Search

Objective: The human gastro-oesophageal junction is exposed to abundant amounts of luminal reactive nitrogen oxide species (RNOS) derived from the enterosalivary recirculation of dietary nitrate. The aim of this study is to investigate the direct effects of luminal RNOS on the adjacent gastric barrier function using an ex vivo chamber model. Methods: A chamber model in which the rat gastric mucosal membrane was mounted between the two halves of a chamber was designed to simulate the microenvironment of the lumen and the adjacent mucosa of the gastro-oesophageal junction. On the mucosal side of the chamber, RNOS were generated by the acidification of physiological concentrations of sodium nitrite. The epithelial barrier function was evaluated by electrophysiological transmembrane resistance, and membrane permeability with [³H]mannitol flux. The expression of occludin was evaluated by immunohistochemistry and immunoblotting. Dinitrosyl dithiolato iron complex (DNIC) was also measured by means of electron paramagnetic resonance spectroscopy to confirm the diffusion of RNOS from the mucosal lumen into the mounted mucosa. Results: The administration of acidified nitrite to the mucosal lumen caused both a decrease in transmembrane resistance and an increase in epithelial permeability, suggesting a disturbance of the gastric barrier function. These changes were accompanied by a derangement of the expression of occludin. The diffusion of luminal RNOS into the mounted membrane was confirmed by showing the generation of DNIC within the tissue. Conclusions: Simulating the microenvironment of the human gastro-oesophageal junction, this study demonstrated that RNOS generated luminally at the human gastro-oesophageal junction can derange the barrier function of the adjacent tissue by disrupting the tight junction. [ABSTRACT FROM AUTHOR]

Background: Bartter syndrome (BS) is a genetic disorder accompanied by hypokalaemic metabolic alkalosis. BS with sensorineural deafness (SND, 0M1M602522) is a newly identified phenotype caused by mutations in the BSNQ gene that encodes barttin, a β-subunit for chloride channel ClC-Ka and CIC-Kb and classified as type IV BS. Type IV BS features the most severe phenotype entailing life-threatening neonatal volume depletion and chronic renal failure developing during infancy. A recent report described a case of BS with SND from a consanguineous family who showed homozygous mutations in the CLCNKA and CLCNKB genes. This case indicated the possibility of the occurrence of digenic inheritance in BS with SND resulting from double mutations in the CLCNKA and CLCNKB genes. Subject and results: The current report concerns a 2- year-old girl from a non-consanguineous family with BS accompanied by SND. In our case, four loss-of-function mutations, consisting of mutations in both parental alleles in both CLCNKA and CLCNKB, were identified. The paternal allele had a nonsense mutation (0260X) in CLCNKA and a splicing site mutation (IVS17+1 g>a) in CLCNKB. The maternal allele had a large deletion mutation (about 12 kbp) extending from CLCNKA to CLCNKB. Our case provides clear evidence that loss-of-function alleles in both alleles of both CLCNKA and CLCNKB results in a phenotype indistinguishable from that of mutations in BSND (type IV BS(. Conclusions: Recent advances in genetics have resulted in a better understanding of many human inherited diseases, but most of them are monogenic disorders and more complex inheritance patterns remain unresolved. Our case provides clear evidence of digenic inheritance outside the scope of Mendelian inheritance disorders. [ABSTRACT FROM AUTHOR]

Background and aims: When saliva, with its high nitrite content derived from the enterosalivary recirculation of dietary nitrate, meets acidic gastric juice, the nitrite is converted to nitrous acid, nitrosative species, and nitric oxide. In healthy volunteers this potentially mutagenic chemistry is focused at the gastric cardia. We have studied the location of this luminal chemistry in Barrett's patients during acid reflux. Methods: Ten Barrett's patients were studied before and after administration of 2 mmol nitrate. Using microdialysis probes we measured nitrite, ascorbic acid, total vitamin C, and thiocyanate concentrations and pH simultaneously in the proximal oesophagus, Barrett's segment, hiatal sac, proximal stomach, and distal stomach. In a subgroup, real time nitric oxide concentrations were also measured. Results: During acid reflux, Barrett's segment was the anatomical site with maximal potential for acid catalysed nitrosation, with its median concentration of nitrite exceeding that of ascorbic acid in two of 10 subjects before nitrate and in four of nine after nitrate. Thiocyanate, which catalyses acid nitrosation, was abundant at all anatomical sites. On entering the acidic Barrett's segment, there was a substantial fall in nitrite and the lowest ascorbic acid to total vitamin C ratio, indicative of reduction of salivary nitrite to nitric oxide at this anatomical site. Episodes of acid reflux were observed to generate nitric oxide concentrations of up to 60 μM within the Barrett's segment. Conclusion: The interaction between acidic gastric refluxate and nitrite rich saliva activates potentially mutagenic luminal nitrosative chemistry within Barrett's oesophagus. [ABSTRACT FROM AUTHOR]

Background: In humans, high concentrations of nitric oxide are generated luminally at the gastro- oesophageal junction through enterosalivary recirculation of dietary nitrate. Aim: To investigate whether luminal nitric oxide can diffuse into the adjacent digestive tissue and alter tissue integrity. Methods: We designed an animal model using Wistar rats in which physiological concentrations of nitrite and acidified ascorbic acid were administered separately so that the two reactants first meet to form nitric oxide at the gastro-oesophageal junction. Luminal and tissue concentrations of nitric oxide were measured with an electrode and an electron paramagnetic resonance spectrometer, respectively. Concentrations of glutathione in the tissue were measured as a marker of nitrosative stress. n; Results: High concentrations of luminal nitric oxide were generated locally at the gastro-oesophageal junction of nitrite administered rats, reproducing a phenomenon observed in humans. High levels of nitric oxide were also detected largely in the superficial epithelium of the gastro-oesophageal junction. The concentration of I tissue glutathione at the gastro-oesophageal junction was significantly lower in nitrite administered rats compared with control rats, whereas that in the distal stomach was similar in the two rat groups. Conclusions: Using an animal model, this study demonstrated that nitric oxide generated in the lumen diffuses into the adjacent gastric tissue to a substantial degree, leading to localised consumption of glutathione in the tissue. Nitrosative stress induced by this mechanism may be involved in the high prevalence of inflammation and metaplasia, and subsequent development of neoplastic disease at this site. [ABSTRACT FROM AUTHOR]

Background: Saliva has a high nitrite concentration, derived from the enterosalivary recirculation of dietary nitrate, and is the main source of nitrite entering the acidic stomach. Acidification of nitrite in the presence of secondary amines or amides generates potentially carcinogenic N-nitroso compounds. The reaction is inhibited by ascorbic acid and catalysed by thiocyanate. Aim: To determine whether there is intragastric regional variation in the chemical conditions promoting luminal nitrosation following nitrate ingestion. Methods: Using microdialysis probes, we measured concentrations of nitrite, ascorbic acid, total vitamin C, and thiocyanate simultaneously in saliva, the distal oesophagus, cardia, and the proximal and distal stomach of 17 healthy volunteers before and following intragastric nitrate (2 mmol) administration. Results: The median pH in the distal oesophagus, cordia, and proximal and distal stomach were 7, 2.6, 1.9, and 1.7, respectively, before, and were similar following nitrate administration. Mean nitrite concentration in the distal oesophagus was similar to that of saliva, being 29.1 µM and 36.7 µM, respectively, before nitrate and increasing to 181.6 µM and 203.3 µM after nitrate ingestion. Within the stomach, mean (SEM) nitrite concentration following nitrate was higher in the cordia (45.5 (12.7) µM) than in the mid (7.8 (3.1)) (p<0.01) or distal (0.8 (0.6)) (p<0.1) stomach, and ascorbic acid concentration was lower at the cordia (13.0 (6.1)) than in the mid (51 (19.2)) (p<0.02) or distal (86 (29)) (p<0.01) stomach. Consequently, the median ascorbic acid to nitrite ratio was lowest at the cardia (0.3) (p<0.01) versus the mid (7.8) or distal (40) stomach. Thiocyanate concentration was similar throughout the stomach. Conclusions: The conditions favouring luminal generation of N-nitroso compounds from dietary nitrate are maximal at the most proximal cordia region of the acidic stomach and may contribute to the high incidence of mutagenesis at this site. [ABSTRACT FROM AUTHOR]

High concentrations of nitric oxide are generated at the gastro-oesophageal (GO) junction due to the reduction of salivary nitrite to nitric oxide by acidic gastric juice containing ascorbic acid. Salivary nitrite is derived from the enterosalivary recirculation of dietary nitrate. Aims: To determine whether nitric oxide generated in the above way will exert nitrosative stress on the adjacent epithelium. Methods: A benchtop model was constructed reproducing the chemistry occurring at the GO junction and incorporating an epithelial compartment maintained at pH 7.4 separated from the lumen by a thin hydrophobic barrier. The secondary amine morpholine was added to each compartment and N-nitrosomorpholine formation at 15 min measured. Results: Adding 100µM nitrite to the acidic (pH 1.5) luminal compartment in the absence of ascorbic acid generated 6.2±2.0 (mean±SE) N-nitrosomorpholine in that compartment and 2.2±0.1 µM in the epithelial compartment. When 100µM nitrite was added to the acidic luminal compartment (pH 1.5) containing ascorbic acid, all the nitrite was immediately converted to nitric oxide and no N-nitrosomorpholine was formed within that compartment. However, the nitric oxide rapidly diffused into the adjacent epithelial compartment (pH 7.4) where it generated very high concentrations of N-nitrosomorpholine (137±5.6µM). The addition of ascorbic acid or glutathione to the epithelial compartment only reduced this nitric oxide induced nitrosation within the epithelial compartment by 40%. Conclusion: Ascorbic acid in gastric juice prevents acid-catalysed nitrosation within the gastric lumen. However, in doing so it generates nitric oxide which exerts a far higher nitrosative stress on the adjacent epithelium. This mechanism may be relevant to the aetiology of mutagenesis at the GO junction. [ABSTRACT FROM AUTHOR]

Background: The lumen of the oesophagus has a high nitrite concentration due to the enterosalivary recirculation of dietary nitrate. On entering the acidic stomach, the nitrite is converted to nitric oxide, which may contribute to mutagenesis at the cardia. Aim: To study luminal nitrite chemistry in patients with Barrett's oesophagus. Methods: Using microdialysis probes, we studied nitrite concentration in saliva, proximal oesophagus within the Barrett's segment, and proximal and distal stomach of 10 Barrett's patients before and following nitrate (2 mmol) administration. Results were compared with 17 healthy controls. In a subgroup of Barrett's patients, we simultaneously monitored oesophageal pH and nitric oxide concentration. Results: In the healthy controls, a high nitrite concentration was present in saliva (mean fasting = 37 µM, after nitrate = 203 µM) and throughout the length of the oesophagus (fasting = 29 µM, after nitrite = 181 µM) but fell by 75% on entering the acidic gastric cardia. The fall in nitrite was associated with a high cardia concentration of nitric oxide (fasting = 2.4 µM, after nitrite = 7.5 µM). In the Barrett's patients, similarly high concentrations of nitrite were present in saliva and in the proximal oesophagus but the nitrite concentration fell by 98% on entering the Barrett's segment exposed to acid reflux. Simultaneous recording of pH and nitric oxide concentration within the Barrett's segment indicated that the reflux of acidic gastric juice into the Barrett's segment resulted in the immediate generation of high concentrations of nitric oxide within the oesophageal lumen (fasting = 13 µM, after nitrate = 20 µM). Conclusions: The lumen of the normal oesophagus contains high concentrations of nitrite derived from the enterosalivary recirculation of dietary nitrate. The reflux of acidic gastric juice into the distal oesophagus immediately converts the nitrite to nitric oxide. The... [ABSTRACT FROM AUTHOR]