SNPexp - A web tool for calculating and visualizing correlation between HapMap genotypes and gene expression levels

<p>Abstract</p> <p>Background</p> <p>Expression levels for 47294 transcripts in lymphoblastoid cell lines from all 270 HapMap phase II individuals, and genotypes (both HapMap phase II and III) of 3.96 million single nucleotide polymorphisms (SNPs) in the same individuals are publicly available. We aimed to generate a user-friendly web based tool for visualization of the correlation between SNP genotypes within a specified genomic region and a gene of interest, which is also well-known as an expression quantitative trait locus (eQTL) analysis.</p> <p>Results</p> <p>SNPexp is implemented as a server-side script, and publicly available on this website: <url>http://tinyurl.com/snpexp</url>. Correlation between genotype and transcript expression levels are calculated by performing linear regression and the Wald test as implemented in PLINK and visualized using the UCSC Genome Browser. Validation of SNPexp using previously published eQTLs yielded comparable results.</p> <p>Conclusions</p> <p>SNPexp provides a convenient and platform-independent way to calculate and visualize the correlation between HapMap genotypes within a specified genetic region anywhere in the genome and gene expression levels. This allows for investigation of both cis and trans effects. The web interface and utilization of publicly available and widely used software resources makes it an attractive supplement to more advanced bioinformatic tools. For the advanced user the program can be used on a local computer on custom datasets.</p

Biliary tree stem cells are involved in the pathogenesis of primary sclerosing cholangitis

Biliary tree stem cells (BTSCs) are multipotent stem cells located in peribiliary glands (PBGs) of extrahepatic and large intrahepatic bile ducts (1). Primary sclerosing cholangitis (PSC) is characterised by fibro-stenosing strictures involving extrahepatic and/or large intrahepatic bile ducts. Mechanisms leading to bile duct injury are poorly understood (2). Our aims are to study the role of BTSC in the pathogenesis of biliary fibrosis in PSC. Specimens containing extrahepatic or large intrahepatic bile ducts were obtained from normal liver (n=6), liver explants from patients with PSC (n=11), and primary biliary cirrhosis (n=6). Specimens were processed for histology, immunohistochemistry and immunofluorescence. In PSC samples, progressive hyperplasia and mucinous metaplasia of PBGs were observed in large ducts with fibrosis, but not in inflamed ducts without fibrosis. PBG hyperplasia was associated with progressive biliary fibrosis and the occurrence of dysplastic lesions. Hyperplasia of PBGs was determined by the expansion of biliary tree stem cells, which sprouted towards the surface epithelium. In PSC, PBGs and myofibroblasts displayed enhanced expression of Hedgehog pathway components. PBGs in ducts with onion skin-like fibrosis expressed epithelial-to-mesenchymal transition traits associated with components of Hedgehog pathway, markers of senescence and autophagy. The biliary tree stem cell compartment is activated in PSC, its activation contributes to biliary fibrosis, and is sustained by the Hedgehog pathway. Our findings suggest a key role for peribiliary glands in the progression of bile duct lesions in PSC and could explain the associated high risk of cholangiocarcinoma. This work was supported by grants from MIUR FIRB 2010 and MIUR PRIN-2009

Electrostatic Modifications of the Human Leukocyte Antigen-DR P9 Peptide-Binding Pocket and Susceptibility to Primary Sclerosing Cholangitis

The strongest genetic risk factors for primary sclerosing cholangitis (PSC) are found in the human leukocyte antigen (HLA) complex at chromosome 6p21. Genes in the HLA class II region encode molecules that present antigen to T lymphocytes. Polymorphisms in these genes are associated with most autoimmune diseases, most likely because they contribute to the specificity of immune responses. The aim of this study was to analyze the structure and electrostatic properties of the peptide-binding groove of HLA-DR in relation to PSC. Thus, four-digit resolution HLA-DRB1 genotyping was performed in 356 PSC patients and 366 healthy controls. Sequence information was used to assign which amino acids were encoded at all polymorphic positions. In stepwise logistic regressions, variations at residues 37 and 86 were independently associated with PSC (P = 1.2 × 10−32 and P = 1.8 × 10−22 in single-residue models, respectively). Three-dimensional modeling was performed to explore the effect of these key residues on the HLA-DR molecule. This analysis indicated that residue 37 was a major determinant of the electrostatic properties of pocket P9 of the peptide-binding groove. Asparagine at residue 37, which was associated with PSC, induced a positive charge in pocket P9. Tyrosine, which protected against PSC, induced a negative charge in this pocket. Consistent with the statistical observations, variation at residue 86 also indirectly influenced the electrostatic properties of this pocket. DRB1*13:01, which was PSC-associated, had a positive P9 pocket and DRB1*13:02, protective against PSC, had a negative P9 pocket. Conclusion: The results suggest that in patients with PSC, residues 37 and 86 of the HLA-DRβ chain critically influence the electrostatic properties of pocket P9 and thereby the range of peptides presented. (Hepatology 2011;53:1967-1976

Imputation of KIR Types from SNP Variation Data.

Large population studies of immune system genes are essential for characterizing their role in diseases, including autoimmune conditions. Of key interest are a group of genes encoding the killer cell immunoglobulin-like receptors (KIRs), which have known and hypothesized roles in autoimmune diseases, resistance to viruses, reproductive conditions, and cancer. These genes are highly polymorphic, which makes typing expensive and time consuming. Consequently, despite their importance, KIRs have been little studied in large cohorts. Statistical imputation methods developed for other complex loci (e.g., human leukocyte antigen [HLA]) on the basis of SNP data provide an inexpensive high-throughput alternative to direct laboratory typing of these loci and have enabled important findings and insights for many diseases. We present KIR∗IMP, a method for imputation of KIR copy number. We show that KIR∗IMP is highly accurate and thus allows the study of KIRs in large cohorts and enables detailed investigation of the role of KIRs in human disease.This work was supported by the Australian National Health and Medical Research Council (NHMRC), Career Development Fellowship ID 1053756 (S.L.); by a Victorian Life Sciences Computation Initiative (VLSCI) grant number VR0240 on its Peak Computing Facility at the University of Melbourne, an initiative of the Victorian Government, Australia (S.L.); by the UK Multiple Sclerosis Society, grant 894/08 (S.S.); and by the Wellcome Trust and the MRC with partial funding from the National Institute of Health Cambridge Biomedical Research Centre (J.T., J.A.T.). Research at the Murdoch Childrens Research Institute was supported by the Victorian Government's Operational Infrastructure Support Program.This is the final version of the article. It first appeared from Elsevier via http://dx.doi.org/10.1016/j.ajhg.2015.09.00

GPR35 promotes glycolysis, proliferation, and oncogenic signaling by engaging with the sodium potassium pump.

The sodium potassium pump (Na/K-ATPase) ensures the electrochemical gradient of a cell through an energy-dependent process that consumes about one-third of regenerated ATP. We report that the G protein-coupled receptor GPR35 interacted with the α chain of Na/K-ATPase and promotes its ion transport and Src signaling activity in a ligand-independent manner. Deletion of Gpr35 increased baseline Ca2+ to maximal levels and reduced Src activation and overall metabolic activity in macrophages and intestinal epithelial cells (IECs). In contrast, a common T108M polymorphism in GPR35 was hypermorphic and had the opposite effects to Gpr35 deletion on Src activation and metabolic activity. The T108M polymorphism is associated with ulcerative colitis and primary sclerosing cholangitis, inflammatory diseases with a high cancer risk. GPR35 promoted homeostatic IEC turnover, whereas Gpr35 deletion or inhibition by a selective pepducin prevented inflammation-associated and spontaneous intestinal tumorigenesis in mice. Thus, GPR35 acts as a central signaling and metabolic pacesetter, which reveals an unexpected role of Na/K-ATPase in macrophage and IEC biology.European Research Council Consolidator Grant n° 648889 to A.K. Scientia Fellowship (FP7-PEOPLE-2013-COFUND) grant agreement n° 609020 to G.S. Addenbrooke’s Charitable Trust (ACT 25/16A) to J.E.E. UniNA and Compagnia Di San Paolo ‘STAR program for young researchers’ fellowship to E.P

Complement C3b contributes to Escherichia coli-induced platelet aggregation in human whole blood

IntroductionPlatelets have essential functions as first responders in the immune response to pathogens. Activation and aggregation of platelets in bacterial infections can lead to life-threatening conditions such as arterial thromboembolism or sepsis-associated coagulopathy.MethodsIn this study, we investigated the role of complement in Escherichia coli (E. coli)-induced platelet aggregation in human whole blood, using Multiplate® aggregometry, flow cytometry, and confocal microscopy.Results and DiscussionWe found that compstatin, which inhibits the cleavage of complement component C3 to its components C3a and C3b, reduced the E. coli-induced platelet aggregation by 42%-76% (p = 0.0417). This C3-dependent aggregation was not C3a-mediated as neither inhibition of C3a using a blocking antibody or a C3a receptor antagonist, nor the addition of purified C3a had any effects. In contrast, a C3b-blocking antibody significantly reduced the E. coli-induced platelet aggregation by 67% (p = 0.0133). We could not detect opsonized C3b on platelets, indicating that the effect of C3 was not dependent on C3b-fragment deposition on platelets. Indeed, inhibition of glycoprotein IIb/IIIa (GPIIb/IIIa) and complement receptor 1 (CR1) showed that these receptors were involved in platelet aggregation. Furthermore, aggregation was more pronounced in hirudin whole blood than in hirudin platelet-rich plasma, indicating that E. coli-induced platelet aggregation involved other blood cells. In conclusion, the E. coli-induced platelet aggregation in human whole blood is partly C3b-dependent, and GPIIb/IIIa and CR1 are also involved in this process

The EASL–<em>Lancet</em> Commission on liver health in Europe: prevention, case-finding, and early diagnosis to reduce liver-related mortality

In December, 2021, the first report from the Lancet Commission on liver disease in Europe, a joint project with the European Association for the Study of the Liver (EASL), was published. 1 The Commission explored the harm to liver health in Europe that results from a combination of increasing obesity, the highest level of alcohol consumption in the world, and delays in viral hepatitis elimination. The Commission emphasised the importance of structural factors that drive risk behaviours and poor outcomes in liver disease, with disproportionate effects on disadvantaged and vulnerable populations. Such structural drivers include the heterogeneous landscape of alcohol policy in Europe, fragmented access to testing and therapy for viral hepatitis, and stigmatisation faced by individuals at risk of liver disease at the societal level and within health-care settings. 2 , 3 This stigma contributes to care avoidance and delayed diagnosis, ultimately leading to a bias in clinical pathways that prioritise managing advanced liver disease rather than early diagnosis and primary and secondary prevention of liver disease. The Commission report included ten recommendations to facilitate a shift towards health promotion, prevention, proactive case-finding, early identification of progressive liver fibrosis, and early management and treatment of liver diseases (figure). 1 The key message of the Commission was paraphrased by Ursula von der Leyen, the President of the European Commission, in her remarks made at the launch event: “in most cases, liver disease can be prevented. Prevention is the best cure that we have.

Activation of the GPR35 pathway drives angiogenesis in the tumour microenvironment

Funder: NIHR Cambridge BRCObjective: Primary sclerosing cholangitis (PSC) is in 70% of cases associated with inflammatory bowel disease. The hypermorphic T108M variant of the orphan G protein-coupled receptor GPR35 increases risk for PSC and ulcerative colitis (UC), conditions strongly predisposing for inflammation-associated liver and colon cancer. Lack of GPR35 reduces tumour numbers in mouse models of spontaneous and colitis associated cancer. The tumour microenvironment substantially determines tumour growth, and tumour-associated macrophages are crucial for neovascularisation. We aim to understand the role of the GPR35 pathway in the tumour microenvironment of spontaneous and colitis-associated colon cancers. Design: Mice lacking GPR35 on their macrophages underwent models of spontaneous colon cancer or colitis-associated cancer. The role of tumour-associated macrophages was then assessed in biochemical and functional assays. Results: Here, we show that GPR35 on macrophages is a potent amplifier of tumour growth by stimulating neoangiogenesis and tumour tissue remodelling. Deletion of Gpr35 in macrophages profoundly reduces tumour growth in inflammation-associated and spontaneous tumour models caused by mutant tumour suppressor adenomatous polyposis coli. Neoangiogenesis and matrix metalloproteinase activity is promoted by GPR35 via Na/K-ATPase-dependent ion pumping and Src activation, and is selectively inhibited by a GPR35-specific pepducin. Supernatants from human inducible-pluripotent-stem-cell derived macrophages carrying the UC and PSC risk variant stimulate tube formation by enhancing the release of angiogenic factors. Conclusions: Activation of the GPR35 pathway promotes tumour growth via two separate routes, by directly augmenting proliferation in epithelial cells that express the receptor, and by coordinating macrophages’ ability to create a tumour-permissive environment

Early and accurate detection of cholangiocarcinoma in patients with primary sclerosing cholangitis by methylation markers in bile

Background and Aims Primary sclerosing cholangitis (PSC) is associated with increased risk of cholangiocarcinoma (CCA). Early and accurate CCA detection represents an unmet clinical need as the majority of patients with PSC are diagnosed at an advanced stage of malignancy. In the present study, we aimed at establishing robust DNA methylation biomarkers in bile for early and accurate diagnosis of CCA in PSC. Approach and Results Droplet digital PCR (ddPCR) was used to analyze 344 bile samples from 273 patients with sporadic and PSC-associated CCA, PSC, and other nonmalignant liver diseases for promoter methylation of cysteine dioxygenase type 1, cannabinoid receptor interacting protein 1, septin 9, and vimentin. Receiver operating characteristic (ROC) curve analyses revealed high AUCs for all four markers (0.77-0.87) for CCA detection among patients with PSC. Including only samples from patients with PSC diagnosed with CCA 36 months) as controls, and remained high (83%) when only including patients with PSC and dysplasia as controls (n = 23). Importantly, the bile samples from the CCA-PSCPeer reviewe

Control of CD1d-restricted antigen presentation and inflammation by sphingomyelin.

Invariant natural killer T (iNKT) cells recognize activating self and microbial lipids presented by CD1d. CD1d can also bind non-activating lipids, such as sphingomyelin. We hypothesized that these serve as endogenous regulators and investigated humans and mice deficient in acid sphingomyelinase (ASM), an enzyme that degrades sphingomyelin. We show that ASM absence in mice leads to diminished CD1d-restricted antigen presentation and iNKT cell selection in the thymus, resulting in decreased iNKT cell levels and resistance to iNKT cell-mediated inflammatory conditions. Defective antigen presentation and decreased iNKT cells are also observed in ASM-deficient humans with Niemann-Pick disease, and ASM activity in healthy humans correlates with iNKT cell phenotype. Pharmacological ASM administration facilitates antigen presentation and restores the levels of iNKT cells in ASM-deficient mice. Together, these results demonstrate that control of non-agonistic CD1d-associated lipids is critical for iNKT cell development and function in vivo and represents a tight link between cellular sphingolipid metabolism and immunity