Ella W. Englander

Ghrelin—not just another stomach hormone

Growth hormone (GH) secretagogues (GHSs) are non-natural, synthetic substances that stimulate GH secretion via a G-protein-coupled receptor called the GHS-receptor (GHS-R). The natural ligand for the GHS-R has been identified recently; it is called ghrelin. Ghrelin and its receptor show a widespread distribution in the body; the greatest expression of ghrelin is in stomach endocrine cells. Administration of exogenous ghrelin has been shown to stimulate pituitary GH secretion, appetite, body growth and fat deposition. Ghrelin was probably designed to be a major anabolic hormone. Ghrelin also exerts several other activities in the stomach. The findings that ghrelin is produced in mucosal endocrine cells of the stomach and intestine, and that ghrelin is measurable in the general circulation indicate its hormonal nature. A maximal expression of ghrelin in the stomach suggests that there is a gastrointestinal hypothalamic-pituitary axis that influences GH secretion, body growth and appetite that is responsive to nutritional and caloric intakes.

Evidence That Growth Hormone Exerts a Feedback Effect on Stomach Ghrelin Production and Secretion

Ghrelin is a recently discovered stomach hormone that stimulates pituitary growth hormone (GH) secretion potently. The purpose of these experiments was to test the hypothesis that a stomach-ghrelin-pituitary-GH axis exists in which either an elevation or reduction in systemic GH levels will exert a negative or positive feedback action, respectively, on stomach ghrelin homeostasis. In rats, GH administration decreased stomach ghrelin mRNA levels and plasma ghrelin levels significantly. In GH-releasing hormone (GHRH) transgenic mice, GHRH overexpression decreased stomach ghrelin peptide levels when compared with control mice. In aged rats (25 months) stomach ghrelin mRNA and peptide levels and plasma ghrelin levels were decreased when compared with young rats (5 months). Because GH secretion is reduced in aged rats, the elevated stomach ghrelin production and secretion may reflect a decreased GH feedback on stomach ghrelin, homeostasis, and secretion. Together, these findings suggest that endogenous pituitary GH exerts a feedback action on stomach ghrelin homeostasis and support the hypothesis that a stomach-ghrelinpituitary GH axis exists.

Nutrient inhibition of ghrelin secretion in the fasted rat

Ghrelin is a recently discovered stomach hormone whose secretion increases with fasting; the fasting-induced elevation is inhibited by refeeding. The aim of this study was to determine whether all nutrient types (i.e., carbohydrates, proteins, fats) and soybean trypsin inhibitor (SBTI), a secretagogue for intestinal cholecystokinin (CCK), given individually into the stomach or intravenously can inhibit ghrelin secretion in the fasted rat. Intragastric (i.g.) administration of intact protein, a protein digest, SBTI, dextrose, or fat decreased plasma ghrelin levels significantly (p<0.05). All nutrients inhibited ghrelin secretion equally. Fat and dextrose given intravenously (i.v.) also reduced ghrelin secretion. These data demonstrate that nutrients can inhibit ghrelin secretion by both the luminal and systemic routes. Additionally, the findings show that all nutrient types given orally are capable of inhibiting ghrelin secretion, and suggest that intestinal CCK may participate in the inhibition of ghrelin secretion following oral intake of nutrients.

Apelin cells in the rat stomach.

Apelin is a recently discovered peptide that is the endogenous ligand for the APJ receptor. Apelin is produced in the central nervous system, heart, lung, mammary gland and gastrointestinal (GI) tract. The aim of this study was to identify by immunohistochemistry (IHC) cell types in the rat stomach that produce apelin peptide. IHC revealed abundant apelin-positive cells, primarily in the neck and upper base regions of the gastric glands in the mucosal epithelium. Apelin is not detected in the muscle layer. Apelin-positive cells were identified as mucous neck, parietal cells, and chief cells. Apelin is also identified in gastric epithelial cells that produce chromogranin A (CGA), a marker of enteroendocrine cells. The findings that apelin is expressed in gastric exocrine and endocrine cells agrees with and extends other data showing that apelin peptide is measurable in the gut lumen and in the systemic circulation by immunoassay.

Rat MYH, a glycosylase for repair of oxidatively damaged DNA, has brain‐specific isoforms that localize to neuronal mitochondria

Mitochondrial genomes are exposed to a heavy load of reactive oxygen species (ROS) that damage DNA. Since in neurons, mitochondrial DNA integrity must be maintained over the entire mammalian life span, neuronal mitochondria most likely repair oxidatively damaged DNA. We show that the Escherichia coli MutY DNA glycosylase homolog (MYH) in rat (rMYH) involved in repair of oxidative damage is abundantly expressed in the rat brain, with isoforms that are exclusive to brain tissue. Confocal microscopy and western analyses reveal localization of rMYH in neuronal mitochondria. To assess involvement of MYH in the neuronal response to oxidative DNA damage, we used a rat model of respiratory hypoxia, in which acutely reduced blood oxygenation leads to generation of superoxide, and formation and subsequent removal of 8‐hydroxy‐2′‐deoxyguanosine (8OHdG). Removal of 8OHdG is accompanied by a spatial increase in rMYH immunoreactivity in the brain and an increase in levels of one of the three mitochondrial MYH isoforms, suggesting that inducible and non‐inducible MYH isoforms exist in the brain. The mitochondrial localization of oxidative DNA damage repair enzymes in neurons may represent a specialized neuronal mechanism that safeguards mitochondrial genomes in the face of routine and accidental exposures to heavy loads of injurious ROS.

Acute Pancreatitis Signals Activation of Apoptosis-Associated and Survival Genes in Mice

In experimental models of acute pancreatitis (AP), acinar cell death occurs by both necrosis and programmed cell death or apoptosis. Apoptosis is an active form of cell death associated with a tightly regulated expression of gene products that are either pro- or antiapoptotic. The aim of this study was to characterize pancreatic mRNA levels by Northern blotting analysis of apoptosis-associated genes used during the course of cerulein-induced AP in mice. Histone H3 mRNA levels were also examined as an indicator of cell proliferation. Acinar cell apoptosis was confirmed histologically. The findings show that AP modifies pancreatic mRNA levels of both pro- and antiapoptotic genes simultaneously. Pancreatic bclXL, bax, and p53 mRNA levels increased significantly in a temporal fashion during induction of AP. Pancreatic bcl-2 mRNA levels were unchanged during AP. Pancreatic mRNA levels of insulin-like growth factor-1 (IGF-1), a mitogen and cell survival factor, and its receptor (IGF-1R) also increased in a temporal fashion during induction of AP. In summary, this study indicates that acinar cell death during cerulein-induced AP in mice can occur by the apoptotic pathway. Since factors promoting and antagonistic for cell survival are activated simultaneously, regulation of acinar cell survival appears complex and dynamic during AP.

Hypoxia‐induced mitochondrial and nuclear DNA damage in the rat brain

In humans, cerebral hypoxia is a common component of severe brain insults, including trauma, stroke, and perinatal asphyxia. Oxidative stress and free radicals incidental to cerebral hypoxia are implicated in damaging macromolecules, leading to collapse of cellular homeostasis and cell death. Neuronal DNA damage, as a direct measurable event, has not been addressed in cerebral hypoxia. Here, we measured hypoxia‐induced damage and repair in nuclear and mitochondrial DNA in rat hippocampus and cortex. Two highly sensitive quantitative polymerase chain reaction (QPCR) assays were used to measure DNA damage. One assay measures the integrity of the entire mitochondrial genome and the other the integrity of nuclear DNA. The latter is a novel assay, developed in our laboratory, which utilizes the high copy number of short interspersed DNA elements (SINEs) residing in introns and untranslated regions of mammalian genes. A unique feature of the SINE‐mediated QPCR is its ability to amplify simultaneously long random segments of DNA. Consequently, the SINE assay offers sufficient sensitivity for detecting DNA damage at levels that are compatible with the cellular capacity for DNA repair, and are likely to be consistent with cellular survival and therefore adequate for studying the DNA damage response in the brain. In rats, we found that exposure to an atmosphere of 4% oxygen for 30 min resulted in induction of DNA damage in nuclear and to a greater extent, in mitochondrial DNA. Following a 3‐hr recovery period in ambient air, dissimilar repair kinetics for nuclear and mitochondrial DNA were measured. J. Neurosci. Res. 58:262–269, 1999.

Increased colonic apelin production in rodents with experimental colitis and in humans with IBD

Apelin and its receptor, the APJ receptor, are expressed in the gastrointestinal tract. The aims of this study were to examine the effects of sodium dextran sulfate (DSS)-induced experimental colitis in rats and mice and inflammatory bowel disease (IBD) in humans on intestinal apelin production, and the influence of exogenous apelin on colonic epithelial cell proliferation in mice. In rodents with experimental colitis, colonic apelin mRNA levels were elevated during the inflammatory reaction as well as during the tissue repair phase that ensues after DSS withdrawal. Fluctuations in colonic apelin expression were paralleled by similar changes in apelin immunostaining. Apelin immunostaining was increased in the surface epithelium, in epithelial cells along the length of the tubular gland and in the stem cell region at the gland base. In ulcerative colitis (UC) and Crohns disease patients, apelin immunostaining revealed a pattern of increased intestinal apelin content similar to that observed in rodents with experimental colitis. Administration of synthetic apelin to mice during the recovery phase of DSS-induced colitis stimulated colonic epithelial cell proliferation significantly. Our observations that colonic apelin production is increased during and after DSS exposure indicate that apelin plays multiple roles during the different stages of colitis. Additionally, the stimulatory action of exogenous apelin on colonic epithelial proliferation suggests that the increased apelin production during intestinal recovery stage may contribute to the repair of the intestinal epithelium in experimental rodent models of colitis and in IBD patients.

Age-associated changes in gene expression patterns in the duodenum and colon of rats

In humans, decreased intestinal motility, compromised nutritional status and increased risk of colon cancer are commonly associated with aging. Here, we used the cDNA microarray analysis to detect age-associated changes in duodenal and colonic gene expression in male Fischer 344 rats. The primary finding of this study is that the magnitude and direction of age-associated changes in gene expression differs in the colon and duodenum. In the colon, 56 genes showed altered expression, whereas expression of only 25 genes was altered in the duodenum. The magnitude of change was greater in the colon than in the duodenum. The direction of change also differed; in the aged colon, expression of 51 genes increased and only five genes decreased. In contrast, in the aged duodenum, only seven genes increased, whereas 18 genes decreased in expression. In the duodenum of aged rats, expression of genes involved in ATP-generating pathways is decreased. In contrast, in the colon of aged rats, expression of genes involved in energy generating pathways and in lipid oxidation is increased. In addition, in the aging colon, an increased expression of genes that show an aberrant regulation in colon cancer, including CD44, ras, and maspin is observed. Collectively, these findings provide clues to molecular events that may be related to compromised intestinal function and the high incidence of colon cancer in the aged population.

A possible role for hypoxia-induced apelin expression in enteric cell proliferation

Apelin is the endogenous ligand for the APJ receptor, and apelin and APJ are expressed in the gastrointestinal (GI) tract. Intestinal inflammation increases intestinal hypoxia-inducible factor (HIF) and apelin expression. Hypoxia and inflammation are closely linked cellular insults. The purpose of these studies was to investigate the influence of hypoxia on enteric apelin expression. Exposure of rat pups to acute hypoxia increased hepatic, stomach-duodenal, and colonic apelin mRNA levels 10-, 2-, and 2-fold, respectively (P < 0.05 vs. controls). Hypoxia also increased colonic APJ mRNA levels, and apelin treatment during hypoxia exposure enhanced colonic APJ mRNA levels further. In vitro hypoxia also increased apelin and APJ mRNA levels. The hypoxia-induced elevation in apelin expression is most likely mediated by HIF, since HIF-activated apelin transcriptional activity is dependent on an intact, putative HIF binding site in the rat apelin promoter. Acute exposure of rat pups to hypoxia lowered gastric and colonic epithelial cell proliferation; hypoxia in combination with apelin treatment increased epithelial proliferation by 50%. In vitro apelin treatment of enteric cells exposed to hypoxia increased cell proliferation. Apelin treatment during normoxia was ineffective. Our studies imply that the elevation in apelin expression during hypoxia and inflammation in the GI tract functions in part to stimulate epithelial cell proliferation.