Differences in the On- and Off-Tumor Microbiota between Right- and Left-Sided Colorectal Cancer.

This study aims to determine differences in the on- and off-tumor microbiota between patients with right- and left-sided colorectal cancer. Microbiome profiling of tumor and tumor-adjacent biopsies from patients with right-sided (n = 17) and left-sided (n = 7) colorectal adenocarcinoma was performed using 16S ribosomal RNA sequencing. Off-tumor alpha and beta diversity were significantly different between right- and left-sided colorectal cancer patients. However, no differences in on-tumor diversity were observed between tumor locations. Comparing the off-tumor microbiota showed the right colon to be enriched with species of the Lachnoclostridium, Selenomonas, and Ruminococcus genera. Whereas the left colon is enriched with Epsilonbacteraeota phylum, Campylobacteria class, and Pasteurellales and Campylobacterales orders, in contrast, the on-tumor microbiota showed relatively fewer differences in bacterial taxonomy between tumor sites, with left tumors being enriched with Methylophilaceae and Vadin BE97 families and Alloprevotella, Intestinibacter, Romboutsia, and Ruminococcus 2 genera. Patients with left-sided colorectal cancer had large taxonomic differences between their paired on- and off-tumor microbiota, while patients with right-sided colorectal cancer showed relatively fewer taxonomic differences. Collectively, this suggests that the right and left colon show distinctive bacterial populations; however, the presence of a colonic tumor leads to a more consistent microbiota between locations.

Iron therapy for preoperative anaemia.

BACKGROUND: Preoperative anaemia is common and occurs in 5% to 76% of patients preoperatively. It is associated with an increased risk of perioperative allogeneic blood transfusion, longer hospital stay, and increased morbidity and mortality. Iron deficiency is one of the most common causes of anaemia. Oral and intravenous iron therapy can be used to treat anaemia. Parenteral iron preparations have been shown to be more effective in conditions such as inflammatory bowel disease, chronic heart failure and postpartum haemorrhage due to rapid correction of iron stores. A limited number of studies has investigated iron therapy for the treatment of preoperative anaemia. The aim of this Cochrane Review is to summarise the evidence for iron supplementation, both enteral and parenteral, for the management of preoperative anaemia. OBJECTIVES: To evaluate the effects of preoperative iron therapy (enteral or parenteral) in reducing the need for allogeneic blood transfusions in anaemic patients undergoing surgery. SEARCH METHODS: We ran the search on 30 July 2018. We searched the Cochrane Injuries Group's Specialised Register, Cochrane Central Register of Controlled Trials (CENTRAL, the Cochrane Library), Ovid MEDLINE(R), Ovid MEDLINE(R) In-Process & Other Non-Indexed Citations, Ovid MEDLINE(R) Daily and Ovid OLDMEDLINE(R), Embase Classic and Embase (Ovid), CINAHL Plus (EBSCO), PubMed, and clinical trials registries, and we screened reference lists. We ran a top-up search on 28 November 2019; one study is now awaiting classification. SELECTION CRITERIA: We included all randomised controlled trials (RCTs) that compared preoperative iron monotherapy to placebo, no treatment, standard care or another form of iron therapy for anaemic adults undergoing surgery. We defined anaemia as haemoglobin values less than 13 g/dL for males and 12 g/dL for non-pregnant females. DATA COLLECTION AND ANALYSIS: Two review authors collected data and a third review author checked all collected data. Data were collected on the proportion of patients who receive a blood transfusion, the amount of blood transfused per patient (units), quality of life, ferritin levels and haemoglobin levels, measured as continuous variables at the following predetermined time points: pretreatment (baseline), preoperatively but postintervention, and postoperatively. We performed statistical analysis using the Cochrane software, Review Manager 5. We summarised outcome data in tables and forest plots. We used the GRADE approach to describe the quality of the body of evidence. MAIN RESULTS: Six RCTs, with a total of 372 participants, evaluated preoperative iron therapy to correct anaemia before planned surgery. Four studies compared iron therapy (either oral (one study) or intravenous (three studies)) with no treatment, placebo or usual care, and two studies compared intravenous iron therapy with oral iron therapy. Iron therapy was delivered over a range of periods that varied from 48 hours to three weeks prior to surgery. The 372 participants in our analysis fall far short of the 819 required - as calculated by our information size calculation - to detect a 30% reduction in blood transfusions. Five trials, involving 310 people, reported the proportion of participants who received allogeneic blood transfusions. Meta-analysis of iron therapy versus placebo or standard care showed no difference in the proportion of participants who received a blood transfusion (risk ratio (RR) 1.21, 95% confidence interval (CI) 0.87 to 1.70; 4 studies, 200 participants; moderate-quality evidence). Only one study that compared oral versus intravenous iron therapy measured this outcome, and reported no difference in risk of transfusion between groups. There was no difference between the iron therapy and placebo/standard care groups for haemoglobin level preoperatively at the end of the intervention (mean difference (MD) 0.63 g/dL, 95% CI -0.07 to 1.34; 2 studies, 83 participants; low-quality evidence). However, intravenous iron therapy produced an increase in preoperative postintervention haemoglobin levels compared with oral iron (MD 1.23 g/dL, 95% CI 0.80 to 1.65; 2 studies, 172 participants; low-quality evidence). Ferritin levels were increased by intravenous iron, both when compared to standard care ((MD 149.00, 95% CI 25.84 to 272.16; 1 study, 63 participants; low-quality evidence) or to oral iron (MD 395.03 ng/mL, 95% CI 227.72 to 562.35; 2 studies, 151 participants; low-quality evidence). Not all studies measured quality of life, short-term mortality or postoperative morbidity. Some measured the outcomes, but did not report the data, and the studies which did report the data were underpowered. Therefore, uncertainty remains regarding these outcomes. The inclusion of new research in the future is very likely to change these results. AUTHORS' CONCLUSIONS: The use of iron therapy for preoperative anaemia does not show a clinically significant reduction in the proportion of trial participants who received an allogeneic blood transfusion compared to no iron therapy. Results for intravenous iron are consistent with a greater increase in haemoglobin and ferritin when compared to oral iron, but do not provide reliable evidence. These conclusions are drawn from six studies, three of which included very small numbers of participants. Further, well-designed, adequately powered, RCTs are required to determine the true effectiveness of iron therapy for preoperative anaemia. Two studies are currently in progress, and will include 1500 randomised participants.

Tight junctions in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer.

Inflammatory bowel diseases are characterised by inflammation that compromises the integrity of the epithelial barrier. The intestinal epithelium is not only a static barrier but has evolved complex mechanisms to control and regulate bacterial interactions with the mucosal surface. Apical tight junction proteins are critical in the maintenance of epithelial barrier function and control of paracellular permeability. The characterisation of alterations in tight junction proteins as key players in epithelial barrier function in inflammatory bowel diseases is rapidly enhancing our understanding of critical mechanisms in disease pathogenesis as well as novel therapeutic opportunities. Here we give an overview of recent literature focusing on the role of tight junction proteins, in particular claudins, in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer.

The role of CD40 expression in dendritic cells in cancer biology; a systematic review.

CD40 is a co-stimulatory molecule belonging to the tumor necrosis factor superfamily and is essential in activation of dendritic cells. Dendritic cells (DCs) are antigen-presenting cells capable of initiating cytotoxic T-lymphocyte immune response against cancer cells. However, there are few studies on the characterization of DCs in cancer, specifically their expression of CD40, despite its implication in cancer immunotherapy. We reviewed available data on the expression of CD40 on DCs in various cancers, and its implications for cancer immunotherapy. A systematic review on CD40 expression on DCs in cancer was performed with reference to preferred reporting items for systematic reviews and meta-analyses (PRISMA). Studies that satisfied the inclusion and exclusion criteria were 21 out of 927. Variations in type and status of the cancers, source of DCs and methodology for detecting CD40 expression amongst the studies resulted in contrasting results. DCs generally expressed low CD40 in tumor infiltrating DCs (tiDCs), in DCs derived by in vitro culture from blood monocytes using cytokine stimulation (MoDCs) and in DCs exposed in vitro to tumor cells lines; the studies suggested that CD40 expression in DCs is impaired in cancer particularly in metastatic disease. However, DCs identified in fresh peripheral blood mononuclear cells (PBMC) expressed higher numbers of CD40 positive cells in some cancer patients, which could be due to tumor-derived factors leading to partially-stimulated DCs. The results provide evidence that some cancer patients may show partial systemic DC activation and expression of increased CD40 in response to the presence of tumor but that such activity may become abortive in the presence of factors produced by the tumor. This review has thus identified key papers on CD40 expression on DCs in various cancers and discusses the limitations and contrasting results of these studies in relation to variations in methodology. The results highlight the need for further studies on the role of CD40-CD40 ligand pathway to inform cancer treatment.

Intestinal dendritic cells: their role in intestinal inflammation, manipulation by the gut microbiota and differences between mice and men.

The intestinal immune system maintains a delicate balance between immunogenicity against invading pathogens and tolerance of the commensal microbiota and food antigens. Dendritic cells (DC) generate primary T-cell responses, and determine whether these responses are immunogenic or tolerogenic. The regulatory role of DC is of particular importance in the gut due to the high antigenic load. Intestinal DC act as sentinels, sampling potentially pathogenic antigens but also harmless antigens including the commensal microbiota. Following antigen acquisition, intestinal DC migrate to secondary lymphoid organs to activate naive T-cells. DC also imprint specific homing properties on T-cells that they stimulate; gut DC specifically induce gut-homing properties on T-cells upon activation, enabling T-cell migration back to intestinal sites. Data regarding properties on gut DC in humans is scarce, although evidence now supports the role of DC as important players in intestinal immunity in humans. Here, we review the role of intestinal DC in shaping mucosal immune responses and directing tissue-specific T-cell responses, with a special focus on the importance of distinguishing DC subsets from macrophages at intestinal sites. We compare and contrast human DC with their murine counterparts, and discuss the ability of the gut microbiota to shape intestinal DC function, and how this may be dysregulated in inflammatory bowel disease (IBD). Lastly, we describe recent advances in the study of probiotics on intestinal DC function, including the use of soluble secreted bacterial products.

Human gut-specific homeostatic dendritic cells are generated from blood precursors by the gut microenvironment.

BACKGROUND: Dendritic cells (DC) dictate not only the type of T-cell immunity, but also homing patterns of T cells in mice. In humans, we characterized normal human gut DC and tested whether gut-specific homeostatic DC could be generated from blood precursors by factors in the gut microenvironment. METHODS: We characterized the phenotype and function of healthy human gut DC compared with blood and skin DC, and studied whether conditioning of blood DC in the presence of colonic biopsy supernatants (Bx-SN) induced gut-like phenotype and functions. RESULTS: Blood DC mostly expressed both gut and skin homing markers, indicating potential to migrate to both major immune surface organs, and induced multi-homing T cells. However, DC within gut or skin did not demonstrate this multi-homing phenotype, were tissue-specific, and induced tissue-specific T cells. Human gut DC were less stimulatory for allogeneic T cells than their dermal and blood counterparts. Human blood DC cultured in vitro lost homing marker expression. Conditioning of human enriched blood DC with colonic Bx-SN from healthy controls induced a gut-homing phenotype and a homeostatic profile. Moreover, Bx-SN-conditioned DC demonstrated a restricted T-cell stimulatory capacity and preferentially induced gut-specific T cells. Retinoic acid and transforming growth factor beta (TGF-ß) mediated the acquisition of the gut-homing and homeostatic properties, respectively, induced by colonic Bx-SN on blood enriched DC. CONCLUSIONS: Tissue-specific factors manipulate immunity via modulating characteristics of DC and may provide tools to generate tissue-specific immunotherapy.

Homing of immune cells: role in homeostasis and intestinal inflammation.

Rather like a satellite navigation system directing a vehicle to a particular destination defined by post-code, immune cells have homing molecules or "immune post-codes" enabling them to be recruited to specific organs, such as the intestine or skin. An efficient system would be designed such that the site of entry of an antigen influences the homing of effector T cells back to the appropriate organ. For example, to mount an immune response against an intestinal pathogen, T cells with a propensity to home to the gut to clear the infection would be induced. In health, there is such a sophisticated and finely tuned system in operation, enabling an appropriate balance of immune activity in different anatomical compartments. In disease states such as inflammatory bowel disease (IBD), which is characterized by intestinal inflammation and often an inflammatory process involving other organs such as skin, joints, liver, and eye, there is accumulating evidence that there is malfunction of this immune cell trafficking system. The clinical importance of dysregulated immune cell trafficking in IBD is reflected in recently proven efficacious therapies that target trafficking pathways such as natalizumab, an α4 integrin antibody, and Traficet-EN, a chemokine receptor-9 (CCR9) antagonist. Here we review the mechanisms involved in the homing of immune cells to different tissues, in particular the intestine, and focus on alterations in immune cell homing pathways in IBD. Unraveling the mechanisms underlying the immune post-code system would assist in achieving the goal of tissue-specific immunotherapy.

Immunosuppressive effects via human intestinal dendritic cells of probiotic bacteria and steroids in the treatment of acute ulcerative colitis.

BACKGROUND: In ulcerative colitis (UC) gut bacteria drive inflammation. Bacterial recognition and T-cell responses are shaped by intestinal dendritic cells (DCs); therapeutic effects of probiotic bacteria may relate to modulation of intestinal DC. The probiotic mixture, VSL#3, increases interleukin (IL)-10 and downregulates IL-12p40 production by DC in vitro. We evaluated in vivo effects of oral VSL#3 and steroids on colonic DC in patients with acute UC. METHODS: Rectal biopsies were obtained from patients with active UC before and after treatment with VSL#3, corticosteroids, or placebo, and from healthy controls. Myeloid colonic DC were studied from freshly isolated lamina propria cells using multicolor flow cytometry. Surface expression of activation markers, CD40, CD86, pattern recognition receptors, Toll-like receptor (TLR)-2 and TLR-4 were assessed. Changed function was measured from ongoing intracellular IL-10, IL-12p40, IL-6, and IL-13 production. RESULTS: Acute UC colonic myeloid DC were producing more IL-10 and IL-12p40 than control DC (P = 0.01). In VSL#3-treated patients DC TLR-2 expression decreased (P < 0.05), IL-10 production increased and IL-12p40 production decreased (P < 0.005); 10/14 patients on VSL#3 showed a clinical response. Corticosteroids also resulted in increased IL-10 and reduced IL-12p40 production by DC. Conversely, in patients on placebo, TLR-2 expression and intensity of staining for IL-12p40 and IL-6 increased (all P < 0.05); 5/14 patients on placebo showed a clinical response (P = NS). CONCLUSIONS: Despite small numbers of human colonic DC available, we showed that treatment of UC patients with probiotic VSL#3 and corticosteroids induced "favorable" intestinal DC function in vivo, increasing regulatory cytokines and lowering proinflammatory cytokines and TLR expression. These effects may contribute to therapeutic benefit.

Adipose tissue of human omentum is a major source of dendritic cells, which lose MHC Class II and stimulatory function in Crohn's disease.

Adipose tissue is reported to contain monocyte-like pre-adipocytes, which may mature into macrophages, contributing to local inflammation. Dendritic cells (DC) can be derived from monocytes and initiate and regulate primary immune responses. We hypothesized, therefore, that adipose tissue may provide DC involved in local immune activity. To test this, we studied cells from human omental adipose tissue samples from 17 patients with benign gynecological disease. The hypothesis that adipose tissue DC are involved in inflammatory disease was tested by comparing these cells with those from 18 patients with Crohn's disease, where hypertrophy of adipose tissue suggests involvement in disease. A high proportion of the 1.33 +/- 0.12 x 10(5) CD45-positive cells/mg, obtained from control omenta, expressed CD11c, CD1a, and CD83; costimulatory molecules CD40, CD80, and CD86; and major histocompatibility complex (MHC) Class II but little CD14, CD16, or CD33. Omental cells showing morphological characteristics of DC were also observed. Metrizamide gradient-enriched DC from these populations were potent stimulators of primary proliferation of allogeneic T cells in mixed leukocyte reactions. Increased numbers of CD45+ cells from omentum of Crohn's patients (4.50+/-1.08 x 10(5) CD45+ cells/mg) contained higher percentages of CD11c+ and CD40+ cells (80.8+/-3.8% vs. 63.4+/-6, P=0.032; 77.9+/-4% vs. 58.8+/-6.5, P=0.029, respectively), but MHC Class II and stimulatory capacity were almost completely lost (P= <0.001), suggesting innate activation but lost capacity to stimulate adaptive immune responses. Granulocytes were also present amongst the omental cells from Crohn's patients. Results indicated that omentum may provide DC, which could "police" local infections and contribute to and/or reflect local inflammatory activity.

Characteristics of intestinal dendritic cells in inflammatory bowel diseases.

BACKGROUND & AIMS: Dendritic cells (DCs) recognize and respond to microbial structures using pattern recognition receptors, including Toll-like receptors (TLRs). In the intestine, DCs are pivotal in tolerance induction and direct the differentiation of T cells. We aimed to identify changes in intestinal DCs that may underlie the dysregulated immune response to enteric bacteria that occurs in patients with inflammatory bowel disease (IBD). METHODS: DCs were identified in freshly isolated lamina propria mononuclear cells by multicolor flow cytometry in patients with IBD and controls. Expression of TLR2, TLR4, and the activation/maturation marker CD40 was assessed by cell surface labeling. Production of cytokines (interleukin [IL]-12, IL-6, and IL-10) was assessed in the absence of exogenous stimulation by intracellular staining of permeabilized cells. RESULTS: In healthy controls, few intestinal DCs expressed TLR2 or TLR4, in contrast to blood DCs. DC expression of both TLRs was significantly enhanced in Crohn's disease and ulcerative colitis. DCs from inflamed tissue of patients with Crohn's disease expressed significantly higher levels of the maturation/activation marker CD40. Elevated levels of CD40 on DCs were decreased after treating patients with anti-tumor necrosis factor alpha. In Crohn's disease, but not ulcerative colitis, more colonic DCs produced IL-12 and IL-6. The number of IL-10-producing DCs did not differ significantly between patients with IBD and controls. CONCLUSIONS: In IBD, DCs are activated, their expression of microbial recognition receptors is up-regulated, and more DCs produce pathologically relevant cytokines. Intestinal DCs are likely to be key initiators or perpetuators of the inflammatory response that characterizes IBD.