Introduction: Proteolysis of vascular endothelial (VE)-cadherin is a biomarker for disruption of endothelial cell (EC) barrier function, with elevated levels of soluble VE (sVE)-cadherin associated with atherosclerosis and sepsis. Factor XI (FXI) plays a regulatory role in inflammation, and while a relationship between FXI activity and VE-cadherin expression levels has been observed, the mechanism by which this process is mediated is unknown. Hypothesis: Activated FXI (FXIa) induces VE-cadherin proteolysis. Methods: Human umbilical vein ECs were stimulated with FXIa (30 and 5 nM) for six hours. Western blot analysis was used to study VE-cadherin proteolysis, disintegrin and metalloproteinase 10 (ADAM10) expression, FXIa-PAI-1 or -very low density lipoprotein receptor (VLDLR) interactions, and cell signaling events. SVE-cadherin in baboons challenged with Staphylococcus aureus was detected by ELISA. Results: FXIa (5 nM) generated a VE-cadherin fragment and increased ECs permeability; the serine protease inhibitor, PPACK, and an inhibitor of ADAM10 reversed these processes. FXIa induced the expression of active ADAM10 on ECs. The receptor-associated protein (RAP) blocked FXIa-induced VE-cadherin proteolysis by preventing FXIa interaction with PAI-1 and VLDLR on ECs. FXIa-VLDLR interaction induced Dab1 and Src kinases phosphorylation. FXIa activated the VEGFR2-PLCgamma1-ERK1/2 signaling pathway. The ERK1/2 inhibitor, LY3214996, and the Src kinases inhibitor, PP2, inhibited VE-cadherin proteolysis by FXIa. PLCgamma1 phosphorylation was also prevented with PP2. The VEGFR2 kinase inhibitor, SU1498, prevented the cleavage of VE-cadherin by FXIa; in contrast, a VEGF blocking antibody had no effect on FXIa-induced VE-cadherin cleavage. Finally, the inhibition of FXI activation in a baboon model of sepsis significantly decreased the levels of sVE-cadherin. Conclusion: The complex formation of FXIa with PAI-1 and VLDLR promoted Src kinase phosphorylation. Src activation triggered the VEGFR2-PLCgamma1-ERK1/2 signaling pathway, which induced the trafficking of active ADAM10 to the EC surfaces inducing the cleavage of VE-cadherin. FXI may be a druggable target to protect EC barrier function in inflammatory diseases.