ApoE4 Promotes Thrombosis via Endothelial Cell ApoER2 and PP2A Activation
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Background
APOE4 , the variant of apolipoprotein E carried by 25% of individuals, is a common genetic risk factor for cardiovascular disease (CVD). Although ApoE classically participates in lipid transport, APOE4 -associated risk goes beyond impact on circulating lipids. Life-threatening CVD events including myocardial infarction and stroke are driven by atherogenesis and thrombosis. In mice ApoE4 increases atherosclerosis severity, but whether other major drivers of CVD events are influenced by ApoE4 is unknown.
Methods
GWAS data for venous thromboembolism (VTE) were analyzed. In humanized APOE3 (hE3) and APOE4 (hE4) mice, thrombosis was assessed by intravital microscopy (IVM) in the mesenteric microcirculation and by inferior vena cava (IVC) partial ligation. Actions of ApoE3 versus ApoE4 on endothelial cells (EC) and their underpinnings were studied in cultured human and mouse aortic EC, interrogating interactomes with immunoprecipitation-mass spectrometry and quantifying the secretion of Von Willebrand Factor (vWF), a critical initiator of thrombosis. Single cell transcriptomics datasets were queried do localize endothelial cell gene expression.
Results
GWAS showed that APOE4 is associated with increased VTE risk, and whereas plasma lipids were similar, both microvascular and venous thrombosis were markedly increased in hE4 compared to hE3 mice. 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 by causing the recruitment of the protein phosphatase 2A (PP2A) catalytic subunit to ApoER2 and the activation of PP2A. PP2A deletion prevented ApoE4-induced eNOS antagonism and vWF secretion by preserving Akt activation, and the NO donor spermine NONOate negated apoE4 stimulation of vWF secretion. PP2A activity was increased in hE4 aortas and IVC, and EC ApoER2 deletion or pharmacologic PP2A inhibition fully prevented exaggerated thrombosis in hE4 mice. In human great saphenous vein ApoER2 is primarily expressed in valvular endothelium.
Conclusions
APOE4 is a risk allele for thrombosis, and ApoE4 is prothrombotic in microvasculature and veins in mice. Mechanistically, the ApoE4-EC ApoER2 tandem enhances vWF secretion by recruiting and activating PP2A and antagonizing eNOS, resulting in exaggerated thrombosis. In human veins ApoER2 is expressed in valvular endothelium, which is the most common site of initiation of venous thrombosis. Targeting these processes may afford protection from both primary thrombotic disorders like VTE and acute CVD events such as myocardial infarction and stroke in 25% of the population.
CLINICAL PERSPECTIVE
What is New?
Apolipoprotein E4 (ApoE4), a variant of ApoE carried by 25% of the population, increases cardiovascular disease risk independent of impact on circulating lipids, but the mechanism is unknown.
This study reveals that through direct actions on endothelial cells, ApoE4 promotes thrombosis, which plays a critical role in acute cardiovascular events like myocardial infarction and stroke.
Our findings demonstrate that ApoE4 uniquely causes ApoE receptor 2 (ApoER2) in endothelial cells to recruit and activate the enzyme protein phosphatase 2A (PP2A), leading to exaggerated secretion of Von Willebrand Factor (vWF), which is a major initiator of thrombosis.
What Are the Clinical Implications?
We demonstrate that ApoE4 promotes both microvascular and venous thrombosis, and that ApoER2 and PP2A are expressed in human leg vein endothelium that are instrumental in the initiation of venous thromboembolism, which has a 30-day case fatality rate of up to 30%.
Until now there have been no interventions available to break the link between ApoE4 and life-threatening vascular disorders, and we demonstrate that pharmacologic inhibition of PP2A fully prevents the exaggerated thrombosis caused by ApoE4.
Targeting the mechanisms initiated by ApoE4 and ApoER2 in the endothelium may afford protection from both primary thrombotic orders like VTE and acute cardiovascular events such as myocardial infarction and stroke in 25% of the population.
