Vitamin C Suppresses TNFα-Induced NFκB Activation by Inhibiting IκBα Phosphorylation

Extracellular stimuli signal for activation of the transcription factor NFκB, leading to gene expression regulating processes involved in immune responses, inflammation, and cell survival. Tumor necrosis factor-α (TNFα) activates NFκB via a well-defined kinase pathways involving NFκB-inducing kinase (NIK), which activates downstream multisubunit IκB kinases (IKK). IKK in turn phosphorylates IκB, the central regulator of NFκB function. We found that intracellular vitamin C inhibits TNFα-induced activation of NFκB in human cell lines (HeLa, monocytic U937, myeloid leukemia HL-60, and breast MCF7) and primary endothelial cells (HUVEC) in a dose-dependent manner. Vitamin C is an important antioxidant, and most cells accumulate ascorbic acid (AA) intracellularly by transporting the oxidized form of the vitamin, dehydroascorbic acid (DHA). Because ascorbic acid is a strong pro-oxidant in the presence of transition metals in vitro, we loaded cells with vitamin C by incubating them with DHA. Vitamin C-loaded cells showed significantly decreased TNFα-induced nuclear translocation of NFκB, NFκB-dependent reporter transcription, and IκBα phosphorylation. Our data point to a mechanism of vitamin C suppression of NFκB activation by inhibiting TNFα-induced activation of NIK and IKKβ kinases independent of p38 MAP kinase. These results suggest that intracellular vitamin C can influence inflammatory, neoplastic, and apoptotic processes via inhibition of NFκB activation.