Abstract
Docosahexaenoic acid (DHA), an ω-3 fatty acid abundant in fish oils, has diverse health beneficial effects, such as anti-oxidative, anti-inflammatory, neuroprotective, and chemopreventive activities. In this study, we found that DHA induced expression of two representative antioxidant/cytoprotective enzymes, heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase (NQO1), in human mammary epithealial (MCF-10A) cells. DHA-induced upregulation of these enzymes was accompanied by enhanced translocation of the redox-sensitive transcription factor Nrf2 into the nucleus and its binding to antioxidant response element. Nrf2 gene silencing by siRNA abolished the DHA-induced expression of HO-1 and NQO1 proteins. When MCF-10A cells were transfected with mutant constructs in which the cysteine 151 or 288 residue of Keap1 was replaced by serine, DHA-induced expression of HO-1 and NQO1 was markedly reduced. Moreover, DHA activated protein kinase C (PKC)δ and induced Nrf2 phosphorylation. DHA-induced phosphorylation of Nrf2 was abrogated by the pharmacological PKCδ inhibitor rottlerin or siRNA knockdown of its gene expression. The antioxidants N-acetyl-l-cysteine and Trolox attenuated DHA-induced activation of PKCδ, phosphorylation of Nrf2, and and its target protein expression. In conclusion, DHA activates Nrf2, possibly through modification of critical Keap1 cysteine 288 residue and PKCδ-mediated phosphorylation of Nrf2, leading to upregulation of HO-1 and NQO1 expression.
Highlights
The two representative omega-3 polyunsaturated fatty acids (n-3 PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), abundant in some fish oils, possess anti-oxidative [1,2], anti-inflammatory [3], neuroprotective [4], and chemopreventive [5] effects
In comparing the effects of EPA and DHA on the expression of heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase-1 (NQO1), we first verified that both n-3 PUFAs elicited no substantial cytotoxicity at a concentration up to 100 μM (Figure 1A)
When MCF-10A cells were incubated with 25 μM DHA, there was a progressive increase in the protein expression of HO-1 and NQO1
Summary
The two representative omega-3 polyunsaturated fatty acids (n-3 PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), abundant in some fish oils, possess anti-oxidative [1,2], anti-inflammatory [3], neuroprotective [4], and chemopreventive [5] effects. A double-blind randomized study with healthy volunteers showed that dietary supplementation with n-3 PUFAs protected against UV-induced DNA damage in peripheral blood lymphocytes [2]. PUFA supplementation during cancer chemotherapy improves patient outcomes [12]. Supplementation with n-3 PUFA increased therapeutic response in patients with advanced non-small cell lung cancer [14]. N-3 PUFAs are very susceptible to free radical attack or oxidation [15], paradoxically their protective effects against oxidative stress and injury have been reported [2,3]. In a mouse model of cigarette smoke (CS)-induced endothelial dysfunction, n-3 PUFA-enriched diet decreased CS-induced production of an oxidative stress marker, 8-isoprostane and significantly reduced the mRNA level of cytochrome P4501A1 [18]. N-3 PUFA-enriched diet protected mice from Concanavalin
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