Dietary intake of zinc and omega-3 fatty acids (DHA) have health benefits for a number of human diseases. However, the molecular basis of these health benefits remains unclear. Recently, we reported that zinc and DHA affect expression levels of histones H3 and H4 in human neuronal M17 cells. Here, using immunoblotting and densitometric analysis, we aimed to investigate the effect of zinc and DHA on post-translational modifications of histone H3 in M17 cells. In response to increase in zinc concentration, we observed increase in deacetylation, methylation and phosphorylation of H3 and decrease in acetylation. We also investigated the role of zinc in apoptosis, and found that zinc reduced the levels of the anti-apoptotic marker Bcl-2 while increasing the apoptotic marker caspase-3 levels, correlating with cell viability assays. Conversely, DHA treatment resulted in increase in acetylation of H3 and Bcl-2 levels and decrease in deacetylation, methylation, phosphorylation of H3 and caspase-3 levels, suggesting that DHA promotes gene expression and neuroprotection. Our novel findings show the opposing effects of zinc and DHA on the epigenetic regulation of human neuronal cells and highlight the potential benefit of dietary intake of DHA for management of neurodegenerative diseases.

Edited by Jesus AvilaKeywords: DHA Omega-3 fatty acid Zinc ZnT3 M17 human neuronal cell Apoptosis a b s t r a c t Docosahexaenoic acid (DHA) is the major polyunsaturated fatty acid in neuronal cell membranes. We hypothesize that DHA induces a decrease in neuronal cell death through reduced ZnT3 expression and zinc uptake. Exposure of M17 cells to DHA-deficient medium increased the levels of active caspase-3, relative to levels in DHA-replete cells, confirming the adverse effects of DHA deficiency in promoting neuronal cell death. In DHA-treated M17 cells, zinc uptake was 65% less and ZnT3 mRNA and protein levels were reduced in comparison with DHA-depleted cells. We propose that the neuroprotective function of DHA is exerted through a reduction in cellular zinc levels that in turn inhibits apoptosis.

Zn and DHA have putative neuroprotective effects and these two essential nutrients are known to interact biochemically. We aimed to identify novel protein candidates that are differentially expressed in human neuronal cell line M17 in response to Zn and DHA that would explain the molecular basis of this interaction. Two-dimensional gel electrophoresis and MS were applied to identify major protein expression changes in the protein lysates of human Ml7 neuronal cells that had been grown in the presence and absence of Zn and DHA. Proteomic findings were further investigated using Western immunoblot and real-time PCR analyses. Four protein spots, which had significant differential expression, were identified and selected for in-gel trypsin digestion followed by matrix-assisted laser desorption ionisation MS analysis. The resultant peptide mass fingerprint for each spot allowed their respective identities to be deduced. Two human histone variants H3 and H4 were identified. Both H3 and H4 were downregulated by Zn in the absence of DHA (Zn effect) and upregulated by DHA (DHA effect) in the presence of Zn (physiological condition). These proteomic findings were further supported by Western immunoblot and real-time PCR analyses using H3-and H4-specific monoclonal antibodies and oligonucleotide primers, respectively. We propose that dietary Zn and DHA cause a global effect on gene expression, which is mediated by histones. Such novel information provides possible clues to the molecular basis of neuroprotection by Zn and DHA that may contribute to the future treatment, prevention and management of neurodegenerative diseases such as Alzheimer's disease.

Omega-3 (ω-3) fatty acids are one of the two main families of long chain polyunsaturated fatty acids (PUFA). The main omega-3 fatty acids in the mammalian body are α-linolenic acid (ALA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). Central nervous tissues of vertebrates are characterized by a high concentration of omega-3 fatty acids. Moreover, in the human brain, DHA is considered as the main structural omega-3 fatty acid, which comprises about 40% of the PUFAs in total. DHA deficiency may be the cause of many disorders such as depression, inability to concentrate, excessive mood swings, anxiety, cardiovascular disease, type 2 diabetes, dry skin and so on. On the other hand, zinc is the most abundant trace metal in the human brain. There are many scientific studies linking zinc, especially excess amounts of free zinc, to cellular death. Neurodegenerative diseases, such as Alzheimer’s disease, are characterized by altered zinc metabolism. Both animal model studies and human cell culture studies have shown a possible link between omega-3 fatty acids, zinc transporter levels and free zinc availability at cellular levels. Many other studies have also suggested a possible omega-3 and zinc effect on neurodegeneration and cellular death. Therefore, in this review, we will examine the effect of omega-3 fatty acids on zinc transporters and the importance of free zinc for human neuronal cells. Moreover, we will evaluate the collective understanding of mechanism(s) for the interaction of these elements in neuronal research and their significance for the diagnosis and treatment of neurodegeneration.