Basic Science and Pathogenesis

Background: In Alzheimer's disease (AD), histone acetylation is disrupted, suggesting loss of transcriptional control. Moreover, converging evidence suggests an age- and AD-dependent loss of transcription controlled by all-trans-retinoic acid (ATRA), the bioactive metabolite of vitamin A (VA). Antioxidant depletion causes oxidative stress (OS). Nrf2-mediated antioxidant defenses are triggered by OS. Here, we investigated roles of VA, histone acetylation, Nrf2, and OS in vitro. Finally, we established a dietary dose of vorinostat that crosses the blood brain barrier and promotes histone acetylation in AD mouse brain.

Method: For in vitro studies, mouse HT22 cells were treated with vorinostat (up to 40 µM), ATRA, and/or H₂O₂. MTT and lipid peroxidation assays were performed. Acetylated histone H3 and Nrf2 expression were examined via western blot (WB) and immunocytochemistry (ICC). For in vivo studies, the hAb-KI AD mice were treated with 0.18 or 0.36 mg vorinostat/gram of diet (125 or 250 mg/kg/week for 2 weeks). HDAC activity in brain tissue was examined via histone H3 acetylation and enzyme activity via WB and colorimetric ELISA.

Result: Vorinostat and ATRA treatment (up to 20 μM) had no significant cytotoxic effects on HT22 cells. H₂O₂ alone (25-50 µM) caused ∼30-40% cell death (p<0.0001). ATRA (5 µM), in combination with vorinostat (0.5 µM), protected against H₂O₂ up to 150 µM. Vorinostat caused gradual and significant (p<0.001) induction in acetylation of histone H3 with 0.5-3.0 µM treatment for 24h, with no change in total histone H3 levels. ROS was significantly reduced. Nuclear translocation of Nrf2 was induced by H₂0₂ and reduced after ATRA and vorinostat treatment. For 0.18 or 0.36 mg vorinostat/gram doses in vivo, histone H3 acetylation was increased, HDAC activity was inhibited (p<0.001 and 0.05, respectively), and peroxidation was reduced.

Conclusion: Doses of vorinostat used in vitro and in vivo increased histone acetylation without cytotoxicity/toxicity. In vivo, we established that 125 mg/kg/week in diet is an optimal tolerable dose. VA, alone and in combination with vorinostat, may exhibit synergistic effects in protecting neuronal cells from oxidative stress. Together, our study provides a possible link between oxidative stress, Nrf2 and HDACs as potential contributors to Alzheimer's disease progression.