幽门螺杆菌感染对延缓乙酸诱导大鼠胃溃疡愈合的实验研究

本工作旨在观察幽门螺秆菌（Ｈｅｌｉｃｏｂａｃｔｅｒｐｙｌｏｒｉ,ＨＰ）感染对大鼠乙酸性胃溃疡愈合的影响。实验发现,大鼠发生乙酸性溃疡后,一次性接种ＨＰ几乎使所有大鼠感染,并引起胃溃疡的自然愈合明显减慢;组织学和生化检测观察到ＨＰ组大鼠胃溃疡病灶处有更多炎症细胞浸润,以中性粒细胞为主,未受溃疡累及的胃窦和胃后壁粘膜也有炎症细胞浸润;免疫组化研究发现ＨＰ组胃粘膜出现较多的ＩＬ－８阳性细胞;此外发现ＨＰ组胃粘膜ＢｒｄＵ标记细胞显著减少,既粘膜上皮的再生减慢。结果提示,ＨＰ感染延缓乙酸性溃疡的愈合,后者与ＨＰ感染后增加ＩＬ－８表达所诱导更强的炎症反应和粘膜细胞再生减慢有关。     

Neutralizing anti-IL-10 antibody blocks the protective effect of tapeworm infection in a murine model of chemically induced colitis

There is increasing evidence that parasitic helminth infection has the ability to ameliorate other disease conditions. In this study the ability of the rat tapeworm, Hymenolepis diminuta, to modulate dinitrobenzene sulfonic acid (DNBS)-induced colitis in mice is assessed. Mice receiving DNBS (3 mg intrarectally) developed colitis by 72 h after treatment. Mice infected 8 days before DNBS with five H. diminuta larvae were significantly protected from the colitis, as gauged by reduced clinical disease, histological damage scores, and myeloperoxidase levels. This anticolitic effect was dependent on a viable infection and helminth rejection, because no benefit was observed in mice given killed larvae or in infected STAT6 knockout mice or rats, neither of which eliminate H. diminuta. The anticolitic effect of H. diminuta was associated with increased colonic IL-10 mRNA and stimulated splenocytes from H. diminuta- plus DNBS-treated mice produced more IL-10 than splenocytes from DNBS-only treated mice. Coadministration of an anti-IL-10 Ab blocked the anticolitic effect of prophylactic H. diminuta infection. Also, mice infected 48 h after DNBS treatment showed an enhanced recovery response. Finally, using a model of OVA hypersensitivity, we found no evidence of concomitant H. diminuta infection enhancing enteric responsiveness to subsequent ex vivo OVA challenge. The data show that a viable infection of H. diminuta in a nonpermissive system exerts a profound anticolitic effect (both prophylactically and as a treatment) that is mediated at least in part via IL-10 and does not predispose to enhanced sensitivity to bystander proteins.     

The effect of the expulsion phase of Trichinella spiralis on Hymenolepis diminuta infection in rats.

The effect of the intestinal changes brought about by the expulsion of Trichinella spiralis in rats was studied in relation to the growth and survival of a concurrent infection with Hymenolepis diminuta, a cestode not normally rejected by the rat in low-level infections. Growth of H. diminuta was stunted in rats given T. spiralis just before, or after, infection with H. diminuta, the stunting being more pronounced when the cestode was given closer to the period of inflammation. There was no loss of the cestode from dual-infected rats and no evidence for destrobilation was found. Lower T. spiralis burdens had a correspondingly weaker effect on growth of H. diminuta, and stunting was abolished by administration of the anti-inflammatory drug cortisone acetate. It is concluded that the stunting of H. diminuta is probably due to the non-specific inflammatory component of the rat's response to T. spiralis infection.     

Study of the effects of insulin on the migration of Hymenolepis diminuta in rats

The effects of insulin on worm (Hymenolepis diminuta) migration was studied. Insulin injection (20 U/kg, s.c.) significantly increased gastric acid output but did not affect the serotonin content of blood, intestinal lumen or worms. The drug produced, dose-dependently, posteriad migration of the worms in rats without pylorus-ligation but ligation of the pylorus prevented this migration. It is concluded that the hypersecretion of gastric acid induced by insulin is responsible for the posteriad migration of H. diminuta in rats.     

Involvement of heat shock proteins in the healing of acetic acid-induced gastric ulcers in rats.

The present study examined the expression of 73-kDa of heat shock cognate protein (HSC70), 72-kDa of heat shock protein (HSP70) and 47-kDa of HSP (HSP47) observed in the ulcer healing process in rats. Gastric ulcers were induced by a luminal application of acetic acid in male Donryu rats. During the ulcer healing process, the expression of HSPs in the ulcerated tissue was determined. A high level of HSC70 expression was observed both in the normal mucosa and ulcerated tissue, but the level did not change upon ulceration and ulcer healing. While HSP70 and HSP47 were markedly expressed in the ulcer base during ulceration, and decreased with ulcer healing. HSP70 expression in the ulcer margin was gradually increased with ulcer healing. Omeprazole accelerated the healing of gastric ulcers with strong inhibition of gastric acid secretion, while indomethactin delayed in ulcer healing despite slight inhibition of gastric acid secretion. Omperazole enhanced the expression of HSP70 both in the ulcer margin and base, but it reduced HSP47 expression in the ulcer base Indomethacin markedly enhanced HSP47 expression only in the ulcer base. In conclusion, the expression of HSP70 and HSP47 is changed during ulcer healing. Furthermore, it was suggested that the enhanced expression of HSP70 is involved in acceleration of ulcer healing, but overexpression of HSP47 is involved in delayed ulcer healing.     

Fish oil-enriched diet is mucosal protective against acetic acid-induced colitis in rats.

Products of arachidonic acid metabolism are elevated in patients with inflammatory bowel disease and this elevation is correlated with disease activity. Eicosapentaenoic acid competes with arachidonic acid and alters eicosanoid biosynthesis. In this experiment, the possibility that eicosapentaenoic acid could be used in the treatment of inflammatory bowel disease was investigated by determining the effect of 6 weeks of a fish oil-supplemented diet, enriched in eicosapentaenoic acid, on colonic and ileal morphology, histology, and in vivo fluid absorption in rats with 4% acetic acid-induced colitis. The results of an eicosapentaenoic acid-enriched diet were compared with results of saturated and polyunsaturated fatty acid-enriched diets. In rats with misoprostol pretreated acetic acid-induced colitis, an eicosapentaenoic acid-enriched diet reversed net colonic fluid secretion to absorption and prevented macroscopic and histologic injury, compared with saturated and poly-unsaturated fatty acid-enriched diets, which did not. The fish oil mucosal protective effect occurred in the presence of a 30-fold enhancement of PGE2 synthesis. In rats with non-misoprostol pretreated acetic acid-induced colitis, an eicosapentaenoic acid-enriched diet returned ileal fluid absorption to control levels, as compared with saturated and polyunsaturated fatty acid-enriched diets, which did not. In conclusion, a fish oil (eicosapentaenoic acid)-enriched diet, but not a saturated- or a polyunsaturated-enriched diet, protected colonic and ileal net fluid absorption in an experimental model of inflammatory bowel disease.     

Effect of adrenomedullin administration on acetic acid-induced colitis in rats

Adrenomedullin (AM) administered intracolonically ameliorated the severity of acetic acid-induced colonic ulceration in rats. Ulcers were induced by subserosal injection of acetic acid into the colon. AM-treated group was administered 0.25–1.0 μg of AM in 0.5 ml of saline intracolonically once a day; the control group received only saline. AM administration dose-dependently and significantly reduced the size of the ulcerative lesions, the associated edema, and the infiltration of the affected area by inflammatory cells. AM also reduced tissue levels of interleukin-6, but not interferon-γ. AM reduces the severity of acetic acid-induced colitis in rats, probably by inhibiting the production and/or release of Th-2 cell-derived factors such as interleukin-6.     

Endothelial nitric oxide synthase modulates gastric ulcer healing in rats

Nitric oxide has been shown to be beneficial for gastric ulcer healing. We determined the relative effects of endothelial and inducible nitric oxide synthases on gastric ulcer healing in rats. Ulcers were induced by serosal application of acetic acid. Ulcer severity, angiogenesis, and nitric oxide synthase expression were assessed 3-10 days later. The effects of inhibitors of nitric oxide synthase were also examined. Inducible nitric oxide synthase mRNA was only detected in ulcerated tissue (maximal at day 3), whereas the endothelial isoform mRNA was detected in normal tissue and increased during ulcer healing. Inducible nitric oxide synthase was expressed in inflammatory cells in the ulcer bed, whereas endothelial nitric oxide synthase was found in the vascular endothelium and in some mucosal cells in both normal and ulcerated tissues. Angiogenesis changed in parallel with endothelial nitric oxide synthase expression. N(6)-(iminoethyl)-L-lysine did not affect angiogenesis or ulcer healing, while N(G)-nitro-L-arginine methyl ester significantly reduced both. In conclusion, endothelial nitric oxide synthase, but not the inducible isoform, plays a significant role in gastric ulcer healing.     

Trikatu,a herbal compound that suppresses monosodium urate crystal‐induced inflammation in rats,an experimental model for acute gouty arthritis

Gout is an inflammatory joint disorder characterized by hyperuricaemia and precipitation of monosodium urate crystals in the joints. In the present study, we aimed to investigate the anti‐inflammatory effect of trikatu, a herbal compound in monosodium urate crystal‐induced inflammation in rats, an experimental model for acute gouty arthritis. Paw volume and levels/activities of lysosomal enzymes, lipid peroxidation, anti‐oxidant status and histopathological examination of ankle joints were determined in control and monosodium urate crystal‐induced rats. In addition, analgesic (acetic acid‐induced writhing response), anti‐pyretic (yeast‐induced pyrexia) and gastric ulceration effects were tested. The levels of lysosomal enzymes, lipid peroxidation and paw volume were significantly increased, and anti‐oxidant status was found to be reduced in monosodium urate crystal‐induced rats, whereas the biochemical changes were reverted to near normal levels upon trikatu (1000 mg/kg b.wt) administration. The trikatu has also been found to exhibit significant analgesic and anti‐pyretic effects with the absence of gastric damage. In conclusion, the present results clearly indicated that trikatu exert a potent anti‐inflammatory effect against monosodium urate crystal‐induced inflammation in rats in association with analgesic and anti‐pyretic effects in the absence of gastrointestinal damage. Copyright © 2013 John Wiley & Sons, Ltd.     

Bacterial colonization and healing of gastric ulcers: the effects of epidermal growth factor

Experimental gastric ulcers are rapidly colonized by various bacteria, resulting in significantly impaired healing. Epidermal growth factor (EGF) is capable of preventing bacterial colonization of the healthy intestinal mucosa. In this study, we examined the possibility that EGF accelerates gastric ulcer healing by reducing bacterial colonization of the ulcer. Gastric ulcers were induced by serosal application of acetic acid. The effect of daily administration of EGF on ulcer healing and bacterial colonization was assessed and compared with the effect of daily treatment with broad-spectrum antibiotics. EGF administration reduced colonization levels and accelerated ulcer healing as effectively as the antibiotic treatment. EGF was without effect on acid secretion or neutrophil infiltration into the ulcer. Bacterial growth was not inhibited in the presence of EGF in vitro. These results demonstrate that EGF reduces bacterial colonization during an established infection of a compromised mucosal surface. This effect may contribute to the ability of EGF to accelerate gastric ulcer healing. This effect is acid independent and not due to an anti-inflammatory effect or to direct bactericidal actions.     

Antiulcer activity of N-phthaloyl GABA--a new GABA mimetic agent

N-phthaloyl GABA (P. GABA) inhibited gastric ulceration induced by 3 hr restraint stress at 4 degrees C (CRS) in albino rats. Antiulcer activity of P. GABA was compared with sodium valproate and cimetidine. P. GABA, sodium valproate and cimetidine showed a dose dependent reduction of gastric ulceration. Pretreatment with GABA antagonists-bicuculline methiodide (0.5 mg/kg, im) or 3 mercaptopropionic acid (2 mg/kg, im) reversed the antiulcerogenic activity of both the drugs (P. GABA and sodium valproate). GABA antagonists as such did not induce gastric ulceration in normal rats.     

Effect of local injection with basic fibroblast growth factor (BFGF) and neutralizing antibody to BFGF on gastric ulcer healing, gastric secretion, angiogenesis and gastric blood flow.

Exogenous administration of bFGF was shown to accelerate tissue repair predominantly due to an increase in the formation of new microvessels (angiogenesis) suggesting that bFGF plays an important role in healing of gastric ulcer. This study was designed: 1) to examine the effect of local application of bFGF with or without neutralizing antibody (NA) to bFGF and 2) to determine the role of gastric secretion, gastric blood flow (GBF) at the ulcer margin and angiogenesis during gastric ulcer healing with or without local application of NA, bFGF or the combination of NA and bFGF. Chronic gastric ulcers were induced in Wistar rats by subserosal application of acetic acid (ulcer area 28 mm2) and gastric secretion during ulcer healing was assessed using animals additionally equipped with chronic gastric fistulas. The bFGF without or with NA to bFGF (10 ng/100 microl]), irrelevant antibodies (rabbit IgG; 10 microg/100 microl) or vehicle (saline) were locally injected into the subserosa immediately upon ulcer induction (day 0) and at day 2. Rats with acetic acid ulcers without subserosal injections served as controls. At day 11, all animals were anaesthetized and GBF was determined at the ulcer base, ulcer margin as well as in intact mucosa using the H2-gas clearance technique and the area of gastric ulcers was measured by planimetry. Gastric mucosa with ulcer was excised and the percentage of area covered with blood vessels, the number of fibroblasts and the percentage of connective tissue at the ulcer edge was assessed by histology. The gastric ulcers were healed spontaneously in control vehicle-treated rats at day 11 and this was accompanied by the significant increase in the GBF and number of microvessels in the ulcer area. The gastric secretion was suppressed immediately after ulcer induction and increased significantly at day 2 and day 11 but failed to return to that recorded in intact animals. In contrast, local application of bFGF inhibited gastric acid and pepsin outputs at each study time intervals tested and this effect was reversed by addition of NA to bFGF. Locally applied bFGF accelerated significantly ulcer healing and this was accompanied by the greater rise in the GBF of ulcer margin and more marked increase in number of microvessels as compared to those in vehicle-treated rats. Subserosal application of NA to bFGF prolonged significantly the ulcer healing and this effect was accompanied by a significant fall in the GBF at the ulcer margin and a decrease in number of capillaries in ulcer bed without significant alteration in gastric acid and pepsin outputs. The ulcer healing effect of bFGF and accompanying increase in the GBF at ulcer margin and in thenumber of microvessels as well as inhibition of gastric acid secretion evoked by bFGF were significantly attenuated by the addition of NA to bFGF. The number of fibroblasts and the distribution of connective tissue did not differ between groups studied. We conclude that; 1) depletion of endogenous bFGF at the ulcer area by specific NA to bFGF delays healing of gastric ulcers, reduces angiogenesis of ulcer bed and impairs the microcirculatory effect of this growth factor at the ulcer margin indicating that the availability of bFGF in the ulcer area plays a crucial role in the ulcer healing through induction of angiogenesis; 2) this prominent antiulcer effect of locally applied bFGF depends, at least in part, upon the inhibition of acid secretion by this peptide.     

Involvement of reactive oxygen species in gastric ulceration: protection by melatonin

Uncontrolled hydrochloric acid secretion and ulceration in the stomach due to various factors are serious global problems today. Although the mechanism of acid secretion from the parietal cell is now fairly known, the mechanism of gastric ulceration is still not clear today. Among various causes of gastric ulceration, lesions caused by stress, alcohol consumption, Helicobacter pylori infection and use of nonsteroidal antiinflammatory drugs have been shown to be mediated largely through the generation of reactive oxygen species especially hydroxyl radical (*OH). A number of excellent drugs have been proved useful in controlling hyperacidity and ulceration but their long term uses are not devoid of disturbing side-effects. Hence, the search is still on to find out a compound possessing antisecretory, antiulcer and antioxidant properties which will serve as a powerful therapeutic agent to cure gastric hyperacidity and ulcer. This article describes the role of reactive oxygen species in gastric ulceration, drugs controlling them with their merits and demerits and, the role of melatonin, a pineal hormone in protecting the gastric lesions with a final commentary on how melatonin research with respect to gastric pathophysiology can be taken forward with a view to projecting this indole as a promising therapeutic agent to control gastric ulceration in humans.     

Alterations of matrix metalloproteinases,gastric mucin and prostaglandin E2 levels by pectic polysaccharide of swallow root (Decalepis hamiltonii) during ulcer healing

Gastric ulcer is a multi-step disease caused due to imbalance between mucosal defense and aggressive factors. Available anti-ulcer drugs although effective at various steps of ulcer pathogenecity, pose adverse effects. Pectic polysaccharide (SRPP) from swallow root (Decalepis hamiltonii) – previously shown to possess ulcer preventive effect against swim stress and ethanol induced gastric ulcers. In the current study, alteration of matrix metalloproteinases, gastric mucin and prostaglandin E₂ levels during polysaccharide mediated ulcer healing was determined in acetic acid induced gastric ulcer model in Wistar albino rats. Results indicated the potential ulcer healing effect of SRPP as evidenced by ∼90% reduction in ulcer index; improvement in the antioxidant defense such as increase of glutathione levels together with significant reduction in lipid and protein oxidation and protection to damaged gastric mucin. Further, histological studies substantiated the result of the recovery of mucin that was eroded during ulceration, rejuvenation of mucosal epithelium and enhancement of high molecular mass mucin as opposed to the degraded ∼55 kDa mucin that appeared only during ulcer condition. Matrix metalloproteinases (MMPs) that are involved in tissue injury was found to be modulated by SRPP treatment in addition to increased cytoprotectivity due to enhanced synthesis of PGE₂ that necessitates the active proliferation of gastric mucin cells. Further, reduction in ∼3 folds of galectin-3, an inflammatory marker suggests gastro protection against acid induced inflammation and gastric wall damages. Overall, studies show the effectiveness of SRPP in inhibiting MMPs and galectin-3 levels which were up-regulated during ulcer conditions. In addition SRPP ensured cytoprotectivity and rejuvenation of mucosal barrier via PGE₂ trigger leading to ulcer healing.     

Study of the damaging effects of acetazolamide on gastric mucosa in rats

The present study demonstrated that acetazolamide (100 and 200 mg/kg, s.c.) induced severe gastric hemorrhagic ulceration in rats. The ulceration was aggravated by oral administration of HCl, but was inhibited by NaHCO3. Furthermore, the severity of ulceration was also decreased by pretreatment with methysergide, chlorpheniramine, or cimetidine. These protective effects were accompanied by an increase in serotonin and histamine released from the stomach. Acetazolamide injection also increased the protein level but reduced the sialic acid content in the gastric secretion, indicating that the gastric mucosal barrier may have been damaged. Prostaglandin E2 content of the gastric mucosa was not affected by the drug; however, carbonic anhydrase activity was markedly reduced in a dose-dependent manner. Thus, it is suggested that the ulceration induced by acetazolamide is mainly due to the inhibition of carbonic anhydrase activity and mucus secretion. The increase in serotonin and histamine release also may have been the contributing factors for gastric ulcer formation.     

Culture-independent analysis of indomethacin-induced alterations in the rat gastrointestinal microbiota

Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly prescribed for a variety of inflammatory conditions; however, the benefits of this class of drugs are accompanied by deleterious side effects, most commonly gastric irritation and ulceration. NSAID-induced ulceration is thought to be exacerbated by intestinal microbiota, but previous studies have not identified specific microbes that contribute to these adverse effects. In this study, we conducted a culture-independent analysis of approximately 1,400 bacterial small-subunit rRNA genes associated with the small intestines and mesenteric lymph nodes of rats treated with the NSAID indomethacin. This is the first molecular analysis of the microbiota of the rat small intestine. A comparison of clone libraries and species-specific quantitative PCR results from rats treated with indomethacin and untreated rats revealed that organisms closely related to Enterococcus faecalis were heavily enriched in the small intestine and mesenteric lymph nodes of the treated rats. These data suggest that treatment of NSAID-induced ulceration may be facilitated by addressing the microbiological imbalances.     

Comparative studies on starvation - and indomethacin - induced ulcerations in albino rats.

Experimental models of chronic and acute peptic ulcerations were produced in the albino rats by means of prolonged starvation and indomethacin administration. In the case of acute indomethacin-induced peptic ulceration, the effects of anticholinergic drugs on the ulcers produced were also studied. Starving the rats for a period of seven days produced gastric ulceration in all the rats used while indomethacin produced gastric ulceration within five hours in all the rats used. Severe ulceration of the degree found in human peptic ulcer disease was produced only by chronic starvation. Anticholinergic drugs ameliorated indomethacin-induced gastric ulceration, partly at least, by reducing intra-gastric acidity.     

The role of gastric and duodenal pH in the cysteamine-induced duodenal ulcer in the rat

Many secretory studies reported an increase in gastric acid secretion by the duodenal ulcerogen cysteamine. A detailed analysis of these experiments, especially the results from rats with chronic gastric fistula suggest that direct stimulation of gastric acid secretion may not be the primary mechanism of the duodenal ulcerogenic action of cysteamine. We used a different approach and measured the pH at the site of ulceration in the proximal duodenum. A duodenal ulcerogenic dose of cysteamine did not change the pH at the anterior or posterior wall of the duodenum during 4 hr. In the same dose and by the same route of administration, cysteamine nevertheless induced duodenal ulcers in 24 hr. These experiments demonstrate that in addition to the effect on gastric acid secretion, other factors are needed to the effect on gastric acid secretion, other factors are needed to explain the early duodenal ulcerogenic action of cysteamine.     

Characterization of the immuno-regulatory response to the tapeworm Hymenolepis diminuta in the non-permissive mouse host

Hymenolepis diminuta is spontaneously expelled from mice; concomitant with worm expulsion was protection against colitis induced by dinitrobenzene sulphonic acid (DNBS). Here we examined the immune response mobilized by Balb/c and C57Bl/6 male mice in response to H. diminuta and assessed the requirement for CD4+ cells (predominantly T cells) in worm expulsion and the anti-colitic effect. Wild-type (CD4+) or CD4 knock-out (CD4-/-) mice received five H. diminuta cysticercoids and segments of jejunum and mesenteric lymph nodes (MLNs), or spleen, were excised 5, 8 and 1l days later for mRNA analysis and cytokine production, respectively. In separate experiments uninfected and infected mice received DNBS by intra-rectal infusion and indices of inflammation were assessed 3 days later (i.e. 11 days p.i.). Infection of Balb/c mice resulted in a time-dependent increase in intestinal mRNA for Foxp3, a marker of natural regulatory T cells, and markers of alternatively activated macrophages (arginase-1, FIZZ1), while concanavalin-A activation of MLN cells revealed a significant increase in T helper 2 (TH2) type cytokines: IL-4, -5, -9, -10, -13. MLN cells showed a reduced ability to induce Foxp3 expression upon stimulation. CD4-/- mice did not display this response to infection, but surprisingly did expel H. diminuta. Moreover, DNBS-induced colitis in CD4-/- mice (wasting, tissue damage, elevated myeloperoxidase) was not reduced by H. diminuta infection, whereas time-matched infected CD4+ C57Bl/6 mice had significantly less DNBS-induced inflammation. In conclusion: (i) in addition to stereotypical TH2 events, H. diminuta-infected Balb/c mice develop a local immuno-regulatory response; and (ii) CD4+ cells are not essential for H. diminuta expulsion from mice but are critical in mediating the anti-colitic effect that accompanies infection in this model.