<p>The human stomach normally contains high levels of bioactive nitric oxide (NO). This NO derives from salivary nitrate (NO₃^-) that is converted to nitrite (NO₂^-) by oral bacteria and thereafter non-enzymatically reduced in the acidic gastric lumen to NO. Nitrate is a common component in vegetables, and after ingestion it is absorbed in the small intestine. Interestingly, circulating nitrate is then concentrated by the salivary glands. Hence, intake of nitrate-rich vegetables results in high levels of NO in the stomach. The physiological effects of the high concentration of NO gas normally present in the gastric lumen have been hitherto unknown, and the present investigations were therefore conducted to address this issue.</p><p>NO produced in the gastric lumen after nitrate ingestion increased gastric mucosal blood flow and the thickness of the firmly adherent mucus layer in the stomach. The blood flow and mucus layer are essential defense mechanisms that protect the mucosa from luminal acid and noxious agents. Nonsteroidal antiinflammatory drugs (NSAID) are commonly prescribed and effective drugs for treating pain and inflammation, but are associated with severe gastrointestinal side effects. We demonstrated that a nitrate-rich diet protects against NSAID-induced gastric damage, as a result of the increased formation of NO in the stomach. We also showed that the gastroprotective effect attributed to nitrate depended completely on conversion of nitrate to nitrite by the bacterial flora colonizing the tongue, and that the oral microflora is therefore important in regulating physiological conditions in the stomach.</p><p>In summary, this thesis challenges the current dogma that nitrate intake is hazardous, and on the contrary suggests that dietary nitrate plays a direct role in regulating gastric homeostasis. It is likely that a sufficient supply of nitrate in the diet together with the oral microflora is essential for preventing pathological conditions in the gastrointestinal tract.</p>

The human stomach normally contains high levels of bioactive nitric oxide (NO). This NO derives from salivary nitrate (NO3-) that is converted to nitrite (NO2-) by oral bacteria and thereafter non-enzymatically reduced in the acidic gastric lumen to NO. Nitrate is a common component in vegetables, and after ingestion it is absorbed in the small intestine. Interestingly, circulating nitrate is then concentrated by the salivary glands. Hence, intake of nitrate-rich vegetables results in high levels of NO in the stomach. The physiological effects of the high concentration of NO gas normally present in the gastric lumen have been hitherto unknown, and the present investigations were therefore conducted to address this issue. NO produced in the gastric lumen after nitrate ingestion increased gastric mucosal blood flow and the thickness of the firmly adherent mucus layer in the stomach. The blood flow and mucus layer are essential defense mechanisms that protect the mucosa from luminal acid and noxious agents. Nonsteroidal antiinflammatory drugs (NSAID) are commonly prescribed and effective drugs for treating pain and inflammation, but are associated with severe gastrointestinal side effects. We demonstrated that a nitrate-rich diet protects against NSAID-induced gastric damage, as a result of the increased formation of NO in the stomach. We also showed that the gastroprotective effect attributed to nitrate depended completely on conversion of nitrate to nitrite by the bacterial flora colonizing the tongue, and that the oral microflora is therefore important in regulating physiological conditions in the stomach. In summary, this thesis challenges the current dogma that nitrate intake is hazardous, and on the contrary suggests that dietary nitrate plays a direct role in regulating gastric homeostasis. It is likely that a sufficient supply of nitrate in the diet together with the oral microflora is essential for preventing pathological conditions in the gastrointestinal tract.

<p>The gastrointestinal mucus gel layer and blood flow are two important mechanisms for protection at the pre-epithelial and sub-epithelial levels, respectively. <i>Helicobacter pylori</i> might circumvent these mechanisms and elicit a chronic inflammatory response with consequent ulcers in the stomach and duodenum. In this thesis, the physical state and properties of the adherent mucus gel layer was studied from the stomach to colon. Furthermore, the acute and chronic effects of <i>H. pylori</i> on the integrity of the mucus gel layer and mucosal blood flow were studied in the anesthetized rat.</p><p>A translucent mucus gel covers all studied segments of the gastrointestinal tract during fasting conditions, with the thickest layers in the colon and ileum. Carefully applied suction revealed that the mucus gel was a multi-layered structure comprising a firmly adherent layer covering the mucosa, impossible to remove, and a loosely adherent upper layer. The firmly adherent layer was thick and continuous in the corpus (80μm), antrum (154μm) and colon (116μm), but thin (<20μm) and discontinuous in the small intestine.</p><p>Following mucus removal, a rapid renewal of the loosely adherent layer ensued. The highest rate was observed in the colon with intermediate values in the small intestine. Mucus renewal in the stomach was attenuated on acute luminal application of water extracts from <i>H. pylori</i> (HPE). In animals with a chronic <i>H. pylori</i> infection the mucus renewal rate was unaffected, but the total gastric mucus gel thickness was reduced and the mucus secretory response to luminal acid (pH1) attenuated in the antrum. </p><p>HPE from type I strains acutely reduced corporal mucosal blood flow, measured with laser-Doppler flowmetry, by approximately 15%. The reduction in blood flow was mediated by a heat stable factor other than VacA and CagA. Inhibition of endogenous nitric oxide production with Nω-nitro-l-arginine augmented the decrease. However, ketotifen, a mast cell stabilizer, completely attenuated the effect of the extract as did the platelet activating factor (PAF) receptor-antagonist, WEB2086, thus depicting a detrimental role for the microvascular actions of PAF.</p>

<p>The stomach is frequently exposed to hazardous agents and to resist this harsh environment, several protective mechanisms exist. Of special interest is the gastric pathogen <i>Helicobacter pylori </i>which causes gastritis, ulcers and cancer but the mechanism leading to these diseases are still unclear. However it is very likely that <i>H. pylori </i>negatively influence the protection mechanisms that exist in the stomach. </p><p>The aims of the present investigation were first to develop an in vivo mouse model in which different protection mechanisms could be studied, and second to investigate the influence of <i>H. pylori</i> on these mechanisms. </p><p>An in vivo preparation of the gastric mucosa in mice was developed. This preparation allows studies of different gastric mucosal variables and can also be applied for studies in other gastro-intestinal organs. </p><p>Mice chronically infected with <i>H. pylori</i>, were shown to have a reduced ability of the mucosa to maintain a neutral pH at the epithelial cell surface. This could be due to the thinner inner, firmly adherent mucus gel layer, and/or to defective bicarbonate transport across the epithelium. The Cl^-/HCO₃^- exchanger SLC26A9 was inhibited by NH₄⁺, which also is produced by <i>H. pylori</i>. The mRNA levels of SLC26A9 were upregulated in infected mice, suggesting a way to overcome the inhibition of the transporter. Furthermore, the hyperemic response to acid pH 2 and 1.5 was abolished in these mice. The mechanisms by which the bacteria could alter the blood flow response might involve inhibition of the epithelial iNOS.</p><p>Water extracts of <i>H. pylori </i>(HPE) reduces the blood flow acutely through an iNOS and nerve-mediated pathway, possibly through the endogenous iNOS inhibitor ADMA. Furthermore, HPE alters the blood flow response to acid as the hyperemic response to acid pH 0.8 is accentuated in mice treated with HPE. </p>

The gastrointestinal mucus gel layer and blood flow are two important mechanisms for protection at the pre-epithelial and sub-epithelial levels, respectively. Helicobacter pylori might circumvent these mechanisms and elicit a chronic inflammatory response with consequent ulcers in the stomach and duodenum. In this thesis, the physical state and properties of the adherent mucus gel layer was studied from the stomach to colon. Furthermore, the acute and chronic effects of H. pylori on the integrity of the mucus gel layer and mucosal blood flow were studied in the anesthetized rat. A translucent mucus gel covers all studied segments of the gastrointestinal tract during fasting conditions, with the thickest layers in the colon and ileum. Carefully applied suction revealed that the mucus gel was a multi-layered structure comprising a firmly adherent layer covering the mucosa, impossible to remove, and a loosely adherent upper layer. The firmly adherent layer was thick and continuous in the corpus (80μm), antrum (154μm) and colon (116μm), but thin (&lt;20μm) and discontinuous in the small intestine. Following mucus removal, a rapid renewal of the loosely adherent layer ensued. The highest rate was observed in the colon with intermediate values in the small intestine. Mucus renewal in the stomach was attenuated on acute luminal application of water extracts from H. pylori (HPE). In animals with a chronic H. pylori infection the mucus renewal rate was unaffected, but the total gastric mucus gel thickness was reduced and the mucus secretory response to luminal acid (pH1) attenuated in the antrum. HPE from type I strains acutely reduced corporal mucosal blood flow, measured with laser-Doppler flowmetry, by approximately 15%. The reduction in blood flow was mediated by a heat stable factor other than VacA and CagA. Inhibition of endogenous nitric oxide production with Nω-nitro-l-arginine augmented the decrease. However, ketotifen, a mast cell stabilizer, completely attenuated the effect of the extract as did the platelet activating factor (PAF) receptor-antagonist, WEB2086, thus depicting a detrimental role for the microvascular actions of PAF.

The stomach is frequently exposed to hazardous agents and to resist this harsh environment, several protective mechanisms exist. Of special interest is the gastric pathogen Helicobacter pylori which causes gastritis, ulcers and cancer but the mechanism leading to these diseases are still unclear. However it is very likely that H. pylori negatively influence the protection mechanisms that exist in the stomach. The aims of the present investigation were first to develop an in vivo mouse model in which different protection mechanisms could be studied, and second to investigate the influence of H. pylori on these mechanisms. An in vivo preparation of the gastric mucosa in mice was developed. This preparation allows studies of different gastric mucosal variables and can also be applied for studies in other gastro-intestinal organs. Mice chronically infected with H. pylori, were shown to have a reduced ability of the mucosa to maintain a neutral pH at the epithelial cell surface. This could be due to the thinner inner, firmly adherent mucus gel layer, and/or to defective bicarbonate transport across the epithelium. The Cl-/HCO3- exchanger SLC26A9 was inhibited by NH4+, which also is produced by H. pylori. The mRNA levels of SLC26A9 were upregulated in infected mice, suggesting a way to overcome the inhibition of the transporter. Furthermore, the hyperemic response to acid pH 2 and 1.5 was abolished in these mice. The mechanisms by which the bacteria could alter the blood flow response might involve inhibition of the epithelial iNOS. Water extracts of H. pylori (HPE) reduces the blood flow acutely through an iNOS and nerve-mediated pathway, possibly through the endogenous iNOS inhibitor ADMA. Furthermore, HPE alters the blood flow response to acid as the hyperemic response to acid pH 0.8 is accentuated in mice treated with HPE.

Inflammatory bowel diseases, (IBD), are a group of chronic disorders of the gastro-intestinal tract, and include Crohn’s disease (CD) and Ulcerative Colitis (UC). The pathogenesis is not known, but involves at least in part a loss of tolerance towards the commensal colonic microbiota. In this thesis, we show in animal models of CD and UC that the colonic mucosal blood flow increased compared to healthy animals. This blood flow increase is due to an up regulation of endothelial nitric oxide synthase (NOS). Further, we show in the UC model that the thickness of the firmly adherent colonic mucus layer increased compared to healthy animals. This increase is due to an up regulation of inducible NOS in the epithelium. Both the blood flow and mucus thickness increase appear to be protective mechanisms.  We demonstrate that the firmly adherent colonic mucus layer acts as a partial barrier towards luminal bacteria. In the UC model, this barrier is destroyed, causing increased bacterial translocation. The adhesion molecule P-selectin was up regulated in the UC model, leading to increased interactions between leukocytes and the endothelium, but also increased interactions between platelets and the endothelium. This indicates that not only leukocytes, but also platelets are involved in colonic inflammation. The addition of the probiotic bacterial strain Lactobacillus reuteri prevented disease by normalizing P-selectin levels and endothelial interactions with leukocytes and platelets. Lactobacillus reuteri also decreased bacterial translocation over the epithelium. In summary, this thesis highlights the importance of colonic barrier functions, and investigates the role of the microbiota in experimental IBD.