and LH release directly and indirectly via its effects on NPY, POMC and ghrelin expression.

Regulation of gastric hormones by systemic rapamycin

The mammalian target of rapamycin (mTOR), an evolutionarily conserved serine–threonine kinase, is an intracellular fuel sensor critical for cellular energy homeostasis. Gastrointestinal endocrine cells play a vital role in the regulation of energy balance by secreting hormones that inform the brain about energy supply. Here we showed the localization of mTOR signaling molecules in more than 90% of gastric ghrelin cells and 36 ± 3% of gastrin cells, while no somatostatin-positive cell showed phospho-S6K1 immunoreactivity. Inhibition of mTOR significantly stimulated expression of gastric ghrelin mRNA and protein, and the concentration of plasma ghrelin (2.06 ± 0.34 ng/ml vs. 12.53 ± 3.9 ng/ml, p < 0.05), inhibited gastrin synthesis and secretion (75.01 ± 6.71 pg/ml vs. 54.04 ± 3.65 pg/ml, p < 0.05), but had no effect on somatostatin production (165.2 ± 25.07 pg/ml vs. 178.9 ± 29.14 pg/ml, p = 0.73). Gastric mTOR is a gastric sensor whose activity is linked to the differential regulation of gastric hormone production and release.

Effect of gsp oncogene on expression of GHSR - 1a and Ghrelin mRNA in human GH - secreting pituitary adenomas

Objective To investigate the relationship between the expression of growth hormone secretagogue receptor type 1a (GHSR- 1a) 、Ghrelin mRNA and gsp oncogene in human GH- secreting pituitary adenomas. Methods The gsp oncogene mutation in 43 cases of GH -secreting pituitary adenomas was detected by PCR - DNA direct sequencing analysis. The expression of GHSR - 1a、Ghrelin mRNA was determined by SYBR green real -time fluorescent quantitative PCR. Then the relationships between the expression of GHSR -1a、Ghrelin mRNA and gsp oncegene were studied with statistical methods. Results There was no significant difference in the expression of ghrelin mRNA between mutation - positive and mutation - negative adenomas. The expression of GHSR - 1a mRNA was significantly higher in gsp - positive adenomas than that in gsp - negative adenomas. There was significant direct correlation between the levels of ghrelin mRNA and GHSR mRNA expression in gsp - positive and gsp - negative adenomas. Conclusions Gsp oncogene may up - regulate the expression of GHSR - 1a mRNA and has no effect on the expression of ghrelin mRNA in human GH - secreting pituitary adenomas. In addition, ghrelin could elevate the expression of GHSR - 1a mRNA in GH - secreting pituitary adenomas with or without gsp mutations. Key words: Gsp oncogene; GHSR - 1a; Ghrelin

Regulation of ghrelin production by gastric mTOR signaling

The role of gastric mTOR signaling in ghrelin production and food intake was examined in this study. Here we showed the reciprocal relationship of gastric mTOR signaling and ghrelin during changes in energy status. mTOR activity was down‐regulated, while gastric preproghrelin and circulating total and acyl ghrelin were increased by fasting. In db/db mice, gastric mTOR signaling was enhanced, whereas gastric preproghrelin and circulating total anc acyl ghrelin were decreased. Inhibition of the gastric mTOR signaling by rapamycin stimulated the expression of gastric preproghrelin and ghrelin mRNA, increased the plasma total and acyl ghrelin in both wildtype and db/db mice. Activation of the gastric mTOR signaling by L‐leucine decreased the expression of gastric preproghrelin and the level of plasma total and acyl ghrelin. Ghrelin receptor antagonist D‐Lys3‐growth hormone releasing peptide‐6 abolished the rapamycin‐induced increment in food intake despite that plasma ghrelin remained elevated. mTOR is therefore a gastric fuel sensor whose activity is linked to the regulation of energy intake through ghrelin.

Ghrelin Gene Expression in Broiler Proventriculus Tissue are Changed by Feed Restriction, Different Dietary Energy and Protein Levels

Problem statement: The aim of this study was to investigate the effects of feed restriction and different energy and protein contents of diet on ghrelin gene expression in broiler chicken. Approach: Feeding programs consisted of ad libitum and feed restriction, two energy levels (3100 and 2800 kcal ME kg-1) and three protein levels (22.3, 19.3 and 16.3% CP). Feed restriction was applied during 22-32 days of age. Proventriculus samples were collected at 21, 32 and 49 days of age. Ghrelin mRNA expression in proventriculus tissue was quantitate using Real Time quantitative PCR. Results: We found that ghrelin gene expression was increased in restricted chicks compared with those fed ad libitum at 32 days of age (p = 0.09) but feed restriction had no effect on ghrelin gene expression at 49 days of age. A positive response in ghrelin gene expression was achieved by decreasing energy level in the diet at 21 days of age (p<0.07) and at 32 days of age (p<0.05). Also, we showed that dietary protein had no effect on ghrelin gene expression, whereas there was a tendency for an increase in ghrelin gene expression as protein decreased at total period. Conclusion: The present study, we investigated the effects of feed restriction and different energy and protein contents of the diet on ghrelin gene expression in broiler chicken. We have characterized chicken ghrelin cDNA in proventriculus tissue in broiler chicken. We also found that ghrelin gene expression is differently suppressed by diet manipulations. Additional studies are necessary to investigate the role of nutrition on ghrelin gene expression in proventriculus tissue in broiler chicken.

Eradication of Helicobacter pylori Increases Ghrelin mRNA Expression in the Gastric Mucosa

It has been suggested that Helicobacter pylori eradication may influence production of some peptides in the stomach, which can affect appetite. This hypothesis is controversial. To verify the hypothesis, we conducted this randomized controlled trial using H. pylori infected subjects without any gastrointestinal symptoms. The treatment group received triple H. pylori eradication therapy for 7 days and the control group received no medication. We measured ghrelin, obestatin and the tumor necrosis factor-α (TNF-α) mRNA levels in endoscopic biopsy specimens and the changes from baseline to follow-up. The plasma active n-octanoyl ghrelin and obestatin levels were measured in both groups. The ghrelin/obestatin ratios in plasma and gastric mRNA expression were calculated at baseline and follow-up. Ghrelin mRNA expression in the fundic mucosa after H. pylori eradication increased significantly compared to the control group (4.47±2.14 vs. 1.79±0.96, P=0.009), independent of inflammatory changes. However, obestatin mRNA expression decreased in the antral mucosa (-0.57±1.06 vs. 0.41±0.72, P=0.028). The treatment group showed a marginal increase (P=0.060) in plasma ghrelin/obestatin ratio. The TNF-α mRNA expression also decreased significantly with treatment. This randomized controlled trial demonstrates that H. pylori eradication increases ghrelin mRNA expression, independent of inflammatory cell changes.

Ghrelin in the human myometrium

BackgroundGhrelin is a 28-amino acid octanolyated peptide, synthesised primarily in the stomach. It stimulates growth hormone release, food intake and exhibits many other diverse effects. Our group have previously determined that ghrelin inhibited human contractility in vitro. The aim of this study therefore, was to investigate the expression of ghrelin, its receptor, the growth hormone secretagogue receptor type 1 (GHS-R1), ghrelin O-acyltransferase (GOAT) which catalyses ghrelin octanoylation, prohormone convertase 1/3 (PC1/3) responsible for pro-ghrelin processing, in human myometrium, during pregnancy prior to labour, during labour and in the non-pregnant state. Modulation of ghrelin and ghrelin receptor expression in cultured myometrial cells was also investigated.MethodsmRNA and protein were isolated from human myometrium and the myometrial smooth muscle cell line hTERT-HM; and real-time fluorescence RT-PCR, western blotting and fluorescence microscopy performed. The effects of β-Estradiol and bacterial lipopolysaccharide (LPS) on hTERT-HM gene expression were evaluated by western blotting.ResultsWe have reported for the first time the expression and processing of ghrelin, GHS-R1, GOAT and PC1/3 expression in human myometrium, and also the down-regulation of ghrelin mRNA and protein expression during labour. Furthermore, GHS-R1 protein expression significantly decreased at labour. Myometrial GOAT expression significantly increased during term non-labouring pregnancy in comparison to both non-pregnant and labouring myometrium. Mature PC1/3 protein expression was significantly decreased at term pregnancy and labour in comparison to non-pregnant myometrium. Ghrelin, GHS-R1, GOAT and PC1/3 mRNA and protein expression was also detected in the hTERT-HM cells. Ghrelin protein expression decreased upon LPS treatment in these cells while β-Estradiol treatment increased GHS-R1 expression.ConclusionsGhrelin processing occurred in the human myometrium at term pregnancy and in the non-pregnant state. GOAT expression which increased during term non-labouring pregnancy demonstrating a similar expression pattern to prepro-ghrelin and GHS-R1, decreased at labour, signifying possible myometrial ghrelin acylation. Moreover, the presence of PC1/3 may contribute to pro-ghrelin processing. These results along with the previous in vitro data suggest that myometrially-produced and processed ghrelin plays a significant autocrine or paracrine role in the maintenance of relaxation in this tissue during pregnancy. Furthermore, the significant uterine modulators LPS and β-Estradiol are involved in the regulation of ghrelin and ghrelin receptor expression respectively, in the human myometrium.

The traditional Japanese medicine Rikkunshito increases the plasma level of ghrelin in humans and mice

Rikkunshito is a traditional Japanese medicine that is widely used for treating upper gastrointestinal symptoms. Our purpose is to clarify the effect of Rikkunshito on the levels of peptide hormones and cytokines in healthy humans and mice. We administered Rikkunshito, 7.5 g per day, to 21 healthy volunteers for 2 weeks and examined the changes in plasma peptide and hormone levels. In mice, we administered free access to water containing 0, 0.7, 1.1, and 1.4% Rikkunshito for 2 weeks and examined the changes in plasma peptide levels and ghrelin mRNA expression levels in the stomach. Plasma acylated ghrelin levels at 0, 2, and 6 weeks after the start of administration were 6.7 +/- 2.8, 11.7 +/- 4.0, and 10.5 +/- 4.4 fmol/ml, respectively. The levels at 2 and 6 weeks were significantly higher than that at 0 time (p < 0.01, paired t test). The levels of interleukin 4, interferon gamma, and granulocyte colony-stimulating factor in plasma decreased significantly (p < 0.01) after the administration of Rikkunshito. In mice, the levels of plasma acylated ghrelin changed in proportion to the concentration of Rikkunshito in drinking water, and it was revealed by quantitative RT-PCR that the ghrelin mRNA expression level in the stomach was up-regulated. Rikkunshito increased the plasma acylated ghrelin level in healthy volunteers and normal mice. Additionally, this change was maintained for at least 4 weeks after the end of administration. In mice, Rikkunshito increased the ghrelin mRNA expression level in the stomach.

Ontogeny of ghrelin mRNA expression and identification of ghrelin-immunopositive cells in the gastrointestinal tract of the Peking duck, Anas platyrhychos

Ghrelin is an acylated peptide and an endogenous ligand for the growth hormone secretagogue receptor (GHS-R), and stimulates growth hormone release and food intake in mammals. Peking duck is a very fast growing species of poultry. Although the sequence and structure of ghrelin have recently been determined, the expression of ghrelin in Peking duck has not been studied. Here, we investigated the tissue expression and distribution of ghrelin by RT-PCR and immunohistochemistry, respectively, in Peking duck at different stages of development. Ghrelin mRNA expression was mainly detected in the proventriculus and proventriculus–gizzard junction. It was first expressed, but weakly, on embryonic day 14 (E14); the expression increased by embryonic day 21 (E21), and was maintained at high levels between post-hatching-day 1 (P1) and post-hatching-day 60 (P60). Weak expression of ghrelin mRNA was also found in the gizzard and duodenum. In the gastrointestinal tract of growing Peking duck in P60, the largest number of ghrelin-ip cells was detected in the epithelium of the compound tubular glands in the proventriculus and the next largest number was in the proventriculus–gizzard junction. Very few ghrelin-ip cells were located in the epithelium of the simple tubular glands adjacent to the gizzard. No ghrelin-ip cells were observed elsewhere in the gastrointestinal tract. Ghrelin-ip cells were found in embryos as early as day E21; at the same time, the compound tubular glands in the proventriculus had formed. The numbers of ghrelin-ip cells on P1 were similar to those of E21 embryos. However, on P60, high numbers of strongly stained ghrelin-ip cells were found to be scattered in the epithelium of the compound tubular glands in the proventriculus. The density of ghrelin-ip cells (cells/mm 2) in the proventriculus on P60 was significantly greater than those of P1 and E21 embryos. These results demonstrate that ghrelin is expressed in the Peking duck gastrointestinal tract, especially in the proventriculus, from mid–late-stage embryos to growing period and suggested an involvement of ghrelin in the development and biology of the gastrointestinal tract of the Peking duck.