Altered physiology of acid secretion in depression-prone Flinders rats results in exacerbated NSAID and stress-induced gastric damage.

BACKGROUND: Flinders Sensitive Line (FSL) rats are characterized by hypersensitivity to cholinergic stimuli and have been extensively used for studying depressive disorders. A link between depression and peptic ulcers has long been established; however, there is a lack of data from animal models. METHODS: We studied the physiology of acid secretion in FSL and Flinders Resistant Line (FRL) rats in vivo and in vitro. We also examined the susceptibility of Flinders rats to water immersion restraint stress (WIRS) or NSAID-induced gastric damage and explored the effect of an anticholinergic agent, atropine, in reversing this effect. KEY RESULTS: Basal acid output was more than twofold greater in FSL compared with FRL rats in vivo, 213.5 and 92.8 µEq/3 h/100 g (P = 0.02), respectively. Carbachol was a more potent secretagog in vitro, and somatostatin was a less potent inhibitory agent, while paradoxically stimulating acid secretion over and above the carbachol response in gastric glands from FSL rats. The FSL rats were more susceptible to indomethacin and WIRS-induced gastric mucosal damage compared with FRL rats. Atropine reduced acid output, which resulted in a reduction in indomethacin and stress-induced gastric damage in FSL rats. CONCLUSIONS & INFERENCES: Our study, for the first time, demonstrates that the altered vagally mediated physiology of acid secretion in depression-prone FSL rats contributes to gastric hypersecretion and, consequently, results in exacerbated stress and NSAID-induced gastric damage. Flinders rats may be a useful animal model for studying acid-related and also gastrointestinal functional disorders in depression.

Spontaneous autoimmune gastritis and hypochlorhydria are manifest in the ileitis-prone SAMP1/YitFcs mice.

SAMP1/YitFcs mice serve as a model of Crohn's disease, and we have used them to assess gastritis. Gastritis was compared in SAMP1/YitFcs, AKR, and C57BL/6 mice by histology, immunohistochemistry, and flow cytometry. Gastric acid secretion was measured in ligated stomachs, while anti-parietal cell antibodies were assayed by immunofluorescence and enzyme-linked immunosorbent spot assay. SAMP1/YitFcs mice display a corpus-dominant, chronic gastritis with multifocal aggregates of mononuclear cells consisting of T and B lymphocytes. Relatively few aggregates were observed elsewhere in the stomach. The infiltrates in the oxyntic mucosa were associated with the loss of parietal cell mass. AKR mice, the founder strain of the SAMP1/YitFcs, also have gastritis, although they do not develop ileitis. Genetic studies using SAMP1/YitFcs-C57BL/6 congenic mice showed that the genetic regions regulating ileitis had comparable effects on gastritis. The majority of the cells in the aggregates expressed the T cell marker CD3 or the B cell marker B220. Adoptive transfer of SAMP1/YitFcs CD4(+) T helper cells, with or without B cells, into immunodeficient recipients induced a pangastritis and duodenitis. SAMP1/YitFcs and AKR mice manifest hypochlorhydria and anti-parietal cell antibodies. These data suggest that common genetic factors controlling gastroenteric disease in SAMP1/YitFcs mice regulate distinct pathogenic mechanisms causing inflammation in separate sites within the digestive tract.

Establishment of T-Helper-2 immune response based gerbil model of enteric infection.

BACKGROUND: The reciprocal antagonism of T-helper-1 (Th-1) and Th-2 type immune responses suggests that helminth parasitic infection may ameliorate disease where a Th-1 type response dominates. The Mongolian gerbil has been useful in the investigation of the pathogenesis of gastric cancer, since long-term infection of gerbils with Helicobacter pylori induces adenocarcinoma. In this study the kinetics of worm expulsion and associated immune responses in gerbils infected with Trichinella spiralis were investigated in an attempt to establish an animal model of parasitic infection that could be helpful when investigating the effect of a Th-2 type response on Th-1-based intestinal disorders. METHODS: Gerbils were infected with various doses of infective T. spiralis larvae and were euthanized on different days after infection to investigate the intestinal worm recovery, goblet cell population, eosinophil response and serum IgG1 responses. RESULTS: The number of worms recovered from the intestine was dependent on the number of larvae used for the infection. Almost all worms were expelled spontaneously by day 26 post-infection, when the gerbils had been infected with 375 or 750 larvae. The number of intestinal goblet cells, eosinophils and the serum IgG1 level significantly increased following infection compared with the control. CONCLUSION: This is the first comprehensive report on the time-course of T. spiralis infection in gerbils. The data indicate that the T. spiralis-infected gerbil could be used as a model of the Th-2-based response to investigate the effect of a parasite-induced Th-2 response on various Th-1-mediated intestinal disorders such as H. pylori-induced gastritis and gastric carcinoma.

Effect of Th1 cytokines on acid secretion in pharmacologically characterised mouse gastric glands.

BACKGROUND AND AIMS: Acid secretion plays an important role in the ecology of Helicobacter species and acid secretory status heralds patterns of gastritis. The presence of inflammatory cells and their products, in close proximity to parietal cells, questions the extent of the effect of cytokines on acid secretion. METHODS: We adopted and extensively characterised the mouse gastric gland preparation and its secretory capacity, which was measured using (14)C-aminopyrine accumulation. Subsequently, we tested the secretory properties of a wide range of species specific cytokines, including those associated with Th1 and Th2 immune responses. RESULTS: (14)C-aminopyrine accumulation in mouse gastric glands was shown to be a very sensitive "in vitro" method of testing classical secretagogues and antisecretory compounds, and provided pharmacological data on acid secretion in the mouse. Only two mouse cytokines, interleukin 2 and interferon gamma, had a direct effect on acid secretion causing dose dependent inhibition. CONCLUSIONS: Both cytokines belong to the Th1 type immune response and consequently their inhibitory effect may play a role in the hyposecretion seen with H pylori infection and colonisation throughout the corpus of the stomach that potentially can lead to gastric atrophy and subsequently, in some cases, cancer.

Effect of purified lipopolysaccharides from strains of Helicobacter pylori and Helicobacter felis on acid secretion in mouse gastric glands in vitro.

As a bacterial product, Helicobacter pylori lipopolysaccharide (LPS) can originate in close proximity to parietal cells, but the role of this uniquely structured endotoxin on acid secretion has not been fully investigated and remains unclear. The purpose of this study was to test the direct effect of purified LPS (tested range, 0.1 to 100 microg/ml) from various strains of H. pylori and from one Helicobacter felis strain on histamine- and carbachol-stimulated acid secretion in vitro using mouse gastric glands and the accumulation of [(14)C]aminopyrine. In addition, we investigated whether H. pylori LPS can interfere with two native antisecretory substances, prostaglandin E(2) (PGE(2)) and somatostatin, which may contribute to bacterial pathogenicity. Except for the LPS from H. pylori SS1 (Sydney strain), which gave a statistically significant increase in both histamine- and carbachol-stimulated acid output (38 and 24%, respectively; P < 0.05), no effect of the tested LPS was observed on acid secretion. H. pylori LPS purified from a patient isolate did not affect the potency or the efficacy of the inhibitory dose response curve to PGE(2) or somatostatin. Bacterial interstrain variation in the direct stimulatory effect of Helicobacter-derived LPS on acid secretion was observed, which probably reflects the molecular structure of LPS and the potential to contribute to virulence. Importantly, the data showed that H. pylori LPS did not have any direct antisecretory properties. It can be speculated that the acid stimulatory properties of LPS from H. pylori SS1 may contribute to the gastric damage observed in the mouse model of H. pylori infection.

Therapeutic effects of the endothelin receptor antagonist Ro 48-5695 in the TNBS/DNBS rat model of colitis.

OBJECTIVE: Endothelins can act as polyfunctional cytokines. It is therefore possible that endothelins could play an active role in gut inflammation. Elevated levels of endothelin-1 have been reported in ulcerative colitis and Crohn's disease. The aim of this study was to establish the therapeutic effect of a 'new' endothelin receptor antagonist Ro 48-5695 in an animal model of inflammatory bowel disease. This study compares the effect of Ro 48-5695 on colonic damage induced by two haptens: trinitrobenzenesulphonic (TNBS) or dinitrobenzenesulphonic acid (DNBS). METHODS: Colitis was induced by intra-rectal administration of TNBS or DNBS. After TNBS/DNBS injury, rats were treated with 10.0, 3.0, 1.0 or 0.3 mg/kg of Ro 48-5695 orally, daily for 5 days. On day 6 post-hapten treatment, colonic tissues were removed and examined in a blinded fashion for macroscopic damage (damage score) and myeloperoxidase (MPO) activity. Stool consistency and adhesions were also measured. RESULTS: Oral administration of Ro 48-5695 almost completely prevented TNBS-induced damage at a dose of 10 mg/kg. The same dose in this model also had a therapeutic effect as measured by MPO and incidence of diarrhoea and adhesions. In DNBS-induced colonic damage, Ro 48-5695 was more potent and at 1.0 and 3.0 mg/kg decreased the damage score by 50 and 60% respectively; also the incidence of adhesions and diarrhoea was significantly reduced. However, MPO activity in this model was affected only by the highest dose of Ro 48-5695 tested (3.0 mg/kg) where it was reduced by 48%. CONCLUSIONS: These data provide evidence for the involvement of endothelins in the pathophysiology of inflammatory bowel disease and support the possibility of exploring a new therapeutic approach.

Anti-ulcerogenic properties of endothelin receptor antagonists in the rat.

BACKGROUND: Endothelins have been implicated in gastric mucosal damage in a variety of animal models. Exogenous ET-1 and ET-3 are causally associated with experimental gastric ulcers. Furthermore, clinical reports also show elevated plasma and gastric mucosal endothelin-1 levels in patients suffering from peptic ulcers. AIM: To study the possibility that endothelin receptor antagonists may have beneficial effects and prevent the development of gastric ulcers. We have tested in rats the orally-active endothelin antagonist bosentan (Ro 47-0203) and Ro 48-5695, which is 10-30 times more potent than bosentan on endothelin receptors. METHODS: Water immersion restrained stress (WIRS) and indomethacin were used to provoke gastric mucosal damage. Endothelin receptor antagonists were administered orally prior to the induction of gastric damage. The gastric lesion index (mm), assessed macroscopically, and myeloperoxidase (MPO) activity were used as markers of the extent of mucosal injury. RESULTS: Bosentan at 100 and 30 mg/kg administered orally caused attenuation of gastric damage in the WIRS model by 58% and 42%, respectively. Bosentan also caused complete reduction of MPO activity. In indomethacin-induced gastric damage, 100 mg/kg bosentan attenuated gastric damage by 45% and 61% as measured by the gastric lesion index and MPO activity respectively. Ro 48-5695 was at least 30 times more potent than bosentan in reducing indomethacin-induced mucosal damage and at 3 mg/kg, caused a decrease of 49% in the gastric lesion index and a reduction in MPO activity of 41%. Bosentan and Ro 48-5695 possess weak antisecretory properties as tested in the mouse gastric gland assay, than cannot, alone, account for their anti-ulcer properties. CONCLUSIONS: Both endothelin receptor antagonists prevented the development of gastric mucosal injury in the rat. Disturbances in the gastric microcirculation are responsible for the development of experimental gastric ulcers. The anti-ulcer properties of these two endothelin antagonists suggest possible new therapeutic approaches to controlling gastric inflammation.

Inhibitory effect of diazepam on human natural killer activity in vitro.

The in vitro effect of diazepam on natural killer (NK) cell activity isolated from human peripheral blood has been investigated. NK cell function was estimated by means of a radioactive chromium (51Cr) assay in which human erythroleukaemia K562 cells were used as the target. Diazepam suppressed NK cell function in a concentration-dependent manner (10(-7)-10(-5) M) and a positive correlation was found between the dose of diazepam and inhibition detected by percentage killing of K562 cells. These data indicate that diazepam influences cytotoxic activity and diminishes anti-viral and anti-tumour defence reactions in humans.

Immunomodulatory properties of diazepam-binding inhibitor: effect on human interleukin-6 secretion, lymphocyte proliferation and natural killer cell activity in vitro.

We have examined the influence of diazepam binding inhibitor (octadecaneuro-peptide, DBI33-50) on cell mediated immune responses including LPS-stimulated monocyte IL-6 secretion, PHA induced lymphocyte proliferation and NK cell function in humans. All studies were performed in vitro on isolated human peripheral blood mononuclear cells in the absence or presence of synthetic DBI33-50. It has been shown that DBI33-50, in concentration between 10(-6)-10(-8) M, enhances the LPS-induced secretion of IL-6, as determined by specific bioassay for this monokine. On the other hand DBI33-50 (10(-6)-10(-12) M), had no significant effect on either PHA-induced lymphocyte proliferation or NK cell function. This data suggests a possible immunomodulatory role for DBI33-50 as an endogenous neuropeptide, which stimulates IL-6 secretion by human monocytes.

Immunomodulatory activities of the somatostatin analogue BIM 23014c: effects on murine lymphocyte proliferation and natural killer activity.

We have examined the effect of somatostatin and its octapeptide analogue BIM 23014c on concanavalin A-induced lymphocyte proliferation and target-specific natural killer activity both in vitro and in vivo. Using Peyer's patches and spleen as a source of lymphocytes, we found that both peptides modulated immunity in a dose-dependent manner. Comparatively, there was no significant difference between the activity of somatostatin or BIM 23014c in the modulation of immunity. Proliferation, both in vitro and in vivo, was significantly inhibited by both peptides in each organ with a higher specificity towards the Peyer's patch lymphocytes. Natural killer activity was also inhibited in both organs in vivo and in vitro. Thus, not only did somatostatin and BIM 23014c have similar effects on proliferation and natural killer activity, but their effect was organ specific. Preliminary data suggest that BIM 23014c works via the same receptor as somatostatin, therefore intimating that these two peptides are both clinically and immunologically similar.

Selective modulation of the natural killer activity of murine intestinal intraepithelial leucocytes by the neuropeptide substance P.

Neuropeptides can influence immune effector cell function at both systemic and mucosal immune sites. We examined the ability of substance P (SP) to modulate the natural killer (NK) activity of intestinal intraepithelial leucocytes (IEL). Yac-1 killing by IEL but not splenic cells was increased after either 18 hr preincubation or 6 hr of co-incubation with SP. We also examined the NK activity of IEL and spleen isolated from mice treated with SP in vivo. The selective increase in NK activity of IEL occurred without any demonstrable change in the number or phenotype of the IEL. The IEL responsive to SP in vivo and induced in vivo by SP were both Thy-1- and did not kill the NK insensitive mastocytoma cell line P815. Lastly, we examined the ability of SP to induce the release of interleukin-2 (IL-2) and IL-4 from IEL after 6 and 18 hr of in vitro culture. No increase in the release of these cytokines was observed, suggesting that IL-2 and IL4 are not involved in the local augmentation of IEL NK activity by SP. These observations suggest that SP has a selective stimulatory effect on intestinal activity and may play a role in the regulation of intestinal cell-mediated immunity.