Disruption of asymmetric dimethylarginine-induced RelA/P65 association with actin in endothelial cells

Asymmetric dimethylarginine (ADMA) activates nuclear factor (NF)-κB in endothelial cells, while actin-stabilizing or -destabilizing drugs prevent ADMA-induced activation of NF-κB. Here we investigated how actin-targeting drugs regulated ADMA-induced NF-κB activation in endothelial cells. Human umbilical vein endothelial cells were treated with ADMA for 24 h in the absence and presence of cytochalasin D or jasplakinolide. Expression levels of proteins and genes were measured by immunoblotting and reverse-transcription polymerase chain reaction, respectively. Chromatin immunoprecipitation was used to detect the binding of NF-κB to the vascular cell adhesion molecule 1 (VCAM-1) promoter. The association of actin with RelA/P65 was detected by immunoprecipitation. It was demonstrated that ADMA induced IκBα degradation, increased nuclear RelA/P65 translocation, and promoted the binding of NF-κB to the VCAM-1 promoter. Consequently, this increased the expression of VCAM-1. In parallel studies, actin-stabilizing and -destabilizing drugs decreased ADMA-induced RelA/P65 nuclear translocation, interfered with NF-κB binding to the VCAM-1 promoter and prevented the expression of VCAM-1. This was independent of total RelA/P65 levels and ADMA-induced IκBα degradation. Most importantly, the association of RelA/P65 with actin was increased after stimulation with ADMA, and impaired after treatment with actin-targeting drugs. In brief, actin-stabilizing or -destabilizing drugs interfere with the ADMA-induced association of RelA/P65 with actin, and consequently disrupt NF-κB activation.