Effect of Administration of an Equal Dose of Selected Dietary Chemicals on Nrf2 Nuclear Translocation in the Mouse Liver

Abstract

Certain dietary chemicals influenced the expression of chemopreventive genes through the Nrf2-Keap1 pathway. However, the difference in Nrf2 activation potency of these chemicals is not well studied. This study is aimed at determining the difference in the potency of liver Nrf2 nuclear translocation induced by the administration of equal doses of selected dietary chemicals in mice. Male ICR white mice were administered 50 mg/kg of sulforaphane, quercetin, curcumin, butylated hydroxyanisole, and indole-3-carbinol for 14 days. On day 15, the animals were sacrificed, and their livers were isolated. Liver nuclear extracts were prepared, and Nrf2 nuclear translocation was detected through Western blotting. To determine the implication of the Nrf2 nuclear translocation on the expression levels of several Nrf2-regulated genes, liver RNA was extracted for qPCR assay. Equal doses of sulforaphane, quercetin, curcumin, butylated hydroxyanisole, and indole-3-carbinol significantly induced the nuclear translocation of Nrf2 with different intensities and subsequently increased the expression of Nrf2-regulated genes with an almost similar pattern as the Nrf2 nuclear translocation intensities (sulforaphane > butylated hydroxyanisole = indole-3-carbinol > curcumin > quercetin). In conclusion, sulforaphane is the most potent dietary chemical that induces the Nrf2 translocation into the nuclear fraction in the mouse liver.

Figure 1

Administration of equal doses (50 mg/kg b.w.) of SUL, CUR, QRC, BHA, and I3C to mice for 14 days induces Nrf2 nuclear translocation in the liver with different potencies. (a) Nuclear fractions (100 μg) from each mouse were separated by electrophoresis, and the expressions of proteins were analyzed by Western blotting. Nrf2 is the protein of interest, while histone H3 is the housekeeping protein for the nuclear fraction. The bands for Nrf2 and histone H3 were densitometrically scanned, and the expression of nuclear Nrf2 (relative to histone) was determined as fold increase of the vehicle-treated (control) animals as shown in the bar chart above. (b) Cytoplasmic fractions (100 μg) from each mouse were separated by electrophoresis, and the expressions of proteins were analyzed by Western blotting. Nrf2 is the protein of interest, while β-actin is the housekeeping protein for the cytoplasmic fraction. The bands for Nrf2 and β-actin were densitometrically scanned, and the expression of Nrf2 in the nuclear fraction (relative to β-actin) was determined as fold increase of the vehicle-treated (control) animals as shown in the bar chart above. Values are expressed as the mean ± SEM (n = 4). Asterisk (∗) indicates a statistically significant difference from all control groups (vehicle 1 control and vehicle 2 control groups) after analysis using the Mann–Whitney test (P < 0.05). SUL: sulforaphane-treated group; CUR: curcumin-treated group; QRC: quercetin-treated group; BHA: butylated hydroxyanisole-treated group; I3C: indole 3 carbinol-treated group; VH1: control group treated with vehicle 1; VH2: control group treated with vehicle 2.

Figure 2

Administration of equal doses (50 mg/kg b.w.) of SUL, CUR, QRC, BHA, and I3C to mice for 14 days did not induce NF-κB nuclear translocation.

Figure 3

Effects of administration of 50 mg/kg SUL, CUR, QRC, BHA, and I3C for 14 days on (a) GCLC, (b) GPX1, and (c) CAT gene expressions in the livers of mice using RT-qPCR. GAPDH was designated as the reference gene. Data were represented as the mean ± SEM of six experiments, n = 6. Asterisk (∗) indicates a statistically significant difference compared to all control groups (vehicle 1 control and vehicle 2 control groups) after analysis using the Mann–Whitney test (P < 0.05). SUL: sulforaphane-treated group; CUR: curcumin-treated group; QRC: quercetin-treated group; BHA: butylated hydroxyanisole-treated group; I3C: indole 3 carbinol-treated group; VH1: control group treated with vehicle 1; VH2: control group treated with vehicle 2.