G. Sica

Aryl hydrocarbon receptor-induced signals up-regulate IL-22 production and inhibit inflammation in the gastrointestinal tract.

BACKGROUND & AIMS The pathogenesis of inflammatory bowel disease (IBD) is believed to involve an altered balance between effector and regulatory T cells. Aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor that mediates the toxicity of dioxins, controls T-cell responses. We investigated the role of AhR in inflammation and pathogenesis of IBD in humans and mouse models. METHODS AhR expression was evaluated in intestinal tissue samples from patients with IBD and controls by real-time polymerase chain reaction (PCR) and flow cytometry. Intestinal lamina propria mononuclear cells (LPMCs) were activated in the presence or absence of the AhR agonist 6-formylindolo(3, 2-b)carbazole (Ficz). Colitis was induced in mice using trinitrobenzene sulfonic acid (TNBS), dextran sulfate sodium (DSS), or T-cell transfer. Mice were given injections of Ficz or the AhR antagonist 2-metyl-2H-pyrazole-3-carboxylic acid; some mice first received injections of a blocking antibody against interleukin (IL)-22. Cytokines were quantified by real-time PCR and flow cytometry. RESULTS Intestine tissue from patients with IBD expressed significantly less AhR than controls. In LPMCs from patients with IBD, incubation with Ficz reduced levels of interferon gamma (IFN)-γ and up-regulated IL-22. Mice injected with Ficz were protected against TNBS-, DSS-, and T-cell transfer-induced colitis; they had marked down-regulation of inflammatory cytokines and induction of IL-22. Mice given AhR antagonist produced more inflammatory cytokines and less IL-22 and developed a severe colitis. Neutralization of endogenous IL-22 disrupted the protective effect of Ficz on TNBS-induced colitis. CONCLUSIONS AhR is down-regulated in intestinal tissue of patients with IBD; AhR signaling, via IL-22, inhibits inflammation and colitis in the gastrointestinal tract of mice. AhR-related compounds might be developed to treat patients with IBDs.

Th17-type cytokines, IL-6 and TNF-α synergistically activate STAT3 and NF-kB to promote colorectal cancer cell growth

Colorectal cancers (CRCs) often show a dense infiltrate of cytokine-producing immune/inflammatory cells. The exact contribution of each immune cell subset and cytokine in the activation of the intracellular pathways sustaining CRC cell growth is not understood. Herein, we isolate tumor-infiltrating leukocytes (TILs) and lamina propria mononuclear cells (LPMCs) from the tumor area and the macroscopically unaffected, adjacent, colonic mucosa of patients who underwent resection for sporadic CRC and show that the culture supernatants of TILs, but not of LPMCs, potently enhance the growth of human CRC cell lines through the activation of the oncogenic transcription factors signal transducer and activator of transcription 3 (STAT3) and nuclear factor-kappa B (NF-kB). Characterization of immune cell complexity of TILs and LPMCs reveals no differences in the percentages of T cells, natural killer T cells, natural killer (NK) cells, macrophages and B cells. However, T cells from TILs show a functional switch compared with those from LPMCs to produce large amounts of T helper type 17 (Th17)-related cytokines (that is, interleukin-17A (IL-17A), IL-17F, IL-21 and IL-22), tumor necrosis factor-α (TNF-α) and IL-6. Individual neutralization of IL-17A, IL-17F, IL-21, IL-22, TNF-α or IL-6 does not change TIL-derived supernatant-driven STAT3 and NF-kB activation, as well as their proproliferative effect in CRC cells. In contrast, simultaneous neutralization of both IL-17A and TNF-α, which abrogates NF-kB signaling, and IL-22 and IL-6, which abrogates STAT3 signaling, reduces the mitogenic effect of supernatants in CRC cells. IL-17A, IL-21, IL-22, TNF-α and IL-6 are also produced in excess in the early colonic lesions in a mouse model of sporadic CRC, associated with enhanced STAT3/NF-kB activation. Mice therapeutically given BP-1-102, an orally bioavailable compound targeting STAT3/NF-kB activation and cross-talk, exhibit reduced colon tumorigenesis and diminished expression of STAT3/NF-kB-activating cytokines in the neoplastic areas. These data suggest that strategies aimed at the cotargeting of STAT3/NF-kB activation and interaction between them might represent an attractive and novel approach to combat CRC.

Wireless capsule endoscopy and small intestine contrast ultrasonography in recurrence of Crohn's disease

Background: The best available tool to assess recurrence of Crohns disease (CD) is ileocolonoscopy (CC). Small intestine contrast ultrasonography (SICUS) and wireless capsule endoscopy (WCE) are noninvasive techniques able to detect small bowel lesions. In a prospective longitudinal study, we aimed to investigate the usefulness of SICUS and WCE for assessing postoperative recurrence of CD 1 year after surgery, using CC as the gold standard. Methods: Twenty‐two patients (11 men, median age 33 years, range 22–67 years) undergoing ileocolonic resection for CD were prospectively followed from July 2003 to May 2006, with the Crohns Disease Activity Index (CDAI) used for clinical assessment every 3 months for 1 year. At 1 year, recurrence was assessed by SICUS and CC, followed by WCE. CD recurrence was assessed by CC (Rutgeerts score). SICUS was performed after ingestion of polyethylene glycol, and WCE was performed with Given M2A equipment. Results: At 1 year, all 22 patients had inactive CD (CDAI < 150). In 5 patients, WCE was not performed because of luminal narrowing or stenosis. Seventeen of the 22 patients had all 3 techniques performed. CC detected recurrence in 21 of 22 patients. Lesions compatible with recurrence were detected by SICUS in all 22 patients (1 false positive). When considering only the 17 patients studied by all 3 techniques, recurrence was detected by CC in 16 of 17 patients, whereas lesions compatible with recurrence were detected by SICUS in all 17 patients (16 true positives [TPs], 1 FP) and by WCE in 16 of 17 patients (16 TPs, 1 true negative). Conclusions: The present findings suggest that SICUS and WCE may be used as noninvasive techniques for the assessment of recurrence of CD in patients being regularly followed up after ileocolonic resection. (Inflamm Bowel Dis 2007)

Autocrine Regulation of IL-21 Production in Human T Lymphocytes

IL-21 has pathologic function in immune-inflammatory diseases. IL-21 mediates its functions through a heterodimeric receptor, composed of a specific subunit, termed IL-21R, and the common γ-chain. IL-21 is mostly produced by CD4+ T cells, but molecular mechanisms that regulate IL-21 synthesis are not fully understood. The fact that CD4+ T cells express high levels of IL-21R and are capable of functionally responding to IL-21 raises the possibility that IL-21 may regulate its own production. We here show that IL-21 enhances IL-21 RNA and protein expression in human peripheral blood CD3+ T cells in a dose- and time-dependent fashion. Additionally, both IL-7 and IL-15, but not IL-4, induce IL-21, thus suggesting that common γ-chain signals are not sufficient to promote IL-21 synthesis. Analysis of molecular mechanisms underlying IL-21 induction reveals that IL-21 activates Stat3 and enhances its recruitment to IL-21 gene promoter. Pharmacologic inhibition and knockdown of Stat3 by small interference RNA largely prevent IL-21 induction in IL-21-treated cells. Consistently, IL-21 is inducible in T cells by IL-6, another cytokine that activates Stat3. Finally, we show that IL-21 positively regulates its own expression in human intestinal CD3+ lamina propria lymphocytes, and blockade of endogenous IL-21 in cultures of CD3+ lamina propria lymphocytes isolated from patients with Crohn’s disease, a chronic inflammatory bowel disease characterized by high IL-21, down-regulates Stat3 activation and IL-21 expression. These data suggest the existence of a positive autocrine loop that could help to amplify and stabilize IL-21-driven, T cell-mediated responses.

Interferon-gamma-expressing cells are a major source of interleukin-21 in inflammatory bowel diseases.

Background: We previously demonstrated that in inflammatory bowel disease (IBD) there is enhanced production of interleukin (IL)‐21, a cytokine that activates multiple pathways that sustain mucosal inflammation. However, the phenotype of IL‐21‐producing cells in IBD, and the cytokine(s) they coproduce, is not known. We here characterized the cell source of IL‐21 and determined which factors regulate IL‐21 in the human gut. Methods: Cytokines were analyzed in CD4+ T intestinal lamina propria lymphocytes (T‐LPL) isolated from IBD patients and controls by flow cytometry. Moreover, IL‐21 was evaluated in mucosal T follicular cells (TFH). To assess the involvement of IL‐12 and IL‐23 in the production of IL‐21, T‐LPL were activated in the presence or absence of IL‐12 or IL‐23. Results: The proportion of IL‐21‐producing CD4+ T‐LPL was increased in IBD compared to controls. The majority of IL‐21‐producing T‐LPL coexpressed interferon (IFN)‐&ggr;, and to a lesser extent IL‐4 or IL‐17A. Activation of CD4+ T‐LPL with IL‐12 but not IL‐23 enhanced the fraction of cells coexpressing IL‐21 and IFN‐&ggr;. TFH cells in LPL were identified by CXCR5 expression and expressed IL‐21 both in IBD and controls; however, the fraction of IL‐21‐positive TFH cells was higher in Crohns disease than in ulcerative colitis and controls. Treatment of CD4+ T‐LPL with IL‐12 enhanced the frequency of CXCR5+ IL‐21‐producing TFH cells. Conclusions: These findings indicate that in IBD IL‐21 is mostly produced by CD4+ T‐LPL coexpressing IFN‐&ggr;, reinforcing the concept that distinct subsets of T cells can produce IL‐21. Inflamm Bowel Dis 2010

Frequency, Pattern, and Risk Factors of Postoperative Recurrence of Crohn’s Disease After Resection Different from Ileo-Colonic

BackgroundThe frequency of recurrence in Crohn’s disease (CD) patients after curative resection different from the ileo-colonic is undefined. We aimed to assess the frequency, pattern, outcome, and risk factors of postoperative recurrence in CD patients under regular follow-up after anastomosis different from ileo-colonic.Materials and MethodsIn a retrospective study, clinical records of 537 CD patients under regular follow-up from January 2001 to August 2007 were reviewed. The outcome after surgery was assessed on the basis of clinical records prospectively recorded.ResultsPrevious resection was observed in 183 of 537 (34%) patients, including the ileo-colon in 145 (79%) and other gastrointestinal (GI) segments in 38 (21%). Recurrence was detected in 16 of 38 (42%) patients (all symptomatic) including five of 14 (35%) with ileostomy, five of five (100%) with ileo-rectal, three of 11 (27%) with ileo-ileal, one or four (25%) with colorectal, and two of three (33%) with duodenum-jejunal anastomosis. Ileo-colonic resection was reported in 145 of 183 (79%) patients, showing recurrence in 128 (88.3%) and symptomatic in 47 (36.7%) patients. The frequency of recurrence was higher in patients with ileo-colonic resection than in patients with other types of resection (128/145, 88% vs 16/38, 42%, p < 0.001). The frequency of symptomatic recurrence was lower in patients with ileo-colonic resection than in those with other resections (47/128, 37% vs 16/16, 100%; p < 0.001). Risk factors for recurrence were comparable in the two subgroups (smoke, odds ratio, OR 1.5 vs 1.4; appendectomy, OR 0.32 vs 0.33; familial inflammatory bowel disease, OR 0.43 vs 1.26). ConclusionsPostoperative recurrence is observed in a high proportion of CD patients after resection different from ileo-colon (including ileostomy), although at a lower frequency than observed after ileo-colonic resection.

Bacterial translocation and intestinal morphological findings in jaundiced rats

The susceptibility to sepsis in obstructive jaundice may be related to bacterial translocation (BT) from the gastrointestinal tract. We evaluated BT to visceral organs and morphological changes of the intestinal mucosa in a rat model of obstructive jaundice. Animals were randomly divided into two groups: in group A the common bile duct was tied and divided, while group B had the bile duct mobilized but not tied. After seven days, peritoneal swabs and liver, spleen, pancreas, lung, mesenteric lymph nodes (MLN), cecum, and terminal ileum biopsies were obtained for cultures. Light and electron microscopy were performed on intestinal samples. The TUNEL assay was performed to detect apoptosis. Data were analyzed using Fisher exact test and Student ttest. Bile duct obliteration resulted in an increased incidence of BT. Seven days after duct obliteration, BT to the peritoneal cavity was evident in 37.5% of the animals in group A and 25% in group B. The respective BT rates for the two groups were: 42.8% vs 37.5% to MLN, 71.4% vs 25% to liver, 42.8% vs 12.5% to spleen, 28.6% vs 0% to pancreas and 14.3% vs 0% to lungs. Despite a trend, this was not statistically significant. Cecal counts did not differ statistically among the groups, while ileal counts were significantly higher in jaundiced rats (P < 0.05). Structural and ultrastructural abnormalities were evident only in the mucosa of the terminal ileum of jaundiced rats. Apoptosis was significantly increased in the terminal ileum of jaundiced rats (P < 0.002). This study suggests the possible association of biliary obstruction and BT. The nonspecific physical injury observed may contribute to breakdown of gastrointestinal barrier function thus promoting BT.

Cancer in Crohn's Disease patients treated with infliximab: A long-term multicenter matched pair study

Background: The long‐term risk of neoplasia in Crohns disease (CD) patients treated with infliximab is undefined. The aim was to assess, in a multicenter, matched‐pair study, whether infliximab use in CD is associated with an increased frequency of neoplasia in the long term. Methods: A multicenter, long‐term, matched‐pair study was conducted in 12 referral inflammatory bowel disease (IBD) centers. An initial cohort of 808 CD patients, including 404 infliximab‐treated (CD‐IFX) and 404 matched CD controls never treated with infliximab (CD‐C) studied from 1999 to 2004, was followed up for an additional 4 years (2004–2008). Cases and controls were matched for: sex, age (±5 years), CD site, follow‐up (±5 years), immunosuppressant use, and CD duration (±5 years). From 1999 to 2008 the frequency and characteristics of neoplasia were compared between CD‐IFX and CD‐C. Results: In 2008, 591 patients (304 CD‐IFX, 287 CD‐C) were in follow‐up. Matched couples included 442 patients: 221 CD‐IFX and 221 CD‐C (median follow‐up, months: 72, range 48–114 versus 75, range 44–114). From 1999 to 2008 the frequency of neoplasia among the 591 patients did not differ between CD‐IFX (12/304; 3.94%) and CD‐C (12/287; 4.19%; P = 0.95). A comparable frequency of neoplasia was also observed between the 221 matched couples (CD‐IFX: 8/221; 3.61% versus CD‐C: 9/221; 4.07%; P = 1). No specific histotype of cancer appeared associated with infliximab use. Conclusions: The frequency of neoplasia was comparable in an adult population of CD patients treated or not with infliximab, matched for clinical variables and followed up for a median of 6 years. (Inflamm Bowel Dis 2011)

Laparoscopic single-port sleeve gastrectomy for morbid obesity: preliminary series.

BACKGROUND Laparoscopic sleeve gastrectomy has been recently proposed as a sole bariatric procedure because of the resulting considerable weight loss in morbidly obese patients. Traditionally, laparoscopic sleeve gastrectomy requires 5-6 skin incisions to allow for placement of multiple trocars. With the introduction of single-incision laparoscopic surgery, multiple abdominal procedures have been performed using a sole umbilical incision, with good cosmetic outcomes. The purpose of our study was to evaluate the feasibility and safety of laparoscopic single incision sleeve gastrectomy for morbid obesity. METHODS A total of 8 consecutive patients underwent laparoscopic single-incision sleeve gastrectomy at the Operative Unit of Bariatric Surgery of the University of Rome Tor Vergata from March 2009 to June 2009. RESULTS Of the 8 patients, 5 were women and 3 were men, with a mean age of 44.4 years. The mean preoperative body mass index was 56.2 kg/m(2). The mean operative time was 128 minutes. The mean postoperative stay was 2.4 days. The mean postoperative body mass index was 49.3 kg/m(2) at a mean follow-up period of 3.6 months. The mean percentage of excess weight loss was 33% for the same period. CONCLUSIONS Laparoscopic single-incision sleeve gastrectomy seems to be safe, technically feasible, and reproducible. A randomized trial comparing single-incision sleeve gastrectomy and conventional sleeve gastrectomy might be needed to evaluate the postoperative results in relation to the development of abdominal wall complications.

A functional role for Smad7 in sustaining colon cancer cell growth and survival.

Initially identified as an inhibitor of transforming growth factor (TGF)-β mainly owing to its ability to bind TGF-β receptor type I and abrogate TGF-β-driven signaling, Smad7 can interact with additional intracellular proteins and regulate TGF-β-independent pathways, thus having a key role in the control of neoplastic processes in various organs. Genome-wide association studies have shown that common alleles of Smad7 influence the risk of colorectal cancer (CRC), even though the contribution of Smad7 in colon carcinogenesis is not fully understood. In this study, we assessed the expression and role of Smad7 in human and mouse models of sporadic CRC. We document a significant increase of Smad7 in human CRC relative to the surrounding nontumor tissues and show that silencing of Smad7 inhibits the growth of CRC cell lines both in vitro and in vivo after transplantation into immunodeficient mice. Knockdown of Smad7 results in enhanced phosphorylation of the cyclin-dependent kinase (CDK)2, accumulation of CRC cells in S phase and enhanced cell death. Smad7-deficient CRC cells have lower levels of CDC25A, a phosphatase that dephosphorylates CDK2, and hyperphosphorylated eukaryotic initiation factor 2 (eIF2)α, a negative regulator of CDC25 protein translation. Consistently, knockdown of Smad7 associates with inactivation of eIF2α, lower CDC25A expression and diminished fraction of proliferating cells in human CRC explants, and reduces the number of intestinal tumors in Apcmin/+ mice. Altogether, these data support a role for Smad7 in sustaining colon tumorigenesis.