Hemodynamic regulation of perivalvular endothelial gene expression prevents deep venous thrombosis
John D. Welsh, Mark H. Hoofnagle, Sharika Bamezai, Michael Oxendine, Lillian Lim, Joshua D. Hall, Jisheng Yang, Susan Schultz, James Douglas Engel, Tsutomu Kume, Guillermo Oliver, Juan M. Jimenez, Mark L. Kahn
John D. Welsh, Mark H. Hoofnagle, Sharika Bamezai, Michael Oxendine, Lillian Lim, Joshua D. Hall, Jisheng Yang, Susan Schultz, James Douglas Engel, Tsutomu Kume, Guillermo Oliver, Juan M. Jimenez, Mark L. Kahn
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Research Article Vascular biology

Hemodynamic regulation of perivalvular endothelial gene expression prevents deep venous thrombosis

Abstract

Deep venous thrombosis (DVT) and secondary pulmonary embolism cause approximately 100,000 deaths per year in the United States. Physical immobility is the most significant risk factor for DVT, but a molecular and cellular basis for this link has not been defined. We found that the endothelial cells surrounding the venous valve, where DVTs originate, express high levels of FOXC2 and PROX1, transcription factors known to be activated by oscillatory shear stress. The perivalvular venous endothelial cells exhibited a powerful antithrombotic phenotype characterized by low levels of the prothrombotic proteins vWF, P-selectin, and ICAM1 and high levels of the antithrombotic proteins thrombomodulin (THBD), endothelial protein C receptor (EPCR), and tissue factor pathway inhibitor (TFPI). The perivalvular antithrombotic phenotype was lost following genetic deletion of FOXC2 or femoral artery ligation to reduce venous flow in mice, and at the site of origin of human DVT associated with fatal pulmonary embolism. Oscillatory blood flow was detected at perivalvular sites in human veins following muscular activity, but not in the immobile state or after activation of an intermittent compression device designed to prevent DVT. These findings support a mechanism of DVT pathogenesis in which loss of muscular activity results in loss of oscillatory shear–dependent transcriptional and antithrombotic phenotypes in perivalvular venous endothelial cells, and suggest that prevention of DVT and pulmonary embolism may be improved by mechanical devices specifically designed to restore perivalvular oscillatory flow.

Authors

John D. Welsh, Mark H. Hoofnagle, Sharika Bamezai, Michael Oxendine, Lillian Lim, Joshua D. Hall, Jisheng Yang, Susan Schultz, James Douglas Engel, Tsutomu Kume, Guillermo Oliver, Juan M. Jimenez, Mark L. Kahn

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Figure 3

Loss of FOXC2 results in loss of THBD and EPCR and gain of vWF in perivalvular venous endothelial cells.

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Loss of FOXC2 results in loss of THBD and EPCR and gain of vWF in periva...
Immunostaining of mouse saphenous veins was performed in animals lacking FOXC2 in perivalvular venous endothelial cells (Foxc2VVKO mice) and littermate controls (Foxc2fl/fl mice). (A and B) Loss of FOXC2 (n = 6 for control and knockout valves) mildly reduces perivalvular endothelial expression of PROX1 (n = 6 for control and knockout valves). (C–E) Loss of FOXC2 results in loss of perivalvular endothelial expression of the antithrombotic proteins THBD (n = 5 control valves, n = 7 knockout valves) and EPCR (n = 8 control valves, n = 9 knockout valves) and gain of the prothrombotic protein vWF (n = 6 control valves, n = 7 knockout valves). (F) Loss of FOXC2 does not significantly alter the expression of the antithrombotic protein TFPI (n = 4 control valves, n = 6 knockout valves). White dashed lines indicate luminal venous endothelial cells, and green dashed lines indicate perivalvular endothelial cells. For all graphs the mean is represented by the bar with dots for each replicate, and error bars indicate SD. Significance for each single comparison was determined by an unpaired 2-tailed Mann-Whitney test. *P < 0.05; **P < 0.01; ***P < 0.005. Scale bars: 50 μm.