Background and Aims: In ulcerative colitis [UC], mucosal damage occurs in areas that are infiltrated with neutrophils. The antimicrobial function of neutrophils relies in part on the formation of extracellular web-like structures, named neutrophil extracellular traps [NETs]. The formation and/or clearance of aberrant NETs have been associated with several immune diseases. Here we investigated the role of NETs in UC-related inflammation. Methods: The expression of NET-associated proteins was evaluated in colonic biopsies of patients with Crohn's disease [CD], UC and in normal controls [NC] by Western blotting, immunofluorescence and immunohistochemistry. Colonic biopsies of UC patients were analysed before and after antitumour necrosis factor α [anti-TNF-α] treatment. The capacity of neutrophils to produce NETs upon activation was tested in vitro. UC lamina propria mononuclear cells [LPMCs] were cultured with NETs in the presence or absence of an extracellular signal-regulated kinase-1/2 [ERK1/2] inhibitor and inflammatory cytokine induction was assessed by real-time polymerase chain reaction and enzyme-linked immunosorbent assay. We also characterized the contribution of NETs in dextran sodium sulfate [DSS]-induced colitis. Results: NET-associated proteins were over-expressed in inflamed colon of UC patients as compared to CD patients and NC. Circulating neutrophils of UC patients produced NETs in response to TNF-α stimulation, and reduced expression of NET-related proteins and diminished NET formation were seen in patients receiving successful treatment with anti-TNF-α. Treatment of UC LPMCs with NETs activated ERK1/2, thus enhancing TNF-α and interleukin-1β [IL-1β] production. NETs were induced in mice with DSS-colitis and in vivo inhibition of NET release attenuated colitis. Conclusions: Our data show that NET release occurs in UC and suggest a role for NETs in sustaining mucosal inflammation in this disorder.

The complement system is a potent component of the innate immune response, promoting inflammation and orchestrating defense against pathogens. However, dysregulation of complement is critical to several autoimmune and inflammatory syndromes. Elevated expression of the proinflammatory cytokine IL-1β is often linked to such diseases. In this study, we reveal the mechanistic link between complement and IL-1β secretion using murine dendritic cells. IL-1β secretion occurs following intracellular caspase-1 activation by inflammasomes. We show that complement elicits secretion of both IL-1β and IL-18 in vitro and in vivo via the NLRP3 inflammasome. This effect depends on the inflammasome components NLRP3 and ASC, as well as caspase-1 activity. Interestingly, sublethal complement membrane attack complex formation, but not the anaphylatoxins C3a and C5a, activated the NLRP3 inflammasome in vivo. These findings provide insight into the molecular processes underlying complement-mediated inflammation and highlight the possibility of targeting IL-1β to control complement-induced disease and pathological inflammation.

P53 is critically important in preventing oncogenesis but its role in inflammation in general and in the function of inflammatory macrophages in particular is not clear. Here, we show that bone marrow-derived macrophages exhibit endogenous p53 activity, which is increased when macrophages are polarized to the M2 (alternatively activated macrophage) subtype. This leads to reduced expression of M2 genes. Nutlin-3a, which destabilizes the p53/MDM2 (mouse double minute 2 homolog) complex, promotes p53 activation and further downregulates M2 gene expression. In contrast, increased expression of M2 genes was apparent in M2-polarized macrophages from p53-deficient and p53 mutant mice. Furthermore, we show, in mice, that p53 also regulates M2 polarization in peritoneal macrophages from interleukin-4-challenged animals and that nutlin-3a retards the development of tolerance to Escherichia coli lipopolysaccharide. P53 acts via transcriptional repression of expression of c-Myc (v-myc avian myelocytomatosis viral oncogene homolog) gene by directly associating with its promoter. These data establish a role for the p53/MDM2/c-MYC axis as a physiological 'brake' to the M2 polarization process. This work reveals a hitherto unknown role for p53 in macrophages, provides further insight into the complexities of macrophage plasticity and raises the possibility that p53-activating drugs, many of which are currently being trialled clinically, may have unforeseen effects on macrophage function. Macrophages have key roles in the response to stress, injury, infection and inflammation. The M1 (classically activated macrophages) are induced by lipopolysaccharide (LPS) and cytokines such as interferon-γ (IFNγ) and characterized by the expression of a wide range of proinflammatory genes. M2 (alternatively activated macrophages) are induced by T helper type 2 (Th2) cytokines such as interleukin-4 (IL4) and IL13 and express high levels of anti-inflammatory and tissue repair marker genes. M2 macrophages perform immunoregulatory functions including defense against infection, promotion of angiogenesis and wound healing. 1 Macrophages exist on a continuum between M1 and M2 subtypes undergoing dynamic changes between these different functional states depending on changes in their microenvironment. This 'plasticity' involves extensive changes in macrophage gene sets and provides the potential to develop drugs to manipulate the macrophage subtype. 2 As such, macrophage polarization, and its molecular basis, has been vigorously researched in recent years.P53 has a crucial role in cancer by controlling the expression of genes involved in apoptosis, cell cycle arrest, metabolism and DNA repair. 3 While inflammation is increasingly recognized as a factor in determining the predisposition to cancer, 4 the role of p53 in inflammation is not clear. Macrophages from p53 − / − mice produce increased quantities of proinflammatory cytokines in response to LPS, 5 while peritoneal macrophage count and susceptibility to lethal septic shock are increased in p53 − ...

Signal transducers and activators of transcription (STATs) mediate essential signaling pathways in different biological processes, including immune responses, hematopoiesis, and neurogenesis. Among the STAT members, STAT3 plays crucial roles in cell proliferation, survival, and differentiation. While STAT3 activation is transient in physiological conditions, STAT3 becomes persistently activated in a high percentage of solid and hematopoietic malignancies (e.g., melanoma, multiple myeloma, breast, prostate, ovarian, and colon cancers), thus contributing to malignant transformation and progression. This makes STAT3 an attractive therapeutic target for cancers. Initial strategies aimed at inhibiting STAT3 functions have focused on blocking the action of its activating kinases or sequestering its DNA binding ability. More recently, the diffusion of proteomic-based techniques, which have allowed for the identification and characterization of novel STAT3-interacting proteins able to modulate STAT3 activity via its subcellular localization, interact with upstream kinases, and recruit transcriptional machinery, has raised the possibility to target such cofactors to specifically restrain STAT3 oncogenic functions. In this article, we summarize the available data about the function of STAT3 interactors in malignant cells and discuss their role as potential therapeutic targets for cancer treatment.

Background & AimsFood additives, such as emulsifiers, stabilizers, or bulking agents, are present in the Western diet and their consumption is increasing. However, little is known about their potential effects on intestinal homeostasis. In this study we examined the effect of some of these food additives on gut inflammation.MethodsMice were given drinking water containing maltodextrin (MDX), propylene glycol, or animal gelatin, and then challenged with dextran sulfate sodium or indomethacin. In parallel, mice fed a MDX-enriched diet were given the endoplasmic reticulum (ER) stress inhibitor tauroursodeoxycholic acid (TUDCA). Transcriptomic analysis, real-time polymerase chain reaction, mucin-2 expression, phosphorylated p38 mitogen-activated protein (MAP) kinase quantification, and H&E staining was performed on colonic tissues. Mucosa-associated microbiota composition was characterized by 16S ribosomal RNA sequencing. For the in vitro experiments, murine intestinal crypts and the human mucus-secreting HT29-methotrexate treated cell line were stimulated with MDX in the presence or absence of TUDCA or a p38 MAP kinase inhibitor.ResultsDiets enriched in MDX, but not propylene glycol or animal gelatin, exacerbated intestinal inflammation in both models. Analysis of the mechanisms underlying the detrimental effect of MDX showed up-regulation of inositol requiring protein 1β, a sensor of ER stress, in goblet cells, and a reduction of mucin-2 expression with no significant change in mucosa-associated microbiota. Stimulation of murine intestinal crypts and HT29-methotrexate treated cell line cells with MDX induced inositol requiring protein 1β via a p38 MAP kinase–dependent mechanism. Treatment of mice with TUDCA prevented mucin-2 depletion and attenuated colitis in MDX-fed mice.ConclusionsMDX increases ER stress in gut epithelial cells with the downstream effect of reducing mucus production and enhancing colitis susceptibility.

Our data indicated that exposure of intestinal mononuclear cells to a high-NaCl diet enhanced effector cytokine production and contributed to the exacerbation of experimental colitis in mice.

IBD (inflammatory bowel disease)-related tissue damage occurs in areas which are massively infiltrated with monocytes/macrophages. These cells respond to inflammatory stimuli with enhanced production of cytokines/chemokines. In the present study, we analysed the expression and role of IL (interleukin)-34, a regulator of monocyte/macrophage differentiation, survival and function, in IBD. A significant increase in IL-34 mRNA and protein expression was seen in inflamed mucosa of patients with CD (Crohn's disease) and patients with UC (ulcerative colitis) compared with the uninvolved areas of the same patients and normal controls. IL-34 was up-regulated in LPMCs (lamina propria mononuclear cells) isolated from normal colon by TNF-α (tumour necrosis factor α) and TLR (Toll-like receptor) ligands and was down-regulated in intestinal biopsies and LPMCs of IBD patients upon treatment with infliximab. Treatment of normal LPMCs with IL-34 increased TNF-α expression in an ERK1/2 (extracellular-signal-regulated kinase 1/2)-dependent fashion and neutralization of IL-34 in IBD mucosal explants reduced TNF-α and IL-6 synthesis. In conclusion, our results indicate that IL-34 is up-regulated in IBD and suggest a role for this cytokine in sustaining the inflammatory responses in this disease.

Growing evidence is unveiling a role for poly(ADP-ribose) polymerase (PARP)-1 in the regulation of inflammatory/immune responses. In the current study, we investigated the effects of PARP-1 deficiency on regulatory T cell differentiation. Increased numbers of regulatory CD4+CD25+/Foxp3+ T cells were found in thymus, spleen, and lymph nodes of PARP-1 knockout (KO) mice compared with wild-type (WT) controls. The increased frequency of regulatory T cells in the periphery resulted in impaired CD4 cell proliferation and IL-2 production, which could be restored by CD25+ cell depletion. Phenotype and inhibitory functions of PARP-1 KO regulatory T cells were similar to WT cells, indicating that PARP-1 affects regulatory T cell differentiation rather than function. Purified naive CD4 cells from PARP-1 KO mice stimulated in vitro expressed forkhead box p3 mRNA at higher levels and generated a greater number of Foxp3+ cells (inducible regulatory T [iTreg] cells) than the WT counterpart. This finding was due to a higher rate of naive CD4 cell to Foxp3+ iTreg cell conversion rather than to higher resistance to apoptosis induction. Interestingly, PARP-1 deficiency did not affect retinoid-related orphan receptor-γt mRNA expression and differentiation of purified naive CD4 cells to Th17 cells. PARP-1 KO iTreg cells showed features similar to WT regulatory T cells, suggesting that modulation of PARP-1 during the immune response might be used to induce greater numbers of functional regulatory T cells. In conclusion, our findings represent the first evidence that PARP-1 can affect regulatory T cell differentiation and open new perspectives on potential targets for modulating immune responses.