Arthur Kaser

XBP1 Links ER Stress to Intestinal Inflammation and Confers Genetic Risk for Human Inflammatory Bowel Disease

Inflammatory bowel disease (IBD) has been attributed to aberrant mucosal immunity to the intestinal microbiota. The transcription factor XBP1, a key component of the endoplasmic reticulum (ER) stress response, is required for development and maintenance of secretory cells and linked to JNK activation. We hypothesized that a stressful environmental milieu in a rapidly proliferating tissue might instigate a proinflammatory response. We report that Xbp1 deletion in intestinal epithelial cells (IECs) results in spontaneous enteritis and increased susceptibility to induced colitis secondary to both Paneth cell dysfunction and an epithelium that is overly reactive to inducers of IBD such as bacterial products (flagellin) and TNFalpha. An association of XBP1 variants with both forms of human IBD (Crohns disease and ulcerative colitis) was identified and replicated (rs35873774; p value 1.6 x 10(-5)) with novel, private hypomorphic variants identified as susceptibility factors. Hence, intestinal inflammation can originate solely from XBP1 abnormalities in IECs, thus linking cell-specific ER stress to the induction of organ-specific inflammation.

Visfatin, an Adipocytokine with Proinflammatory and Immunomodulating Properties

Adipocytokines are mainly adipocyte-derived cytokines regulating metabolism and as such are key regulators of insulin resistance. Some adipocytokines such as adiponectin and leptin affect immune and inflammatory functions. Visfatin (pre-B cell colony-enhancing factor) has recently been identified as a new adipocytokine affecting insulin resistance by binding to the insulin receptor. In this study, we show that recombinant visfatin activates human leukocytes and induces cytokine production. In CD14+ monocytes, visfatin induces the production of IL-1β, TNF-α, and especially IL-6. Moreover, it increases the surface expression of costimulatory molecules CD54, CD40, and CD80. Visfatin-stimulated monocytes show augmented FITC-dextran uptake and an enhanced capacity to induce alloproliferative responses in human lymphocytes. Visfatin-induced effects involve p38 as well as MEK1 pathways as determined by inhibition with MAPK inhibitors and we observed activation of NF-κB. In vivo, visfatin induces circulating IL-6 in BALB/c mice. In patients with inflammatory bowel disease, plasma levels of visfatin are elevated and its mRNA expression is significantly increased in colonic tissue of Crohn’s and ulcerative colitis patients compared with healthy controls. Macrophages, dendritic cells, and colonic epithelial cells might be additional sources of visfatin as determined by confocal microscopy. Visfatin can be considered a new proinflammatory adipocytokine.

Gut microbiome, obesity, and metabolic dysfunction

The prevalence of obesity and related disorders such as metabolic syndrome has vastly increased throughout the world. Recent insights have generated an entirely new perspective suggesting that our microbiota might be involved in the development of these disorders. Studies have demonstrated that obesity and metabolic syndrome may be associated with profound microbiotal changes, and the induction of a metabolic syndrome phenotype through fecal transplants corroborates the important role of the microbiota in this disease. Dietary composition and caloric intake appear to swiftly regulate intestinal microbial composition and function. As most findings in this field of research are based on mouse studies, the relevance to human biology requires further investigation.

Adiponectin and its receptors in non-alcoholic steatohepatitis

Background: Adiponectin, an adipocyte derived polypeptide, has been shown to alleviate steatosis and inflammation in mice with non-alcoholic fatty liver disease. Aim: In the present study, we wished to define liver expression of adiponectin and its receptors in morbidly obese patients undergoing bariatric surgery. Patients with non-alcoholic steatohepatitis (NASH) or simple steatosis were investigated to test whether dysregulation of this system might be involved in these disorders. Patients and methods: Liver mRNA expression of adiponectin and its recently cloned receptors RI and RII (adipoRI and adipoRII) were analysed by fluorescence based real time polymerase chain reaction in 13 patients with NASH and nine with simple steatosis. Adiponectin and adipoRII protein expression were assessed by immunohistochemistry in a subgroup of patients. Results: Adiponectin and adipoRII mRNA expression were significantly reduced in liver biopsies of patients with NASH compared with simple steatosis while no difference was found in adipoRI mRNA expression. In NASH, adipoRII mRNA expression was negatively correlated with serum aspartate aminotransferase levels, serum alanine aminotransferase levels, and grade of fibrosis. Liver adiponectin protein expression was mainly found in endothelial cells of portal vessels and liver sinusoids whereas adipoRII expression was seen in hepatocytes only. Adiponectin and adipoRII staining were lower in biopsies of subjects with NASH compared with simple steatosis. Conclusion: Reduced hepatic expression of adiponectin and adipoRII might be of pathophysiological relevance in non-alcoholic fatty liver diseases.

Paneth cells as a site of origin for intestinal inflammation

The recognition of autophagy related 16-like 1 (ATG16L1) as a genetic risk factor has exposed the critical role of autophagy in Crohn’s disease. Homozygosity for the highly prevalent ATG16L1 risk allele, or murine hypomorphic (HM) activity, causes Paneth cell dysfunction. As Atg16l1HM mice do not develop spontaneous intestinal inflammation, the mechanism(s) by which ATG16L1 contributes to disease remains obscure. Deletion of the unfolded protein response (UPR) transcription factor X-box binding protein-1 (Xbp1) in intestinal epithelial cells, the human orthologue of which harbours rare inflammatory bowel disease risk variants, results in endoplasmic reticulum (ER) stress, Paneth cell impairment and spontaneous enteritis. Unresolved ER stress is a common feature of inflammatory bowel disease epithelium, and several genetic risk factors of Crohn’s disease affect Paneth cells. Here we show that impairment in either UPR (Xbp1ΔIEC) or autophagy function (Atg16l1ΔIEC or Atg7ΔIEC) in intestinal epithelial cells results in each other’s compensatory engagement, and severe spontaneous Crohn’s-disease-like transmural ileitis if both mechanisms are compromised. Xbp1ΔIEC mice show autophagosome formation in hypomorphic Paneth cells, which is linked to ER stress via protein kinase RNA-like endoplasmic reticulum kinase (PERK), elongation initiation factor 2α (eIF2α) and activating transcription factor 4 (ATF4). Ileitis is dependent on commensal microbiota and derives from increased intestinal epithelial cell death, inositol requiring enzyme 1α (IRE1α)-regulated NF-κB activation and tumour-necrosis factor signalling, which are synergistically increased when autophagy is deficient. ATG16L1 restrains IRE1α activity, and augmentation of autophagy in intestinal epithelial cells ameliorates ER stress-induced intestinal inflammation and eases NF-κB overactivation and intestinal epithelial cell death. ER stress, autophagy induction and spontaneous ileitis emerge from Paneth-cell-specific deletion of Xbp1. Genetically and environmentally controlled UPR function within Paneth cells may therefore set the threshold for the development of intestinal inflammation upon hypomorphic ATG16L1 function and implicate ileal Crohn’s disease as a specific disorder of Paneth cells.

Genome-Wide Association Analysis in Primary Sclerosing Cholangitis

BACKGROUND & AIMS We aimed to characterize the genetic susceptibility to primary sclerosing cholangitis (PSC) by means of a genome-wide association analysis of single nucleotide polymorphism (SNP) markers. METHODS A total of 443,816 SNPs on the Affymetrix SNP Array 5.0 (Affymetrix, Santa Clara, CA) were genotyped in 285 Norwegian PSC patients and 298 healthy controls. Associations detected in this discovery panel were re-examined in independent case-control panels from Scandinavia (137 PSC cases and 368 controls), Belgium/The Netherlands (229 PSC cases and 735 controls), and Germany (400 cases and 1832 controls). RESULTS The strongest associations were detected near HLA-B at chromosome 6p21 (rs3099844: odds ratio [OR], 4.8; 95% confidence interval [CI], 3.6-6.5; P = 2.6 x 10(-26); and rs2844559: OR, 4.7; 95% CI, 3.5-6.4; P = 4.2 x 10(-26) in the discovery panel). Outside the HLA complex, rs9524260 at chromosome 13q31 showed significant associations in 3 of 4 study panels. Lentiviral silencing of glypican 6, encoded at this locus, led to the up-regulation of proinflammatory markers in a cholangiocyte cell line. Of 15 established ulcerative colitis susceptibility loci, significant replication was obtained at chromosomes 2q35 and 3p21 (rs12612347: OR, 1.26; 95% CI, 1.06-1.50; and rs3197999: OR, 1.22; 95% CI, 1.02-1.47, respectively), with circumstantial evidence supporting the G-protein-coupled bile acid receptor 1 and macrophage-stimulating 1, respectively, as the likely disease genes. CONCLUSIONS Strong HLA associations and a subset of genes involved in bile homeostasis and other inflammatory conditions constitute key components of the genetic architecture of PSC.

Gut, inflammation and osteoporosis: basic and clinical concepts

Chronic inflammatory disorders such as inflammatory bowel diseases (IBD) affect bone metabolism and are frequently associated with the presence of osteoporosis. Bone loss is regulated by various mediators of the immune system such as the pro-inflammatory cytokines tumour necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), IL-6, or interferon-gamma. TNF-α, a master cytokine in human IBD, causes bone erosions in experimental models and these effects are exerted by osteoclasts. Other TNF-related cytokines such as receptor activator of nuclear factor kappa B (RANK), its ligand, RANKL, and osteoprotegerin are important mediators in inflammatory processes in the gut and are critically involved in the pathophysiology of bone loss. The awareness and early diagnosis of osteoporosis in states of chronic inflammation, together with applied therapies such as bisphosphonates, may be beneficial in inflammation-associated osteoporosis. Although several mechanisms may contribute to osteoporosis in patients with IBD and coeliac disease, inflammation as an important factor has so far been neglected. As key inflammatory mediators in IBD such as TNF-α are involved in the disease process both in gut and bone, we hypothesise that neutralisation of TNF-α could prove an efficient strategy in the treatment of inflammation-related osteoporosis in the future.

Anti-tumor necrosis factor-alpha monoclonal antibody therapy in severe alcoholic hepatitis

BACKGROUND/AIMS Severe alcoholic hepatitis (AH) is associated with high mortality. Tumor necrosis factor-alpha (TNFalpha) has been demonstrated to play an important role in its pathophysiology. METHODS Twelve patients with biopsy-confirmed AH and a Maddrey discriminant factor >32 were treated with a single infusion of the anti-TNF monoclonal antibody Infliximab at a dose of 5mg/kg body weight. Serial measurements were made for various cytokines using specific enzyme-linked immunoassays (ELISA). In four patients, liver biopsy samples were available pretreatment and on day+28 of therapy. RESULTS Ten of the 12 patients are alive at a median of 15 (12-20) months. Two patients died within 30 days from septicemia. Serum bilirubin levels, Maddrey score, neutrophil count and C-reactive protein fell significantly within the first month. There was an early, though not significant, decrease in plasma levels of proinflammatory cytokines (interleukins (IL)-1beta, IL-6, IL-8, interferon-gamma), whereas plasma levels of TNFalpha remained near the sensitivity limit of the assay throughout the treatment course. While TNFalpha mRNA expression in the liver did not change, expression of IL-8, a cytokine regulated mainly by TNFalpha, was almost absent on day+28. CONCLUSIONS Our data suggest that randomized controlled trials of anti-TNF antibody in severe AH are warranted.

Treatment of Crohn's disease with recombinant human interleukin 10 induces the proinflammatory cytokine interferon gamma

Background: Interleukin 10 (IL-10) exerts anti-inflammatory actions by counteracting many biological effects of interferon γ (IFN-γ). Aims: To investigate this in humans, we studied the effects of human recombinant IL-10 administration on IFN-γ production by patient leucocytes. Furthermore, we assessed the IFN-γ inducible molecule neopterin and nitrite/nitrate serum levels, which are indicative of endogenous nitric oxide formation. Methods: As part of two placebo controlled double blind studies, we analysed patients with chronic active Crohns disease (CACD) who received either subcutaneous recombinant human IL-10 (n=44) or placebo (n=10) daily for 28 days, and patients with mild to moderate Crohns disease (MCD) treated with either subcutaneous IL-10 (n=52) or placebo (n=16) daily for 28 days. Neopterin and nitrite/nitrate concentrations were measured in serum, and ex vivo IFN-γ formation by lipopolysaccharide or phytohaemagglutinin (PHA) stimulated whole blood cells were investigated before, during, and after IL-10 therapy. Results: In patients with CACD, the highest dose of 20 μg/kg IL-10 caused a significant increase in serum neopterin on days +15 and +29 of therapy compared with pretreatment levels. No changes were observed for nitrite/nitrate levels under either condition. In MCD, treatment with 20 μg/kg IL-10 resulted in a significant increase in PHA induced IFN-γ production. Conclusions: High doses of IL-10 upregulate the production of IFN-γ and neopterin. This phenomenon may be responsible for the lack of efficacy of high doses of IL-10 in the treatment of CACD and MCD.

Hepatitis B virus-induced lipid alterations contribute to natural killer T cell-dependent protective immunity

In most adult humans, hepatitis B is a self-limiting disease leading to life-long protective immunity, which is the consequence of a robust adaptive immune response occurring weeks after hepatitis B virus (HBV) infection. Notably, HBV-specific T cells can be detected shortly after infection, but the mechanisms underlying this early immune priming and its consequences for subsequent control of viral replication are poorly understood. Using primary human and mouse hepatocytes and mouse models of transgenic and adenoviral HBV expression, we show that HBV-expressing hepatocytes produce endoplasmic reticulum (ER)-associated endogenous antigenic lipids including lysophospholipids that are generated by HBV-induced secretory phospholipases and that lead to activation of natural killer T (NKT) cells. The absence of NKT cells or CD1d or a defect in ER-associated transfer of lipids onto CD1d results in diminished HBV-specific T and B cell responses and delayed viral control in mice. NKT cells may therefore contribute to control of HBV infection through sensing of HBV-induced modified self-lipids.