Antibiotic Therapy for Active Crohn's Disease Targeting Pathogens: An Overview and Update.

Crohn's disease (CD) is a multifactorial chronic disorder that involves a combination of factors, including genetics, immune response, and gut microbiota. Therapy includes salicylates, immunosuppressive agents, corticosteroids, and biologic drugs. International guidelines do not recommend the use of antibiotics for CD patients, except in the case of septic complications. Increasing evidence of the involvement of gut bacteria in this chronic disease supports the rationale for using antibiotics as the primary treatment for active CD. In recent decades, several pathogens have been reported to be involved in the development of CD, but only Escherichia coli (E. coli) and Mycobacterium avium paratubercolosis (MAP) have aroused interest due to their strong association with CD pathogenesis. Several meta-analyses have been published concerning antibiotic treatment for CD patients, but randomized trials testing antibiotic treatment against E. coli and MAP have not shown prolonged benefits and have generated conflicting results; several questions are still unresolved regarding trial design, antibiotic dosing, the formulation used, the treatment course, and the outcome measures. In this paper, we provide an overview and update of the trials testing antibiotic treatment for active CD patients, taking into account the role of pathogens, the mechanisms by which different antibiotics act on harmful pathogens, and antibiotic resistance. Finally, we also present new lines of study for the future regarding the use of antibiotics to treat patients with active CD.

Pathogens in Crohn's Disease: The Role of Adherent Invasive Escherichia coli.

In Crohn's disease (CD), gut dysbiosis is marked by the prevalence of pathogenic bacterial species. Although several microbes have been reported as risk factors or causative agents of CD, it is not yet clear which is the real trigger of the disease. Thirty years ago, a new pathovar of Escherichia coli strain was isolated in the ileal mucosa of CD patients. This strain, called adherent invasive E. coli (AIEC), for its ability to invade the intestinal mucosa, could represent the causative agent of the disease. Several authors studied the mechanisms by which the AIEC penetrate and replicate within macrophages, and release inflammatory cytokines sustaining inflammation. In this review we will discuss about the role of AIEC in the pathogenesis of CD, the virulence factors mediating adhesion and invasion of AIEC in mucosal tissue, the environmental conditions improving AIEC survival and replication within macrophages. Finally, we will also give an overview of the new strategies developed to limit AIEC overgrowth.

Cell-type-specific gene expression profile by laser capture microdissection on mirror sections.

Immuno-laser capture microdissection (Immuno-LCM) has been used to analyze cell-specific gene expression profiles. However, the usefulness of such a technique is frequently limited by RNA degradation. We, therefore, developed a rapid protocol of LCM on mirror sections, which allows for preserving RNA integrity. With such a procedure, we investigated cell-type-specific gene expression of γδ intraepithelial lymphocytes (IELs) in untreated celiac disease (CD). An increase in TGF-ß mRNA expression levels was observed in γδ + IELs compared to intestinal enterocytes (IEs), whereas anti-inflammatory IL-10 mRNA production from γδ + IELs was lower compared to IEs. In untreated CD patients, the production of anti-inflammatory cytokines by γδ + IELs is suggestive of a regulatory function, thus playing a critical role in limiting inflammation. This work underscores the importance of LCM on mirror sections as a valuable tool to perform cell-type-specific molecular analysis in tissue.

Adult autoimmune enteropathy in autoimmune hepatitis patient. Case report and literature review.

Autoimmune enteropathy (AIE) is a rare disease characterized by prolonged diarrhea, vomiting and weight loss; although it is mainly a rare pediatric disease, over the years a number of adults have also been found to be affected. In this study, we present a case report of a 73-year-old woman with a history of autoimmune hepatitis, antinuclear (ANA) and positive anti-enterocyte antibodies (AEA), who has suffered two months of intractable diarrhea, nausea, anorexia and severe weight loss. The histological examination of the endoscopic duodenal mucosa biopsies revealed severe shortening and flattening of the villi, resulting in mucosal atrophy. The immunohistochemical study revealed a polymorphic lymphoid population, exhibiting a B cell (CD20+) phenotype in follicles and a T cell phenotype (CD3+) in the diffuse component within the lamina propria. Our patient had a complete recovery after two weeks of taking prednisone and following a gluten-rich diet. To our knowledge this is the first case of autoimmune enteropathy in adults with ANA and AEA 7 years after a diagnosis of autoimmune hepatitis. To date, the patient is still in clinical remission on a low dose of orally administered predinisone without any additional immunosuppression.

Molecular Mechanisms of Helicobacter pylori Pathogenesis.

Helicobacter pylori infects 50% of mankind. The vast majority of H. pylori infection occurs in the developing countries where up to 80% of the middle-aged adults may be infected. Bacterial infection causes an inflammatory response that proceeds through a series of intermediated stages of precancerous lesions (gastritis, atrophy, intestinal metaplasia, and dysplasia). Among infected individuals, approximately 10% develops severe gastric lesions such as peptic ulcer disease, 1-3% progresses to gastric cancer (GC) with a low 5-year survival rate, and 0.1% develops mucosa-associated lymphoid tissue (MALT). GC is one of the most common cancer and the third leading cause of cancer-related deaths worldwide. In this review, we have summarized the most recent papers about molecular mechanisms of H. pylori pathogenesis. The main important steps of H. pylori infection such as adhesion, entry in epithelial gastric cells, activation of intracellular pathways until epigenetic modifications have been described.

The Helicobacter pylori protein HspB interferes with Nrf2/Keap1 pathway altering the antioxidant response of Ags cells.

BACKGROUND: Helicobacter pylori infection causes chronic oxidative stress on gastric mucosa, thereby causing mucosal damage and increasing the risk of gastric adenocarcinoma. Nrf2 is an important transcription factor, regulating the antioxidant response in the cells. Nrf2 signaling is repressed by Keap1 at basal condition and induced by oxidative stress. The aim of our study was to analyze whether the H. pylori proteins interfered in the Nrf2/Keap1 pathway. MATERIAL AND METHODS: Gene expression in AGS cells transiently and stably transfected was analyzed by real-time PCR. Immunoprecipitation and immunofluorescence assays were performed to investigate the ability of H. pylori proteins to interfere with the Nrf2 pathway. RESULTS: We demonstrated that the H. pylori HspB protein interferes with Nrf2/Keap1 pathway. When HspB was transiently transfected in AGS cells, a significant increase in Keap1 gene expression was induced. The same result was observed when AGS cells were HspB stably transfected. In this case, the increase in Keap1 was associated with reduced gene expression of Nrf2, and of the antioxidant enzymes superoxide dismutase, hemeoxygenase-1, and phase II detoxifying enzyme NAD(P)H:quinone oxidoreductase-1. Immunoprecipitation and immunofluorescence assays confirmed the ability of HspB protein to interfere with the Nrf2 pathway. Lastly, in HspB-transfected AGS cells, sustained activation of IL-8, COX2, MMP3, and MMP7 was demonstrated. CONCLUSION: The results here reported suggest that inhibited nuclear translocation of Nrf2, associated with induced inflammation and increased production of MMPs, might represent a condition enhancing the risk of gastric adenocarcinoma.

Capsule endoscopy is useful and safe for small-bowel surveillance in familial adenomatous polyposis.

BACKGROUND: Duodenal cancer and ampullary cancer are major causes of death after a prophylactic colectomy in patients with familial adenomatous polyposis (FAP). Forward-viewing endoscopy and side-viewing endoscopy are recommended in patients with FAP for surveillance of periampullary and duodenal polyposis. The study of polyps distal to the duodenum in FAP is limited. A capsule endoscopy (CE) allows visualization of the mucosa of the entire small bowel. OBJECTIVE: The objective was to detect whether CE has clinical value or any utility for the surveillance of small-bowel polyps in patients with FAP and to evaluate whether there are genotypic factors that predict which patients are at a lower risk of small-bowel polyps. SETTING: Two Italian tertiary-referral centers. PATIENTS: Twenty-three patients with FAP who presented for a CE. MAIN OUTCOME MEASUREMENTS: Patients with FAP were examined by CE to assess the location, size, and number of small-bowel polyps. Patient age at CE, sex, years of observation after surgery, type of surgery, duodenal adenomas, and colorectal cancer at surgery were analyzed. All patients were selected for mutation analysis, and the germline adenomatous polyposis coli (APC) gene mutation was detected. RESULTS: Eleven of 23 patients with FAP had duodenal polyps. During CE, jejunal-ileal polyps were detected in 7 of 23 FAPs, with a total number of 15 polyps in the ileum. The presence of duodenal adenomas was the only clinical feature predictive of small-bowel polyps. Identification of the ampulla of Vater was not achieved with CE; duodenal polyps were only seen in 4 of 11 patients identified endoscopically, with an underestimation of polyp numbers. APC mutations between codons 499 and 805 were associated with the absence of small-bowel polyps. CONCLUSIONS: CE is useful and safe for the surveillance of jejunal-ileal polyps in selected patients with FAP. CE is not useful in the surveillance of the duodenum where the majority of small-bowel cancers occur.

Effects of Helicobacter pylori infection on cell cycle progression and the expression of cell cycle regulatory proteins.

Helicobacter pylori lives in the stomach lumen adhering and specifically interacting with gastric epithelial cells. H. pylori infection can cause a broad range of diseases. Although most infected individuals only develop a chronic inflammation of the stomach, some patients progress to chronic gastritis, duodenal ulceration, or, rarely, cancer. H. pylori is able to send and to receive signals from the gastric epithelium, allowing host and bacteria to become linked in a dynamic equilibrium. Several studies have demonstrated that H. pylori infection induces morphological changes of gastric epithelial cells other than cell proliferation, increase of mitosis and mutations. It has also been demonstrated that H. pylori may predispose to cancer by altering gastric epithelial cell turnover acting specifically on transcription factors. Although H. pylori is able to induce several host responses, it specifically perturbs the delicate balance of those factors that usually help to maintain cell homeostasis. The study of mechanisms of interaction between the bacterium and gastric cells will surely help to prevent the increase and diffusion of malignancies all over the world.

Coexpression of Helicobacter pylori's proteins CagA and HspB induces cell proliferation in AGS gastric epithelial cells, independently from the bacterial infection.

Adenocarcinoma of the stomach is the second most common cause of cancer mortality in the world. The purpose of this study was to evaluate the potential role in carcinogenesis of two secreted Helicobacter pylori's proteins, CagA and HspB, both shown to increase the risk of gastric carcinoma in patients infected with H. pylori-positive strain. The effects of these two proteins on cell kinetics and the ability to selectively affect the expression of cell cycle-related proteins by transfection of a human gastric epithelial cell line (AGS) were analyzed. Using a genomic library of H. pylori, we isolated and cloned CagA and HspB. The effects of the overexpression of these proteins on cell growth were analyzed in AGS cells by immunoblots, proliferation assay, and flow cytometry. Coexpression of CagA and HspB in AGS cells in the first 48 h caused an increase of the level of E2F transcription factor, cyclin D3, and phosphorylated retinoblastoma protein, all involved in the G(1)-S checkpoint of the cell cycle. Consistently, an increase of cell proliferation, corresponding to an augment of the fraction of the cells in the S-G(2)-M phase of the cell cycle, was also demonstrated. Moreover, an increase of c-jun protein levels, but not of c-fos, was also found after coexpression of CagA and HspB. All these data suggest that CagA and HspB, independently from the bacterial infection, have a direct effect on the cell growth of the gastric cells acting on the G(1)-S checkpoint of the cell cycle.