Abstract 163: NFkB Promotes Oxidative Stress-induced Necrosis and Ischemia Reperfusion Injury Through NRF2-ARE Pathway

Abstract

Objective: Despite the well-studied pro-survival function of nuclear factor-κB (NFκB), recent studies suggest that NFκB may also play a pathogenic role in myocardial ischemia injury and adverse remodeling. This study aims to define a new pro-cell death role of NFκB in response to oxidative stress and the functional implications in ischemia reperfusion (I/R) injury. Methods and Results: We identified an unexpected pro-cell death role of NFκB in oxidative stress-induced necrosis, and provide new mechanistic evidence that NFκB, in cooperation with HDAC3, negatively regulates NRF2-ARE anti-oxidative signaling through transcriptional silencing. Genetic deletion of NFκB-p65 inhibits, whereas activation of NFκB promotes, oxidative stress-induced cell death and HMGB1 release, a biomarker of necrosis. Moreover, simulated ischemia reperfusion (sI/R) and doxorubicin (Dox) treatment both induce NFκB-luciferase activity in cardiomyocytes, and inhibition of NFκB diminishes sI/R- and Dox- induced necrosis. Importantly, NFκB negatively regulates NRF2-ARE activity and the expression of anti-oxidant proteins. Mechanistically, co-immunoprecipitation reveals that p65 is required for the association between NRF2 and HDAC3 and transcriptional silencing of NRF2-ARE activity. Further, the ability of HDAC3 to repress NRF2-ARE activity is lost in p65-/- cells. The HADC inhibitor TSA and NFκB inhibitor BMS-345541 both increase NRF2-ARE activity and promote cell survival following sI/R. In vivo , NFκB transcriptional activity in the heart is significantly elevated after I/R injury, which is abolished by cardiac-specific deletion of p65. Moreover, ablation of p65 using p65 fl/fl-Nkx-Cre mice reduces myocardial infarct size after acute I/R, and prevents chronic remodeling and contractile dysfunction after myocardial infarction. Conclusions: Our results identified NFκB as a key regulator of oxidative stress-induced necrosis by suppressing the NRF2-ARE anti-oxidant pathway through an HDAC3-dependent mechanism. Ablation of NFκB-p65 attenuates oxidative stress-induced necrosis and I/R injury, suggesting a new pathogenic role of the NFκB pathway, and thus a therapeutic target, in myocardial ischemic injury and remodeling.