Background Pre-statin trials reported positive effects of omega-3 fatty acids (n-3 PUFA) in cardiovascular disease (CVD), whereas recent studies and meta-analyses have not reproduced these results. The effect of n-3 PUFA in patients with familial hypercholesterolemia (FH), a group with particularly high risk of CVD, is not well established. Objective We investigated the effect of n-3 PUFA on the early stage of atherosclerosis in FH patients by evaluating in vivo (peripheral arterial tonometry (PAT)) and in vitro (plasma asymmetric dimethylarginine (ADMA) and E-selectin) endothelial function. Methods This was a double blind, placebo-controlled crossover study with 34 FH patients on statin treatment (mean age 46.6 years). In random order, all individuals were treated for three months with high dose n-3 PUFA (2g x2) and three months placebo (olive oil, 2g x2), separated by a three months washout period. Anthropometric data, blood samples and PAT were collected at four time points. Results There were no significant changes in reactive hyperemia index (RHI) measured by PAT after n-3 PUFA compared to placebo, median RHI after n-3 PUFA was 1.98 and after placebo 1.96, p=0.51. No significant changes were detected in the soluble endothelial marker ADMA (in two different assays) when comparing n-3 PUFA and placebo (p=0.92 and 0.14, respectively). Finally, the level of E-selectin did not change significantly during the trial, p=0.26. Conclusion Addition of n-3 PUFA to standard lipid-lowering treatment in genetically verified FH patients did not affect the in vivo endothelial function or soluble endothelial markers.

Background: It remains uncertain whether activation of the complement system, assessed by the soluble terminal C5b-9 complement complex (plasma TCC), is associated with future risk of incident venous thromboembolism (VTE). Objectives:To investigate the association between plasma levels of TCC and future risk of incident VTE in a nested case-control study, and to explore genetic variants associated with TCC using protein quantitative trait loci analysis of exome sequencing data. Methods:We sampled 415 VTE cases and 848 age-and sex-matched controls from a population-based cohort, the Tromsø study. Logistic regression models were used to calculate odds ratios with 95% confidence intervals for VTE across quartiles of plasma levels of TCC. Whole exome sequencing was conducted using the Agilent SureSelect 50 Mb capture kit.Results: The risk of VTE increased across increasing quartiles of plasma TCC, particularly for unprovoked VTE. Participants with TCC in the highest quartile (>1.40 complement arbitrary units/mL) had an odds ratio for unprovoked VTE of 1.74 (95% confidence interval: 1.10-2.78) compared with those with TCC in the lowest quartile (≤0.80 complement arbitrary units/mL) in analyses adjusted for age, sex, and body mass index. A substantially higher risk for VTE was observed in samples taken shortly before VTE event. We found no association between genome-wide or complementrelated gene variants and plasma levels of TCC. Conclusions:We found that high levels of plasma TCC were associated with VTE risk, and unprovoked events in particular. There was no genome-wide association between gene variants and plasma levels of TCC. K E Y W O R D S complement system, protein quantitative trait loci analysis, terminal complement complex, venous thromboembolism, whole exome sequencing | 935 HØILAND et AL. S U PP O RTI N G I N FO R M ATI O N Additional supporting information may be found online in the Supporting Information section at the end of the article. How to cite this article: Høiland II, Liang RA, Braekkan SK, Pettersen K, Ludviksen JK, Latysheva N, et al. Complement activation assessed by the plasma terminal complement complex and future risk of venous thromboembolism. J Thromb Haemost. 2019;17:934-943. https ://doi.

Dysfunctional complement activation and Toll-like receptor signaling immediately after trauma are associated with development of trauma-induced coagulopathy and multiple organ dysfunction syndrome. We assessed the efficacy of the combined inhibition therapy of complement factor C5 and the TLR co-receptor CD14 on thrombo-inflammation and organ damage in an exploratory 72-h polytrauma porcine model, conducted under standard surgical and intensive care management procedures. Twelve male pigs were subjected to polytrauma, followed by resuscitation (ATLS® guidelines) and operation of the femur fracture (intramedullary nailing technique). The pigs were allocated to combined C5 and CD14 inhibition therapy group (n=4) and control group (n=8). The therapy group received intravenously C5 inhibitor (RA101295) and anti-CD14 antibody (rMil2) 30 min post-trauma. Controls received saline. Combined C5 and CD14 inhibition reduced the blood levels of the terminal complement complex (TCC) by 70% (p=0.004), CRP by 28% (p=0.004), and IL-6 by 52% (p=0.048). The inhibition therapy prevented the platelet consumption by 18% and TAT formation by 77% (p=0.008). Moreover, the norepinephrine requirements in the treated group were reduced by 88%. The inhibition therapy limited the organ damage, thereby reducing the blood lipase values by 50% (p=0.028), LDH by 30% (p=0.004), AST by 33%, and NGAL by 30%. Immunofluorescent analysis of the lung tissue revealed C5b-9 deposition on blood vessels in five from the untreated, and in none of the treated animals. In kidney and liver, the C5b-9 deposition was similarly detected mainly the untreated as compared to the treated animals. Combined C5 and CD14 inhibition limited the inflammatory response, the organ damage, and reduced the catecholamine requirements after experimental polytrauma and might be a promising therapeutic approach.

IntroductionAir embolism may complicate invasive medical procedures. Bubbles trigger complement C3-mediated cytokine release, coagulation, and platelet activation in vitro in human whole blood. Since these findings have not been verified in vivo, we aimed to examine the effects of air embolism in pigs on thromboinflammation.MethodsForty-five landrace pigs, average 17 kg (range 8.5-30), underwent intravenous air infusion for 300 or 360 minutes (n=29) or served as sham (n=14). Fourteen pigs were excluded due to e.g. infections or persistent foramen ovale. Blood was analyzed for white blood cells (WBC), complement activation (C3a and terminal C5b-9 complement complex [TCC]), cytokines, and hemostatic parameters including thrombin-antithrombin (TAT) using immunoassays and rotational thromboelastometry (ROTEM). Lung tissue was analyzed for complement and cytokines using qPCR and immunoassays. Results are presented as medians with interquartile range.ResultsIn 24 pigs receiving air infusion, WBC increased from 17×109/L (10-24) to 28 (16-42) (p<0.001). C3a increased from 21 ng/mL (15-46) to 67 (39-84) (p<0.001), whereas TCC increased only modestly (p=0.02). TAT increased from 35 µg/mL (28-42) to 51 (38-89) (p=0.002). ROTEM changed during first 120 minutes: Clotting time decreased from 613 seconds (531-677) to 538 (399-620) (p=0.006), clot formation time decreased from 161 seconds (122-195) to 124 (83-162) (p=0.02) and α-angle increased from 62 degrees (57-68) to 68 (62-74) (p=0.02). In lungs from pigs receiving air compared to sham animals, C3a was 34 ng/mL (14-50) versus 4.1 (2.4-5.7) (p<0.001), whereas TCC was 0.3 CAU/mL (0.2-0.3) versus 0.2 (0.1-0.2) (p=0.02). Lung cytokines in pigs receiving air compared to sham animals were: IL-1β 302 pg/mL (190-437) versus 107 (66-120), IL-6 644 pg/mL (358-1094) versus 25 (23-30), IL-8 203 pg/mL (81-377) versus 21 (20-35), and TNF 113 pg/mL (96-147) versus 16 (13-22) (all p<0.001). Cytokine mRNA in lung tissue from pigs receiving air compared to sham animals increased 12-fold for IL-1β, 121-fold for IL-6, and 17-fold for IL-8 (all p<0.001).ConclusionVenous air embolism in pigs activated C3 without a corresponding C5 activation and triggered thromboinflammation, consistent with a C3-dependent mechanism. C3-inhibition might represent a therapeutic approach to attenuate this response.

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