Nrf2 transcription factor modulates endothelial inflammatory response activated by TNF-α

Atherosclerosis is a disease characterized by hardening and thickening of the arterial system, and is a major cause of mortality worldwide. The progression of the disease is generally believed to have both oxidative and inflammatory components that ultimately lead to artery calcification and plaque formation. This process is triggered by proinflammatory mediators (eg TNF-9), which induce the expression of inflammatory biomarkers, such as adhesion molecules, on the surface of the activated endothelial cells. Increased circulating levels of these adhesion molecules are considered as predictive of cardiovascular disease risk because they indicate a proinflammatory state in the vasculature. Oxidative signals play important roles in the pathogenesis of chronic inflammatory diseases by mediating expression of inflammatory genes. Under oxidative stresses, to defend against the deleterious consequences of ROS, cells upregulate multiple antioxidant enzymes to scavenge these toxic species. Such an adaptive response results from the activation of a redox-sensitive gene regulatory network mediated by nuclear factor E2-related factor 2 (Nrf2). Based on the function of Nrf2 target genes (eg. HO-1, NQO1, g-GCS), one can easily conclude that its activation may protect cells from various stresses imposed by toxic exposure. Because oxidative stress contributes to the pathogenesis of atherosclerosis by stimulating inflammatory gene expression, we have hypothesized that the enhanced expression of the Nrf2/ARE-regulated cytoprotective proteins might suppress redox-sensitive inflammatory gene expression so contributing to the atheroprotective and anti-inflammatory phenotype and protecting against oxidant-mediated injury in endothelial cells. To demonstrate this hypothesis we investigated the potential of GSH depletion (induced by the GSH synthesis inhibitor buthionine sulfoximine, BSO) to elicit an adaptive response in human umbilical vein endothelial cells (HUVECs), exposed or not to TNF-9, a well-established inducer of adhesion molecules and of leukocyte adhesion to the endothelium (Speciale et al., 2010). Herein we demonstrated that, in this experimental model, GSH depletion does not affect, by itself, physiological gene expression of adhesion molecules but is able to reduce their mRNAlevels which appeared overexpressed following TNF-9 challenge. Furthermore, GSH depletion induced Nrf2 nuclear translocation in HUVECs exposed or not to TNF-9 exposure, whereas TNF-9 alone is not able to activate this pathway. Activation of an adaptive response was also confirmed by ARE gene expression (HO-1 and NQO-1) that shows a similar trend. Moreover, we established the involvement of MAP kinases ERK1/2 in Nrf2 nuclear translocation activated by BSO, by using a specific ERK1/2 inhibitor PD98059. These findings suggest that BSO reduces endothelial sensitivity to TNF-9 induced activation through the activation of the Nrf2/ARE adaptive response pathway. In conclusion, since the inducible expression of several inflammatory genes plays a crucial role in the initiation and progression of atherosclerosis, the coordinate induction of endogenous cytoprotective proteins through activation of the Nrf2/ARE pathway might serve as a new therapeutic approach for the treatment of atherosclerosis as well as of other inflammatory diseases.