Olivia Morelli

NCX-1000, a NO-releasing derivative of ursodeoxycholic acid, selectively delivers NO to the liver and protects against development of portal hypertension

Portal hypertension resulting from increased intrahepatic resistance is a common complication of chronic liver diseases and a leading cause of death in patients with liver cirrhosis, a scarring process of the liver that includes components of both increased fibrogenesis and wound contraction. A reduced production of nitric oxide (NO) resulting from an impaired enzymatic function of endothelial NO synthase and an increased contraction of hepatic stellate cells (HSCs) have been demonstrated to contribute to high intrahepatic resistance in the cirrhotic liver. 2-(Acetyloxy) benzoic acid 3-(nitrooxymethyl) phenyl ester (NCX-1000) is a chemical entity obtained by adding an NO-releasing moiety to ursodeoxycholic acid (UDCA), a compound that is selectively metabolized by hepatocytes. In this study we have examined the effect of NCX-1000 and UDCA on liver fibrosis and portal hypertension induced by i.p. injection of carbon tetrachloride in rats. Our results demonstrated that although both treatments reduced liver collagen deposition, NCX-1000, but not UDCA, prevented ascite formation and reduced intrahepatic resistance in carbon tetrachloride-treated rats as measured by assessing portal perfusion pressure. In contrast to UDCA, NCX-1000 inhibited HSC contraction and exerted a relaxing effect similar to the NO donor S-nitroso-N-acetylpenicillamine. HSCs were able to metabolize NCX-1000 and release nitrite/nitrate in cell supernatants. In aggregate these data indicate that NCX-1000, releasing NO into the liver microcirculation, may provide a novel therapy for the treatment of patients with portal hypertension.

NO-Aspirin Protects From T Cell-Mediated Liver Injury by Inhibiting Caspase-Dependent Processing of Th1-like Cytokines

BACKGROUND & AIMS Concanavalin A (con A)-induced hepatitis is an immunomediated disease in which assembly of CD4(+) T cells and T helper (Th)1-like cytokines causes Fas-mediated liver cell death. Nitric oxide (NO) modulates Th1 response in vitro. NCX-4016 is an NO-aspirin derivative that spares the gastrointestinal tract and shares molecular targets with NO. The aim of this study was to investigate whether this NO-aspirin modulates Th1-like response induced by con A. METHODS BALB/c mice were injected with 0.3 mg con A per mouse alone or in combination with NO-aspirin (18-100 mg/kg) or aspirin (10-55 mg/kg). RESULTS NO-aspirin, but not aspirin, caused a dose-dependent protection against liver damage induced by con A. At a dose of 100 mg/kg, NO-aspirin caused a 40%-80% reduction of interleukin (IL)-1beta, IL-12, IL-18, interferon (IFN)-gamma, and tumor necrosis factor alpha production without affecting cytokine messenger RNA expression. NO-aspirin prevented Fas, Fas ligand, and IL-2 receptor up-regulation on spleen lymphocytes and Fas ligand on hepatocytes and caused the S-nitrosylation/inhibition of IL-1beta-converting enzyme-like cysteine proteases (caspases) involved in the processing and maturation of IL-1beta and IL-18. IL-18 immunoneutralization prevented IFN-gamma release and protected from liver injury induced by con A. In contrast to a selective caspase 1 inhibitor, zVAD.FMK, a pancaspase inhibitor, prevented IFN-gamma release and protected the liver from injury. CONCLUSIONS Th1-like response induced by con A is mediated by IL-18 and requires activation of multiple caspases. NCX-4016 causes the S-nitrosylation/inhibition of caspases involved in cytokine production. Inhibition of Th1-like response is a new anti-inflammatory mechanism of action of NO-aspirin.

Nitric oxide-releasing NSAIDs inhibit interleukin-1beta converting enzyme-like cysteine proteases and protect endothelial cells from apoptosis induced by TNFalpha.

: Nitric oxide (NO)‐releasing NSAIDs are a new class of NSAID derivatives with markedly reduced gastrointestinal toxicity. Although it has been demonstrated that NO‐NSAIDs spare gastric mucosal blood flow, molecular determinants involved in this effect are unknown.

TNFalpha processing enzyme inhibitors prevent aspirin-induced TNFalpha release and protect against gastric mucosal injury in rats.

Although previous studies indicate that prevention of tumour necrosis factor α (TNFα) release protects against NSAID‐induced gastric mucosal injury, intracellular pathways by which aspirin causes TNFα release are unknown. TNFα is synthesized as a precursor which is proteolytically cleaved by a specific converting enzyme, TACE, to release the mature cytokine. TACE inhibitors prevent TNFα release and protect against TNFα‐mediated disease.

Twenty-four-hour manometric study of colonic propulsive activity in patients with diarrhea due to inflammatory (ulcerative colitis) and non-inflammatory (irritable bowel syndrome) conditions.

BackgroundLittle is known concerning colonic motility and almost nothing is known concerning propulsive activity in pathological conditions characterized by diarrhea of both inflammatory and non-inflammatory origin.AimsThe purpose of the present study was to investigate colonic propulsive activity in ulcerative colitis and diarrhea-predominant irritable bowel syndrome (IBS) patients.Patients and methodsSeven patients with active, moderate ulcerative colitis and nine diarrhea-predominant IBS patients entered the study. Sixteen healthy volunteers were recruited as a control group. In all subjects, 24-h colonic motility was recorded by a colonoscopically positioned manometric catheter. Both high- (mass movements) and low-amplitude propagated contractions were analyzed.ResultsHigh-amplitude propagated contractions were significantly increased in ulcerative colitis with respect to controls; no significant differences were found with respect to IBS patients, and between IBS and controls. Concerning amplitude, no significant difference was found between groups, although IBS patients approached the statistical difference with respect to controls. Low-amplitude propagated contractions were significantly increased in ulcerative colitis with respect to controls; no significant differences were found compared with IBS patients. The latter, however, displayed a trend toward an increase with respect to controls that approached but did not reach statistical significance.ConclusionsBoth inflammatory and non-inflammatory diarrheal conditions are characterized by an overall increase of colonic propulsive activity. This observation may be useful for a better understanding of the pathophysiologic mechanisms of these disorders.

Small intestinal bacterial overgrowth and warfarin dose requirement variability

The dose of warfarin needed to obtain a therapeutic anticoagulation level varies widely among patients and can undergo abrupt changes for unknown reasons. Drug interactions and genetic factors may partially explain these differences. Intestinal flora produces vitamin K2 (VK2) and patients with small intestinal bacterial overgrowth (SIBO) rarely present reduced INR values due to insufficient dietary vitamin K. The present study was undertaken to investigate whether SIBO occurrence may affect warfarin dose requirements in anticoagulated patients. Based on their mean weekly dose of warfarin while on stable anticoagulation, 3 groups of 10 patients each were defined: low dose (LD, <or=17.5 mg/wk of warfarin); high dose (HD, from 35-70 mg/wk); and very high dose (VHD>or=70 mg/wk). Each patient underwent a lactulose breath test to diagnose SIBO. Plasma levels of warfarin and vitamin K-analogues were also assessed. Patients with an altered breath test were 50% in the VHD group, 10% in the HD group, and none in the LD group (P=0.01). Predisposing factors to SIBO were more frequent in the VHD group, while warfarin interfering variables were not. VHD patients were younger and had a higher plasma vitamin K1 (VK1) concentration (P>0.05). On the contrary, the plasma VK2 levels tended to be lower. This pilot study suggests that SIBO may increase a patients warfarin dose requirement by increasing dietary VK1 absorption through the potentially damaged intestinal mucosa rather than increasing intestinal VK2 biosynthesis. Larger studies are needed to confirm these preliminary data and to evaluate the effects of SIBO decontamination on warfarin dosage.

Ultrasonographic assessment of colonic wall in moderate–severe ulcerative colitis: Comparison with endoscopic findings

BACKGROUND Bowel ultrasound has been shown to be a useful tool to evaluate patients with inflammatory bowel disease, especially Crohns disease. However, such data are still scarce in ulcerative colitis patients. AIMS To establish the value of bowel ultrasound in moderate to severe ulcerative colitis patients, and compare these data with endoscopic findings. PATIENTS AND METHODS Endoscopic, ultrasound and C-reactive protein data from 51 patients with moderate to severe ulcerative colitis observed during a 3-year period were retrospectively obtained and analysed. RESULTS All patients displayed pathological thickness (>4 mm) of the colon wall. This value strongly correlated with C-reactive protein values (p=0.0001) and the endoscopic score (p<0.0001). Also, a strong correlation (p<0.0001) was found between CRP values and endoscopic score. CONCLUSIONS Bowel ultrasound, in expert hands, may represent a useful adjunctive (or first line) tool for the evaluation of patients with moderate to severe ulcerative colitis.

Acute appendicitis due to Cytomegalovirus in an apparently immunocompetent patient: a case report

IntroductionIn healthy subjects, Cytomegalovirus infection can be asymptomatic or manifest as mononucleosis syndrome, but organ disease has also been reported. However, in immunocompromised patients this infection can lead to its most significant and severe disease and even mortality. When Cytomegalovirus causes a gastrointestinal tract infection, it more commonly manifests with luminal tract disease and is usually characterized by ulcerative lesions. Appendicitis is a rare manifestation, and has been reported mainly in human immunodeficiency virus-infected patients or patients with other causes of immunocompromise.Case presentationThe authors report on a case of acute primary Cytomegalovirus infection complicated with acute appendicitis due to Cytomegalovirus in an apparently immunocompetent 24-year-old Caucasian man also suffering from primary sclerosing cholangitis and ulcerative colitis. Diagnosis was based on clinical manifestations, serology results, as well as microbiological and histological findings. Treatment consisted of surgery and anti-Cytomegalovirus therapy.ConclusionsCytomegalovirus should be included among the etiologic agents of acute appendicitis in patients with primary sclerosing cholangitis and ulcerative colitis. Currently, there are no definitive data regarding the frequency of Cytomegalovirus appendicitis and the role of anti-Cytomegalovirus treatment in human immunodeficiency virus-negative and apparently immunocompetent subjects.

Prevention of gastric damage by no-releasing aspirin (NCX-4016) is mediated by inhibition of ICE/-like proteases in gastric mucosa and endothelial cells

Background. Tumor necrosis factor-ct (TNFtx), a proinflammatory cytokine, is released in vivo and in vitro after exposure to non-steroidal anti-inflammatory drugs (NSAIDs). TNFct receptor binding leads to activation of multiple interleukin-1 converting enzyme (ICE)-like proteases (caspases) and endothelial cell death. Endothelial cell dysfunction is an early step in the process of NSAID-gastropathy. Nilric oxide(NO)-releasing NSAIDs, are a new class of NSAID derivatives with markedly reduced gastrointestinal toxicity. NO-releasing agents (sodium nitroprusside [SNP] and L-Arginine) prevent TNF-induced ICE/caspase 1 activation in human endothelial ceils (HUVECs). Aims. To assess whether NO-NSAIDs modulate ICE-like proteases in vivo and in vitro and protect endothelial cells from TNFct-induced apoptosis. Methods. Macrophages, prepared from mouse spleen, were used to investigate TNFet release in vitro. Gastric damage was induced in rat by oral administration of 150 mg/kg aspirin or an equimolar dose (249 mg/kg) of NCX-4016 (NicOx, Milan, Italy). ICE-like activity was measured in cell Iysates and gastric homogenates using YVAD.AMC as substrate in a fluorimetric assay. HUVEC apoptosis was assessed by FACS-scan analysis. Results. In vitro studies demonstrated that aspirin, but not NCX-4016, released TNFct from mouse macrophages. Co-incubation with 1-100 laM SNP resulted in a concentrationdependent inhibition of aspirin-induced TNFct-release. In viw) aspirin administration caused a time-dependent increase in gastric mucosal damage and gastric mucosa ICE/caspase activity. Pretreating rats with 0.1 mM L-Arginine, SNP and Z.VAD.fmk, a pan-ICE/caspase inhibitor, prevented both gastric mucosal damage and ICE/caspase upregulation induced by aspirin. No gastric damage was detectable with any dose of NCX-4016 tested. Like aspirin NCX-4016 was largely metabolized to salicylate. In contrast to aspirin NCX-416 significantly increased plasma nitrite/nitrate concentrations and inhibited the aspirin-induced upregulation of ICE/caspase activity into the gastric mucosa. Although aspirin had no effect on HUVEC apoptosis induced by TNFcq NCX-4016 and SNP caused a concentrationdependent inhibition of apoptosis and prevented ICEqike pmteases activation as demonstrated by Westem blot analysis and fluorimetric assay. NCX-4016 inhibited IL-113 release from endotoxin-stimulated macrophages. Conclusions. 1) Activation of ICE/caspase pmteases is a key step in the process of NSAID-gastropathy; 2) NCX-4016 spares the gastric mucosa and protects endothelial cells from TNFetinduced apoptosis. The potential of NO-releasing NSAIDs to modulate ICE-like pmteases and IL-113 release might help to explain the anti-inflammatory activity of these compounds.

A simplified method for anal ultrasonography: preliminary report.

Background Anal endosonography (AES) is able to reliably visualize and identify anal sphincter abnormalities. However, dedicated probes are quite expensive. Aim We describe a simple and less costly method to perform AES in a unit that already has echoendoscopes available by inserting the endoscope through a disposable anoscope filled with standard ultrasound gel. Patients Subjects without anal abnormalities and patients with anal disease (abscesses, fistulas) were evaluated. Results Good-quality images were obtained in both controls and patients, with optimal visualization of the anatomic structures and pathologic features. The latter (abscesses, fistulas) were always confirmed by magnetic resonance imaging. Conclusions This simple and less costly method allows to perform good-quality AES in units having echoendoscopes availability, without the need of a more expensive dedicated probe.