NOX4 Inhibit the eNOS Protein folding With Aging

Abstract

Increased expression of Nox4 has seen in aged cells which is responsible for ROS generation. We have selected HUVECs because of predominant NOX4 expression, suitability for aging and vascular pathological studies. eNOS activity is considered as the marker for vascular aging and beta galactosidase is a subsequent classical marker for aging. We have selected HUVEC young (6th passage), middle (9th and 14thPassage) and aged (17th passage) for the analysis. Rise in Nox4 expression, ROS accumulation and ER stress have seen in aged passages, along with low NO bioavailability. ROS accumulation in aged passage caused mitochondrial dysfunction, alteration in morphology and energy crisis. For energy compensations, ER cause to increase uptake of glucose, rise in intra‐luminal NOX4 level and ROS accumulation. Redox imbalance cause ER stress, affect the chaperonic machinery and ultimate result in eNOS protein misfolding. So, accumulation of ROS interact eNOS both misfolding (in ER) and further rise in uncoupling in cytoplasm. For further confirmation of our hypothesis, we intended to treat the cells with GKT137831 (Nox4 inhibitor), TUDCA (ER stress inhibitor) and ascorbate. eNOS activity, beta galactosidase expression, ROS and ER stress was reduced with increase in passage number, whereas nitric oxide level was improved. Cellular ROS depletion, ER stress inhibition and PKC activity reduction and proper folding by ERO‐1‐PDI machinery, inhibited the effect of aging on eNOS activity, NO release and ER stress. We concluded that NOX4 is the main contributing enzyme for the aging of cell. We may delay the aging process by minimizing NOX4 level in a cell to have proper folding of eNOS by chaperonic activity of ERO‐1‐PDI machinery.