Abstract Body: Background: APOE4 (which encodes apolipoprotein E4) is a common genetic risk factor for cardiovascular disease (CVD). Although apoE classically participates in lipid transport, APOE4-associated risk likely goes beyond changes in circulating lipids. Life-threatening CVD events including myocardial infarction and stroke are driven by atherogenesis and thrombosis. Whereas studies in mice have shown that apoE4 increases atherosclerosis severity, it is unknown if and how apoE4 influences thrombosis.
Hypothesis: ApoE4 promotes thrombosis.
Methods: GWAS data for venous thromboembolism (VTE) in 9 combined cohorts were analyzed. In humanized APOE3 (hE3) and APOE4 (hE4) mice thrombosis was assessed by intravital microscopy (IVM) in the mesenteric microcirculation or by inferior vena cava (IVC) partial ligation. Direct actions of apoE3 versus apoE4 on endothelial cells (EC) and their underpinnings were studied in cultured human aortic EC, quantifying the secretion of Von Willebrand Factor (vWF), a critical initiator of thrombosis.
Results: GWAS showed that APOE4 is associated with increased VTE risk, with P = 0.002. IVM revealed that compared to hE3 mice, hE4 developed larger thrombi and more rapid occlusion in both arterioles and venules. In parallel, in the IVC model hE4 mice had more frequent thrombosis and increased thrombus size. hE3 and hE4 had similar plasma lipids. In cultured EC, whereas apoE3 attenuated vWF secretion, it was enhanced by apoE4, and both processes were mediated by ApoE receptor 2 (ApoER2). ApoE4, but not apoE3, suppressed VEGF eNOS activation and NO production. In ApoER2 interactome studies apoE4 uniquely caused recruitment of the protein phosphatase 2A (PP2A) catalytic subunit. In cultured EC apoE4 also caused PP2A activation via ApoER2, and PP2A activity was increased in hE4 aortas. PP2A deletion prevented apoE4 antagonism of eNOS by preserving Akt activation, and the NO donor spermine NONOate negated apoE4 stimulation of vWF secretion. Showing that the prothrombotic actions of apoE4 are endothelial in nature, EC ApoER2 deletion fully prevented exaggerated thrombosis in hE4 mice.
Conclusions: APOE4 is a risk allele for thrombosis. Mechanistically, the apoE4-EC ApoER2 tandem enhances vWF secretion by activating PP2A and antagonizing eNOS, resulting in exaggerated thrombosis. Targeting these processes may afford protection from both thrombotic disorders and acute CVD events such as myocardial infarction and stroke in 15 to 20% of the population.