A comprehensive analysis of shared loci between systemic lupus erythematosus (SLE) and sixteen autoimmune diseases reveals limited genetic overlap

In spite of the well-known clustering of multiple autoimmune disorders in families, analyses of specific shared genes and polymorphisms between systemic lupus erythematosus (SLE) and other autoimmune diseases (ADs) have been limited. Therefore, we comprehensively tested autoimmune variants for association with SLE, aiming to identify pleiotropic genetic associations between these diseases. We compiled a list of 446 non–Major Histocompatibility Complex (MHC) variants identified in genome-wide association studies (GWAS) of populations of European ancestry across 17 ADs. We then tested these variants in our combined Caucasian SLE cohorts of 1,500 cases and 5,706 controls. We tested a subset of these polymorphisms in an independent Caucasian replication cohort of 2,085 SLE cases and 2,854 controls, allowing the computation of a meta-analysis between all cohorts. We have uncovered novel shared SLE loci that passed multiple comparisons adjustment, including the VTCN1 (rs12046117, P = 2.02×10⁻⁰⁶) region. We observed that the loci shared among the most ADs include IL23R, OLIG3/TNFAIP3, and IL2RA. Given the lack of a universal autoimmune risk locus outside of the MHC and variable specificities for different diseases, our data suggests partial pleiotropy among ADs. Hierarchical clustering of ADs suggested that the most genetically related ADs appear to be type 1 diabetes with rheumatoid arthritis and Crohn''s disease with ulcerative colitis. These findings support a relatively distinct genetic susceptibility for SLE. For many of the shared GWAS autoimmune loci, we found no evidence for association with SLE, including IL23R. Also, several established SLE loci are apparently not associated with other ADs, including the ITGAM-ITGAX and TNFSF4 regions. This study represents the most comprehensive evaluation of shared autoimmune loci to date, supports a relatively distinct non–MHC genetic susceptibility for SLE, provides further evidence for previously and newly identified shared genes in SLE, and highlights the value of studies of potentially pleiotropic genes in autoimmune diseases.     

Autoimmune diseases association study with the KIAA1109–IL2–IL21 region in a Tunisian population

Autoimmune diseases (ADs) share several genetic factors resulting in similarity of disease mechanisms. For instance polymorphisms from the KIAA1109–interleukin 2 (IL2)–IL21 block in the 4q27 chromosome, has been associated with a number of autoimmune phenotypes. Here we performed a haplotype-based analysis of this AD related region in Tunisian patients. Ten single nucleotide polymorphisms (rs6534347, rs11575812, rs2069778, rs2069763, rs2069762, rs6852535, rs12642902, rs6822844, rs2221903, rs17005931) of the block were investigated in a cohort of 93 systemic lupus erythematosus (SLE), 68 ulcerative colitis (UC), 39 Crohn’s disease (CD) patients and 162 healthy control subjects of Tunisian origin. In SLE population, haplotypes AGCAGGGTC, AGAAGAGTC, AGAAGGGTC and AGCCGAGTC provided significant evidence of association with SLE risk (p = 0.013, 0.028, 0.018 and 0.048, respectively). In the UC population, haplotype AGCCGGGTC provided a susceptibility effect for UC (p = 0.025). In the CD population, haplotype CAGGCC showed a protective effect against the development of CD (p = 0.038). Haplotype AAGGTT provided significant evidence to be associated with CD risk (p = 0.007). Our results support the existence of the associations found in the KIAA1109/IL2/IL21 gene region with ADs, thus confirms that the 4q27 locus may contribute to the genetic susceptibility of ADs in the Tunisian population.     

Association study on IL4, IL13 and IL4RA polymorphisms in mite-sensitized persistent allergic rhinitis in a Chinese population

Background

The IL4, IL13, and IL4 receptor α chain (IL4RA) genes are candidate genes for atopic diseases. We hypothesized that the polymorphisms in these genes are associated with persistent allergic rhinitis (PER).

Objective

To investigate the association of the potential functional polymorphisms in IL4, IL13, and IL4RA with PER induced by house dust mites in a Chinese population.

Methods

Using the TaqMan method, we genotyped six single nucleotide polymorphisms (SNPs) including C-590T in IL4, C-1055T and Arg130Gln in IL13, and Ile50Val, Ser478Pro and Gln551Arg in IL4RA, in a case-control study of 265 patients with PER and 275 healthy controls.

Results

We found that the CT/CC genotypes in IL4 C-590T were associated with a significantly decreased risk of mite-sensitized PER [adjusted odds ratio (OR)  = 0.64, 95% confidence interval (CI) 0.45–0.92], compared to the TT genotype. Furthermore, PER patients with CT/CC genotypes had significantly lower serum levels of total IgE than those with TT genotype (P = 0.001). However, there was no significant association of the IL13 and IL4RA polymorphisms with mite-sensitized PER (P>0.05).

Conclusions

Our results suggest that the C-590T polymorphism in IL4 may contribute to the susceptibility to mite-sensitized PER in a Chinese population.     

Apolipoprotein E region molecular signatures of Alzheimer's disease

Although the APOE region is the strongest genetic risk factor for Alzheimer's diseases (ADs), its pathogenic role remains poorly understood. Elucidating genetic predisposition to ADs, a subset of age‐related diseases characteristic for postreproductive period, is hampered by the undefined role of evolution in establishing molecular mechanisms of such diseases. This uncertainty is inevitable source of natural‐selection–free genetic heterogeneity in predisposition to ADs. We performed first large‐scale analysis of linkage disequilibrium (LD) structures characterized by 30 polymorphisms from five genes in the APOE 19q13.3 region (BCAM, NECTIN2, TOMM40, APOE, and APOC1) in 2,673 AD‐affected and 16,246 unaffected individuals from five cohorts. Consistent with the undefined role of evolution in age‐related diseases, we found that these structures, being highly heterogeneous, are significantly different in subjects with and without ADs. The pattern of the difference represents molecular signature of AD comprised of single nucleotide polymorphisms (SNPs) from all five genes in the APOE region. Significant differences in LD in subjects with and without ADs indicate SNPs from different genes likely involved in AD pathogenesis. Significant and highly heterogeneous molecular signatures of ADs provide unprecedented insight into complex polygenetic predisposition to ADs in the APOE region. These findings are more consistent with a complex haplotype than with a single genetic variant origin of ADs in this region.     

Identification of BACE2 as an avid ?-amyloid-degrading protease

Background

Glutathione S-transferase omega-1 and 2 genes (GSTO1, GSTO2), residing within an Alzheimer and Parkinson disease (AD and PD) linkage region, have diverse functions including mitigation of oxidative stress and may underlie the pathophysiology of both diseases. GSTO polymorphisms were previously reported to associate with risk and age-at-onset of these diseases, although inconsistent follow-up study designs make interpretation of results difficult. We assessed two previously reported SNPs, GSTO1 rs4925 and GSTO2 rs156697, in AD (3,493 ADs vs. 4,617 controls) and PD (678 PDs vs. 712 controls) for association with disease risk (case-controls), age-at-diagnosis (cases) and brain gene expression levels (autopsied subjects).

Results

We found that rs156697 minor allele associates with significantly increased risk (odds ratio = 1.14, p = 0.038) in the older ADs with age-at-diagnosis > 80 years. The minor allele of GSTO1 rs4925 associates with decreased risk in familial PD (odds ratio = 0.78, p = 0.034). There was no other association with disease risk or age-at-diagnosis. The minor alleles of both GSTO SNPs associate with lower brain levels of GSTO2 (p = 4.7 × 10^-11-1.9 × 10^-27), but not GSTO1. Pathway analysis of significant genes in our brain expression GWAS, identified significant enrichment for glutathione metabolism genes (p = 0.003).

Conclusion

These results suggest that GSTO locus variants may lower brain GSTO2 levels and consequently confer AD risk in older age. Other glutathione metabolism genes should be assessed for their effects on AD and other chronic, neurologic diseases.     

iNKT Cells Are Responsible for the Apoptotic Reduction of Basophils That Mediate Th2 Immune Responses Elicited by Papain in Mice Following γPGA Stimulation

Recent studies have demonstrated that Bacillus subtilis-derived poly-gamma glutamic acid (γPGA) treatment suppresses the development of allergic diseases such as atopic dermatitis (AD). Although basophils, an innate immune cell, are known to play critical roles in allergic immune responses and repeated long-term administration of γPGA results in decreased splenic basophils in an AD murine model, the underlying mechanisms by which γPGA regulates basophil frequency remain unclear. To investigate how γPGA modulates basophils, we employed basophil-mediated Th2 induction in vivo model elicited by the allergen papain protease. Repeated injection of γPGA reduced the abundance of basophils and their production of IL4 in mice, consistent with our previous study using NC/Nga AD model mice. The depletion of basophils by a single injection of γPGA was dependent on the TLR4/DC/IL12 axis. CD1d-dependent Vα14 TCR invariant natural killer T (iNKT) cells are known to regulate a variety of immune responses, such as allergy. Because iNKT cell activation is highly sensitive to IL12 produced by DCs, we evaluated whether the effect of γPGA on basophils is mediated by iNKT cell activation. We found that in vivo γPGA treatment did not induce the reduction of basophils in iNKT cell-deficient CD1d KO mice, suggesting the critical role of iNKT cells in γPGA-mediated basophil depletion at the early time points. Furthermore, increased apoptotic basophil reduction triggered by iNKT cells upon γPGA stimulation was mainly attributed to Th1 cytokines such as IFNγ and TNFα, consequently resulting in inhibition of papain-induced Th2 differentiation via diminishing basophil-derived IL4. Taken together, our results clearly demonstrate that γPGA-induced iNKT cell polarization toward the Th1 phenotype induces apoptotic basophil depletion, leading to the suppression of Th2 immune responses. Thus, elucidation of the crosstalk between innate immune cells will contribute to the design and development of new therapeutics for Th2-mediated immune diseases such as AD.     

Distinct gene expression profiles of human type 1 and type 2 T helper cells

Background

The development and activation of CD4⁺ helper T cell (Th) subsets with distinct patterns of unbalanced production of cytokines play an important part in infectious, allergic and autoimmune diseases. Human neonatal cord blood CD4⁺ Th cells can be polarized into type 1 or type 2-like effector cells in vitro by culturing them in the presence of interleukin (IL)-12 or IL-4, respectively. We have exploited this experimental system to identify marker genes that are differentially expressed by polarized Th1 and Th2 cells. An oligonucleotide microarray specifically designed to screen for inflammation-related candidate genes was used and the differential expression was further validated with a quantitative real-time RT-PCR method.

Results

In addition to the previously described marker genes of Th cells, we report subtle changes in the expression of several other genes that represent growth factors, receptors and other signaling molecules in polarized Th1 and Th2 cell subsets. Additionally, we describe a novel set of genes as Th1/Th2 differentiation markers for cells activated by anti-CD3 and anti-CD28 antibodies.

Conclusions

This study demonstrates the power of the targeted use of microarrays in combination with quantitative real-time RT-PCR in identifying and validating new marker genes for gene expression studies.     

Inherited Inflammatory Response Genes Are Associated with B-Cell Non-Hodgkin’s Lymphoma Risk and Survival

Background

Malignant B-cell clones are affected by both acquired genetic alterations and by inherited genetic variations changing the inflammatory tumour microenvironment.

Methods

We investigated 50 inflammatory response gene polymorphisms in 355 B-cell non-Hodgkin’s lymphoma (B-NHL) samples encompassing 216 diffuse large B cell lymphoma (DLBCL) and 139 follicular lymphoma (FL) and 307 controls. The effect of single genes and haplotypes were investigated and gene-expression analysis was applied for selected genes. Since interaction between risk genes can have a large impact on phenotype, two-way gene-gene interaction analysis was included.

Results

We found inherited SNPs in genes critical for inflammatory pathways; TLR9, IL4, TAP2, IL2RA, FCGR2A, TNFA, IL10RB, GALNT12, IL12A and IL1B were significantly associated with disease risk and SELE, IL1RN, TNFA, TAP2, MBL2, IL5, CX3CR1, CHI3L1 and IL12A were, associated with overall survival (OS) in specific diagnostic entities of B-NHL. We discovered noteworthy interactions between DLBCL risk alleles on IL10 and IL4RA and FL risk alleles on IL4RA and IL4. In relation to OS, a highly significant interaction was observed in DLBCL for IL4RA (rs1805010) * IL10 (rs1800890) (HR = 0.11 (0.02–0.50)). Finally, we explored the expression of risk genes from the gene-gene interaction analysis in normal B-cell subtypes showing a different expression of IL4RA, IL10, IL10RB genes supporting a pathogenetic effect of these interactions in the germinal center.

Conclusions

The present findings support the importance of inflammatory genes in B-cell lymphomas. We found association between polymorphic sites in inflammatory response genes and risk as well as outcome in B-NHL and suggest an effect of gene-gene interactions during the stepwise oncogenesis.     

Rule-Based Models of the Interplay between Genetic and Environmental Factors in Childhood Allergy

Both genetic and environmental factors are important for the development of allergic diseases. However, a detailed understanding of how such factors act together is lacking. To elucidate the interplay between genetic and environmental factors in allergic diseases, we used a novel bioinformatics approach that combines feature selection and machine learning. In two materials, PARSIFAL (a European cross-sectional study of 3113 children) and BAMSE (a Swedish birth-cohort including 2033 children), genetic variants as well as environmental and lifestyle factors were evaluated for their contribution to allergic phenotypes. Monte Carlo feature selection and rule based models were used to identify and rank rules describing how combinations of genetic and environmental factors affect the risk of allergic diseases. Novel interactions between genes were suggested and replicated, such as between ORMDL3 and RORA, where certain genotype combinations gave odds ratios for current asthma of 2.1 (95% CI 1.2-3.6) and 3.2 (95% CI 2.0-5.0) in the BAMSE and PARSIFAL children, respectively. Several combinations of environmental factors appeared to be important for the development of allergic disease in children. For example, use of baby formula and antibiotics early in life was associated with an odds ratio of 7.4 (95% CI 4.5-12.0) of developing asthma. Furthermore, genetic variants together with environmental factors seemed to play a role for allergic diseases, such as the use of antibiotics early in life and COL29A1 variants for asthma, and farm living and NPSR1 variants for allergic eczema. Overall, combinations of environmental and life style factors appeared more frequently in the models than combinations solely involving genes. In conclusion, a new bioinformatics approach is described for analyzing complex data, including extensive genetic and environmental information. Interactions identified with this approach could provide useful hints for further in-depth studies of etiological mechanisms and may also strengthen the basis for risk assessment and prevention.     

Unifying candidate gene and GWAS Approaches in Asthma

The first genome wide association study (GWAS) for childhood asthma identified a novel major susceptibility locus on chromosome 17q21 harboring the ORMDL3 gene, but the role of previous asthma candidate genes was not specifically analyzed in this GWAS. We systematically identified 89 SNPs in 14 candidate genes previously associated with asthma in >3 independent study populations. We re-genotyped 39 SNPs in these genes not covered by GWAS performed in 703 asthmatics and 658 reference children. Genotyping data were compared to imputation data derived from Illumina HumanHap300 chip genotyping. Results were combined to analyze 566 SNPs covering all 14 candidate gene loci. Genotyped polymorphisms in ADAM33, GSTP1 and VDR showed effects with p-values <0.0035 (corrected for multiple testing). Combining genotyping and imputation, polymorphisms in DPP10, EDN1, IL12B, IL13, IL4, IL4R and TNF showed associations at a significance level between p = 0.05 and p = 0.0035. These data indicate that (a) GWAS coverage is insufficient for many asthma candidate genes, (b) imputation based on these data is reliable but incomplete, and (c) SNPs in three previously identified asthma candidate genes replicate in our GWAS population with significance after correction for multiple testing in 14 genes.     

Different Genetic Associations of the IgE Production among Fetus,Infancy and Childhood

Elevation of serum IgE levels has long been associated with allergic diseases. Many genes have been linked to IgE production, but few have been linked to the developmental aspects of genetic association with IgE production. To clarify developmental genetic association, we investigated what genes and gene-gene interactions affect IgE levels among fetus, infancy and childhood in Taiwan individuals. A birth cohort of 571 children with completion of IgE measurements from newborn to 1.5, 3, and 6 years of age was subject to genetic association analysis on the 384-customized SNPs of 159 allergy candidate genes. Fifty-three SNPs in 37 genes on innate and adaptive immunity, and stress and response were associated with IgE production. Polymorphisms of the IL13, and the HLA-DPA1 and HLA-DQA1 were, respectively, the most significantly associated with the IgE production at newborn and 6 years of age. Analyses of gene-gene interactions indentified that the combination of NPSR1, rs324981 TT with FGF1, rs2282797 CC had the highest risk (85.7%) of IgE elevation at 1.5 years of age (P = 1.46×10⁻⁴). The combination of IL13, CYFIP2 and PDE2A was significantly associated with IgE elevation at 3 years of age (P = 5.98×10⁻⁷), and the combination of CLEC2D, COLEC11 and CCL2 was significantly associated with IgE elevation at 6 years of age (P = 6.65×10⁻⁷). Our study showed that the genetic association profiles of the IgE production among fetus, infancy and childhood are different. Genetic markers for early prediction and prevention of allergic sensitization may rely on age-based genetic association profiles.     

CCR3, CCR5, CCR8 and CXCR3 expression in memory T helper cells from allergic rhinitis patients, asymptomatically sensitized and healthy individuals

Background

Interleukin 8 (IL8) belongs to the family of chemokines. It mediates the activation and migration of neutrophils from peripheral blood into tissue and hereby plays a pivotal role in the initiation of inflammation. Thus it is important in inflammatory lung diseases like bronchial asthma or severe infections by Respiratory Syncytial Virus (RSV). IL8 acts through binding to the IL8-Receptor alpha (IL8RA). For both genes association with asthma has been described. In addition, IL8 has been found in association with RSV bronchiolitis. The aim of our study was to test both genes for association with asthma and severe RSV infections. In addition we were interested in whether a common genetic background of both diseases exists in regards to these genes.

Methods

We genotyped the two IL8 promotor polymorphisms -251A/T and -781C/T and the three amino acid variants M31R, S276T and R335C in IL8RA on 322 children with asthma, 131 infants with severe RSV associated diseases and 270 controls. Statistical analyses made use of the Armitage's trend test for single polymorphisms and FAMHAP for calculations of haplotypes.

Results

We found association of the IL8 polymorphism -781C/T as well as IL8 haplotypes with asthma (p = 0.011 and p = 0.036, respectively). In addition, direct comparison of the asthmatic population with the RSV population revealed significant differences, both for -781C/T alone (p = 0.034) and IL8 haplotypes (p = 0.005). The amino acid variants in IL8RA were evenly distributed in between all three populations.

Conclusion

We conclude from our data that IL8 might play a role in the genetic predisposition to asthma and that these effects are different or even opposite to the effects on severe RSV diseases. Furthermore, IL8RA is unlikely to play a major role in the genetics of either disease.     

Th2 Cytokines Augment IL-31/IL-31RA Interactions via STAT6-dependent IL-31RA Expression

Interleukin 31 receptor α (IL-31RA) is a novel Type I cytokine receptor that pairs with oncostatin M receptor to mediate IL-31 signaling. Binding of IL-31 to its receptor results in the phosphorylation and activation of STATs, MAPK, and JNK signaling pathways. IL-31 plays a pathogenic role in tissue inflammation, particularly in allergic diseases. Recent studies demonstrate IL-31RA expression and signaling in non-hematopoietic cells, but this receptor is poorly studied in immune cells. Macrophages are key immune-effector cells that play a critical role in Th2-cytokine-mediated allergic diseases. Here, we demonstrate that Th2 cytokines IL-4 and IL-13 are capable of up-regulating IL-31RA expression on both peritoneal and bone marrow-derived macrophages from mice. Our data also demonstrate that IL-4Rα-driven IL-31RA expression is STAT6 dependent in macrophages. Notably, the inflammation-associated genes Fizz1 and serum amyloid A (SAA) are significantly up-regulated in M2 macrophages stimulated with IL-31, but not in IL-4 receptor-deficient macrophages. Furthermore, the absence of Type II IL-4 receptor signaling is sufficient to attenuate the expression of IL-31RA in vivo during allergic asthma induced by soluble egg antigen, which may suggest a role for IL-31 signaling in Th2 cytokine-driven inflammation and allergic responses. Our study reveals an important counter-regulatory role between Th2 cytokine and IL-31 signaling involved in allergic diseases.     

Bee venom ameliorates ovalbumin induced allergic asthma via modulating CD4+CD25+ regulatory T cells in mice

Asthma is a potentially life-threatening inflammatory disease of the lung characterized by the presence of large numbers of CD4⁺ T cells. These cells produce the Th2 and Th17 cytokines that are thought to orchestrate the inflammation associated with asthma. Bee venom (BV) has traditionally been used to relieve pain and to treat chronic inflammatory diseases. Recent reports have suggested that BV might be an effective treatment for allergic diseases. However, there are still unanswered questions related to the efficacy of BV therapy in treating asthma and its therapeutic mechanism. In this study, we evaluated whether BV could inhibit asthma and whether BV inhibition of asthma could be correlated with regulatory T cells (Treg) activity. We found that BV treatment increased Treg populations and suppressed the production of Th1, Th2 and Th17-related cytokines in an in vitro culture system, including IL2, IL4, and IL17. Interestingly, production of IL10, an anti-inflammatory cytokine secreted by Tregs, was significantly augmented by BV treatment. We next evaluated the effects of BV treatment on allergic asthma in an ovalbumin (OVA)-induced mouse model of allergic asthma. Cellular profiling of the bronchoalveolar lavage (BAL) and histopathologic analysis demonstrated that peribronchial and perivascular inflammatory cell infiltrates were significantly lowered following BV treatment. BV also ameliorated airway hyperresponsiveness, a hallmark symptom of asthma. In addition, IL4 and IL13 levels in the BAL fluid were decreased in the BV treated group. Surprisingly, the beneficial effects of BV treatment on asthma were eradicated following Treg depletion by anti-CD25 antibody injection, suggesting that the major therapeutic targets of BV were Tregs. These results indicate that BV efficiently diminishes bronchial inflammation in an OVA-induced allergic asthma murine model, and that this effect might correlate with Tregs, which play an important role in maintaining immune homeostasis and suppressing the function of other T cells to limit the immune response. These results also suggest that BV has potential therapeutic value for controlling allergic asthma responses.     

Lack of Clinical Manifestations in Asymptomatic Dengue Infection Is Attributed to Broad Down-Regulation and Selective Up-Regulation of Host Defence Response Genes

Objectives

Dengue represents one of the most serious life-threatening vector-borne infectious diseases that afflicts approximately 50 million people across the globe annually. Whilst symptomatic infections are frequently reported, asymptomatic dengue remains largely unnoticed. Therefore, we sought to investigate the immune correlates conferring protection to individuals that remain clinically asymptomatic.

Methods

We determined the levels of neutralizing antibodies (nAbs) and gene expression profiles of host immune factors in individuals with asymptomatic infections, and whose cognate household members showed symptoms consistent to clinical dengue infection.

Results

We observed broad down-regulation of host defense response (innate, adaptive and matrix metalloprotease) genes in asymptomatic individuals as against symptomatic patients, with selective up-regulation of distinct genes that have been associated with protection. Selected down-regulated genes include: TNF α (TNF), IL8, C1S, factor B (CFB), IL2, IL3, IL4, IL5, IL8, IL9, IL10 and IL13, CD80, CD28, and IL18, MMP8, MMP10, MMP12, MMP15, MMP16, and MMP24. Selected up-regulated genes include: RANTES (CCL5), MIP-1α (CCL3L1/CCL3L3), MIP-1β (CCL4L1), TGFβ (TGFB), and TIMP1.

Conclusion

Our findings highlight the potential association of certain host genes conferring protection against clinical dengue. These data are valuable to better explore the mysteries behind the hitherto poorly understood immunopathogenesis of subclinical dengue infection.