Genetic Profile of Endotoxemia Reveals an Association With Thromboembolism and Stroke

Abstract

Background Translocation of lipopolysaccharide from gram-negative bacteria into the systemic circulation results in endotoxemia. In addition to acute infections, endotoxemia is detected in cardiometabolic disorders, such as cardiovascular diseases and obesity. Methods and Results We performed a genome-wide association study of serum lipopolysaccharide activity in 11 296 individuals from 6 different Finnish study cohorts. Endotoxemia was measured by limulus amebocyte lysate assay in the whole population and by 2 other techniques (Endolisa and high-performance liquid chromatography/tandem mass spectrometry) in subpopulations. The associations of the composed genetic risk score of endotoxemia and thrombosis-related clinical end points for 195 170 participants were analyzed in FinnGen. Lipopolysaccharide activity had a genome-wide significant association with 741 single-nucleotide polymorphisms in 5 independent loci, which were mainly located at genes affecting the contact activation of the coagulation cascade and lipoprotein metabolism and explained 1.5% to 9.2% of the variability in lipopolysaccharide activity levels. The closest genes included KNG1, KLKB1, F12, SLC34A1, YPEL4, CLP1, ZDHHC5, SERPING1, CBX5, and LIPC. The genetic risk score of endotoxemia was associated with deep vein thrombosis, pulmonary embolism, pulmonary heart disease, and venous thromboembolism. Conclusions The biological activity of lipopolysaccharide in the circulation (ie, endotoxemia) has a small but highly significant genetic component. Endotoxemia is associated with genetic variation in the contact activation pathway, vasoactivity, and lipoprotein metabolism, which play important roles in host defense, lipopolysaccharide neutralization, and thrombosis, and thereby thromboembolism and stroke.

Keywords: coagulation; contact activation; endotoxin; gene; genome‐wide association study; lipopolysaccharide.

Figure 1. Manhattan and QQ plots of genome‐wide association study (GWAS) results combined in fixed‐effects meta‐analysis.

We performed a GWAS of endotoxemia, measured by limulus amebocyte lysate assay in 11 296 individuals with Finnish ancestry. The horizontal red line represents genome‐wide significance (P<5×10⁻⁸). Single‐nucleotide polymorphisms in 5 independent loci available in all cohorts passed the genome‐wide significance threshold. Inflation of the P values is presented in the QQ plot (λ_qc=1.049).

Figure 2. Forest plots of the lead single‐nucleotide polymorphisms of loci reaching genome‐wide significance in meta‐analysis of the genome‐wide association study for endotoxemia.

The meta‐analysis included 11 296 samples. Presented point size is proportional to the inverse of SE of the estimate. DinnDiane: rest, FinnDiane cohort of individuals with unclassified diabetes; FinnDiane: T1D, FinnDiane cohort of participants with type 1 diabetes; FR indicates FINRISK; and FT16 and VpEpi, subpopulations of Finnish Twin Cohort.

Figure 3. Hypothesized connection between intrinsic pathway of coagulation and genetic variants associating with endotoxemia.

Genetic associations connect endotoxemia to increased coagulation: via gene expression (expression quantitative trait loci [eQTL]), protein expression, and protein activity. Two possible mechanisms are hypothesized to explain the association. Bradykinin is cleaved from kininogen and can affect intestinal permeability by allowing increased microbial leakage from the gut. Immunothrombosis, a mechanism proposed to be involved in normal immunology, is hypothesized to alter both immunological defense and formation of thrombosis when dysregulated. Genetic risk score and Mendelian randomization analysis connect endotoxemia to venous thromboembolism, deep vein thrombosis, and stroke. GWAS indicates genome‐wide association study.

Figure 4. Associations of the cardiovascular disease end points with 5 endotoxemia‐associated single‐nucleotide polymorphisms (SNPs) and the composed genetic risk score (GRS).

A, Significant associations of the lead SNPs with stroke events in the Megastroke population. P value for logistic regression analysis is presented on the right side. The asterisk marking represents P<0.01. B, GRS of endotoxemia (LPS‐GRS) is calculated from the 5 lead SNP genotypes, based on stepwise conditional regression (GCTA–conditional and joint analysis [COJO]) of lipopolysaccharide genome‐wide associated study results weighted with effect sizes. False discovery rate (accounting for total of 46 end points tested) is presented on the right side. Population consists of 195 170 individuals from the FinnGen study. Full list of analyzed phenotypes is in Table S8. C, Association between endotoxemia and prothrombotic end points is analyzed by Mendelian randomization in Megastroke (for ischemic stroke) and UK Biobank populations using MR‐base platform. The 5 lead SNPs, based on GCTA‐COJO, were used in the analysis. DVT indicates deep vein thrombosis; OR, odds ratio; and TOAST, classification of 5 subtypes of ischemic stroke.