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The objective was to identify the main factors influencing micronutrient policies in the opinion of policy actors in ten European countries. Study was carried out during Jan-Nov 2010 in European countries: the Czech Republic, Denmark, England, Germany, Greece, Italy, the Netherlands, Norway, Poland and Spain. Semi-structured qualitative interviews were conducted with representatives of stakeholders involved in the vitamin D, folate and iodine policy making process. Fifty eight key informants representing mainly scientific advisory bodies (n=24) and governmental organisations (n=19) participated in the study. The remaining interviewees represented non-governmental organisations (n=6), industry (n=4) or were independent academic or health professional experts (n=5). Data were analysed by theoretical interpretative thematic analysis. Insights from interviewees on the development of micronutrient policies were grouped using the Public Health Nutrition Policy-making model. The main factors influencing the micronutrient policies were: systematic monitoring of nutrition and health, causal relationships between consumers’ diet-related behaviours and health outcomes, scientific recommendations from national bodies (Science area); scientific recommendations from international authorities and experiences of other countries, EU legislation, cultural factors (Wider context) and political environment, national capacity to deal with the problem, national legislation, economics, stakeholder engagement, relationships between stakeholders (Policy and institutions area). The spectrum and weight of the factors influencing nutritional policy depends on nutrient, country and degree of its “advanced status” within nutrition policy, political environment, culture and socio-economic conditions as well as the point of view (who is expressing the opinion).

Objective: Low Glycemic Index (GI) diets improve cardiometabolic risk (CMR) specifically in those with insulin resistance. However, the prospective association between pasta (a low GI staple) consumption and CMR is unclear. We evaluated the longitudinal association of pasta consumption with CMR (after 2 y: body weight, body mass index (BMI), waist circumference (WC), blood pressure (BP); after 1 y: fasting blood glucose, HbA1c, HDL-cholesterol and triglycerides) in ∼6000 older adults (50% women) at high CMR., Methods: Consumption of pasta and other staples were determined as the cumulative average of reported intakes at baseline and annual follow-up visits from food frequency questionnaires and defined as energy-adjusted (residuals) and the number of daily servings. Longitudinal association between pasta consumption and CMR was assessed in PREDIMED-Plus participants (Trail registry number: ISRCTN89898870 )., Results: Mean (SD) dry pasta intake was 9(7) g/d at Year 1 and 8(6) g/d at Year 2. In linear regression models, higher pasta intake was associated with greater 2 y decreases in body weight, BMI and WC. When fully adjusted, every additional serving of pasta was associated with significantly greater 2 y decreases in body weight (-2.23(-3.47, -0.98 kg), BMI (-0.86(-1.27, -0.34 kg/m 2 ) and WC (-1.92 (-3.46, -0.38 cm). There was no evidence of association with other outcomes. Additionally, substituting equivalent servings of pasta for white bread or white rice or potato was significantly associated with greater 2 y decreases in body weight and BMI. Replacing white bread with pasta was associated with higher 2 y reductions in WC. Replacing potato with pasta was associated with improvements in diastolic BP and HDL-cholesterol. Conclusions: Equivalent serving substitutions of white bread/white rice/potato with pasta may help reduce CMR in older Mediterranean adults with overweight/obesity. While such substitutions are feasible where pasta consumption aligns with the local gastronomic culture, the feasibility and potential CMR benefit of such interventions should be confirmed in other populations.

Uncontrolled regeneration leads to neoplastic transformation 1-3 . The intestinal epithelium requires precise regulation during continuous homeostatic and damage-induced tissue renewal to prevent neoplastic transformation, suggesting that pathways unlinking tumour growth from regenerative processes must exist. Here, by mining RNA-sequencing datasets from two intestinal damage models 4,5 and using pharmacological, transcriptomics and genetic tools, we identified liver X receptor (LXR) pathway activation as a tissue adaptation to damage that reciprocally regulates intestinal regeneration and tumorigenesis. Using single-cell RNA sequencing, intestinal organoids, and gain- and loss-of-function experiments, we demonstrate that LXR activation in intestinal epithelial cells induces amphiregulin (Areg), enhancing regenerative responses. This response is coordinated by the LXR-ligand-producing enzyme CYP27A1, which was upregulated in damaged intestinal crypt niches. Deletion of Cyp27a1 impaired intestinal regeneration, which was rescued by exogenous LXR agonists. Notably, in tumour models, Cyp27a1 deficiency led to increased tumour growth, whereas LXR activation elicited anti-tumour responses dependent on adaptive immunity. Consistently, human colorectal cancer specimens exhibited reduced levels of CYP27A1, LXR target genes, and B and CD8 T cell gene signatures. We therefore identify an epithelial adaptation mechanism to damage, whereby LXR functions as a rheostat, promoting tissue repair while limiting tumorigenesis., Competing Interests: Competing interests: E.J.V. has received research grants from F. Hoffmann-La Roche and is a founder of PaperVids. S.D. works as a consultant for Cellphi Biotechnology. The other authors declare no competing interests., (© 2024. The Author(s).)

Aims: Pannexin-1 (PANX1) is a hemichannel that releases ATP upon opening, initiating inflammation, cell proliferation, and migration. However, the role of PANX1 channels in colon cancer remains poorly understood, thus constituting the focus of this study., Main Methods: PANX1 mRNA expression was analyzed using multiple cancer databases. PANX1 protein expression and distribution were evaluated by immunohistochemistry on primary tumor tissue and non-tumor colonic mucosa from colon cancer patients. PANX1 inhibitors (probenecid or 10 Panx) were used to assess colon cancer cell lines viability. To study the role of PANX1 in vivo, a subcutaneous xenograft model using HCT116 cells was performed in BALB/c NOD/SCID immunodeficient mice to evaluate tumor growth under PANX1 inhibition using probenecid., Key Findings: PANX1 mRNA was upregulated in colon cancer tissue compared to non-tumor colonic mucosa. Elevated PANX1 mRNA expression in tumors correlated with worse disease-free survival. PANX1 protein abundance was increased on tumor cells compared to epithelial cells in paired samples, in a cancer stage-dependent manner. In vitro and in vivo experiments indicated that blocking PANX1 reduced cell viability and tumor growth., Significance: PANX1 can be used as a biomarker of colon cancer progression and blocking PANX1 channel opening could be used as a potential therapeutic strategy against this disease., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Improving patient selection and development of biological therapies such as vedolizumab in IBD requires a thorough understanding of the mechanism of action and target binding, thereby providing individualised treatment strategies. We aimed to visualise the macroscopic and microscopic distribution of intravenous injected fluorescently labelled vedolizumab, vedo-800CW, and identify its target cells using fluorescence molecular imaging (FMI)., Design: Forty three FMI procedures were performed, which consisted of macroscopic in vivo assessment during endoscopy, followed by macroscopic and microscopic ex vivo imaging. In phase A, patients received an intravenous dose of 4.5 mg, 15 mg vedo-800CW or no tracer prior to endoscopy. In phase B, patients received 15 mg vedo-800CW preceded by an unlabelled (sub)therapeutic dose of vedolizumab., Results: FMI quantification showed a dose-dependent increase in vedo-800CW fluorescence intensity in inflamed tissues, with 15 mg (153.7 au (132.3-163.7)) as the most suitable tracer dose compared with 4.5 mg (55.3 au (33.6-78.2)) (p=0.0002). Moreover, the fluorescence signal decreased by 61% when vedo-800CW was administered after a therapeutic dose of unlabelled vedolizumab, suggesting target saturation in the inflamed tissue. Fluorescence microscopy and immunostaining showed that vedolizumab penetrated the inflamed mucosa and was associated with several immune cell types, most prominently with plasma cells., Conclusion: These results indicate the potential of FMI to determine the local distribution of drugs in the inflamed target tissue and identify drug target cells, providing new insights into targeted agents for their use in IBD., Trial Registration Number: NCT04112212., Competing Interests: Competing interests: GD received research grants from Royal DSM, Takeda and Janssen Pharmaceuticals and speaker fees from AbbVie, Pfizer, Takeda and Janssen Pharmaceuticals. EAMF is supported by a ZonMW Clinical Fellowship grant (project number 90719075) and has received an unrestricted research grant from Takeda. MD and SJ are employees and shareholders of Regeneron Pharmaceuticals., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

This study investigates the anti-inflammatory effects of pectins with different degrees of methyl esterification (DM) on intestinal epithelial cells (IECs) expressing low and high levels of TLR2. It also studies the influence of soluble TLR2 (sTLR2) which may be enhanced in patients with inflammatory bowel syndrome on the inflammation-attenuating effects of pectins. Also, it examines the impact of pectins on tight junction gene expression in IECs. Lemon pectins with DM18 and DM88 were characterized, and their effects on TLR2-1-induced IL8 gene expression and secretion were investigated in low-TLR2 expressing Caco-2 and high-TLR2 expressing DLD-1 cells. The results demonstrate that both DM18 and DM88 pectins can counteract TLR2-1-induced IL-8 expression and secretion, with more pronounced effects observed in DLD-1 cells expressing high levels of TLR2. Furthermore, the presence of sTLR2 does not interfere with the attenuating effects of low DM18 pectin and may even support its anti-inflammatory effects in Caco-2 cells. The impact of pectins and sTLR2 on tight junction gene expression also demonstrates cell-type-dependent effects. Overall, these findings suggest that low DM pectins possess potent anti-inflammatory properties and may influence tight junction gene expression in IECs, thereby contributing to the maintenance of gut homeostasis.

From the venom of the Bothrops pictus snake, an endemic species from Peru, we recently have described toxins that inhibited platelet aggregation and cancer cell migration. In this work, we characterize a novel P-III class snake venom metalloproteinase, called pictolysin-III (Pic-III). It is a 62 kDa proteinase that hydrolyzes dimethyl casein, azocasein, gelatin, fibrinogen, and fibrin. The cations Mg 2+ and Ca 2+ enhanced its enzymatic activity, whereas Zn 2+ inhibited it. In addition, EDTA and marimastat were also effective inhibitors. The amino acid sequence deduced from cDNA shows a multidomain structure that includes a proprotein, metalloproteinase, disintegrin-like, and cysteine-rich domains. Additionally, Pic-III reduces the convulxin- and thrombin-stimulated platelet aggregation and in vivo, it has hemorrhagic activity (DHM = 0.3 µg). In epithelial cell lines (MDA-MB-231 and Caco-2) and RMF-621 fibroblast, it triggers morphological changes that are accompanied by a decrease in mitochondrial respiration, glycolysis, and ATP levels, and an increase in NAD(P)H, mitochondrial ROS, and cytokine secretion. Moreover, Pic-III sensitizes to the cytotoxic BH3 mimetic drug ABT-199 (Venetoclax) in MDA-MB-231 cells. To our knowledge, Pic-III is the first SVMP reported with action on mitochondrial bioenergetics and may offer novel opportunities for promising lead compounds that inhibit platelet aggregation or ECM-cancer-cell interactions.

Inflammatory Bowel Disease (IBD) is characterized by a loss of intestinal barrier function caused by an aberrant interaction between the immune response and the gut microbiota. In IBD, imbalance in cholesterol homeostasis and mitochondrial bioenergetics have been identified as essential events for activating the inflammasome-mediated response. Mitochondrial alterations, such as reduced respiratory complex activities and reduced production of tricarboxylic acid (TCA) cycle intermediates (e.g., citric acid, fumarate, isocitric acid, malate, pyruvate, and succinate) have been described in in vitro and clinical studies. Under inflammatory conditions, mitochondrial architecture in intestinal epithelial cells is dysmorphic, with cristae destruction and high dynamin-related protein 1 (DRP1)-dependent fission. Likewise, these alterations in mitochondrial morphology and bioenergetics promote metabolic shifts towards glycolysis and down-regulation of antioxidant Nuclear erythroid 2-related factor 2 (Nrf2)/Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) signaling. Although the mechanisms underlying the mitochondrial dysfunction during mucosal inflammation are not fully understood at present, metabolic intermediates and cholesterol may act as signals activating the NLRP3 inflammasome in IBD. Notably, dietary phytochemicals exhibit protective effects against cholesterol imbalance and mitochondrial function alterations to maintain gastrointestinal mucosal renewal in vitro and in vivo conditions. Here, we discuss the role of cholesterol and mitochondrial metabolism in IBD, highlighting the therapeutic potential of dietary phytochemicals, restoring intestinal metabolism and function., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Astorga, Gasaly, Dubois-Camacho, De la Fuente, Landskron, Faber, Urra and Hermoso.)

Adherent-invasive E. coli (AIEC) is a pathotype associated with the etiopathogenesis of Crohn's disease (CD), albeit with an as-yet unclear role. The main pathogenic mechanisms described for AIEC are adherence to epithelial cells, invasion of epithelial cells, and survival and replication within macrophages. A few virulence factors have been described as participating directly in these phenotypes, most of which have been evaluated only in AIEC reference strains. To date, no molecular markers have been identified that can differentiate AIEC from other E. coli pathotypes, so these strains are currently identified based on the phenotypic characterization of their pathogenic mechanisms. The identification of putative AIEC molecular markers could be beneficial not only from the diagnostic point of view but could also help in better understanding the determinants of AIEC pathogenicity. The objective of this study was to identify molecular markers that contribute to the screening of AIEC strains. For this, we characterized outer membrane protein (OMP) profiles in a group of AIEC strains and compared them with the commensal E. coli HS strain. Notably, we found a set of OMPs that were present in the AIEC strains but absent in the HS strain. Moreover, we developed a PCR assay and performed phylogenomic analyses to determine the frequency and distribution of the genes coding for these OMPs in a larger collection of AIEC and other E. coli strains. As result, it was found that three genes ( chuA , eefC , and fitA ) are widely distributed and significantly correlated with AIEC strains, whereas they are infrequent in commensal and diarrheagenic E. coli strains (DEC). Additional studies are needed to validate these markers in diverse strain collections from different geographical regions, as well as investigate their possible role in AIEC pathogenicity.

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) and can be treated with glucocorticoids (GC), although some patients are unresponsive to this therapy. The transcription factor LRH-1/ NR5A2 is critical to intestinal cortisol production (intestinal steroidogenesis), being reduced in UC patients. However, the relationship between LRH-1 expression and distribution with altered corticosteroid responses is unknown. To address this, we categorized UC patients by their steroid response. Here, we found that steroid-dependent and refractory patients presented reduced glucocorticoid receptor (GR)-mediated intestinal steroidogenesis compared to healthy individuals and responder patients, possibly related to increased colonic mucosa GR isoform beta (GRβ) content and cytoplasmic LRH-1 levels in epithelial and lamina propria cells. Interestingly, an intestinal epithelium-specific GR-induced knockout (GR iKO ) dextran sodium sulfate (DSS)-colitis mice model presented decreased epithelial LRH-1 expression, whilst it increased in the lamina propria compared to DSS-treated control mice. Mechanistically, GR directly induced NR5A2 gene expression in CCD841CoN cells and human colonic organoids. Furthermore, GR bound to two glucocorticoid-response elements within the NR5A2 promoter in dexamethasone-stimulated CCD841CoN cells. We conclude that GR contributes to intestinal steroidogenesis by inducing LRH-1 in epithelial cells, suggesting LRH-1 as a potential marker for glucocorticoid-impaired response in UC. However, further studies with a larger patient cohort will be necessary to confirm role of LRH-1 as a therapeutic biomarker.

The prognosis of severe COVID-19 patients has motivated research communities to uncover mechanisms of SARS-CoV-2 pathogenesis also on a regional level. In this work, we aimed to understand the immunological dynamics of severe COVID-19 patients with different degrees of illness, and upon long-term recovery. We analyzed immune cellular subsets and SARS-CoV-2-specific antibody isotypes of 66 COVID-19 patients admitted to the Hospital Clínico Universidad de Chile, which were categorized according to the WHO ten-point clinical progression score. These included 29 moderate patients (score 4-5) and 37 severe patients under either high flow oxygen nasal cannula (18 patients, score 6), or invasive mechanical ventilation (19 patients, score 7-9), plus 28 convalescent patients and 28 healthy controls. Furthermore, six severe patients that recovered from the disease were longitudinally followed over 300 days. Our data indicate that severe COVID-19 patients display increased frequencies of plasmablasts, activated T cells and SARS-CoV-2-specific antibodies compared to moderate and convalescent patients. Remarkably, within the severe COVID-19 group, patients rapidly progressing into invasive mechanical ventilation show higher frequencies of plasmablasts, monocytes, eosinophils, Th1 cells and SARS-CoV-2-specific IgG than patients under high flow oxygen nasal cannula. These findings demonstrate that severe COVID-19 patients progressing into invasive mechanical ventilation show a distinctive type of immunity. In addition, patients that recover from severe COVID-19 begin to regain normal proportions of immune cells 100 days after hospital discharge and maintain high levels of SARS-CoV-2-specific IgG throughout the study, which is an indicative sign of immunological memory. Thus, this work can provide useful information to better understand the diverse outcomes of severe COVID-19 pathogenesis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 García-González, Tempio, Fuentes, Merino, Vargas, Simon, Ramirez-Pereira, Rojas, Tobar, Landskron, Araya, Navarrete, Bastias, Tordecilla, Varas, Maturana, Marcoleta, Allende, Naves, Hermoso, Salazar-Onfray, Lopez, Bono and Osorio.)

Background/aim: The functions of interleukin 33 (IL-33) in cholangiocarcinoma (CCA) are unclear. This study aimed to evaluate the roles of IL-33 in CCA progression., Materials and Methods: The effect of intracellular IL-33 using shIL-33 knocked down KKU-055 (IL-33KD-KKU-055) compared to parental (Pa) KKU-055 and extracellular IL-33 using recombinant human IL-33 (rhIL-33) treatment on the proliferation and invasion of CCA cells grown in 3D cultures was studied. Relevant markers were determined by western blot or ELISA., Results: IL-33KD-KKU-055 cells showed increased proliferation and invasion in 3D cultures compared to Pa-KKU-055 cells, with NF-κB and IL-6 up-regulation. Treatment with 2 ng/ml rhIL-33 promoted Pa-KKU-055 cell proliferation by inducing NF-κB and IL-6 expressions. Upon GSK-3β inactivation and increased nuclear full-length IL-33 (flIL-33), 20 ng/ml rhIL-33 had no effect on proliferation. Both 2 and 20 ng/ml rhIL-33 induced proliferation and invasion of IL-33-negative KKU-213 cells in 3D cultures, as well as NF-κB and IL-6 up-regulation., Conclusion: Intracellular and extracellular IL-33 have distinct roles in the mechanisms of CCA progression., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Signals from the tumor microenvironment (TME) have a profound influence on the maintenance and progression of cancers. Chronic inflammation and the infiltration of immune cells in breast cancer (BC) have been strongly associated with early carcinogenic events and a switch to a more immunosuppressive response. Cancer-associated fibroblasts (CAFs) are the most abundant stromal component and can modulate tumor progression according to their secretomes. The immune cells including tumor-infiltrating lymphocytes (TILs) (cytotoxic T cells (CTLs), regulatory T cells (Tregs), and helper T cell (Th)), monocyte-infiltrating cells (MICs), myeloid-derived suppressor cells (MDSCs), mast cells (MCs), and natural killer cells (NKs) play an important part in the immunological balance, fluctuating TME between protumoral and antitumoral responses. In this review article, we have summarized the impact of these immunological players together with CAF secreted substances in driving BC progression. We explain the crosstalk of CAFs and tumor-infiltrating immune cells suppressing antitumor response in BC, proposing these cellular entities as predictive markers of poor prognosis. CAF-tumor-infiltrating immune cell interaction is suggested as an alternative therapeutic strategy to regulate the immunosuppressive microenvironment in BC., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2021 Jarupa Soongsathitanon et al.)

Gut microbiota is known to be transferred from the mother to their offspring. This study determines whether the innate microbiota of rats selectively bred for generations as high alcohol drinkers play a role in their alcohol intake. Wistar-derived high-drinker UChB rats (intake 10-g ethanol/kg/day) administered nonabsorbable oral antibiotics before allowing access to alcohol, reducing their voluntary ethanol intake by 70%, an inhibition that remained after the antibiotic administration was discontinued. Oral administration of Lactobacillus rhamnosus Gorbach-Goldin (GG) induced the synthesis of FGF21, a vagal β-Klotho receptor agonist, and partially re-invoked a mechanism that reduces alcohol intake. The vagus nerve constitutes the main axis transferring gut microbiota information to the brain ("microbiota-gut-brain" axis). Bilateral vagotomy inhibited rat alcohol intake by 75%. Neither antibiotic treatment nor vagotomy affected total fluid intake. A microbiota-mediated marked inflammatory environment was observed in the gut of ethanol-naïve high-drinker rats, as gene expression of proinflammatory cytokines (TNF-α; IL-6; IL-1β) was significantly reduced by nonabsorbable antibiotic administration. Gut cytokines are known to activate the vagus nerve, while vagal activation induces pro-rewarding effects in nucleus accumbens. Both alcoholics and alcohol-preferring rats share a marked preference for sweet tastes-likely an evolutionary trait to seek sweet fermented fruits. Saccharin intake by UChB rats was inhibited by 75%-85% by vagotomy or oral antibiotic administration, despite saccharin-induced polydipsia. Overall, data indicate that the mechanisms that normally curtail heavy drinking are inhibited in alcohol-preferring animals and inform a gut microbiota origin. Whether it applies to other mammals and humans merits further investigation., (© 2021 Society for the Study of Addiction.)

Inflammatory bowel diseases (IBDs), which include ulcerative colitis (UC) and Crohn's disease (CD), cause chronic inflammation of the gut, affecting millions of people worldwide. IBDs have been frequently associated with an alteration of the gut microbiota, termed dysbiosis, which is generally characterized by an increase in abundance of Proteobacteria such as Escherichia coli , and a decrease in abundance of Firmicutes such as Faecalibacterium prausnitzii (an indicator of a healthy colonic microbiota). The mechanisms behind the development of IBDs and dysbiosis are incompletely understood. Using samples from colonic biopsies, we studied the mucosa-associated intestinal microbiota in Chilean and Spanish patients with IBD. In agreement with previous studies, microbiome comparison between IBD patients and non-IBD controls indicated that dysbiosis in these patients is characterized by an increase of pro-inflammatory bacteria (mostly Proteobacteria) and a decrease of commensal beneficial bacteria (mostly Firmicutes). Notably, bacteria typically residing on the mucosa of healthy individuals were mostly obligate anaerobes, whereas in the inflamed mucosa an increase of facultative anaerobe and aerobic bacteria was observed. We also identify potential co-occurring and mutually exclusive interactions between bacteria associated with the healthy and inflamed mucosa, which appear to be determined by the oxygen availability and the type of respiration. Finally, we identified a panel of bacterial biomarkers that allow the discrimination between eubiosis from dysbiosis with a high diagnostic performance (96% accurately), which could be used for the development of non-invasive diagnostic methods. Thus, this study is a step forward towards understanding the landscapes and alterations of mucosa-associated intestinal microbiota in patients with IBDs., Competing Interests: Conflict of Interest: Funding institutions did not influence the study design or interpretation of the result. Moreover, the authors declare that they have no competing interests., (Copyright: © 2021 Chamorro et al.)

The diverse and dynamic microbial community of the human gastrointestinal tract plays a vital role in health, with gut microbiota supporting the development and function of the gut immune barrier. Crosstalk between microbiota-gut epithelium and the gut immune system determine the individual health status, and any crosstalk disturbance may lead to chronic intestinal conditions, such as inflammatory bowel diseases (IBD) and celiac disease. Microbiota-derived metabolites are crucial mediators of host-microbial interactions. Some beneficially affect host physiology such as short-chain fatty acids (SCFAs) and secondary bile acids. Also, tryptophan catabolites determine immune responses, such as through binding to the aryl hydrocarbon receptor (AhR). AhR is abundantly present at mucosal surfaces and when activated enhances intestinal epithelial barrier function as well as regulatory immune responses. Exogenous diet-derived indoles (tryptophan) are a major source of endogenous AhR ligand precursors and together with SCFAs and secondary bile acids regulate inflammation by lowering stress in epithelium and gut immunity, and in IBD, AhR expression is downregulated together with tryptophan metabolites. Here, we present an overview of host microbiota-epithelium- gut immunity crosstalk and review how microbial-derived metabolites contribute to host immune homeostasis. Also, we discuss the therapeutic potential of bacterial catabolites for IBD and celiac disease and how essential dietary components such as dietary fibers and bacterial tryptophan catabolites may contribute to intestinal and systemic homeostasis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Gasaly, de Vos and Hermoso.)

Colorectal cancer (CRC) is one of the most common cancers worldwide. As with other cancers, CRC is a multifactorial disease due to the combined effect of genetic and environmental factors. Most cases are sporadic, but a small proportion is hereditary, estimated at around 5-10%. In both, the tumor interacts with heterogeneous cell populations, such as endothelial, stromal, and immune cells, secreting different signals (cytokines, chemokines or growth factors) to generate a favorable tumor microenvironment for cancer cell invasion and metastasis. There is ample evidence that inflammatory processes have a role in carcinogenesis and tumor progression in CCR. Different profiles of cell activation of the tumor microenvironment can promote pro or anti-tumor pathways; hence they are studied as a key target for the control of cancer progression. Additionally, the intestinal mucosa is in close contact with a microorganism community, including bacteria, bacteriophages, viruses, archaea, and fungi composing the gut microbiota. Aberrant composition of this microbiota, together with alteration in the diet-derived microbial metabolites content (such as butyrate and polyamines) and environmental compounds has been related to CRC. Some bacteria, such as pks+ Escherichia coli or Fusobacterium nucleatum , are involved in colorectal carcinogenesis through different pathomechanisms including the induction of genetic mutations in epithelial cells and modulation of tumor microenvironment. Epithelial and immune cells from intestinal mucosa have Pattern-recognition receptors and G-protein coupled receptors (receptor of butyrate), suggesting that their activation can be regulated by intestinal microbiota and metabolites. In this review, we discuss how dynamics in the gut microbiota, their metabolites, and tumor microenvironment interplays in sporadic and hereditary CRC, modulating tumor progression., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Hanus, Parada-Venegas, Landskron, Wielandt, Hurtado, Alvarez, Hermoso, López-Köstner and De la Fuente.)

This review describes current evidence supporting butyrate impact in the homeostatic regulation of the digestive ecosystem in health and inflammatory bowel diseases (IBDs). Butyrate is mainly produced by bacteria from the Firmicutes phylum. It stimulates mature colonocytes and inhibits undifferentiated malignant and stem cells. Butyrate oxidation in mature colonocytes (1) produces 70-80% of their energetic requirements, (2) prevents stem cell inhibition by limiting butyrate access to crypts, and (3) consumes oxygen, generating hypoxia and maintaining luminal anaerobiosis favorable to the microbiota. Butyrate stimulates the aryl hydrocarbon receptor (AhR), the GPR41 and GPR109A receptors, and inhibits HDAC in different cell types, thus stabilizing the gut barrier function and decreasing inflammatory processes. However, some studies indicate contrary effects according to butyrate concentrations. IBD patients exhibit a lower abundance of butyrate-producing bacteria and butyrate content. Additionally, colonocyte butyrate oxidation is depressed in these subjects, lowering luminal anaerobiosis and facilitating the expansion of Enterobacteriaceae that contribute to inflammation. Accordingly, gut dysbiosis and decreased barrier function in IBD seems to be secondary to the impaired mitochondrial disturbance in colonic epithelial cells.

Initially described as Th2 promoter cytokine, more recently, IL-33 has been recognized as an alarmin, mainly in epithelial and endothelial cells. While localized in the nucleus acting as a gene regulator, it can be also released after injury, stress or inflammatory cell death. As proinflammatory signal, IL-33 binds to the surface receptor ST2, which enhances mast cell, Th2, regulatory T cell, and innate lymphoid cell type 2 functions. Besides these Th2 roles, free IL-33 can activate CD8 + T cells during ongoing Th1 immune responses to potentiate its cytotoxic function. Celiac Disease (CD) is a chronic inflammatory disorder characterized by a predominant Th1 response leading to multiple pathways of mucosal damage in the proximal small intestine. By immunofluorescence and western blot analysis of duodenal tissues, we found an increased expression of IL-33 in duodenal mucosa of active CD (ACD) patients. Particularly, locally digested IL-33 releases active 18/21kDa fragments which can contribute to expand the proinflammatory signal. Endothelial (CD31 + ) and mesenchymal, myofibroblast and pericyte cells from microvascular structures in villi and crypts, showed IL-33 nuclear location; while B cells (CD20 + ) showed a strong cytoplasmic staining. Both ST2 forms, ST2L and sST2, were also upregulated in duodenal mucosa of CD patients. This was accompanied by increased number of CD8 + ST2 + T cells and the expression of T-bet in some ST2 + intraepithelial lymphocytes and lamina propria cells. IL-33 and sST2 mRNA levels correlated with IRF1, an IFN induced factor relevant in responses to viral infections and interferon mediated proinflammatory responses highly represented in duodenal tissues in ACD. These findings highlight the potential contribution of IL-33 and its fragments to exacerbate the proinflammatory circuit and potentiate the cytotoxic activity of CD8 + T cells in CD pathology., (Copyright © 2020 Perez, Ruera, Miculan, Carasi, Dubois-Camacho, Garbi, Guzman, Hermoso and Chirdo.)

Interleukin 33 (IL-33) promotes cholangiocarcinoma (CCA) genesis in a mouse model, however, its function in human CCA has not been clearly understood. This study was aimed to investigate IL-33 level in CCA tissues and its clinicopathological correlations. The results revealed that IL-33 was found in both cancer cells and stromal cancer-associated fibroblast (CAFs) staining patterns which were divided into high (CH) and low level (CL) in cancer cells; and presence (FP) and absence (FA) in CAFs. Kaplan-Meier analysis showed that patients in the CL group were significantly correlated with a short 2-year survival time ( P = 0.027). The CL/FP group had a shorter survival time compared to the other groups with statistical significance for 2-year ( P = 0.030) and 5-year ( P = 0.023) survivals. In contrast, CH/FP patients had significantly greater 2-year ( P = 0.003) and 5-year ( P = 0.003) survivals. Univariate and multivariate analysis confirmed that CL/FP was a significantly independent risk factor whereas CH/FP was a significant protective factor in CCA patients. High IL-33 expressing CCA cells had low migration, but they showed increased migration when IL-33 expression was knocked down. The low level of recombinant human IL-33 (rhIL-33) (0.002 - 2 ng/ml) could promote CCA cell migration, in contrast to the suppressive effect at a high dose (20 - 200 ng/ml). In conclusion, the combination of high IL-33 level in cancer cells and CAFs is a potentially good prognosis marker in CCA patients. The in vitro migration suppressive effect of IL-33 may be the potential mechanism supporting its role as a good prognostic marker in CCA patients. The obtained results strengthen IL-33 as a promising predictor and therapeutic target for CCA., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

A subset of oral carcinomas is etiologically related to high-risk human papillomavirus (HR-HPV) infection, with HPV16 being the most frequent HR-HPV type found in these carcinomas. The oncogenic role of HR-HPV is strongly dependent on the overexpression of E6 and E7 oncoproteins, which, in turn, induce p53 and pRb degradation, respectively. Additionally, it has been suggested that HR-HPV oncoproteins are involved in the regulation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), inducing cancer progression and metastasis. Previously, we reported that HPV16 E7 oncoprotein promotes Pirin upregulation resulting in increased epithelial-mesenchymal transition (EMT) and cell migration, with Pirin being an oxidative stress sensor and activator of NF-κB. In this study, we demonstrate the mechanism by which HPV16 E7-mediated Pirin overexpression occurs by promoting EGFR/PI3K/AKT1/NRF2 signaling, thus causing PIR/NF-κB activation in oral tumor cells. Our results demonstrate a new mechanism by which E7 contributes to oral cancer progression, proposing PIR as a potential new therapeutic target.

Objectives: Xerostomia in SS patients has been associated with low quality and quantity of salivary mucins, which are fundamental for the hydration and protection of the oral mucosa. The aim of this study was to evaluate if cytokines induce aberrant mucin expression and whether tauroursodeoxycholic acid (TUDCA) is able to counteract such an anomaly., Methods: Labial salivary glands from 16 SS patients and 15 control subjects, as well as 3D acini or human submandibular gland cells stimulated with TNF-α or IFN-γ and co-incubated with TUDCA, were analysed. mRNA and protein levels of Mucin 1 (MUC1) and MUC7 were determined by RT-qPCR and western blot, respectively. Co-immunoprecipitation and immunofluorescence assays for mucins and GRP78 [an endoplasmic reticulum (ER)-resident protein] were also performed. mRNA levels of RelA/p65 (nuclear factor-κB subunit), TNF-α, IL-1β, IL-6, SEL1L and EDEM1 were determined by RT-qPCR, and RelA/p65 localization was evaluated by immunofluorescence., Results: MUC1 is overexpressed and accumulated in the ER of labial salivary gland from SS patients, while MUC7 accumulates throughout the cytoplasm of acinar cells; however, MUC1, but not MUC7, co-precipitated with GRP78. TUDCA diminished the overexpression and aberrant accumulation of MUC1 induced by TNF-α and IFN-γ, as well as the nuclear translocation of RelA/p65, together with the expression of inflammatory and ER stress markers in 3D acini., Conclusion: Chronic inflammation alters the secretory process of MUC1, inducing ER stress and affecting the quality of saliva in SS patients. TUDCA showed anti-inflammatory properties decreasing aberrant MUC1 accumulation. Further studies are necessary to evaluate the potential therapeutic effect of TUDCA in restoring glandular homeostasis in SS patients., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Glucocorticoids are potent endogenous anti-inflammatory molecules, and their cognate receptor, glucocorticoid receptor (GR), is expressed in nearly all immune cells. Macrophages are heterogeneous immune cells having a central role in both tissue homeostasis and inflammation and also play a role in the pathogenesis of some inflammatory diseases. Paradoxically, glucocorticoids have only a limited efficacy in controlling the resolution of these macrophage-related diseases. Here, we report that the transcriptomes of monocyte-like THP-1 cells and macrophage-like THP-1 cells (THP1-MΦ) have largely conserved gene expression patterns. In contrast, the differentiation to THP1-MΦ significantly altered the sensitivity of gene transcription to glucocorticoids. Among glucocorticoid-regulated genes, we identified the exopeptidase dipeptidyl peptidase-4 ( DPP4 ) as a critical glucocorticoid-responsive gene in THP1-MΦ. We found that GR directly induces DPP4 gene expression by binding to two glucocorticoid-responsive elements (GREs) within the DPP4 promoter. Additionally, we show that glucocorticoid-induced DPP4 expression is blocked by the GR antagonist RU-486 and by GR siRNA transfection and that DPP4 enzyme activity is reduced by DPP4 inhibitors. Of note, glucocorticoids highly stimulated macrophage mobility; unexpectedly, DPP4 mediated the glucocorticoid-induced macrophage migration, and siRNA-mediated knockdowns of GR and DPP4 blocked dexamethasone-induced THP1-MΦ migration. Moreover, glucocorticoid-induced DPP4 activation was also observed in proinflammatory M1-polarized murine macrophages, as well as peritoneal macrophages, and was associated with increased macrophage migration. Our results indicate that glucocorticoids directly up-regulate DPP4 expression and thereby induce migration in macrophages, potentially explaining why glucocorticoid therapy is less effective in controlling macrophage-dominated inflammatory disorders.

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) characterized by mucosa damage associated with an uncontrolled inflammatory response. This immunological impairment leads to altered inflammatory mediators such as IL-33, which is shown to increase in the mucosa of active UC (aUC) patients. MicroRNAs present a distorted feature in inflamed colonic mucosa and are potential IL-33 regulating candidates in UC. Therefore, we studied the microRNA and mRNA profiles in inflamed colonic samples of UC patients, evaluating the effect of a microRNA (selected by in silico analysis and its expression in UC patients), on IL-33 under inflammatory conditions. We found that inflamed mucosa ( n = 8) showed increased expression of 40 microRNAs and 2,120 mRNAs, while 49 microRNAs and 1,734 mRNAs were decreased, as determined by microarrays. In particular, IL-33 mRNA showed a 3.8-fold increase and eight members of a microRNA family (miR-378), which targets IL-33 mRNA in the 3'UTR, were decreased (-3.9 to -3.0 times). We selected three members of the miR-378 family (miR-378a-3p, miR-422a, and miR-378c) according to background information and interaction energy analysis, for further correlation analyses with IL-33 expression through qPCR and ELISA, respectively. We determined that aUC ( n = 24) showed high IL-33 levels, and decreased expression of miR-378a-3p and miR-422a compared to inactive UC ( n = 10) and controls ( n = 6). Moreover, both microRNAs were inversely correlated with IL-33 expression, while miR-378c does not show a significant difference. To evaluate the effect of TNFα on the studied microRNAs, aUC patients with anti-TNF therapy were compared to aUC receiving other treatments. The levels of miR-378a-3p and miR-378c were higher in aUC patients with anti-TNF. Based on these findings, we selected miR-378a-3p to exploring the molecular mechanism involved by in vitro assays, showing that over-expression of miR-378a-3p decreased the levels of an IL-33 target sequence β-gal-reporter gene in HEK293 cells. Stable miR-378a-3p over-expression/inhibition inversely modulated IL-33 content and altered viability of HT-29 cells. Additionally, in an inflammatory context, TNFα decreased miR-378a-3p levels in HT-29 cells enhancing IL-33 expression. Together, our results propose a regulatory mechanism of IL-33 expression exerted by miR-378a-3p in an inflammatory environment, contributing to the understanding of UC pathogenesis., (Copyright © 2019 Dubois-Camacho, Diaz-Jimenez, De la Fuente, Quera, Simian, Martínez, Landskron, Olivares-Morales, Cidlowski, Xu, Gao, Xie, Chnaiderman, Soto-Rifo, González, Calixto and Hermoso.)

In colorectal cancer (CRC), cancer-associated fibroblasts (CAFs) are the most abundant component from the tumor microenvironment (TM). CAFs facilitate tumor progression by inducing angiogenesis, immune suppression and invasion, thus altering the organization/composition of the extracellular matrix (i.e., desmoplasia) and/or activating epithelial-mesenchymal transition (EMT). Soluble factors from the TM can also contribute to cell invasion through secretion of cytokines and recently, IL-33/ST2 pathway has gained huge interest as a protumor alarmin, promoting progression to metastasis by inducing changes in TM. Hence, we analyzed IL-33 and ST2 content in tumor and healthy tissue lysates and plasma from CRC patients. Tissue localization and distribution of these molecules was evaluated by immunohistochemistry (using localization reference markers α-smooth muscle actin or α-SMA and E-cadherin), and clinical/histopathological information was obtained from CRC patients. In vitro experiments were conducted in primary cultures of CAFs and normal fibroblasts (NFs) isolated from tumor and healthy tissue taken from CRC patients. Additionally, migration and proliferation analysis were performed in HT29 and HCT116 cell lines. It was found that IL-33 content increases in left-sided CRC patients with lymphatic metastasis, with localization in tumor epithelia associated with abundant desmoplasia. Although ST2 content showed similarities between tumor and healthy tissue, a decreased immunoreactivity was observed in left-sided tumor stroma, associated to metastasis related factors (advanced stages, abundant desmoplasia, and presence of tumor budding). A principal component analysis (including stromal and epithelial IL-33/ST2 and α-SMA immunoreactivity with extent of desmoplasia) allowed us to distinguish clusters of low, intermediate and abundant desmoplasia, with potential to develop a diagnostic signature with benefits for further therapeutic targets. IL-33 transcript levels from CAFs directly correlated with CRC cell line migration induced by CAFs conditioned media, with rhIL-33 inducing a mesenchymal phenotype in HT29 cells. These results indicate a role of IL-33/ST2 in tumor microenvironment, specifically in the interaction between CAFs and epithelial tumor cells, thus contributing to invasion and metastasis in left-sided CRC, most likely by activating desmoplasia.

Ulcerative colitis (UC) and Crohn's disease (CD), collectively known as Inflammatory Bowel Diseases (IBD), are caused by a complex interplay between genetic, immunologic, microbial and environmental factors. Dysbiosis of the gut microbiome is increasingly considered to be causatively related to IBD and is strongly affected by components of a Western life style. Bacteria that ferment fibers and produce short chain fatty acids (SCFAs) are typically reduced in mucosa and feces of patients with IBD, as compared to healthy individuals. SCFAs, such as acetate, propionate and butyrate, are important metabolites in maintaining intestinal homeostasis. Several studies have indeed shown that fecal SCFAs levels are reduced in active IBD. SCFAs are an important fuel for intestinal epithelial cells and are known to strengthen the gut barrier function. Recent findings, however, show that SCFAs, and in particular butyrate, also have important immunomodulatory functions. Absorption of SCFAs is facilitated by substrate transporters like MCT1 and SMCT1 to promote cellular metabolism. Moreover, SCFAs may signal through cell surface G-protein coupled receptors (GPCRs), like GPR41, GPR43, and GPR109A, to activate signaling cascades that control immune functions. Transgenic mouse models support the key role of these GPCRs in controlling intestinal inflammation. Here, we present an overview of microbial SCFAs production and their effects on the intestinal mucosa with specific emphasis on their relevance for IBD. Moreover, we discuss the therapeutic potential of SCFAs for IBD, either applied directly or by stimulating SCFAs-producing bacteria through pre- or probiotic approaches.

Here, we determined the 5-hydroxymethylcytosine (5hmC), 5-methylcytosine (5mC), Ten Eleven Translocation (TETs), and DNA methyltransferases (DNMTs) levels in epithelial and inflammatory cells of labial salivary glands (LSG) from Sjögren's syndrome (SS)-patients and the effect of cytokines on HSG cells. LSG from SS-patients, controls and HSG cells incubated with cytokines were analysed. Levels of 5mC, 5hmC, DNMTs, TET2 and MeCP2 were assessed by immunofluorescence. In epithelial cells from SS-patients, an increase in TET2, 5hmC and a decrease in 5mC and MeCP2 were observed, additionally, high levels of 5mC and DNMTs and low levels of 5hmC were detected in inflammatory cells. Cytokines increased TET2 and 5hmC and decreased 5mC levels. Considering that the TET2 gene.promoter contains response elements for transcription factors activated by cytokines, together to in vitro results suggest that changes in DNA hydroxymethylation, resulting from altered levels of TET2 are likely to be relevant in the Sjögren's syndrome etiopathogenesis., (Copyright © 2018 Elsevier Inc. All rights reserved.)

For many years, researchers in the field of autoimmunity have focused on the role of the immune components in the etiopathogenesis of autoimmune diseases. However, some studies have demonstrated the importance of target tissues in their pathogenesis and the breach of immune tolerance. The immune system as well as target tissue cells (plasmatic, β-pancreatic, fibroblast-like synoviocytes, thyroid follicular and epithelial cells of the lachrymal glands, salivary glands, intestine, bronchioles and renal tubules) share the characteristic of secretory cells with an extended endoplasmic reticulum (ER). The function of these cells depends considerably on a normal ER function and calcium homeostasis, so they can produce and secrete their main components, which include glycoproteins involved in antigenic presentation such as major histocompatibility complex (MHC) class I and II. All these proteins are synthesized and modified in the ER, and for this reason disturbances in the normal functions of this organelle such as protein folding, protein quality control, calcium homeostasis and redox balance, promote accumulation of unfolded or misfolded proteins, a condition known as ER stress. Autoimmune diseases are characterized by inflammation, which has been associated with an ER stress condition. Interestingly, patients with these diseases contain circulating auto-antibodies against chaperone proteins (such as Calnexin and GRP94), thus affecting the folding and assembly of MHC class I and II glycoproteins and their loading with peptide. The main purpose of this article is to review the involvement of the protein quality control and unfolded protein response (UPR) in the ER protein homeostasis (proteostasis) and their alterations in autoimmune diseases. In addition, we describe the interaction between ER stress and inflammation and evidences are shown of how autoimmune diseases are associated with an ER stress condition, with a special emphasis on the second most prevalent autoimmune rheumatic disease, Sjögren's syndrome., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objectives: Labial salivary glands (LSGs) of SS patients show alterations related to endoplasmic reticulum stress. Glandular dysfunction could be partly the consequence of an altered inositol-requiring enzyme 1α (IRE1α)/X box-binding protein 1 (XBP-1) signalling pathway of the unfolded protein response, which then regulates genes involved in biogenesis of the secretory machinery. This study aimed to determine the expression, promoter methylation and localization of the IRE1α/XBP-1 pathway components in LSGs of SS patients and also their expression induced by IFN-γ in vitro., Methods: IRE1α, XBP-1 and glucose-regulated protein 78 (GRP78) mRNA and protein levels were measured by qPCR and western blot, respectively, in LSGs of SS patients (n = 47) and control subjects (n = 37). Methylation of promoters was evaluated by methylation-sensitive high resolution melting, localization was analysed by immunofluorescence and induction of the IRE1α/XBP-1 pathway components by IFN-γ was evaluated in 3D acini., Results: A significant decrease of IRE1α, XBP-1u, XBP-1s, total XBP-1 and GRP78 mRNAs was observed in LSGs of SS patients, which was correlated with increased methylation levels of their respective promoters, and consistently the protein levels for IRE1α, XBP-1s and GRP78 were observed to decrease. IFN-γ decreased the mRNA and protein levels of XBP-1s, IRE1α and GRP78, and increased methylation of their promoters. Significant correlations were also found between IRE1α/XBP-1 pathway components and clinical parameters., Conclusion: Decreased mRNA levels for IRE1α, XBP-1 and GRP78 can be partially explained by hypermethylation of their promoters and is consistent with chronic endoplasmic reticulum stress, which may explain the glandular dysfunction observed in LSGs of SS patients. Additionally, glandular stress signals, including IFN-γ, could modulate the expression of the IRE1α/XBP-1 pathway components.

Crohn's disease (CD) is a chronic inflammatory bowel disorder characterized by deregulated inflammation triggered by environmental factors. Notably, adherent-invasive Escherichia coli (AIEC), a bacterium with the ability to survive within macrophages is believed to be one of such factors. Glucocorticoids are the first line treatment for CD and to date, it is unknown how they affect bactericidal and inflammatory properties of macrophages against AIEC. The aim of this study was to evaluate the impact of glucocorticoid treatment on AIEC infected macrophages. First, THP-1 cell-derived macrophages were infected with a CD2-a AIEC strain, in the presence or absence of the glucocorticoid dexamethasone (Dex) and mRNA microarray analysis was performed. Differentially expressed mRNAs were confirmed by TaqMan-qPCR. In addition, an amikacin protection assay was used to evaluate the phagocytic and bactericidal activity of Dex-treated macrophages infected with E. coli strains (CD2-a, HM605, NRG857c, and HB101). Finally, cytokine secretion and the inflammatory phenotype of macrophages were evaluated by ELISA and flow cytometry, respectively. The microarray analysis showed that CD2-a, Dex, and CD2-a + Dex-treated macrophages have differential inflammatory gene profiles. Also, canonical pathway analysis revealed decreased phagocytosis signaling by Dex and anti-inflammatory polarization on CD2-a + Dex macrophages. Moreover, amikacin protection assay showed reduced phagocytosis upon Dex treatment and TaqMan-qPCR confirmed Dex inhibition of three phagocytosis-associated genes. All bacteria strains induced TNF-α, IL-6, IL-23, CD40, and CD80, which was inhibited by Dex. Thus, our data demonstrate that glucocorticoids impair phagocytosis and induce anti-inflammatory polarization after AIEC infection, possibly contributing to the survival of AIEC in infected CD patients.

Inflammatory bowel diseases (IBDs), such as ulcerative colitis and Crohn's disease, are chronic pathologies associated with a deregulated immune response in the intestinal mucosa, and they are triggered by environmental factors in genetically susceptible individuals. Exogenous glucocorticoids (GCs) are widely used as anti-inflammatory therapy in IBDs. In the past, patients with moderate or severe states of inflammation received GCs as a first line therapy with an important effectiveness in terms of reduction of the disease activity and the induction of remission. However, this treatment often results in detrimental side effects. This downside drove the development of second generation GCs and more precise (non-systemic) drug-delivery methods. Recent clinical trials show that most of these new treatments have similar effectiveness to first generation GCs with fewer adverse effects. The remaining challenge in successful treatment of IBDs concerns the refractoriness and dependency that some patients encounter during GCs treatment. A deeper understanding of the molecular mechanisms underlying GC response is key to personalizing drug choice for IBDs patients to optimize their response to treatment. In this review, we examine the clinical characteristics of treatment with GCs, followed by an in depth analysis of the proposed molecular mechanisms involved in its resistance and dependence associated with IBDs. This thorough analysis of current clinical and biomedical literature may help guide physicians in determining a course of treatment for IBDs patients and identifies important areas needing further study., Competing Interests: Conflict-of-interest statement: No potential conflicts of interest.

Background: Different strains of invasive Escherichia coli (E. coli), isolated from intestinal mucosa of patients, are related to the pathogenesis of inflammatory bowel diseases (IBD)., Aim: To evaluate an association between intracellular E. coli and IBD; its clinical characteristics and use of steroids., Material and Methods: Sixty one patients with Crohn's disease and 83 with ulcerative colitis were studied. To determine the intracellular E. coli content, colonoscopy biopsies of these patients and 29 control subjects were processed using the gentamicin protection assay. Differences in the bacterial content between patient groups were evaluated using Mann-Whitney test, while the association between presence of E. coli with endoscopic activity, location/extension and use of corticosteroid as anti-inflammatory treatment were evaluated with Fisher's exact test or Chi-square test., Results: E. coli strains were detected in 36.1, 39.3 and 10.3% of patients with ulcerative colitis, Crohn's disease and controls, respectively. The number of bacteria per biopsy in Crohn's disease and ulcerative colitis was significantly higher than in controls (p < 0.01 between patients and controls). In ulcerative colitis, significant associations were found between the presence of bacteria and disease location and use of corticosteroids. In Crohn's disease, no association was found., Conclusions: IBD are associated with the presence of intracellular E. coli strains in the intestinal mucosa, suggesting an alteration in the microbiota or loss of integrity of the epithelial barrier. The association of intracellular E. coli with clinical features and the use of corticosteroids in ulcerative colitis suggests that different factors could promote colonization or proliferation of these bacteria.

The ST2/IL33 signalling pathway has been associated with ulcerative colitis (UC). ST2, encoded by the IL1RL1 gene, is expressed as both a membrane-anchored receptor (ST2L) activated by IL33 and as a soluble receptor (sST2) with anti-inflammatory properties. In UC patients, sST2 is further increased by corticosteroid treatment; however, the glucocorticoid-mediated molecular regulation remains unknown. We therefore tested whether genetic variants in the IL1RL1 distal promoter are involved in UC and affect glucocorticoid-mediated ST2 expression. Serum ST2 levels and genetic variants in the IL1RL1 distal promoter were examined by ELISA and PCR sequencing in UC patients receiving corticosteroids. Glucocorticoid-mediated ST2 production was evaluated in intestinal mucosa cultures. Molecular regulation of glucocorticoid-mediated ST2 was assessed by RT-qPCR, ChIP assay and luciferase reporter assay. Dexamethasone effect on ST2 transcript expression was analyzed in leukocytes and related to IL1RL1 variants. Sequencing of a distal IL1RL1 promoter region demonstrated that SNPs rs6543115(C) and rs6543116(A) are associated with increased sST2 in UC patients on corticosteroids. Dexamethasone up-regulated sST2 transcription through interaction with the glucocorticoid-response element (GRE) carrying rs6543115(C) variant. Our data indicate that IL1RL1 SNPs rs6543115(C) confer susceptibility to UC and is contained in the GRE, which may modulate glucocorticoid-induced sST2 expression.

Background: The ST2/IL-33 pathway has been related to ulcerative colitis (UC), and soluble ST2 (sST2), to disease severity. We tested the potential usefulness of sST2 as a predictive marker of treatment response and patients' outcome., Methods: Twenty-six patients with active UC were prospectively recruited and grouped according to an endoscopic score and therapy response. Colonoscopic biopsies were collected at baseline and 6 months or when patients showed clinical activity. The protocol was reinitiated in patients requiring rescue therapy. Blood and stool were collected at baseline, 1, 3, 6 and 12 months. Serum and mucosal ST2, and fecal calprotectin (FC) content were determined by ELISA and correlated to Mayo clinical and endoscopic subscore. Intestinal ST2 was evaluated by immunofluorescence. Wilcoxon signed rank test and Spearman correlations (Rs) were applied (p <0.05)., Results: Follow-up was completed in 24 patients. sST2 levels (median and range) varied from 173.5 [136.6-274.0] to 86.5 [54.6-133.2] in responders (p < 0.05), and 336.3 [211.0-403.2] to 385.3 pg/mL [283.4-517.3] in non-responders at baseline and 6 months, respectively. sST2 levels correlated with Mayo clinical and endoscopic subscore, mucosal ST2 and FC (Rs = 0.57, 0.66, 0.74 and 0.42, respectively; p < 0.0001) and showed a trend similar to that of FC in responders. Non-responders revealed an increased ST2 content, restricted to the lamina propria's cellular infiltrate., Conclusions: Consecutive sST2 measurement to follow changes in inflammatory activity of UC patients who respond or not to treatment identifies sST2, like FC, as a useful biomarker in predicting clinical outcome of UC patients.

Unlabelled: Cardiac inflammation can be produced by pathogen-associated molecular patterns (PAMPs), from parasitic, bacterial or viral origin; or by danger-associated molecular patterns (DAMPs), released from dead cells after cardiac tissue damage, for example by cardiac infarction. Both, PAMPS and DAMPS activate TLR4 on resident immune cells and heart tissue cells, triggering an inflammatory process necessary to begin the wound healing process. Cardiac fibroblasts (CF) are the most abundant cells in the heart and are critical to wound healing, along with cardiac myofibroblasts (CMF), which are differentiated from CF through a TGF-β1-mediated process. While TLR4 and the inflammasome complex are known to play important roles in CF function, the effects of TGF-β1 on TLR4 and inflammasome expression and activity remain unknown. To elucidate this important point, we evaluated the effect of TGF-β1 on TLR4, and the inflammasome protein expression and activity through activation by LPS, mimicking a myocarditis condition by bacterial origin. We found that TGF-β1 increased TLR4 expression in CF and that the process was mediated by the TGFβRI and p38 signaling pathways. In both CF and CMF, LPS triggered ERK1/2, PI3K-Akt, and p65-NF-κB phosphorylation. All of these effects were blocked by TAK-242, a TLR4 signaling pathway inhibitor. LPS increased pro-IL-1β levels, which were dependent on the ERK1/2, PI3K-Akt, and NF-κB signaling pathways, and levels were higher in CF than CMF. NLRP3 and ASC levels were similar in CF and CMF, while pro-caspase-1 levels and caspase-1 activity were higher in CMF. LPS+ATP treatment induced inflammasome complex assembly and activation, triggering the release of IL-1β in both CMF and CF. Finally, the unsecreted pro-IL-1β in the CF was degraded by autophagy., Conclusion: TGF-β1 increases TLR4 expression in CF. Despite different pro-IL-1β and caspase-1 activity levels in CF versus CMF, the two cell types secreted similar levels of IL-1β after LPS+ATP treatment. These findings suggest that both cell types are active participants in inflammation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

IL-33, an IL-1 family member, is expressed by many cell types and can regulate gene transcription. IL-33 is released upon cell necrosis and the precursor form is enzymatically processed, and then drives inflammation as a damage-associated molecular pattern. The IL-33 receptor ST2, encoded by IL1RL1, is expressed as both a membrane-anchored receptor (ST2L) activated by IL-33, and as a soluble variant (sST2) that exhibits anti-inflammatory properties. The IL-33/ST2 axis is involved in the pathogenesis of atopic and autoimmune diseases, cancer, and central nervous system disorders. Here, we review recent findings on the role of the IL-33/ST2 axis in health and disease., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Asp-Glu-Ala-Asp (DEAD)-box polypeptide 3, or DDX3, belongs to the DEAD-box family of ATP-dependent RNA helicases and is known to play different roles in RNA metabolism ranging from transcription to nuclear export, translation, and assembly of stress granules. In addition, there is growing evidence that DDX3 is a component of the innate immune response against viral infections. As such, DDX3 has been shown to play roles both upstream and downstream of I-kappa beta kinase ε (IKKε)/TANK-binding kinase 1, leading to IFN-β production. Interestingly, several RNA viruses, including human threats such as HIV-1 and hepatitis C virus, hijack DDX3 to accomplish various steps of their replication cycles. Thus, it seems that viruses have evolved to exploit DDX3's functions while threatening the innate immune response. Understanding this interesting dichotomy in DDX3 function will help us not only to improve our knowledge of virus-host interactions but also to develop novel antiviral drugs targeting the multifaceted roles of DDX3 in viral replication., (Copyright © 2015 John Wiley & Sons, Ltd.)

Objectives: A hallmark characteristic of SS patients is the ectopic presence of the mucins MUC5B and MUC7 in the extracellular matrix of salivary glands that have lost apical-basolateral acinar-cell polarity. This study aims to determine whether exogenous salivary mucins induce gene expression of pro-inflammatory cytokines, as well as to evaluate whether the Toll-like receptor-4 (TLR4) pathway is involved in this response., Methods: Differentiated human submandibular gland (HSG) cells were stimulated with mucins or oligosaccharide residues at different concentrations and for different periods of time. The expression of pro-inflammatory cytokines and their receptors was determined by semi-quantitative real time PCR (sqPCR). TLR4-mediated responses induced by mucin were evaluated with the Toll-IL-1 receptor domain containing adaptor protein (TIRAP) inhibitory peptide or using anti-hTLR4 blocking antibody. TLR4-receptor expression was also determined in SS patients, controls and HSG cells., Results: Mucins induced a significant increase in CXCL8, TNF-α, IFN-α, IFN-β, IL-6 and IL-1β, but not B cell activating factor (BAFF). Cytokine induction was mediated by TLR4, as shown using TIRAP or using anti-hTLR4 antibody. Sugar residues present in MUC5B, such as sulpho-Lewis (SO3-3Galβ1-3GlcNAc), also induced cytokines. Unexpectedly, mucins induced MUC5B, but not MUC7 expression., Conclusion: Salivary mucins were recognized by TLR4 in epithelial cells initiating a pro-inflammatory response that could attract inflammatory cells to amplify and perpetuate inflammation and thereby contribute to the development of a chronic state characteristic of SS. The ectopic localization of MUC5B and MUC7 in the salivary gland extracellular matrix from SS patients and the current results reveal the importance of salivary epithelial cells in innate immunity, as well as in SS pathogenesis., (© The Author 2015. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Perinatal asphyxia (PA) is a leading cause of neuronal damage in newborns, resulting in long-term neurological and cognitive deficits, in part due to impairment of mesostriatal and mesolimbic neurocircuitries. The insult can be as severe as to menace the integrity of the genome, triggering the overactivation of sentinel proteins, including poly (ADP-ribose) polymerase-1 (PARP-1). PARP-1 overactivation implies increased energy demands, worsening the metabolic failure and depleting further NAD(+) availability. Using a global PA rat model, we report here evidence that hypoxia increases PARP-1 activity, triggering a signalling cascade leading to nuclear translocation of the NF-κB subunit p65, modulating the expression of IL-1β and TNF-α, pro-inflammatory molecules, increasing apoptotic-like cell death in mesencephalon of neonate rats, monitored with Western blots, qPCR, TUNEL and ELISA. PARP-1 activity increased immediately after PA, reaching a maximum 1-8 h after the insult, while activation of the NF-κB signalling pathway was observed 8 h after the insult, with a >twofold increase of p65 nuclear translocation. IL-1β and TNF-α mRNA levels were increased 24 h after the insult, together with a >twofold increase in apoptotic-like cell death. A single dose of the PARP-1 inhibitor nicotinamide (0.8 mmol/kg, i.p.), 1 h post delivery, prevented the effect of PA on PARP-1 activity, p65 translocation, pro-inflammatory cytokine expression and apoptotic-like cell death. The present study demonstrates that PA leads to PARP-1 overactivation, increasing the expression of pro-inflammatory cytokines and cell death in mesencephalon, effects prevented by systemic neonatal nicotinamide administration, supporting the idea that PARP-1 inhibition represents a therapeutic target against the effects of PA.

Toll-like receptor (TLR) 2, a type I membrane receptor that plays a key role in innate immunity, recognizes conserved molecules in pathogens, and triggering an inflammatory response. It has been associated with inflammatory and autoimmune diseases. Soluble TLR2 (sTLR2) variants have been identified in human body fluids, and the TLR2 ectodomain can negatively regulate TLR2 activation by behaving as a decoy receptor. sTLR2 generation does not involve alternative splicing mechanisms, indicating that this process might involve a post-translational modification of the full-length receptor; however, the specific mechanism has not been studied. Using CD14+ peripheral human monocytes and the THP-1 monocytic leukemia-derived cell line, we confirm that sTLR2 generation increases upon treatment with pro-inflammatory agents and requires a post-translational mechanism. We also find that the constitutive and ligand-induced release of sTLR2 is sensitive to pharmacological metalloproteinase activator and inhibitors leading us to conclude that metalloproteinase TLR2 shedding contributes to soluble receptor production. By expressing human TLR2 in ADAM10- or ADAM17-deficient MEF cells, we find both enzymes to be implicated in TLR2 ectodomain shedding. Moreover, using a deletion mutant of the TLR2 juxtamembrane region, we demonstrate that this domain is required for sTLR2 generation. Functional analysis suggests that sTLR2 generated by metalloproteinase activation inhibitsTLR2-induced cytokine production by this monocytic leukemia-derived cell line. The identification of the mechanisms involved in regulating the availability of soluble TLR2 ectodomain and cell surface receptors may contribute further research on TLR2-mediated processes in innate immunity and inflammatory disorders.

Crohn's 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., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

Acute inflammation is a response to an alteration induced by a pathogen or a physical or chemical insult, which functions to eliminate the source of the damage and restore homeostasis to the affected tissue. However, chronic inflammation triggers cellular events that can promote malignant transformation of cells and carcinogenesis. Several inflammatory mediators, such as TNF-α, IL-6, TGF-β, and IL-10, have been shown to participate in both the initiation and progression of cancer. In this review, we explore the role of these cytokines in important events of carcinogenesis, such as their capacity to generate reactive oxygen and nitrogen species, their potential mutagenic effect, and their involvement in mechanisms for epithelial mesenchymal transition, angiogenesis, and metastasis. Finally, we will provide an in-depth analysis of the participation of these cytokines in two types of cancer attributable to chronic inflammatory disease: colitis-associated colorectal cancer and cholangiocarcinoma.

The most difficult component in our understanding of human autoimmunity remains a rigorous dissection of etiological events. Indeed, the vast literature on autoimmune diseases focuses on the inflammatory response, with the hope of developing drugs that reduce inflammation. However, there is increasing recognition that understanding the immunobiology of target tissues will also have direct relevance to disease natural history, including breach of tolerance. Sjögren's syndrome is essentially an epitheliitis and there are major changes to normal architectural salivary organization. We propose that loss of homeostasis is the initial event that precipitates inflammation and that such inflammatory response includes not only the adaptive response, but also an intense innate immune/bystander response. To understand these events this review focuses on the architecture, phenotype, function and epithelial cell organization. We further submit that there are several critical issues that must be defined to fully understand epithelial cell immunobiology in Sjögren's syndrome, including defining epithelial cell polarity, cell-cell and cell to extracellular matrix interactions and a variety of chemical and mechanical signals. We also argue that disruption of tight junctions induces disorganization of the apical pole of salivary acinar cells in Sjögren's syndrome. In addition, there will be a critical role of inflammatory cytokines in the apico-basal relocation of tight junction proteins. Further, the altered disorganization and relocation of proteins that participate in secretory granule formation are also dysregulated in Sjögren's syndrome and will contribute to abnormalities of mucins within the extracellular matrix. Our ability to understand Sjögren's syndrome and develop viable therapeutic options will depend on defining these events of epithelial cell biology., (Copyright © 2013 Elsevier Ltd. All rights reserved.)