A diet rich in olive oil phenolics reduces oxidative stress in the heart of SAMP8 mice by induction of Nrf2-dependent gene expression

Abstract

A Mediterranean diet rich in olive oil has been associated with health benefits in humans. It is unclear if and to what extent olive oil phenolics may mediate these health benefits. In this study, we fed senescence-accelerated mouse-prone 8 (SAMP8, n=11 per group) semisynthetic diets with 10% olive oil containing either high (HP) or low amounts of olive oil phenolics (LP) for 4.5 months. Mice consuming the HP diet had significantly lower concentrations of the oxidative damage markers thiobarbituric acid-reactive substances and protein carbonyls in the heart, whereas proteasomal activity was similar in both groups. Nrf2-dependent gene expression may be impaired during the aging process. Therefore, we measured Nrf2 and its target genes glutathione-S-transferase (GST), γ-glutamyl cysteine synthetase (γ-GCS), nicotinamide adenine dinucleotide phosphate [NAD(P)H]:quinone oxidoreductase (NQO1), and paraoxonase-2 (PON2) in the hearts of these mice. Nrf2 as well as GST, γ-GCS, NQO1, and PON2 mRNA levels were significantly higher in heart tissue of the HP as compared to the LP group. The HP-fed mice had significantly higher PON1 activity in serum compared to those receiving the LP diet. Furthermore, HP feeding increased relative SIRT1 mRNA levels. Additional mechanistic cell culture experiments were performed, and they suggest that the olive oil phenolic hydroxytyrosol present in the HP oil may be responsible for the induction of Nrf2-dependent gene expression and the increase in PON activity. In conclusion, a diet rich in olive oil phenolics may prevent oxidative stress in the heart of SAMP8 mice by modulating Nrf2-dependent gene expression.

FIG. 1.

Relative messenger RNA (mRNA) expression (normalized for glyceraldehyde 3-phosphate dehydrogenase [GAPDH]) of Nrf2 (A), glutathione-S-transferase (GST) (B), γ-glutamyl cysteine synthetase (γ-GCS) (C), nicotinamide adenine dinucleotide phosphate [NAD(P)H]:quinone oxidoreductase 1 (NQO1) (D), paraoxonase-2 (PON2) (E), and SIRT-1 (F) in hearts of senescence-accelerated mouse-prone 8 (SAMP8) mice fed for 4.5 months a Western type diet with 0.15% cholesterol and 20% fat, in which 10% of fat was from olive oil containing either low or high amount of phenolics. Mice were killed at 7 months of age. Values are expressed as mean±standard error of the mean (SEM) (n=10). * indicates statistical significant differences between groups.

FIG. 1.

Relative messenger RNA (mRNA) expression (normalized for glyceraldehyde 3-phosphate dehydrogenase [GAPDH]) of Nrf2 (A), glutathione-S-transferase (GST) (B), γ-glutamyl cysteine synthetase (γ-GCS) (C), nicotinamide adenine dinucleotide phosphate [NAD(P)H]:quinone oxidoreductase 1 (NQO1) (D), paraoxonase-2 (PON2) (E), and SIRT-1 (F) in hearts of senescence-accelerated mouse-prone 8 (SAMP8) mice fed for 4.5 months a Western type diet with 0.15% cholesterol and 20% fat, in which 10% of fat was from olive oil containing either low or high amount of phenolics. Mice were killed at 7 months of age. Values are expressed as mean±standard error of the mean (SEM) (n=10). * indicates statistical significant differences between groups.

FIG. 2.

Mean (±standard error of the mean [SEM]) paraoxonase-1 (PON1) activity in serum of senescence-accelerated mouse-prone 8 (SAMP8) mice fed a Western type diet with 0.15% cholesterol and 20% fat, in which 10% of fat was from olive oil containing either low or high amount of phenolics (n=9). *indicates statistical significant differences between groups.

FIG. 3.

Representative western blot (including densitometry) of Nrf2 in NIH 3T3 nuclear cell extracts in response to hydroxytyrosol following 6 hr of incubation with increasing concentrations of the test compound. Sulforaphane (SFN) was used as positive and TATA-binding protein (TBP) as a loading control. In densitometric analysis, the control equals one arbitrary unit.

FIG. 4.

Induction of PON1 transactivation by hydroxytyrosol in stably transfected HuH7 liver cells. Values are expressed as mean±standard error of the mean (SEM) of three independent experiments performed in triplicate. * indicates statistical significant differences between treatment and control (p<0.05).