Manuel Alvarez-Lobos

Crohn's Disease Patients Carrying Nod2/CARD15 Gene Variants Have an Increased and Early Need for First Surgery due to Stricturing Disease and Higher Rate of Surgical Recurrence

Objective:To study the predictive value of Nod2/CARD15 gene variants along with disease phenotypic characteristics for requirement of initial surgery and for surgical recurrence in Crohns disease (CD). Summary Background Data:Nod2/CARD15 gene variants play an important role in the susceptibility to CD. Studies of genotype-phenotype relationship suggest that these variants are associated with development of intestinal strictures. Preliminary reports analyzing the association between these variants and need for surgery have produced inconsistent results. Methods:A total of 170 CD patients were included prospectively in the study and followed up regularly for a mean of 7.4 ± 6.1 years. Clinical characteristics of CD, time and indication for surgery, and recurrence were registered. Nod2/CARD15 gene variants were determined by DNA sequencing analysis. Results:Surgery for stricturing disease was significantly more frequent in patients with Nod2/CARD15 variants in the univariate analysis (odds ratio [OR], 3.63; 95% confidence interval [CI], 1.42–9.27), and it was required at an earlier time (P = 0.004). Only Nod2/CARD15 variants (OR, 3.58; 95% CI, 1.21–10.5) and stricturing phenotype at diagnosis of CD (OR, 9.34; 95% CI, 2.56–33.3) were independent predictive factors of initial surgery for stricturing lesions in the multivariate analysis. Among 70 patients that required surgery, postoperative recurrence was also more frequent in patients with Nod2/CARD15 variants in the univariate and multivariate analysis (OR, 3.29; 95% CI, 1.13–9.56), and reoperation was needed at an earlier time (P = 0.03). Conclusion:Nod2/CARD15 variants are associated with early initial surgery due to stenosis and with surgical recurrence in Crohns disease. Patients with these variants could benefit from preventive and/or early therapeutic strategies.

Soluble ST2: A new and promising activity marker in ulcerative colitis

AIM To correlate circulating soluble ST2 (sST2) levels with the severity of ulcerative colitis (UC) and serum levels of pro-inflammatory cytokines, and to demonstrate the predictive power of sST2 levels for differentiation between active and inactive UC. METHODS We recruited 153 patients: 82 with UC, 26 with Crohns disease (CD) and 43 disease controls [non-inflammatory bowel disease (IBD)]. Subjects were excluded if they had diagnosis of asthma, autoimmune diseases or hypertension. The serum levels of sST2 and pro-inflammatory cytokines [pg/mL; median (25th-75th)] as well as clinical features, endoscopic and histological features, were subjected to analyses. The sST2 performance for discrimination between active and inactive UC, non-IBD and healthy controls (HC) was determined with regard to sensitivity and specificity, and Spearmans rank correlation coefficient (r). To validate the method, the area under the curve (AUC) of receiver-operator characteristic (ROC) was determined (AUC, 95% CI) and the total ST2 content of the colonic mucosa in UC patients was correlated with circulating levels of sST2. RESULTS The serum sST2 value was significantly higher in patients with active [235.80 (90.65-367.90) pg/mL] rather than inactive UC [33.19 (20.04-65.32) pg/mL], based on clinical, endoscopic and histopathological characteristics, as well as compared with non-IBD and HC (P < 0.001). The median level of sST2 in CD patients was 54.17 (35.02-122.0) pg/mL, significantly higher than that of the HC group only (P < 0.01). The cutoff was set at 74.87 pg/mL to compare active with inactive UC in a multicenter cohort of patients. Values of sensitivity, specificity, and ability to correctly classify UC, according to activity, were 83.33%, 83.33% and 83.33%, respectively. The AUC of the ROC curve to assess the ability of this molecule to discriminate between active vs inactive UC was 0.92 (0.86-0.97, P < 0.0001). The serum levels of sST2 in patients with UC significantly correlated with endoscopic and histopathological scores (r = 0.76 and r = 0.67, P < 0.0001, respectively), and with the pro-inflammatory cytokine, tumor necrosis factor-α (r = 0.69 and r = 0.61, respectively, P < 0.0001). Interestingly, we found a direct correlation between total intestinal ST2 content and serum levels of sST2, adjusted to endoscopic activity score in patients with mild (r = 0.44, P = 0.004), moderate (r = 0.59, P = 0.002) and severe disease (r = 0.82, P = 0.002). Only patients with inactive UC showed no significant correlation (r = 0.45, P = 0.267). CONCLUSION sST2 levels correlated with disease severity and inflammatory cytokines, are able to differentiate active from inactive UC and might have a role as a biomarker.

Escherichia coli isolates from inflammatory bowel diseases patients survive in macrophages and activate NLRP3 inflammasome.

Crohns disease (CD) is a multifactorial pathology associated with the presence of adherent-invasive Escherichia coli (AIEC) and NLRP3 polymorphic variants. The presence of intracellular E. coli in other intestinal pathologies (OIP) and the role of NLRP3-inflammasome in the immune response activated by these bacteria have not been investigated. In this study, we sought to characterize intracellular strains isolated from patients with CD, ulcerative colitis (UC) and OIP, and analyze NLRP3-inflammasome role in the immune response and bactericidal activity induced in macrophages exposed to invasive bacteria. For this, intracellular E. coli isolation from ileal biopsies, using gentamicin-protection assay, revealed a prevalence and CFU/biopsy of E. coli higher in biopsies from CD, UC and OIP patients than in controls. To characterize bacterial isolates, pulsed-field gel electrophoresis (PFGE) patterns, virulence genes, serogroup and phylogenetic group were analyzed. We found out that bacteria isolated from a given patient were closely related and shared virulence factors; however, strains from different patients were genetically heterogeneous. AIEC characteristics in isolated strains, such as invasive and replicative properties, were assessed in epithelial cells and macrophages, respectively. Some strains from CD and UC demonstrated AIEC properties, but not strains from OIP. Furthermore, the role of NLRP3 in pro-inflammatory cytokines production and bacterial elimination was determined in macrophages. E. coli strains induced IL-1β through NLRP3-dependent mechanism; however, their elimination by macrophages was independent of NLRP3. Invasiveness of intracellular E. coli strains into the intestinal mucosa and IL-1β production may contribute to CD and UC pathogenesis.

Interleukin-10 plays a key role in the modulation of neutrophils recruitment and lung inflammation during infection by Streptococcus pneumoniae.

Streptococcus pneumoniae is a major aetiological agent of pneumonia worldwide, as well as otitis media, sinusitis, meningitis and sepsis. Recent reports have suggested that inflammation of lungs due to S. pneumoniae infection promotes bacterial dissemination and severe disease. However, the contribution of anti‐inflammatory molecules to the pathogenesis of S. pneumoniae remains unknown. To elucidate whether the production of the anti‐inflammatory cytokine interleukin‐10 (IL‐10) is beneficial or detrimental for the host during pneumococcal pneumonia, we performed S. pneumoniae infections in mice lacking IL‐10 (IL‐10−/− mice). The IL‐10−/− mice showed increased mortality, higher expression of pro‐inflammatory cytokines, and an exacerbated recruitment of neutrophils into the lungs after S. pneumoniae infection. However, IL‐10−/− mice showed significantly lower bacterial loads in lungs, spleen, brain and blood, when compared with mice that produced this cytokine. Our results support the notion that production of IL‐10 during S. pneumoniae infection modulates the expression of pro‐inflammatory cytokines and the infiltration of neutrophils into the lungs. This feature of IL‐10 is important to avoid excessive inflammation of tissues and to improve host survival, even though bacterial dissemination is less efficient in the absence of this cytokine.

Achieving the best bowel preparation for colonoscopy

Bowel preparation is a core issue in colonoscopy, as it is closely related to the quality of the procedure. Patients often find that bowel preparation is the most unpleasant part of the examination. It is widely accepted that the quality of cleansing must be excellent to facilitate detecting neoplastic lesions. In spite of its importance and potential implications, until recently, bowel preparation has not been the subject of much study. The most commonly used agents are high-volume polyethylene glycol (PEG) electrolyte solution and sodium phosphate. There has been some confusion, even in published meta-analyses, regarding which of the two agents provides better cleansing. It is clear now that both PEG and sodium phosphate are effective when administered with proper timing. Consequently, the timing of administration is recognized as one of the central factors to the quality of cleansing. The bowel preparation agent should be administered, at least in part, a few hours in advance of the colonoscopy. Several low volume agents are available, and either new or modified schedules with PEG that usually improve tolerance. Certain adjuvants can also be used to reduce the volume of PEG, or to improve the efficacy of other agents. Other factors apart from the choice of agent can improve the quality of bowel cleansing. For instance, the effect of diet before colonoscopy has not been completely clarified, but an exclusively liquid diet is probably not required, and a low-fiber diet may be preferable because it improves patient satisfaction and the quality of the procedure. Some patients, such as diabetics and persons with heart or kidney disease, require modified procedures and certain precautions. Bowel preparation for pediatric patients is also reviewed here. In such cases, PEG remains the most commonly used agent. As detecting neoplasia is not the main objective with these patients, less intensive preparation may suffice. Special considerations must be made for patients with inflammatory bowel disease, including safety and diagnostic issues, so that the most adequate agent is chosen. Identifying neoplasia is one of the main objectives of colonoscopy with these patients, and the target lesions are often almost invisible with white light endoscopy. Therefore excellent quality preparation is required to find these lesions and to apply advanced methods such as chromoendoscopy. Bowel preparation for patients with lower gastrointestinal bleeding represents a challenge, and the strategies available are also reviewed here.

Epidemic Clostridium difficile Ribotype 027 in Chile

To the Editor: The increased severity of Clostridium difficile infection is primarily attributed to the appearance of an epidemic strain characterized as PCR ribotype 027 (1). The only report that identified epidemic C. difficile ribotype 027 in an American country outside of North America comes from Costa Rica, raising the possibility that strains 027 might also be present in other countries of Latin America (2). Several studies between 2001 and 2009 have been conducted in South American countries to detect the incidence of C. difficile infection in hospitalized patients, but they did not identify which C. difficile strains were causing these infections (3).

Increased production of soluble TLR2 by lamina propria mononuclear cells from ulcerative colitis patients.

Toll-like receptor 2 (TLR2) is a type I pattern recognition receptor that has been shown to participate in intestinal homeostasis. Its increased expression in the lamina propria has been associated with the pathogenesis in inflammatory bowel disease (IBD), such as ulcerative colitis (UC) and Crohns disease (CD). Recently, soluble TLR2 (sTLR2) variants have been shown to counteract inflammatory responses driven by the cognate receptor. Despite the evident roles of TLR2 in intestinal immunity, no study has elucidated the production and cellular source of sTLR2 in IBD. Furthermore, an increase in the population of activated macrophages expressing TLR2 that infiltrates the intestine in IBD has been reported. We aimed first to assess the production of the sTLR2 by UC and CD organ culture biopsies and lamina propria mononuclear cells (LPMCs) as well as the levels of sTLR2 in serum, and then characterize the cell population from lamina propria producing the soluble protein. Mucosa explants, LPMCs and serum were obtained from UC, CD patients and control subjects. The level of sTLR2 was higher in conditioned media from organ culture biopsies and LPMCs from UC patients in comparison to CD and controls. Moreover, an inverse correlation between the content of intestinal and serum sTLR2 levels was observed in UC patients. Additionally, when characterizing the cellular source of the increased sTLR2 by LPMCs from UC patients, an increase in TLR2(+)/CD33(+) cell population was found. Also, these cells expressed CX3CR1, which was related to the increased levels of intestinal FKN in UC patients, suggesting that a higher proportion of TLR2(+) mononuclear cells infiltrate the lamina propria. The increased production of sTLR2 suggests that a differential regulating factor of the innate immune system is present in the intestinal mucosa of UC patients.

Infecciones causadas por Clostridium difficile: una visión actualizada

Resumen de manejo de la infeccion asociada a C. dif fi cile (IACD) durante el primer episodio y recurrencias. SNG: sonda nasogastrica. TMF: trasplante de microbiota fecal. IV: intravenoso.Cuadro clinico compatible y estudio de laboratorio positivoCategorizar segun gravedadLeve o moderado Grave ComplicadoMetronidazol oral 500 mg c/8 horasVancomicina oral 125 mg c/6 horas Vancomicina oral o por SNG 500 mg c/6 horas + Metronidazol 500 mg c/8 horas ivSi existe ileo: agregar vancomicina intracolonica 500 mg c/4 a 6 horasConsiderar colectomiaRecurrencia de IACD (1a recurrencia)Tratar segun gravedad de episodio actualRecurrencia de IACD (2a recurrencia)Vancomicina en pulso o dosis decrecienteRecurrencia de IACD (3a recurrencia)Considerar TMF de acuerdo aceptabilidad del paciente, inmunoglobulina iv, rifaximina o fidaxomicina Tratamiento de IACD El tratamiento de IACD dependera de si se trata del primer, segundo o tercer episodio de IACD y de su gra-vedad, la cual se defi ne segun los parametros senalados previamente (Figura 4)

Prospective comparison of a commercial multiplex real-time polymerase chain reaction and an enzyme immunoassay with toxigenic culture in the diagnosis of Clostridium difficile–associated infections ☆

Clostridium difficile infections (CDI) is a leading cause of nosocomial infections worldwide. The changes in the epidemiology of CDI during the past years, including the appearance of new epidemic strains of C. difficile that cause CDI episodes with increased severity, have led to the development of molecular methods with improved sensitivity and specificity. This study was designed to compare the performances of one antigen assay (Vidas, bioMérieux) and one molecular assay (GeneXpert, Cepheid). Fecal specimens from hospitalized patients (n = 230) suspected of having CDI were tested by both assays. Eleven specimens were positive and 202 were negative for both methods. After discrepant analysis by C. difficile toxigenic culture with broth enrichment and neutralization assay, the total numbers of stool specimens classified as positive and negative for toxigenic C. difficile were 23 (10%) and 206 (89.6%), respectively. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value for GeneXpert were 91.7%, 99%, 91.7%, and 99%, and for Vidas were 48%, 99%, 84.6%, and 94.5%, respectively. The sensitivity and PPV of polymerase chain reactoin GeneXpert assay far exceeded those of the EIA Vidas assay. The clinical characteristics of concordant and discrepant study patients were similar with the exception of the number of previous CDI episodes, which were higher in the concordant study patients; the clinical characteristics of both groups were similar. In conclusion, due to the appearance of more virulent strains of C. difficile during the last years that have produced dramatic changes in the epidemiology of C. difficile, we recommend that toxin enzyme immunoassays be replaced with rapid molecular-based tests for toxigenic C. difficile.

Modulation of antigen processing by haem-oxygenase 1. Implications on inflammation and tolerance.

Haem‐oxygenase‐1 (HO‐1) is an enzyme responsible for the degradation of haem that can suppress inflammation, through the production of carbon monoxide (CO). It has been shown in several experimental models that genetic and pharmacological induction of HO‐1, as well as non‐toxic administration of CO, can reduce inflammatory diseases, such as endotoxic shock, type 1 diabetes and graft rejection. Recently, it was shown that the HO‐1/CO system can alter the function of antigen‐presenting cells (APCs) and reduce T‐cell priming, which can be beneficial during immune‐driven inflammatory diseases. The molecular mechanisms by which the HO‐1 and CO reduce both APC‐ and T‐cell‐driven immunity are just beginning to be elucidated. In this article we discuss recent findings related to the immune regulatory capacity of HO‐1 and CO at the level of recognition of pathogen‐associated molecular patterns and T‐cell priming by APCs. Finally, we propose a possible regulatory role for HO‐1 and CO over the recently described mitochondria‐dependent immunity. These concepts could contribute to the design of new therapeutic tools for inflammation‐based diseases.