Rory W. Blackler

Gastrointestinal-Sparing Effects of Novel NSAIDs in Rats with Compromised Mucosal Defence

Nonsteroidal anti-inflammatory drugs are among the most commonly used prescription and over-the-counter medications, but they often produce significant gastrointestinal ulceration and bleeding, particularly in elderly patients and patients with certain co-morbidities. Novel anti-inflammatory drugs are seldom tested in animal models that mimic the high risk human users, leading to an underestimate of the true toxicity of the drugs. In the present study we examined the effects of two novel NSAIDs and two commonly used NSAIDs in models in which mucosal defence was expected to be impaired. Naproxen, celecoxib, ATB-346 (a hydrogen sulfide- and naproxen-releasing compound) and NCX 429 (a nitric oxide- and naproxen-releasing compound) were evaluated in healthy, arthritic, obese, and hypertensive rats and in rats of advanced age (19 months) and rats co-administered low-dose aspirin and/or omeprazole. In all models except hypertension, greater gastric and/or intestinal damage was observed when naproxen was administered in these models than in healthy rats. Celecoxib-induced damage was significantly increased when co-administered with low-dose aspirin and/or omeprazole. In contrast, ATB-346 and NCX 429, when tested at doses that were as effective as naproxen and celecoxib in reducing inflammation and inhibiting cyclooxygenase activity, did not produce significant gastric or intestinal damage in any of the models. These results demonstrate that animal models of human co-morbidities display the same increased susceptibility to NSAID-induced gastrointestinal damage as observed in humans. Moreover, two novel NSAIDs that release mediators of mucosal defence (hydrogen sulfide and nitric oxide) do not induce significant gastrointestinal damage in these models of impaired mucosal defence.

Hydrogen Sulfide Protects from Colitis and Restores Intestinal Microbiota Biofilm and Mucus Production

Background:Microbiota dysbiosis and impaired barrier function are among the most prominent features of inflammatory bowel disease. In the gastrointestinal tract, hydrogen sulfide (H2S) is an important regulator of mucosal homeostasis. We hypothesized that H2S promotes resolution of colonic inflammation through actions on microbiota biofilm and the mucus barrier. Methods:We used mice genetically deficient for a key enzyme for H2S production (cystathionine &ggr;-lyase) and pharmacologically inhibited that enzyme during colitis in wild-type mice. We tested the effects of administering an H2S donor (diallyl disulfide) to rodents during hapten-induced colitis. Colonic microbiota biofilm was visualized by fluorescent in situ hybridization, and mucus granules were quantified with periodic acid–alcian blue staining. We exposed human microbiota biofilms and planktonic bacteria to H2S donors ex vivo to determine changes in their growth, viability, and biomass. Results:Intestinal microbiota formed linear biofilms in the colon of healthy rodents. During colitis, microbiota biofilms were fragmented and mucus granule production decreased. Endogenous production of H2S had beneficial effects on establishment of microbiota biofilms and colonic mucus production. Therapeutic delivery of H2S into the colon reduced inflammation, restored the microbiota biofilm, and increased the production of mucus granules. In ex vivo human microbiota, H2S not only promoted biofilm formation but also reduced growth of planktonic bacteria. Conclusions:Our results suggest that H2S donors could be used therapeutically during colitis, facilitating correction of microbiota biofilm dysbiosis and mucus layer reconstitution.

Impaired hydrogen sulfide synthesis and IL-10 signaling underlie hyperhomocysteinemia-associated exacerbation of colitis.

Significance Inflammatory bowel diseases (IBDs) are debilitating conditions with no known cure. Recent evidence suggests that elevated intestinal hydrogen sulfide (H2S) synthesis promotes healing and reduces inflammation. H2S is synthesized from cysteine largely via vitamin B6-dependent enzymes. People with IBD are also at increased risk of hyperhomocysteinemia, a condition that is often caused by vitamin B deficiency. Dietary induction of vitamin B deficiency markedly increased serum homocysteine levels and worsened colitis in rodents. The latter was due to the absence of the typical injury-induced elevation of H2S synthesis. Interleukin-10 plays a key role in increasing H2S synthesis, attenuating the severity of colitis, and reducing serum homocysteine levels. The H2S–interleukin 10 axis may be a viable target for therapy of IBD. Vitamin B deficiencies, which can lead to hyperhomocysteinemia (Hhcy), are commonly reported in patients with inflammatory bowel disease (IBD) and may be a causative underlying factor. However, the mechanism for this effect is not known. Hydrogen sulfide (H2S) is a gaseous mediator that promotes tissue repair and resolution of inflammation. In experimental colitis, a marked increase in colonic H2S synthesis drives ulcer healing and resolution of inflammation. Because H2S synthesis is in part dependent upon enzymes that require vitamin B6 as a cofactor, we tested the hypothesis that Hhcy in rodent models would increase the susceptibility to colitis. In all three models tested, diet-induced Hhcy significantly exacerbated colitis. The usual elevation of colonic H2S synthesis after induction of colitis was absent in all three models of colitis. Administration of an H2S donor to Hhcy rats significantly decreased the severity of colitis. Compared with wild-type mice, interleukin (IL) 10-deficient mice on a normal diet had decreased levels of colonic H2S synthesis, a 40% increase in serum homocysteine, and a phenotype similar to wild-type mice with Hhcy. IL-10–deficient mice fed the vitamin B-deficient diet exhibited more severe colonic inflammation, but the normal elevation of colonic H2S synthesis was absent. Administration of IL-10 to the IL-10–deficient mice restored colonic H2S synthesis and significantly decreased serum homocysteine levels. These results suggest that the exacerbation of colitis in Hhcy is due in part to impaired colonic H2S synthesis. Moreover, IL-10 plays a novel role in promoting H2S production and homocysteine metabolism, which may have therapeutic value in conditions characterized by Hhcy.

NSAID-gastroenteropathy: new aspects of pathogenesis and prevention

Nonsteroidal anti-inflammatory drugs (NSAIDs) remain among the most commonly used medications because of their effectiveness in reducing pain and inflammation. Inhibitors of gastric acid secretion can substantially reduce the damaging effects of NSAIDs in the stomach and duodenum. However, there are no proven effective preventative or curative treatments for NSAID-induced enteropathy. In recent years, substantial progress has been made in better understanding the pathogenesis of NSAID-enteropathy, and in particular the interplay of enteric bacteria, bile and the enterohepatic recirculation of the NSAIDs. Moreover, it is becoming clear that suppression of gastric acid secretion significantly worsens NSAID-enteropathy.

Enhanced chemopreventive effects of a hydrogen sulfide-releasing anti-inflammatory drug (ATB-346) in experimental colorectal cancer

Regular use of nonsteroidal anti-inflammatory drugs is associated with a significantly lower incidence of several types of cancer, particularly those affecting the gastrointestinal tract. However, the propensity of these drugs to cause ulcers and bleeding in the stomach and small intestine limits their utility for chemoprevention of cancer. In the present study, we evaluated the effectiveness of a novel hydrogen sulfide-releasing derivative of naproxen in reducing the incidence of pre-cancerous lesions (aberrant crypt foci) in mice treated with the carcinogen azoxymethane. Weekly administration of azoxymethane over a 4-week period resulted in formation of an average of ∼50 aberrant crypt foci in the colon. Twice-daily treatment with naproxen at high doses significantly reduced the number of aberrant crypt foci. However, a significantly greater effect was observed with ATB-346 (H2S-releasing naproxen) and it was also effective at much lower doses, where naproxen was ineffective. The H2S-releasing moiety of ATB-346 did not significantly affect the number of aberrant crypt foci, suggesting that both the inhibition of cyclooxygenase activity and release of H2S were necessary for the enhanced chemopreventative effect. ATB-346 suppressed colonic prostaglandin synthesis and whole blood thromboxane synthesis as effectively as naproxen, but did not induce any gastrointestinal injury. These results demonstrate that ATB-346 exerts superior chemopreventive effects to those of naproxen, while sparing the gastrointestinal tract of the injury normally associated with use of the parent drug. ATB-346 may therefore be an attractive agent for chemoprevention of colon cancer, and possibly of cancers in other tissues.

Hydrogen sulphide protects against NSAID-enteropathy through modulation of bile and the microbiota.

Hydrogen sulphide is an important mediator of gastrointestinal mucosal defence. The use of non‐steroidal anti‐inflammatory drugs (NSAIDs) is significantly limited by their toxicity in the gastrointestinal tract. Particularly concerning is the lack of effective preventative or curative treatments for NSAID‐induced intestinal damage and bleeding. We evaluated the ability of a hydrogen sulphide donor to protect against NSAID‐induced enteropathy.

NSAID enteropathy and bacteria: a complicated relationship

The clinical significance of small intestinal damage caused by nonsteroidal anti-inflammatory drugs (NSAIDs) remains under-appreciated. It occurs with greater frequency than the damage caused by these drugs in the upper gastrointestinal tract, but is much more difficult to diagnose and treat. Although the pathogenesis of NSAID enteropathy remains incompletely understood, it is clear that bacteria, bile, and the enterohepatic circulation of NSAIDs are all important factors. However, they are also interrelated with one another. Bacterial enzymes can affect the cytotoxicity of bile and are essential for enterohepatic circulation of NSAIDs. Gram-negative bacteria appear to be particularly important in the pathogenesis of NSAID enteropathy, possibly through release of endotoxin. Inhibitors of gastric acid secretion significantly aggravate NSAID enteropathy, and this effect is due to significant changes in the intestinal microbiome. Treatment with antibiotics can, in some circumstances, reduce the severity of NSAID enteropathy, but published results are inconsistent. Specific antibiotic-induced changes in the microbiota have not been causally linked to prevention of intestinal damage. Treatment with probiotics, particularly Bifidobacterium, Lactobacillus, and Faecalibacteriaum prausnitzii, has shown promising effects in animal models. Our studies suggest that these beneficial effects are due to colonization by the bacteria, rather than to products released by the bacteria.

Acetylsalicylic Acid Daily vs Acetylsalicylic Acid Every 3 Days in Healthy Volunteers: Effect on Platelet Aggregation, Gastric Mucosa, and Prostaglandin E2 Synthesis

Substantial platelet inhibition was observed 3 days after a single administration of acetylsalicylic acid 81 mg to healthy volunteers. Here we investigate prostaglandin E2 (PGE2) antrum concentrations and gastrointestinal symptoms in two treatment groups: one receiving losartan and acetylsalicylic acid every day and the other receiving losartan every day and acetylsalicylic acid every 3 days. Twenty‐eight healthy volunteers from both sexes received either 50 mg losartan and acetylsalicylic acid 81 mg daily or 50 mg losartan and acetylsalicylic acid 81 every 3 days with placebo on the other days. Therapy was delivered for 30 days for both groups. Gastric endoscopy was performed before and after treatment period. Biopsies were collected for PGE2 quantification. Platelet function tests were carried out before and during treatment and TXB2 release on platelet rich plasma was measured. The every 3 day low‐dose acetylsalicylic acid regimen produced complete inhibition of platelet aggregation compared to the daily treatment. Thromboxane B2 release was substantially abolished for both groups during treatment. There was no significant difference on the endoscopic score of both treatment groups after the 30‐day treatment (P = .215). There was over 50% suppression of antrum PGE2 content on volunteers receiving acetylsalicylic acid daily (P = .0016), while for the every 3 day dose regimen there was no significant difference between pre and post‐treatment antrum PGE2 dosages (P = .4193). Since PGE2 is involved in gastric healing, we understand that this new approach could be safer and as efficient as the standard daily therapy on a long‐term basis.