Aldehyde stress and up-regulation of Nrf2-mediated antioxidant systems accompany functional adaptations in cardiac mitochondria from mice fed n-3 polyunsaturated fatty acids

Abstract

Diets replete with n-3 PUFAs (polyunsaturated fatty acids) are known to have therapeutic potential for the heart, although a specifically defined duration of the n-3 PUFA diet required to achieve these effects remains unknown, as does their mechanism of action. The present study was undertaken to establish whether adaptations in mitochondrial function and stress tolerance in the heart is evident following short- (3 weeks) and long- (14 weeks) term dietary intervention of n-3 PUFAs, and to identify novel mechanisms by which these adaptations occur. Mitochondrial respiration [mO2 (mitochondrial O2)], H2O2 emission [mH2O2 (mitochondrial H2O2)] and Ca2+-retention capacity [mCa2+ (mitochondrial Ca2+)] were assessed in mouse hearts following dietary intervention. Mice fed n-3 PUFAs for 14 weeks showed significantly lower mH2O2 and greater mCa2+ compared with all other groups. However, no significant differences were observed after 3 weeks of the n-3 PUFA diet, or in mice fed on an HFC (high-fat control) diet enriched with vegetable shortening, containing almost no n-3 PUFAs, for 14 weeks. Interestingly, expression and activity of key enzymes involved in antioxidant and phase II detoxification pathways, all mediated by Nrf2 (nuclear factor E2-related factor 2), were elevated in hearts from mice fed the n-3 PUFA diet, but not hearts from mice fed the HFC diet, even at 3 weeks. This increase in antioxidant systems in hearts from mice fed the n-3 PUFA diet was paralleled by increased levels of 4-hydroxyhexenal protein adducts, an aldehyde formed from peroxidation of n-3 PUFAs. The findings of the present study demonstrate distinct time-dependent effects of n-3 PUFAs on mitochondrial function and antioxidant response systems in the heart. In addition, they are the first to provide direct evidence that non-enzymatic oxidation products of n-3 PUFAs may be driving mitochondrial and redox-mediated adaptations, thereby revealing a novel mechanism for n-3 PUFA action in the heart.

Figure 1

Dietary n-3 PUFA intake causes time-dependent adaptations in mitochondrial function in mouse heart, resulting in decreased mH₂O₂ emission (A) and increased tolerance to mCa²⁺ overload (B) by 14 weeks. Shown are mean ± S.E.M. in N = 7 for 3 week diet and N = 5 for 14 week diet; * P<0.05 vs. Std Chow

Figure 2

Time-dependent effects of dietary n-3 PUFAs on levels of total glutathione GSH_t (A), oxidized glutathione GSSG (B) and GSH/GSSG ratio (C) in mouse hearts. Data are mean ± S.E.M. in N = 7 for 3 week diet and N = 6 for 14 week diet; * P<0.05 vs. Std Chow

Figure 3

Rates of glutathione peroxidase (A) and glutathione reductase (B) activity in mouse heart after dietary n-3 PUFA treatment for 3 and 14 weeks. Data are mean ± S.E.M. in N = 7 for 3 week diet and N = 6 for 14 week diet; * P<0.05 vs. Std Chow

Figure 4

Immunoblots of protein adducts with 4-hydroxyhexenal, a n-3 PUFA-derived aldehyde, in mouse heart following n-3 PUFA diet are shown in representative blots (A) and quantified by densitometry (B). Shown in (C) are quantified levels of protein adducts with 4-hydroxynonenal, a n-6 PUFA-derived aldehyde. Data are mean ± S.E.M. in N = 6 for 3 week diet and N = 4 for 14 week diet; * P<0.05 vs. Std Chow