NADPH oxidase‐derived reactive oxygen species (ROS) have been demonstrated to mediate the activation of podocyte NLRP3 inflammasomes in response to elevated levels of homocysteine (Hcys). We have also previously provided evidence that the guanine nucleotide exchange factor Vav2 exhibits high specificity to Rac1‐mediated NADPH oxidase activation by elevated Hcys. The precise role of Vav2 in NLRP3 inflammasome activation remains unknown. The present study was designed to test whether Vav2 contributes to Hcys‐induced NLRP3 inflammasome activation through activation of NADPH oxidase. In our experiments, podocytes were transfected with either a scramble, constitutively active form of Vav2 (oncoVav2) or Vav2 shRNA plasmid directly to the nucleus via nucleofection. Confocal microscopic analysis showed Hcys treatment as well as oncoVav2 transfection alone increased colocalization between inflammasome proteins NLRP3 with ASC and NLRP3 with caspase‐1, suggesting NLRP3 inflammasome formation. Additionally, both Hcys treatment as well as oncoVav2 transfection alone resulted in increased caspase‐1 activation and IL‐1β production, signifying NLRP3 inflammasome activation. These Hcys‐induced measures of inflammasome formation and activation were all inhibited by Vav2 shRNA transfection, observed in decreased colocalization of inflammasome components, caspase‐1 activity, and IL‐1β production in podocytes. Furthermore, Vav2 shRNA transfection also prevented podocyte damage by restoring impaired VEGF secretion as well as decreased podocyte marker podocin expression seen in Hcys injured podocytes. Together, these findings suggest that NADPH oxidase activation by Vav2 overexpression by oncoVav2 was sufficient to activate NLRP3 inflammasomes, independent of Hcys treatment. However, Vav2 inhibition was able to inhibit Hcys‐induced NLRP3 inflammasome activation and podocyte injury, further substantiating the role of Vav2 in this mechanism.Grant Funding Source: Supported by NIH grants DK54927 and 1F31AG043289