Immune-Mediated Inflammatory Diseases Awareness and Management among Physicians Treating Patients with Inflammatory Bowel Disease: An IG-IBD Survey.

(1) Background: Inflammatory bowel disease (IBD) is frequently associated to other immune-mediated inflammatory diseases (IMIDs). This study aims at assessing physicians' awareness of the issue and the current status of IMID management. (2) Methods: A web-based survey was distributed to all 567 physicians affiliated to IG-IBD. (3) Results: A total of 249 (43.9%) physicians completed the survey. Over 90% of the responding physicians were gastroenterology specialists, primarily working in public hospitals. About 51.0% of the physicians had access to an integrated outpatient clinic, where gastroenterologists collaborated with rheumatologists and 28.5% with dermatologists. However, for 36.5% of physicians, integrated ambulatory care was not feasible. Designated appointment slots for rheumatologists and dermatologists were accessible to 72.2% and 58.2% of physicians, respectively, while 20.1% had no access to designated slots. About 5.2% of physicians report investigating signs or symptoms of IMIDs only during the initial patient assessment. However, 87.9% inquired about the presence of concomitant IMIDs at the initial assessment and actively investigated any signs or symptoms during subsequent clinical examination. (4) Conclusions: While Italian physicians recognize the importance of IMIDs associated with IBD, organizational challenges impede the attainment of optimal multidisciplinary collaboration. Efforts should be directed toward enhancing practical frameworks to improve the overall management of these complex conditions.

Defining Comprehensive Disease Control for Use as a Treatment Target for Ulcerative Colitis in Clinical Practice: International Delphi Consensus Recommendations.

BACKGROUND AND AIMS: Treatment of ulcerative colitis [UC] requires a patient-centric definition of comprehensive disease control that considers improvements in aspects not typically captured by classical landmark trial endpoints. In an international initiative, we reviewed aspects of UC that affect patients and/or indicate mucosal inflammation, to achieve consensus on which aspects to combine in a definition of comprehensive disease control, using a modified Delphi process. METHODS: The Delphi panel comprised 12 gastroenterologists and one patient advocate. Two gastroenterologists were elected as chairs and did not vote. To inform statements, we asked 18 patients and the panel members about their experiences of remission and reviewed published literature. Panel members voted on statements anonymously in three rounds, with a live discussion before Round 3. Consensus was met if ≥67% of the panel agreed. Statements without consensus in Rounds 1 and 2 were revised or discarded after Round 3. RESULTS: The panel agreed to measure individual patient benefit using a definition of comprehensive disease control that combines aspects currently measured in trials [rectal bleeding, stool frequency, disease-related quality of life, endoscopy, histological inflammatory activity, inflammatory biomarkers, and corticosteroid use] with additional patient-reported symptoms [bowel urgency, abdominal pain, extraintestinal manifestations, fatigue, and sleep disturbance]. The panel agreed on scoring systems and thresholds for many aspects. CONCLUSIONS: Using a robust methodology, we defined comprehensive disease control in UC. Next, we will combine the measurement and scoring of these aspects into a multicomponent tool and will adopt comprehensive disease control as a treatment target in clinical practice and trials.

An Objective Comparison of Vedolizumab and Ustekinumab Effectiveness in Crohn's Disease Patients' Failure to TNF-Alpha Inhibitors.

INTRODUCTION: The use of ustekinumab and vedolizumab as second-line therapies in patients with Crohn's disease (CD) in which tumour necrosis factor alpha inhibitors (TNFi) failed is still debated. The aim of this study was to compare, in a large multicenter observational retrospective cohort, the effectiveness of ustekinumab and vedolizumab as second-line therapies, as assessed by clinical and objective outcomes including endoscopy and gastrointestinal imaging. METHODS: Clinical response, remission, and steroid-free remission at weeks 26 and 52 were evaluated in a retrospective propensity score-weighted and propensity score-matched cohort of patients in which TNFi failed. Objective response and remission were evaluated by 1 or more techniques among endoscopy, magnetic resonance/computed tomography enteroclysis, and small bowel ultrasound. RESULTS: A total of 470 patients with CD (239 treated with ustekinumab and 231 treated with vedolizumab) were included in the study. At week 26, clinical outcomes were similar between the 2 groups. At week 52, clinical remission (ustekinumab 42.5% vs vedolizumab 55.5%, P = 0.01) and steroid-free remission (ustekinumab 40.6% vs vedolizumab 51.1%, P = 0.038) rates were significantly higher in vedolizumab-treated patients. Three hundred two patients (hundred thirty-five treated with ustekinumab and hundred sixty-seven treated with vedolizumab) had an objective evaluation of disease activity at baseline and week 52. At week 52, objective response and remission rates were similar between the 2 groups. Clinical response at week 26 predicted steroid-free remission at week 52 in both ustekinumab-treated and vedolizumab-treated patients. Safety profiles were similar between the 2 groups. DISCUSSION: In patients with CD in which TNFi failed, both ustekinumab and vedolizumab showed similar clinical effectiveness after 26 weeks of treatment. At 1 year, vedolizumab was associated with a higher rate of clinical remission when compared with ustekinumab. However, no difference was observed between the 2 groups when objective outcomes were investigated at this time point.

IL10 Secretion Endows Intestinal Human iNKT Cells with Regulatory Functions Towards Pathogenic T Lymphocytes.

BACKGROUND AND AIMS: Invariant natural killer T [iNKT] cells perform pleiotropic functions in different tissues by secreting a vast array of pro-inflammatory and cytotoxic molecules. However, the presence and function of human intestinal iNKT cells capable of secreting immunomodulatory molecules such as IL-10 has never been reported so far. Here we describe for the first time the presence of IL10-producing iNKT cells [NKT10 cells] in the intestinal lamina propria of healthy individuals and of Crohn's disease [CD] patients. METHODS: Frequency and phenotype of NKT10 cells were analysed ex vivo from intestinal specimens of Crohn's disease [n = 17] and controls [n = 7]. Stable CD-derived intestinal NKT10 cell lines were used to perform in vitro suppression assays and co-cultures with patient-derived mucosa-associated microbiota. Experimental colitis models were performed by adoptive cell transfer of splenic naïve CD4+ T cells in the presence or absence of IL10-sufficient or -deficient iNKT cells. In vivo induction of NKT10 cells was performed by administration of short chain fatty acids [SCFA] by oral gavage. RESULTS: Patient-derived intestinal NKT10 cells demonstrated suppressive capabilities towards pathogenic CD4+ T cells. The presence of increased proportions of mucosal NKT10 cells associated with better clinical outcomes in CD patients. Moreover, an intestinal microbial community enriched in SCFA-producing bacteria sustained the production of IL10 by iNKT cells. Finally, IL10-deficient iNKT cells failed to control the pathogenic activity of adoptively transferred CD4+ T cells in an experimental colitis model. CONCLUSIONS: These results describe an unprecedentd IL10-mediated immunoregulatory role of intestinal iNKT cells in controlling the pathogenic functions of mucosal T helper subsets and in maintaining the intestinal immune homeostasis.

Quercetin and its derivates as antiviral potentials: A comprehensive review.

Quercetin, widely distributed in fruits and vegetables, is a flavonoid known for its antioxidant, antiviral, antimicrobial, and antiinflammatory properties. Several studies highlight the potential use of quercetin as an antiviral, due to its ability to inhibit the initial stages of virus infection, to be able to interact with proteases important for viral replication, and to reduce inflammation caused by infection. Quercetin could also be useful in combination with other drugs to potentially enhance the effects or synergistically interact with them, in order to reduce their side effects and related toxicity. Since there is no comprehensive compilation about antiviral activities of quercetin and derivates, the aim of this review is providing a summary of their antiviral activities on a set of human viral infections along with mechanisms of action. Thus, the following family of viruses are examined: Flaviviridae, Herpesviridae, Orthomyxoviridae, Coronaviridae, Hepadnaviridae, Retroviridae, Picornaviridae, Pneumoviridae, and Filoviridae.

Immune system and gut microbiota senescence in elderly IBD patients.

In inflammatory bowel disease (IBD), the loss of immune tolerance against gut microbiota causes chronic inflammation and the progressive accumulation of organ damage in genetically susceptible individuals. In the elderly, IBD is often characterized by a different disease behaviour when compared with paediatric and young adult disease. Besides disease behaviour, another aspect of the multifaceted impact of age on elderly IBD course is increased susceptibility to infections. In this context, age-of-onset-dependent IBD behaviour and clinical course are two major contributors to immune system senescence and change of gut microbiota in older subjects. Here, we review the available literature linking immunosenescence and age-dependent changes in the gut microbiota composition to IBD pathogenesis speculating on their possible implications in disease expression in this age class.

A Pharmacological Batch of Mongersen that Downregulates Smad7 is Effective as Induction Therapy in Active Crohn's Disease: A Phase II, Open-Label Study.

BACKGROUND: A recent phase III trial did not confirm the previous clinical and endoscopic improvements seen in patients with Crohn's disease (CD) receiving Mongersen, an oral Smad7 antisense oligonucleotide. Factors accounting for such a discrepancy are unknown. OBJECTIVE: Our objective was to further assess whether Mongersen was effective as induction therapy in active CD and evaluate the in vitro inhibitory effect of various batches of Mongersen used in the previous and present trials on Smad7 expression. METHODS: In a phase II, open-label study, 18 patients with active CD (Crohn's Disease Activity Index [CDAI] score > 220 and evidence of endoscopic lesions) received Mongersen 160 mg/day for 12 weeks. The rates of clinical remission, defined as CDAI < 150, and clinical response, defined as a CDAI score decrease ≥ 100, were evaluated at week 4, 8, and 12. The fraction of circulating CCR9-expressing leukocytes was assessed by flow cytometry. Smad7 expression was evaluated in the human colorectal cancer cell line HCT-116 transfected with different batches of Mongersen using real-time polymerase chain reaction (PCR) and Western blotting, RESULTS: The proportions of patients experiencing clinical remission were 38.9%, 55.6%, and 50.0% at week 4, 8, and 12, respectively. At the same time points, the rates of clinical response were 72.2%, 77.8%, and 77.8%, respectively. Mongersen reduced the percentages of CCR9-expressing CD45+ cells. The batch of Mongersen used in this study, but not two batches used in the phase III study, inhibited Smad7 expression in HCT-116 cells. CONCLUSIONS: The present findings support the clinical benefit of Mongersen in active CD and show that various batches manufactured during the GED0301 program differ in their ability to inhibit in vitro Smad7. TRIAL REGISTRATION NUMBER: NCT02685683; EudraCT 2015-001693-18.

Activities related to inflammatory bowel disease management during and after the coronavirus disease 2019 lockdown in Italy: How to maintain standards of care.

BACKGROUND AND AIMS: Restructuring activities have been necessary during the lockdown phase of the coronavirus disease 2019 (COVID-19) pandemic. Few data are available on the post-lockdown phase in terms of health-care procedures in inflammatory bowel disease (IBD) care, and no data are available specifically from IBD units. We aimed to investigate how IBD management was restructured during the lockdown phase, the impact of the restructuring on standards of care and how Italian IBD units have managed post-lockdown activities. METHODS: A web-based online survey was conducted in two phases (April and June 2020) among the Italian Group for IBD affiliated units within the entire country. We investigated preventive measures, the possibility of continuing scheduled visits/procedures/therapies because of COVID-19 and how units resumed activities in the post-lockdown phase. RESULTS: Forty-two referral centres participated from all over Italy. During the COVID-19 lockdown, 36% of first visits and 7% of follow-up visits were regularly done, while >70% of follow-up scheduled visits and 5% of first visits were done virtually. About 25% of scheduled endoscopies and bowel ultrasound scans were done. More than 80% of biological therapies were done as scheduled. Compared to the pre-lockdown situation, 95% of centres modified management of outpatient activity, 93% of endoscopies, 59% of gastrointestinal ultrasounds and 33% of biological therapies. Resumption of activities after the lockdown phase may take three to six months to normalize. Virtual clinics, implementation of IBD pathways and facilities seem to be the main factors to improve care in the future. CONCLUSION: Italian IBD unit restructuring allowed quality standards of care during the COVID-19 pandemic to be maintained. A return to normal appears to be feasible and achievable relatively quickly. Some approaches, such as virtual clinics and identified IBD pathways, represent a valid starting point to improve IBD care in the post-COVID-19 era.

The Role of Gut Microbiota Biomodulators on Mucosal Immunity and Intestinal Inflammation.

Alterations of the gut microbiota may cause dysregulated mucosal immune responses leading to the onset of inflammatory bowel diseases (IBD) in genetically susceptible hosts. Restoring immune homeostasis through the normalization of the gut microbiota is now considered a valuable therapeutic approach to treat IBD patients. The customization of microbe-targeted therapies, including antibiotics, prebiotics, live biotherapeutics and faecal microbiota transplantation, is therefore considered to support current therapies in IBD management. In this review, we will discuss recent advancements in the understanding of host-microbe interactions in IBD and the basis to promote homeostatic immune responses through microbe-targeted therapies. By considering gut microbiota dysbiosis as a key feature for the establishment of chronic inflammatory events, in the near future it will be suitable to design new cost-effective, physiologic, and patient-oriented therapeutic strategies for the treatment of IBD that can be applied in a personalized manner.

Response Assessed by Ultrasonography as Target of Biological Treatment for Crohn's Disease.

BACKGROUND & AIMS: Mucosal healing, determined by ileocolonoscopy, is a goal for treatment of Crohn's disease (CD), but this is an invasive assessment procedure. We investigated whether response to tumor necrosis factor (TNF) antagonists, determined by small-intestine contrast ultrasonography, associates with long-term outcomes. METHODS: We performed observational study of 80 patients with CD treated with anti-TNF agents for at least 1 year who underwent serial small intestine contrast ultrasonography (SICUS) at the University of Rome, in Italy. SICUS was used to evaluate disease site (based on bowel wall thickness), extent of lesions, and presence of complications. Inclusion criteria required pre-therapy SICUS with follow-up SICUS after 18 months. At second SICUS, patients were assigned to categories of complete or partial responder or non-responder. CD-related outcomes (corticosteroid need, hospitalization, and surgery) were assessed at 1 year from the second SICUS, using multivariate models, and were analyzed after long term follow up (5 years) using Kaplan-Meier survival analysis. RESULTS: Based on SICUS, after a median of 18 months, 36 patients (51%) were complete responders, 30 were partial responders (34%), and 13 were non-responders (15%). At 1 year from the second SICUS, no patients with a complete response, based on ultrasonography, underwent surgery, in comparison to partial responders (P = .0003) or non-responders (P = .001). Complete responders used smaller amounts of corticosteroids than partial responders (P = .0001) or non-responders (P < .0001). Complete responders required fewer hospitalizations than non-responders (P = .001). Kaplan-Meier survival analysis of long-term follow up data demonstrated a lower cumulative probability of need for surgery, hospitalization, and need for steroids among SICUS-categorized complete responders (P < .0001, P = .003 and P = .0001 respectively) than SICUS-categorized non-responders. CONCLUSIONS: In patients with CD, response to anti-TNF agents, determined by SICUS, is associated with better long-term outcomes than partial or no response. Ultrasonographic assessment therefore provides a relatively non-invasive method for monitoring response to treatment in patients with CD.

Smad7 in intestinal CD4+ T cells determines autoimmunity in a spontaneous model of multiple sclerosis.

Environmental triggers acting at the intestinal barrier are thought to contribute to the initiation of autoimmune disorders. The transforming growth factor beta inhibitor Smad7 determines the phenotype of CD4+ T cells. We hypothesized that Smad7 in intestinal CD4+ T cells controls initiation of opticospinal encephalomyelitis (OSE), a murine model of multiple sclerosis (MS), depending on the presence of gut microbiota. Smad7 was overexpressed or deleted in OSE CD4+ T cells to determine the effect on clinical progression, T cell differentiation, and T cell migration from the intestine to the central nervous system (CNS). Smad7 overexpression worsened the clinical course of OSE and increased CNS inflammation and demyelination. It favored expansion of intestinal CD4+ T cells toward an inflammatory phenotype and migration of intestinal CD4+ T cells to the CNS. Intestinal biopsies from MS patients revealed decreased transforming growth factor beta signaling with a shift toward inflammatory T cell subtypes. Smad7 in intestinal T cells might represent a valuable therapeutic target for MS to achieve immunologic tolerance in the intestine and suppress CNS inflammation.

Tbet Expression in Regulatory T Cells Is Required to Initiate Th1-Mediated Colitis.

In normal conditions gut homeostasis is maintained by the suppressive activity of regulatory T cells (Tregs), characterized by the expression of the transcription factor FoxP3. In human inflammatory bowel disease, which is believed to be the consequence of the loss of tolerance toward antigens normally contained in the gut lumen, Tregs have been found to be increased and functionally active, thus pointing against their possible role in the pathogenesis of this immune-mediated disease. Though, in inflammatory conditions, Tregs have been shown to upregulate the T helper (Th) type 1-related transcription factor Tbet and to express the pro-inflammatory cytokine IFNγ, thus suggesting that at a certain point of the inflammatory process, Tregs might contribute to inflammation rather than suppress it. Starting from the observation that Tregs isolated from the lamina propria of active but not inactive IBD patients or uninflamed controls express Tbet and IFNγ, we investigated the functional role of Th1-like Tregs in the dextran sulfate model of colitis. As observed in human IBD, Th1-like Tregs were upregulated in the inflamed lamina propria of treated mice and the expression of Tbet and IFNγ in Tregs preceded the accumulation of conventional Th1 cells. By using a Treg-specific Tbet conditional knockout, we demonstrated that Tbet expression in Tregs is required for the development of colitis. Indeed, Tbet knockout mice developed milder colitis and showed an impaired Th1 immune response. In these mice not only the Tbet deficient Tregs but also the Tbet proficient conventional T cells showed reduced IFNγ expression. However, Tbet deficiency did not affect the Tregs suppressive capacity in vitro and in vivo in the adoptive transfer model of colitis. In conclusion here we show that Tbet expression by Tregs sustains the early phase of the Th1-mediated inflammatory response in the gut.

Nonalcoholic fatty liver disease in patients with inflammatory bowel disease: Beyond the natural history.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is a frequently reported condition in patients with inflammatory bowel disease (IBD). Both intestinal inflammation and metabolic factors are believed to contribute to the pathogenesis of IBD-associated NAFLD. AIM: To evaluate the prevalence of steatosis and liver fibrosis (LF) in a cohort of IBD patients and the identification of metabolic- and IBD-related risk factors for NAFLD and LF. METHODS: IBD patients were consecutively enrolled from December 2016 to January 2018. Demographic, anthropometric and biochemical data were collected so as eating habits. Abdominal ultrasound and transient elastography were performed to evaluate the presence of NAFLD and LF respectively. RESULTS: A total of 178 consecutive patients were enrolled and included in the analysis (95 Ulcerative colitis, 83 Crohn's disease). NAFLD was detected by imaging in 72 (40.4%) patients. Comparison between patients with and without NAFLD showed no significant differences in terms of IBD severity, disease duration, location/extension, use of IBD-related medications (i.e., steroids, anti-TNFs, and immunomodulators) and surgery. NAFLD was significantly associated with the presence of metabolic syndrome [MetS; odds ratio (OR): 4.13, P = 0.001] and obesity defined by body mass index (OR: 9.21, P = 0.0002). IBD patients with NAFLD showed higher caloric intake and lipid consumption than those without NAFLD, regardless disease activity. At the multivariate analysis, male sex, advanced age and high lipid consumption were independent risk factors for the development of NAFLD. An increased liver stiffness was detected in 21 patients (16%) and the presence of MetS was the only relevant factor associated to LF (OR: 3.40, P = 0.01). CONCLUSION: In this study, we demonstrate that risk factors for NAFLD and LF in the IBD population do not differ from those in the general population.

The Resolution of Intestinal Inflammation: The Peace-Keeper's Perspective.

The uncontrolled activation of the immune system toward antigens contained in the gut lumen in genetically predisposed subjects is believed to be the leading cause of inflammatory bowel disease (IBD). Two not mutually exclusive hypotheses can explain the pathogenic process leading to IBD. The first and mostly explored hypothesis states that the loss of tolerance toward gut microbiota antigens generates an aberrant inflammatory response that is perpetuated by continuous and unavoidable exposure to the triggering antigens. However, the discovery that the resolution of inflammation is not the mere consequence of clearing inflammatory triggers and diluting pro-inflammatory factors, but rather an active process in which molecular and cellular elements are involved, implies that a defect in the pro-resolving mechanisms might cause chronic inflammation in different immune-mediated diseases, including IBD. Here we review data on pro-resolving and counter-regulatory mechanisms involved in the resolution of inflammation, aiming to identify their possible involvement in the pathogenesis of IBD.

Heligmosomoides polygyrus bakeri Infection Decreases Smad7 Expression in Intestinal CD4+ T Cells, Which Allows TGF-ß to Induce IL-10-Producing Regulatory T Cells That Block Colitis.

Helminthic infections modulate host immunity and may protect their hosts from developing immunological diseases like inflammatory bowel disease. Induction of regulatory T cells (Tregs) may be an important part of this protective process. Heligmosomoides polygyrus bakeri infection also promotes the production of the regulatory cytokines TGF-ß and IL-10 in the gut. In the intestines, TGF-ß helps induce regulatory T cells. This study used Foxp3/IL-10 double reporter mice to investigate the effect of TGF-ß on the differentiation of colon and mesenteric lymph node-derived murine Foxp3- IL-10- CD4+ T cells into their regulatory phenotypes. Foxp3- IL-10- CD4+ T cells from H. polygyrus bakeri-infected mice, as opposed to T cells from uninfected animals, cultured in vitro with TGF-ß and anti-CD3/CD28 mAb differentiated into Foxp3+ and/or IL-10+ T cells. The IL-10-producing T cells nearly all displayed CD25. Smad7 is a natural inhibitor of TGF-ß signaling. In contrast to gut T cells from uninfected mice, Foxp3- IL10- CD4+ T cells from H. polygyrus bakeri-infected mice displayed reduced Smad7 expression and responded to TGF-ß with Smad2/3 phosphorylation. The TGF-ß-induced Tregs that express IL-10 blocked colitis when transferred into the Rag/CD25- CD4+ T cell transfer model of inflammatory bowel disease. TGF-ß had a greatly diminished capacity to induce Tregs in H. polygyrus bakeri-infected transgenic mice with constitutively high T cell-specific Smad7 expression. Thus, infection with H. polygyrus bakeri causes down-modulation in Smad7 expression in intestinal CD4+ T cells, which allows the TGF-ß produced in response to the infection to induce the Tregs that prevent colitis.

Alternative Splice Forms of CYLD Mediate Ubiquitination of SMAD7 to Prevent TGFB Signaling and Promote Colitis.

BACKGROUND & AIMS: The CYLD lysine 63 deubiquitinase gene (CYLD) encodes tumor suppressor protein that is mutated in familial cylindromatosus, and variants have been associated with Crohn disease (CD). Splice forms of CYLD that lack exons 7 and 8 regulate transcription factors and functions of immune cells. We examined the expression of splice forms of CYLD in colon tissues from patients with CD and their effects in mice. METHODS: We performed immunohistochemical analyses of colon tissues from patients with untreated CD and patients without inflammatory bowel diseases (controls). We obtained mice that expressed splice forms of CYLD (sCYLD mice) without or with SMAD7 (sCYLD/SMAD7 mice) from transgenes and CYLD-knockout mice (with or without transgenic expression of SMAD7) and performed endoscopic analyses. Colitis was induced in Rag1-/- mice by transfer of CD4+ CD62L+ T cells from C57/Bl6 or transgenic mice. T cells were isolated from mice and analyzed by flow cytometry and quantitative real-time polymerase chain reaction and intestinal tissues were analyzed by histology and immunohistochemistry. CYLD forms were expressed in mouse embryonic fibroblasts, primary T cells, and HEK293T cells, which were analyzed by immunoblot, mobility shift, and immunoprecipitation assays. RESULTS: The colonic lamina propria from patients with CD was infiltrated by T cells and had higher levels of sCYLD (but not full-length CYLD) and SMAD7 than tissues from controls. Incubation of mouse embryonic fibroblasts and T cells with transforming growth factor ß increased their production of sCYLD and decreased full-length CYLD. Transgenic expression of sCYLD and SMAD7 in T cells prevented the differentiation of regulatory T cells and T-helper type 17 cells and increased the differentiation of T-helper type 1 cells. The same effects were observed in colon tissues from sCYLD/SMAD7 mice but not in those from CYLD-knockout SMAD7 mice. The sCYLD mice had significant increases in the numbers of T-helper type 1 cells and CD44high CD62Llow memory-effector CD4+ T cells in the spleen and mesenteric lymph nodes compared with wild-type mice; sCYLD/SMAD7 mice had even larger increases. The sCYLD/SMAD7 mice spontaneously developed severe colitis, with infiltration of the colon by dendritic cells, neutrophils, macrophages, and CD4+ T cells and increased levels of Ifng, Il6, Il12a, Il23a, and Tnf mRNAs. Co-transfer of regulatory T cells from wild-type, but not from sCYLD/SMAD7, mice prevented the induction of colitis in Rag1-/- mice by CD4+ T cells. We found increased levels of poly-ubiquitinated SMAD7 in sCYLD CD4+ T cells. CYLD formed a nuclear complex with SMAD3, whereas sCYLD recruited SMAD7 to the nucleus, which inhibited the expression of genes regulated by SMAD3 and SMAD4. We found that sCYLD mediated lysine 63-linked ubiquitination of SMAD7. The sCYLD-SMAD7 complex inhibited transforming growth factor ß signaling in CD4+ T cells. CONCLUSIONS: Levels of the spliced form of CYLD are increased in colon tissues from patients with CD. sCYLD mediates ubiquitination and nuclear translocation of SMAD7 and thereby decreases transforming growth factor ß signaling in T cells. This prevents immune regulatory mechanisms and leads to colitis in mice.

RORγt-Expressing Tregs Drive the Growth of Colitis-Associated Colorectal Cancer by Controlling IL6 in Dendritic Cells.

Chronic inflammation drives colitis-associated colorectal cancer (CAC) in inflammatory bowel disease (IBD). FoxP3+ regulatory T cells (Treg) coexpressing the Th17-related transcription factor RORγt accumulate in the lamina propria of IBD patients, where they are thought to represent an intermediate stage of development toward a Th17 proinflammatory phenotype. However, the role of these cells in CAC is unknown. RORγt+FoxP3+ cells were investigated in human samples of CAC, and their phenotypic stability and function were investigated in an azoxymethane/dextran sulfate sodium model of CAC using Treg fate-mapping reporter and Treg-specific RORγt conditional knockout mice. Tumor development and the intratumoral inflammatory milieu were characterized in these mice. The functional role of CTLA-4 expressed by Tregs and FoxO3 in dendritic cells (DC) was studied in vitro and in vivo by siRNA-silencing experiments. RORγt expression identified a phenotypically stable population of tumor-infiltrating Tregs in humans and mice. Conditional RORγt knockout mice showed reduced tumor incidence, and dysplastic cells exhibited low Ki67 expression and STAT3 activation. Tumor-infiltrating DCs produced less IL6, a cytokine that triggers STAT3-dependent proliferative signals in neoplastic cells. RORγt-deficient Tregs isolated from tumors overexpressed CTLA-4 and induced DCs to have elevated expression of the transcription factor FoxO3, thus reducing IL6 expression. Finally, in vivo silencing of FoxO3 obtained by siRNA microinjection in the tumors of RORγt-deficient mice restored IL6 expression and tumor growth. These data demonstrate that RORγt expressed by tumor-infiltrating Tregs sustains tumor growth by leaving IL6 expression in DCs unchecked. Cancer Immunol Res; 6(9); 1082-92. ©2018 AACR.

Update on the Therapeutic Efficacy of Tregs in IBD: Thumbs up or Thumbs down?

Crohn's disease and ulcerative colitis, the 2 major forms of inflammatory bowel disease (IBD) in humans, arise in genetically predisposed individuals because of an abnormal immune response direct against constituents of the gut flora. Defects in counter-regulatory mechanisms are supposed to amplify and maintain the IBD-associated mucosal inflammation. Therefore, restoring the balance between inflammatory and anti-inflammatory pathways in the gut could contribute to halt the IBD-associated tissue-damaging immune response. Various suppressive T cell (Tregs) subsets have been characterized phenotypically and functionally and over the last decade, there has been enormous effort for optimizing the procedures for the in vitro expansion/generation of these cells for therapeutic purposes. Here we review the mechanisms of action and functional relevance of Tregs in the maintenance of gut inflammation and analyze the available data about the use of these cells in the treatment of IBD patients.

Smad7 knockdown activates protein kinase RNA-associated eIF2α pathway leading to colon cancer cell death.

Upregulation of Smad7, an inhibitor of transforming growth factor-ß1 (TGF-ß1), occurs in sporadic colorectal cancer (CRC) and knockdown of Smad7 inhibits CRC cell growth, a phenomenon that associates with decreased expression of cell division cycle 25 homolog A and arrest of cells in the S phase of the cell cycle. These findings occur in CRC cells unresponsive to TGF-ß1, thus suggesting the existence of a Smad7-mediated TGF-ß1-independent mechanism that controls CRC cell behavior. Here we show that Smad7 inhibition with a specific Smad7 antisense oligonucleotide upregulates eukaryotic translation initiation factor 2α (eIF2α) phosphorylation, a transcription factor involved in the regulation of cell cycle arrest and induction of cell death, and induces activating transcription factor 4 (ATF4) and CCAAT/enhancer binding protein homology protein (CHOP), two downstream targets of eIF2α. Among the upstream kinases that control eIF2α phosphorylation, the serine-threonine protein kinase RNA (PKR), but not general control non-derepressible 2 (GCN2) and protein kinase RNA-like endoplasmic reticulum kinase (PERK), is activated by Smad7 knockdown. PKR silencing abolishes Smad7 antisense-induced eIF2α phosphorylation and ATF4/CHOP induction, thereby preventing Smad7 antisense-driven cell death. Smad7 inhibition diminishes interaction of PKR with protein kinase inhibitor p58 (p58IPK), a cellular inhibitor of PKR, but does not change the expression and/or activity of other factors involved in the control of PKR activation. These findings delineate a novel mechanism by which Smad7 knockdown promotes CRC cell death.