3.4 Involvement of the NRF2 pathway in the suppressive effects
of β-damascone on DCs
Some substances derived from plants, especially phytochemicals (e.g.,
sulforaphane), often activate the NRF2 pathway by directly acting on
Keap1, resulting in the induction of antioxidant and anti-inflammatory
responses 11. To clarify whether β-damascone activates
the NRF2 pathway in DCs, we determined the NRF2 protein and Hmox1mRNA levels in DB-treated DCs. As shown in Figure 4A , a Western
blot analysis revealed that the amount of NRF2 protein was increased in
DCs in the presence of β-damascone and peaked at 1 h after the addition
of DB to the culture medium. The mRNA levels of Hmox1 , which is a
target gene of NRF2, were markedly upregulated by β-damascone much
higher than those induced by LPS (Figure 4B ). These results
showing the increase in NRF2 protein and Hmox1 mRNA levels in
β-damascone-treated DCs suggest the possibility that β-damascone
activates the NRF2 pathway in DCs. Then, we investigated the roles of
NRF2 in DB-mediated modification of DC function by usingNrf2-/- DCs. First, it was confirmed that the
β-damascone-induced increase in Hmox1 mRNA levels in DCs was
almost abolished by NRF2 deficiency (Figure 4C ). When
OVA-pulsed Nrf2-/- BMDCs were cocultured with
OT-II CD4+ T cells under a Th1-polalizing conditions,
the suppression of Th1 development by β-damascone was not observed,
whereas control (Nrf2+/- ) DC-dependent Th1
development was significantly suppressed in the presence of β-damascone
(Figure 4D ), as was the case for WT BMDCs (Figure 1E ).
Furthermore, IL-12p40 release from LPS-stimulated DCs tended to increase
by Nrf2 deficiency in DCs, and the suppressive effect of
β-damascone on IL-12p40 production was markedly reduced inNrf2-/- BMDCs compared with that inNrf2+/- BMDCs (Figure 4E ). These
results demonstrate that DC functions, including Th1 induction and IL-12
production, were suppressed by β-damascone depending on activation of
the NRF2 pathway in DCs.