To fully examine PM-mediated vascular hyperpermeability, we explored effector targets such as tight junction and adherens junction proteins, which are known to be critical in EC barrier maintenance. We demonstrated that PM exposure induced the relocation of tight junction protein Zona occludens-1 (ZO-1) from the cell periphery, which was accompanied by a significant reduction in the level of ZO-1 protein but not in the levels of adherens junction proteins (VE-cadherin and β-catenin). PM also mediated intracellular calcium mobilization via the transient receptor potential cation channel M2 (TRPM2), in a ROS-dependent manner with subsequent activation of the Ca 2+ -dependent protease calpain. PM-activated calpain is responsible for ZO-1 degradation and EC barrier disruption. These observations not only provide new information as to how PM disrupts endothelial tight junctions, but also represent the first evidence establishing the critical role of calpain signaling in modulating endothelial cell barrier function under oxidative stress. These results increase our understanding of PM-induced adverse cardiopulmonary outcomes. Moreover, as the newly characterized signaling cascade of ROS/TRPM2/Calpain/ ZO-1 likely has fundamental roles in regulating the cytoskeleton under oxidative stress, these novel observations may have broad applicability to vascular pathophysiology in a variety of cell types.