Ghrelin O-acyltransferase mRNA Research Articles

Characterization of Ghrelin O-acyltransferase mRNA in channel catfish

Food intake is regulated by the reaction between hypothalamic neurotransmitters and peripheral signals. Ghrelin, known as a hunger hormone, stimulates food intake. Ghrelin O-acyltransferase (GOAT) catalyzes the acylation of medium-chain fatty acids to ghrelin to modify ghrelin activity. GOAT is found in all vertebrates, including fish, and is critical for modifying the activity of ghrelin. We hypothesized that GOAT is critical for regulating food intake because it modifies the activity of ghrelin in channel catfish. However, very little is known about GOAT in channel catfish, including the tissue distribution of its mRNA. We identified the nucleotide sequence that corresponds to the GOAT mRNA in GenBank. The sequencing alignment analysis showed that the sequence similarity between channel catfish GOAT mRNA and those of other fish species was high (>65%) in the region that corresponded to the open reading frame of the channel catfish GOAT mRNA. Examination of the tissue distribution of GOAT mRNA, including the brain, liver, muscle, kidneys, spleen, heart, and small and large intestines, revealed that GOAT mRNA expression was detectable in the brain, small intestine, and muscles. We are currently verifying the results of tissue distribution analysis using quantitative real-time PCR. We are also examining the relationship between changes in food intake and the expression of GOAT mRNA in the brain and muscle using real-time PCR. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The suppressive effect of REVERBs on ghrelin and GOAT transcription in gastric ghrelin-producing cells

Ghrelin is a multifunctional gut peptide with a unique structure, which is modified by a medium chain fatty acid at the third serine by ghrelin O-acyl transferase (GOAT). It is well known that the major source of plasma ghrelin is the stomach, but the transcriptional regulation of gastric ghrelin and GOAT is incompletely understood. Here, we studied the involvement of the nuclear receptors REV-ERBα and REV-ERBβ on ghrelin and GOAT gene expression in vivo and in vitro. Reverse-transcriptase polymerase chain reaction analysis showed that REV-ERBα and REV-ERBβ mRNAs were expressed in the stomach and a stomach-derived ghrelin cell line (SG-1 cells). In vivo experiments with mice revealed the circadian rhythm of ghrelin, GOAT, and REV-ERBs. The peak expression of ghrelin and GOAT mRNAs occurred at Zeitgeber time (ZT) 4, whereas that of REV-ERBα and REV-ERBβ was observed at ZT8 and ZT12, respectively. Treatment of SG-1 cells with SR9009, a REV-ERB agonist, led to a significant reduction in ghrelin and GOAT mRNA levels. Overexpression of REV-ERBα and REV-ERBβ decreased ghrelin and GOAT mRNA levels in SG-1 cells. In contrast, small-interfering RNA (siRNA)-mediated double-knockdown of REV-ERBα and REV-ERBβ in SG-1 cells led to the upregulation in the expression of ghrelin and GOAT mRNAs. These results suggest that REV-ERBs suppress ghrelin and GOAT mRNA expression.

Gastric ghrelin cells in obese patients are hyperactive.

Distribution and activity of ghrelin cells in the stomach of obese subjects are controversial. We examined samples from stomachs removed by sleeve gastrectomy in 49 obese subjects (normoglycemic, hyperglycemic and diabetic) and quantified the density of ghrelin/chromogranin endocrine cells by immunohistochemistry. Data were compared with those from 13 lean subjects evaluated by gastroscopy. In 44 cases (11 controls and 33 obese patients) a gene expression analysis of ghrelin and its activating enzyme ghrelin O-acyl transferase (GOAT) was performed. In 21 cases (4 controls and 17 obese patients) the protein levels of unacylated and acylated-ghrelin were measured by ELISA tests. In 18 cases (4 controls and 14 obese patients) the morphology of ghrelin-producing cells was evaluated by electron microscopy. The obese group, either considered as total population or divided into subgroups, did not show any significant difference in ghrelin cell density when compared with control subjects. Inter-glandular smooth muscle fibres were increased in obese patients. In line with a positive trend of the desacylated form found by ELISA, Ghrelin and GOAT mRNA expression in obese patients was significantly increased. The unique ghrelin cell ultrastructure was maintained in all obese groups. In the hyperglycemic obese patients, the higher ghrelin expression matched with ultrastructural signs of endocrine hyperactivity, including expanded rough endoplasmic reticulum and reduced density, size and electron-density of endocrine granules. A positive correlation between ghrelin gene expression and glycemic values, body mass index and GOAT was also found. All obese patients with type 2 diabetes recovered from diabetes at follow-up after 5 months with a 16.5% of weight loss. Given the known inhibitory role on insulin secretion of ghrelin, these results suggest a possible role for gastric ghrelin overproduction in the complex architecture that takes part in the pathogenesis of type 2 diabetes.

Metabolic stress leads to divergent changes in the ghrelinergic system in goldfish (Carassius auratus) gonads

Various endocrine factors that regulate energy homeostasis are also implicated in the reproductive physiology of mammals. However, the hormonal link between metabolism and reproduction in fish is poorly understood. Ghrelin is a multifunctional hormone with both metabolic and reproductive roles in vertebrates. Post-translational acylation by ghrelin-O-acyltransferase (GOAT) is critical for its biological actions. The expression of ghrelin, ghrelin or growth hormone secretagogue receptor (GHSR), and GOAT (which forms the ghrelinergic system) in fish under metabolic stress remains unclear. In this research, we used RT-qPCR and Western blot analysis to determine the expression of the ghrelinergic system in goldfish (during the reproductively active phase) hypothalamus and gonads under 7 and 28 days of fasting. We found a significant increase in preproghrelin mRNA expresson in the ovary, and GOAT mRNA expression in the testis of goldfish deprived of food for 7 days. In fish deprived of food for 28 days, preproghrelin, GHSR and GOAT mRNA expression was significantly increased in the hypothalamus of male goldfish. Such differences were not observed in the hypothalamus of female fish, and in the testis of 28 days fasted fish. Meanwhile, preproghrelin, GHSR, and GOAT expression (both mRNA and protein) was significantly increased in the ovary of female fish fasted for 28 days. Ghrelin has been shown to suppress oocyte maturation in fish. The upregulation of a system that has ovarian inbititory roles suggests a role for ghrelin in maintaining reduced reproductive capability during metabolically challenging periods.

Ghrelin acylation by ghrelin- O-acyltransferase can occur in healthy part of oncological liver in humans.

Activation of ghrelin is controlled by the enzyme ghrelin- O-acyl transferase (GOAT). In humans, localization of this acylation is poorly understood. The aim of this study is to explore GOAT localization and activation in the human liver by evaluating both bioactive and non-bioactive ghrelin in the bloodstream entering and leaving the liver and to simultaneously evaluate GOAT mRNA expression in the liver. A healthy part of oncologic hepatic tissue collected from nine patients undergoing hepatectomy was used to evaluate GOAT mRNA expression by quantitative real-time polymerase chain reaction (RT-qPCR). Simultaneously, blood from the portal vein, the suprahepatic vein, the subclavicular vein, and the radial artery was also sampled to assay total and acylated ghrelin. Acylated ghrelin level was significantly increased in the suprahepatic vein compared with the portal vein level (385 ± 42 ng/ml vs. 268 ± 24 ng/ml, P = 0.04). Suprahepatic-to-portal vein ratio for acylated ghrelin (acylation ratio) is 1.4 ± 0.1. Mean expression of GOAT mRNA in the liver, expressed as 2-∆Ct·µg total RNA-1·1 µl of liver tissue-1 was at 0.042 ± 0.021 arbitrary units. GOAT mRNA expression in the liver was correlated with acylated-to-total ghrelin ratio in the suprahepatic vein ( P = 0.016, R = 0.75) and with the acylation liver ratio ( P = 0.05, R = 0.61). Blood concentration of acylated ghrelin was found significantly increased after its passage through the liver, suggesting that acylation can occur in the liver. RT-qPCR data confirmed the presence of GOAT in the liver, with a positive correlation between GOAT expression and acylated ghrelin liver ratio. This study strongly suggests that the liver is a site of ghrelin acylation in humans. NEW & NOTEWORTHY Although the activation of ghrelin by the enzyme ghrelin- O-acyl transferase (GOAT) is yet well demonstrated, its localization, especially in humans, remains poorly understood. We explored GOAT localization and activation in the human liver by simultaneously evaluating both bioactive and non-bioactive ghrelin in the bloodstream entering and leaving the liver and also GOAT mRNA expression in the liver. We therefore showed for the first time, to our knowledge, that GOAT localized in the liver is active and takes part in ghrelin activation.

The effect of ghrelin O-acyltransferase inhibitor on gastric H+–K+-ATPase activity and GOAT/ghrelin system in gastric mucosal cells in vitro

Ghrelin is implicated in the regulation of gastric functional development. The octanoylation of ghrelin is critical for its physiological functions which dependent upon ghrelin O-acyltransferase (GOAT) catalyzation. To investigate the effect of GOAT on gastric acid secretion and expression of ghrelin in vitro. Primary cultures of gastric mucosal cells were challenged with 1.5 × 10−5, 1.5 × 10−4 and 1.5 × 10−3 mol/mL GO-CoA-Tat (The GOAT inhibitor), respectively, for 24 h in order to further clarify the effect of GOAT on H+–K+-ATPase activity. In vitro, GO-CoA-Tat significantly increased ghrelin and GOAT mRNA expression at 1.5 × 10−5, 1.5 × 10−4 and 1.5 × 10−3 mol/mL, and augmented cell total ghrelin secretion at 1.5 × 10−3 mol/mL. But cell acylated ghrelin secretion was reduced at 1.5 × 10−3 mol/mL GO-CoA-Tat (P < 0.05). And cell acylated ghrelin synthesis was reduced at 1.5 × 10−4 and 1.5 × 10−3 mol/mL GO-CoA-Tat (P < 0.05). In accordance with acylated ghrelin level, H+–K+-ATPase activity were decreased with 1.5 × 10−4 and 1.5 × 10−3 mol/mL GO-CoA-Tat (P < 0.05). These results indicated that GOAT inhibitor decreases the acylated ghrelin level and H+–K+-ATPase activity in vitro.

Ghrelin, ghrelin-O-acyl transferase, nucleobindin-2/nesfatin-1 and prohormone convertases in the pancreatic islets of Sprague Dawley rats during development

Ghrelin and nesfatin-1 are regulators of blood glucose and energy balance. Prohormone convertases (PCs) enable processing of ghrelin and nesfatin-1 from its precursors. An acylation, enabled by ghrelin O-acyl transferase (GOAT) is critical for many of the biological actions of ghrelin. To date, there is no research on the developmental expression of GOAT, and the co-expression of both NUCB2/nesfatin-1 and prohormone convertases in the pancreas. The objective of this research was to immunolocalize ghrelin, GOAT, NUCB2/nesfatin-1, PC1/3 and PC2 in the pancreas during fetal and postnatal periods of Sprague Dawley (SD) rats using immunohistochemical analysis. GOAT mRNA in the rat pancreas during development was also determined. In the pancreas, not all islet cells immunopositive for GOAT are immunoreactive for ghrelin on postnatal (P) days 20, 27 and adult. GOAT mRNA expression in the pancreas at P27 was higher than the expression levels in rest of the developmental stages tested. Both PC1/3 and PC2 are co-expressed with NUCB2/nesfatin-1 on embryonic (E) day E21, P13, P20. While similar co-localization was also found in P27 for NUCB2/nesfatin-1 and PC1/3, NUCB2/nesfatin-1 and PC2 were found in distinct populations of cells in P27. Some ghrelin and GOAT positive cells stained for nesfatin-1 as well. Meanwhile, no co-localization of somatostatin and glucaon with nesfatin-1 was found in the pancreas of SD rats. Our findings suggest that the endocrine pancreas can produce and process precursors of ghrelin and nesfatin-1 to make bioactive forms of both peptides.

RNAi-mediated Ghrelin affects gastric H+–K+-ATPase activity and expression of GOAT–Ghrelin system in vitro

Ghrelin has been implicated in the regulation of gastric functional development, and its physiological functions are mediated by Ghrelin-O-acyltransferase (GOAT) which is capable of generating the active form of this polypeptide hormone. However, whether and how ghrelin gene silencing may modify gastric acid secretion and GOAT–Ghrelin system is yet to be explored. The study was performed in gastric mucosal cells from weanling piglets in vitro. We evaluated the effect of ghrelin on gastric acid secretion, gene expression of GOAT and ghrelin as well as ghrelin levels by RNA interference assay. shGhrelin triggered the down-regulation of ghrelin mRNA expression (P<0.05) via an RNAi mechanism, as observed by real-time RT-PCR. In addition, shGhrelin showed reduced total ghrelin production and secretion (P<0.05) using ELISA in vitro. We also detected that GOAT mRNA expression was reduced in shGhrelin group (P<0.05), compared with control groups. In accordance with the GOAT expression, acylated ghrelin production and secretion were reduced in gastric mucosal cells and culture medium (P<0.05). Silencing of ghrelin gene achieved by RNAi-mediation inhibited the activity of H+–K+-ATPase and pepsin (P<0.05) in gastric mucosal cells. These results indicated that RNAi of Ghrelin gene inhibited the gastric acid secretion with decreased GOAT mRNA and acylated Ghrelin in gastric mucosal cells.

Altered intestinal neuroendocrine gene expression in humans with obesity.

Gastrointestinal hormones are critically involved in the regulation of food intake and body weight. Previous studies support an interplay between gastrointestinal hormones and the serotonergic system. This study explored intestinal neuroendocrine expression patterns in humans with obesity versus nonobese humans. Jejunum samples were collected from 164 humans with obesity (120 women; BMI (mean ± SD): 43.5 ± 6.6 kg/m(2) ) while they underwent Roux-en-Y gastric bypass surgery and from 18 nonobese humans (7 women; BMI: 23.5 ± 3.0 kg/m(2) ) undergoing distinct intestinal surgeries. mRNA expression of cholecystokinin (CCK), peptide YY3-36 (PYY), nesfatin1, ghrelin, ghrelin O-acyltransferase (GOAT), leptin, leptin receptor (leptinR), glucagon-like-peptide 1 receptor (GLP1R), serotonin transporter (SERT), tryptophan hydroxylase 1 (TPH1), and serotonin receptor 3A (5HT3A R) was determined with qRT-PCR. Ghrelin and GOAT protein expression was quantified using immunohistological stainings. Statistical analyses were performed with SPSS. Jejunum samples from humans with obesity showed a higher expression of GOAT (mRNA and protein), TPH1, and SERT mRNA compared with the nonobese humans (all P < 0.05). Positive correlations were observed between TPH1, CCK, PYY, and nesfatin1 in nonobese and GOAT, ghrelin, TPH1, SERT, CCK, and PYY in humans with obesity (all P < 0.01). Our top-down approach substantiates the dysregulation of jejunal neuroendocrine hormones in obesity.

Ghrelin O-Acyl Transferase in Zebrafish Is an Evolutionarily Conserved Peptide Upregulated During Calorie Restriction.

Ghrelin is a multifunctional orexigenic hormone with a unique acyl modification enabled by ghrelin O-acyl transferase (GOAT). Ghrelin is well-characterized in nonmammals, and GOAT sequences of several fishes are available in the GenBank. However, endogenous GOAT in non-mammals remains poorly understood. In this research, GOAT sequence comparison, tissue-specific GOAT expression, and its regulation by nutrient status and exogenous ghrelin were studied. It was found that the bioactive core of zebrafish GOAT amino acid sequence share high identity with that of mammals. GOAT mRNA was most abundant in the gut. GOAT-like immunoreactivity (i.r.) was found colocalized with ghrelin in the gastric mucosa. Food deprivation increased, and feeding decreased GOAT and preproghrelin mRNA expression in the brain and gut. GOAT and ghrelin peptides in the gut and brain showed corresponding decrease in food-deprived state. Intraperitoneal injection of acylated fish ghrelin caused a significant decrease in GOAT mRNA expression, suggesting a feedback mechanism regulating its abundance. Together, these results provide the first in-depth characterization of GOAT in a non-mammal. Our results demonstrate that endogenous GOAT expression is responsive to metabolic status and availability of acylated ghrelin, providing further evidences for GOAT in the regulation of feeding in teleosts.

Knockdown of central ghrelin O-acyltransferase by vivo-morpholino reduces body mass of rats fed a high-fat diet

The enzyme ghrelin O-acyltransferase (GOAT) activates the orexigenic peptide ghrelin by transferring an acyl group from fatty acids to the serine-3 residue of the ghrelin molecule. This allows ghrelin to bind to its only known receptor, the growth hormone secretagogue receptor type 1a (GHSR1a). While studies have examined the hypothalamic transcriptional response of GOAT to metabolic challenge in mice, little has been examined in the rat hypothalamus. Furthermore, it has not been possible to identify the role of central GOAT separate from that of the periphery, since previous studies either knocked out GOAT system-wide or administered a GOAT inhibitor intraperitoneally. To determine if central GOAT expression is modulated by changes in energy state, we subjected rats to either forty-eight hours of food deprivation or three weeks of food restriction and found that GOAT mRNA increases significantly in both the hypothalamus and the stomach fundus in response to both metabolic challenges. We also found increases in hypothalamic ghrelin mRNA and stomach GHSR1a mRNA in response to food deprivation, as well as increases in hypothalamic GHSR1a mRNA in response to food restriction. We then conducted a second study where we continuously infused amorpholino antisense oligonucleotide into the lateral ventricles of rats to knock-down GOAT centrally while the animals were exposed to a high fat diet. Our results show that rats receiving the GOAT antisense gained less weight, and decreased their caloric efficiency when eating a high fat diet compared to control animals. These data suggest that central GOAT plays a role in modulating metabolism in rats.

Acylated ghrelin is not required for the surge in pituitary growth hormone observed in pregnant mice

Objective: Ghrelin is produced by the stomach, hypothalamus and pituitary. It circulates as acylated ghrelin (AG, which stimulates growth hormone (GH) secretion) and unacylated ghrelin (UAG). Acylation is mediated by the enzyme ghrelin O-acyltransferase (GOAT). In mice, pregnancy is associated with a marked increase in circulating pituitary GH. We investigated the role of AG and UAG in the surge of plasma GH concentrations in pregnant mice at the end of pregnancy. Design: Using a mouse model generated on a C57BL/6 background (wild type, WT) in which the GOAT gene has been deleted (KO), we measured plasma AG, UAG and GH concentrations and tissue (stomach, pituitary and hypothalamus) preproghrelin and GOAT mRNA in non-pregnant (NP) and pregnant (P), WT and KO mice. Results: GOAT deletion was associated with undetectable concentrations of AG. UAG concentrations were similar in all groups. In both WT and KO animals, mean GH concentrations increased 30 to 50 times during pregnancy. There was a tendency toward lower median GH concentrations in KO (301ng/mL) compared to WT (428ng/mL) mice (p=0.059). Preproghrelin expression was not affected by GOAT deletion or by pregnancy in the stomach. In contrast, pituitary and hypothalamic ghrelin gene expression were lower in KO-NP and KO-P mice compared to their WT counterparts. Conclusion: The complete absence of ghrelin acylation, which is associated with undetectable AG concentrations, does not prevent the marked increase in pituitary GH concentrations observed in pregnant mice, suggesting that AG is not the major mediator of GH secretion during pregnancy.

Tissue distribution and developmental changes of ghrelin and GOAT expression in broiler chickens during embryogenesis

Ghrelin plays important roles, such as regulating growth hormone release and energy metabolism, but little is known about its developmental changes in the proventriculi of chicken embryos. This study was designed to elucidate the distributions and developmental changes of ghrelin and ghrelin-O-acyltransferase (GOAT) expression in broiler embryos using qRT-PCR and immunohistochemistry. Our results demonstrated the following: (1) on E18, ghrelin and GOAT are ubiquitously expressed in every tissue examined. The expression level of ghrelin mRNA was the highest in the proventriculus, reaching a level that was 50-fold higher than that in the hypothalamus, while GOAT mRNA expression was low in the proventriculus and it was only 67.6% as high as that of hypothalamus; (2) ghrelin and GOAT mRNA expression were detected in the proventriculus on E9, but only at 1.9% and 1.7% of the level expressed on E18, respectively, and their expression levels increased rapidly from E18 to E21. There was similar developmental pattern in the ghrelin and GOAT mRNA expression; and (3) ghrelin-immunopositive cells were first detected in the proventriculus on E15, were located only in the compound tubular glands of the proventriculus, and were of the closed-cell type. The density of ghrelin-immunopositive cells increased significantly from E15 to E21. These results suggest that ghrelin may be an important regulating factor that plays a vital role during the development of chicken embryos.

Resveratrol Prevents Hypertension and Cardiac Hypertrophy in Hypertensive Rodents

s / Can J Diabetes 37 (2013) S13eS84 S23 and unacylated ghrelin (UAG). Acylation is mediated by ghrelin Oacyltransferase (GOAT). Maintenance of euglycemia during calorie restriction (CR) in pregnancy is key for optimal outcome. Hypothesis: Upregulation of GOAT/ghrelin, increased AG and GH concentrations are part of the hormonal response to hypoglycemia during CR. Methods: Wild-type (WT) and GOAT-KO (KO), pregnant (P) and non-pregnant (NP) mice were freely fed (FF) or submitted to 50% CR for 7 days. In P mice, CR occurred between days 11 to 18 (1⁄4 sacrifice) of pregnancy. Blood glucose (BG), plasma AG, UAG and GH were measured by EIA and GOAT/ghrelin expression by RT-PCR. Results (Table 1): 1. In WT mice, CR did not affect BG in NP mice but caused lower BG in P mice. This was associated with higher AG and GH (from placenta) concentrations and lower stomach GOAT and ghrelin mRNA. 2. In KO mice, where plasma AG is by definition absent, CR caused severe hypoglycemia (causing systematic P termination) in P mice and lower BG in NP mice. The latter was associated with higher UAG and lower stomach ghrelin mRNA. Conclusion: 1. GOAT-KO mice are more prone to hypoglycemia, suggesting a role for AG that is independent from GH secretion; 2. The AG increase during CR not due to increased stomach production but likely decreased degradation. 3. AG is not necessary for GH production (by the placenta) in P mice.

Ghrelin O-acyltransferase (GOAT) is expressed in prostate cancer tissues and cell lines and expression is differentially regulated in vitroby ghrelin

BackgroundGhrelin is a 28 amino acid peptide hormone that is expressed in the stomach and a range of peripheral tissues, where it frequently acts as an autocrine/paracrine growth factor. Ghrelin is modified by a unique acylation required for it to activate its cognate receptor, the growth hormone secretagogue receptor (GHSR), which mediates many of the actions of ghrelin. Recently, the enzyme responsible for adding the fatty acid residue (octanoyl/acyl group) to the third amino acid of ghrelin, GOAT (ghrelin O-acyltransferase), was identified.MethodsWe used cell culture, quantitative real-time reverse transcription (RT)-PCR and immunohistochemistry to demonstrate the expression of GOAT in prostate cancer cell lines and tissues from patients. Real-time RT-PCR was used to demonstrate the expression of prohormone convertase (PC)1/3, PC2 and furin in prostate cancer cell lines. Prostate-derived cell lines were treated with ghrelin and desacyl ghrelin and the effect on GOAT expression was measured using quantitative RT-PCR.ResultsWe have demonstrated that GOAT mRNA and protein are expressed in the normal prostate and human prostate cancer tissue samples. The RWPE-1 and RWPE-2 normal prostate-derived cell lines and the LNCaP, DU145, and PC3 prostate cancer cell lines express GOAT and at least one other enzyme that is necessary to produce mature, acylated ghrelin from proghrelin (PC1/3, PC2 or furin). Finally, ghrelin, but not desacyl ghrelin (unacylated ghrelin), can directly regulate the expression of GOAT in the RWPE-1 normal prostate derived cell line and the PC3 prostate cancer cell line. Ghrelin treatment (100nM) for 6 hours significantly decreased GOAT mRNA expression two-fold (P < 0.05) in the PC3 prostate cancer cell line, however, ghrelin did not regulate GOAT expression in the DU145 and LNCaP prostate cancer cell lines.ConclusionsThis study demonstrates that GOAT is expressed in prostate cancer specimens and cell lines. Ghrelin regulates GOAT expression, however, this is likely to be cell-type specific. The expression of GOAT in prostate cancer supports the hypothesis that the ghrelin axis has autocrine/paracrine roles. We propose that the RWPE-1 prostate cell line and the PC3 prostate cancer cell line may be useful for investigating GOAT regulation and function.

Plasma ghrelin isoforms and gastric ghrelin O-acyltransferase expression are influenced by Helicobacter pylori status

ObjectiveHelicobacter pylori is known to affect the host’s nutritional status. This study was performed to elucidate the relationship between H. pylori status and the dynamics of the ghrelin system, in the context of ghrelin O-acyltransferase (GOAT) expression. MethodsWe conducted a clinical study of 30 subjects focusing on the following points: 1) the effects of H. pylori infection on the concentrations of circulating ghrelin isoforms and on ghrelin and GOAT mRNA expression in the gastric mucosa, and 2) the effects of H. pylori eradication on the same parameters. ResultsThe plasma acyl-ghrelin and desacyl-ghrelin concentrations of 16 H. pylori positive participants were significantly lower than those of 14 H. pylori negative controls. The acyl-ghrelin/desacyl-ghrelin ratio was not significantly different between the H. pylori positive and H. pylori negative participants. The levels of ghrelin and GOAT mRNA in the gastric mucosa were significantly lower in the H. pylori positive participants than in the H. pylori negative controls. In 11 subjects in whom H. pylori eradication was successful, their plasma acyl-ghrelin levels tended to increase after H. pylori eradication, but the difference was not significant; however, their plasma desacyl-ghrelin levels were significantly reduced. Although gastric ghrelin mRNA expression increased significantly after H. pylori eradication, gastric GOAT mRNA expression tended to increase but was not significantly altered. ConclusionH. pylori status might affect the host’s nutritional status through changes in the plasma levels of ghrelin isoforms and the gastric expression levels of ghrelin and GOAT mRNA.

The expression of ghrelin O-acyltransferase (GOAT) in human tissues

Ghrelin is a circulating growth hormone-releasing and appetite-inducing brain-gut peptide. It needs to be acylated on its serine-3 with octanoate for its endocrine actions. The acyl-transferase that catalyses ghrelin octanoylation has recently been identified and named as GOAT (ghrelin O-acyltransferase); GOAT enzyme is coded by the MBOAT4 gene. This study aimed to investigate GOAT expression in the human. The distribution of GOAT mRNA expression was studied in various human tissues using classical and real-time reverse transcription and polymerase chain reaction. GOAT expression was found in all tissues studied (stomach, adrenal cortex, breast, right and left colon, duodenum, jejunum, ileum, fat, Fallopian tube, gallbladder, lymph node, lymphocyte cell line, kidney, liver, lung, muscle, myocardium, pituitary, oesophagus, pancreas, ovary, placenta, prostate, testis, spleen and thyroid). The widespread expression of GOAT corresponds to the widespread distribution of ghrelin expression. GOAT expression was high in stomach and gut, the major ghrelin-secreting tissues, and in the pituitary, in which ghrelin is known to show autocrine and paracrine effects. Identification of GOAT expression in various tissues support the concept that in addition to the important endocrine effect of acylated ghrelin, the paracrine effects of locally synthetised and acylated ghrelin may be important.

Lipopolysaccharide differentially decreases plasma acyl and desacyl ghrelin levels in rats: Potential role of the circulating ghrelin-acylating enzyme GOAT

Bacterial lipopolysaccharide (LPS) in rodents is an established model for studying innate immune responses to gram-negative bacteria and mimicking symptoms of infections including reduced food intake associated with decreased circulating total ghrelin levels. The ghrelin-acylating enzyme, ghrelin- O-acyltransferase (GOAT) involved in the formation of acyl ghrelin (AG) was recently identified. We investigated changes in circulating AG, desacyl ghrelin (DG) and GOAT induced by intraperitoneal LPS (100 μg/kg) and associated changes in food intake. Plasma AG and total ghrelin were assessed by radioimmunoassay, GOAT protein by Western blot and mRNA by RT-qPCR. DG was derived from total minus AG. Plasma AG and DG were decreased at 2, 5 and 7 h ( p < 0.01) post-injection compared to vehicle and recovered at 24 h. At 2 h there was a significantly greater decrease of AG (−53%) than DG (−28%) resulting in a decreased AG/DG ratio (1:5, p < 0.01), which thereafter returned to pre-injection values (1:3). This altered ratio was associated with a 38% decrease in plasma GOAT protein compared to vehicle ( p < 0.001), whereas gastric GOAT protein was slightly increased by 10% ( p < 0.05). GOAT mRNA expression was unchanged. Food intake was reduced by 58% measured during the 1.5–2 h period post-LPS injection. Decreased plasma AG and DG preceded the rise in rectal temperature and blood glucose that peaked at 7 h. These data indicate that LPS induces a long-lasting reduction of AG and DG levels that may have a bearing with the decrease in food intake. The faster drop in AG than DG within 2 h is associated with reduced circulating GOAT.

Age, sex, and lactating status regulate ghrelin secretion and GOAT mRNA levels from isolated rat stomach

Ghrelin is a stomach derivate peptide involved in energy homeostasis regulation, and ghrelin O-acyltransferase (GOAT) is the enzyme responsible for ghrelin acylation. Puberty is a period characterized by profound changes in the metabolic requirements and notable variations of sexual hormone levels. On the other hand, the weaning process is a fundamental modification of the diet, which implicates several adaptations of the gastrointestinal tract physiology. Until now the direct secretion of ghrelin by the stomach in these conditions, without interferences from other organs, has never been studied. The main objective of this article was to investigate how the stomach modulates ghrelin production and secretion as well as GOAT expression on these periods of life. Gastric ghrelin secretion is regulated through postnatal life in an independent way of gastric expression and circulating levels of this hormone. The present work shows a strong regulation of gastric ghrelin secretion by estrogens. The weaning strongly regulates gastric ghrelin secretion. Animals subjected to delayed weaning present a lower body weight than the corresponding controls. For the first time, it is shown that a noticeable decrease in circulating levels of testosterone and estrogens is associated with delay of weaning. GOAT mRNA levels in the stomach are strongly regulated by age, breastfeeding, and testosterone. In conclusion, the stomach itself regulates ghrelin and GOAT production to adapt the organism to the metabolic requirements demanded through each stage of life.

Long-term dietary restriction influences plasma ghrelin and GOAT mRNA level in rats

The objective of this study was to examine the effect of chronic dietary restriction on the physical characteristics of the intestine and gut-derived satiety hormone production. Male Wistar rats (8 weeks) were randomized to ad libitum (AL) or 35% dietary restriction (DR) for 5 months. At the end of the study, physical measurements were made on the intestine and satiety hormone secretion and mRNA expression determined. A comparison group of young, growing AL rats (5 weeks) was also examined. The adult DR rats gained less weight over 5 months and had lower fat mass than adult AL rats ( p < 0.05). The weight of the small intestine as a percentage of total body weight was greater in adult DR compared to adult AL but lower than young AL rats. Compared to AL, DR down-regulated proglucagon and cholecystokinin mRNA in the duodenum and ghrelin mRNA in the stomach of adult rats but was not different from young AL. Ghrelin-O-acyltransferase (GOAT) mRNA in the stomach was up-regulated 21-fold in adult AL rats compared to young AL and 14-fold compared to adult DR rats. Total and des-acyl ghrelin was approximately 50% higher in adult DR and young AL rats compared to adult AL. Plasma leptin and insulin were lower in adult DR and young AL rats compared to adult AL. Our findings suggest that long-term energy deficits continue to drive up ghrelin levels which may have profound implications for practical implementation of DR as an anti-aging or anti-obesity strategy in humans.