Fig. 2

Figure 2. Relationship between equol and the Keap1-Nrf2 pathway. (A) Antioxidant effects of equol in Nrf2-mutant larvae. Wild-type larvae (WT) or Nrf2-homozygous mutant larvae (nfe2l2a^fh318) at 3.5 dpf were pretreated with equol at concentrations of 0 µM (gray, dotted), 12.5 µM (orange), 25 µM (red), or 50 µM (pink). After pretreatment for 12 h, the solution was replaced with 1.9 mM (WT) or 1.4 mM (nfe2l2a^fh318) sodium arsenite (NaAsO₂), and survival was measured every 12 h for 48 h. P values of <0.01 were considered to indicate statistical significance. (B) Expression of estrogen receptor-target genes in equol-treated larvae. Larvae at 3.5 dpf were treated with or without 25 µM equol (EQ) for 12 h, and the expression of vtg1, foxc1a, and esr1 was analyzed using qRT-PCR. The expression of untreated larvae (Ctr) was normalized to 1. Each experiment was conducted at least three times with duplicate samples. (C) Equol-induced expression of gstp1.2. Larvae at 3.5 dpf were treated with or without 25 µM equol for 12 h, and the expression of gstp1.2 was analyzed using qRT-PCR. (D) Venn diagrams showing upregulated (left) and downregulated genes (right) in 4-dpf larvae treated with 25 µM equol for 12 h identified using RNA-seq analysis. (E) List of Nrf2-target genes induced by the equol treatment in RNA-seq analysis. It should be noted that the induction of these genes was not canceled by the nfe2l2a^fh318 mutation.