Figure 2. The Nrf2 promoter is epigenetically repressed in cortical neurons.

(a) ChIP analysis of acetylated histone H3 (Ac-H3) occupancy at the Nrf2 (left) and β-actin (right) promoters in neurons and astrocytes, normalized to input and expressed relative to each other. *P<0.05, NS=no significant difference (n=5). (b) Effect of TSA treatment (8 h) on Ac-H3 levels at the Nrf2 and β-actin promoters in neurons. *P<0.05 (n=5). (c,d) Effect of TSA treatment on Nrf2 expression in mouse cortical neurons (c) and human H9 ESC-derived neurons (d), normalized to Rpl13a. *P<0.05 (n=3). (e) Adult mice were subjected to intra-peritoneal injection of TSA (10 mg kg⁻¹) or vehicle (Veh.) and at 8 h culled and cortical neurons obtained by enzymic dissociation of the cortex followed by FAC-sorting of NeuN⁺ cells. RNA was extracted immediately and Nrf2 expression studied. *P<0.05, (n=7 (Veh.), 4 (TSA)). (f) Pre-treating cultured cortical neurons with TSA renders the neuronal Nrf2 pathway amenable to activation by tBHQ. Neurons were treated as indicated in the schematic, and expression of Nrf2 and Nrf2 target gene Srxn1 analysed by qRT–PCR. (P<0.05 analysis of variance plus Tukey's post-hoc test, *significant difference compared with control untreated cells; ^#significant effect of tBHQ+TSA compared with TSA alone condition). ‘NS' emphasizes non-significant effect of tBHQ versus untreated neurons (n=4 except control (no TSA, no tBHQ, n=5)).