The role of zinc in modulating oxidative stress has recently been recognized. Oxidative stress is an important contributing factor in several chronic human diseases, such as atherosclerosis and related vascular diseases, mutagenesis and cancer, neurodegeneration, immunologic disorders, and the aging process (20–22). Together O2·−, H2O2, and ·OH are known as reactive oxygen species (ROS), and these are produced continuously in vivo under aerobic conditions. The NADPH oxidases are a group of plasma membrane associated enzymes, which catalyze the production of O2·− from oxygen by using NADPH as the electron donor. Zinc is an inhibitor of this enzyme. The dismutation of O2·− to H2O2 is catalyzed by an enzyme super oxide dismutase (SOD), which contains both copper and zinc. Zinc is known to induce the production of metallothionein, which is very rich in cysteine, and is an excellent scavenger of ·OH (6). Iron and copper ions catalyze the production of ·OH from H2O2. Zinc is known to compete with both iron and copper for binding to cell membrane, thus decreasing the production of ·OH (6).
A few investigators have reported that inflammatory cytokines such as TNF-α (tumor necrosis factor-α) and IL-1β, generated by activated monocytes-macrophages, also are known to produce increased amounts of ROS (23,24). Increases in these cytokines are associated with decreased zinc status in patients.
NF-κB is involved in the expression of a variety of responsive specific genes and is activated by several stimuli such as cytokines, radiation, and oxidative stress. In vitro activation of NF-κB by TNF-α in MNC has been shown to be an excellent model of oxidative stresssensitive transactivating factor, and has been used to evaluate the efficacy of compounds in protecting cells from oxidative stress (21). Zinc has been shown to inhibit NF-κB activation in prostate cancer cells, thus enhancing anti-cancer therapy (25), bovine cerebral epithelial cells (26), as well as reducing increased levels of activated NF-κB in diabetic CD1 mice (27) and zinc deficient cultured human hepatocellular carcinoma-derived cell line (28).
The induction of NF-κB activation pathway appears to be cell specific and is counterbalanced by concomitant activation of NF-κB activation inhibitors. One such inhibitor of NF-κB activation is A20, a zinc finger-transactivating factor which also binds to DNA, producing the A20 protein which inhibits TNF-α-induced NF-κB activation (29–33). A20 plays an important role in reducing IL-1β- and TNF-α-induced NF-κB activation (29–33).
Our data showed that zinc supplementation to normal healthy subjects a) lowers the oxidative stress-related byproducts MDA (malondialdehyde), 4-hydroxyalkenals (HAE), and 8-hydroxy deoxyguanine (8-OHdG) generated by cells and released into the plasma, b) inhibits the induction of TNF-α and IL-1β mRNA in MNCs, and c) exhibits a protective effect against TNF-α-induced NF-κB activation in isolated MNCs (34). In addition, we provided evidence to show that, in the human pro-myelocytic leukemia cell line HL-60 which differentiates to the monocyte-macrophage phenotype by PMA, zinc increased the expression of A20 and the binding of A20 transactivating factor to DNA, thereby enhancing inhibition of induced NF-κB activation (34).
The role of zinc in regulation of the gene expression of IL-1β and TNF-α has not been defined. The zinc finger protein A20 has been shown to inhibit NF-κB signaling by TNF-α and IL-1β via TNF-receptor gene (TRAF pathways) in endothelial cells (31,35). A20 is expressed in various types of cells in response to a number of stimuli such as TNF-α, IL-1β, LPS (lipopolysaccharide), PMA, Epstein-Barr virus latent membrane protein, as well as other stimuli (32). A20 expression primarily protects cells from TNF-α-induced cytotoxicity by decreasing the activation of NF-κB, which leads to decreased IL-1β and TNF-α gene expression as has been demonstrated in endothelial cells. We propose that a similar effect of zinc supplementation on the A20 pathway occurs in primary cells. Further studies are ongoing in our laboratory to aid in the understanding the mechanisms of zinc action.
Our study provides molecular evidence for an anti-oxidant effect of zinc in human subjects and shows that zinc supplementation in vivo protected MNC against oxidative stress (34). Although there are several possible biochemical mechanisms by which zinc may decrease oxidative stress in cells, our study shows that zinc negatively regulates gene expression of inflammatory cytokines such as TNF-α and IL-1β, which are known to generate ROS and this may be one additional mechanism by which zinc may be functioning as an antioxidant in humans. Thus, our study provides rationale for use of zinc in therapeutic trials either alone or in conjunction with other modalities in chronic diseases, including chemo-prevention of cancer in which oxidative stress is known to play an important role.
The immunological hallmarks of zinc deficiency in humans and higher animals include thymic atrophy, lymphopenia, and compromised cell- and antibody-mediated responses that result in increased incidences of infections (36). In chronic zinc deficiency, a reprogramming of the immune system occurs, beginning with the activation of the stress axis and chronic production of glucocorticoids that accelerate apoptosis of pre-B and pre-T cells (36), resulting in reduced lymphopoeisis and atrophy of thymus. In contrast, myelopoesis is preserved. Changes in the gene expression for cytokines, DNA repair enzymes, zinc transporters, and signaling molecules, suggest that the cells of the immune system are attempting to adapt to the stress of suboptimal zinc (36).
We have observed that, as a result of zinc deficiency, the macrophages-monocytes are stressed and they generate inflammatory cytokines such as TNF-α and IL-1β (14,34). In a recent study, we reported that, in comparison to the younger adults, the elderly subjects had lower plasma zinc, increased oxidative markers, and increased generation of inflammatory cytokines (14). Following zinc supplementation to the elderly subjects, the plasma zinc increased, oxidative stress markers decreased, and generation of inflammatory cytokine decreased, in comparison to the placebo group.
In a large study organized by the National Eye Institute, NIH, (Bethesda, MD, USA) it was reported that zinc and antioxidants (vitamin C, vitamin E, and β carotene) significantly reduced the odds of developing advanced age related macular degeneration (AMD), and prevented blindness in the high-risk group of elderly subjects (37). Also, it was reported that longevity was increased in the zinc supplemented group (38). Although the mechanism of zinc effect was not defined, one may hypothesize that zinc reduced the oxidative stress and thus was beneficial in AMD.