Vorinostat, a Pan Histone Deacetylase Inhibitor, Stimulates Neuritic Outgrowth in PC12 Cells via Activation of MAPK and PI3K Signal Transduction Pathways

Degeneration of neurons is a major cause of neurodegenerative diseases. Neurotrophic actions and neuroprotection leading to neuroplasticity and neurite outgrowth are important markers for neuroregenration. Neuritogenesis is a fundamental process in the differentiation of neurons and formation of synapses. Highly regulated signal transduction pathways control these processes. Recent studies have shown neuroprotective effects of histone deacetylase (HDAC) inhibitors. The aim of this study was to investigate whether Vorinostat, (suberoylanilide hydroxamic acid-SAHA) the first pan HDAC inhibitor approved for clinical use, induces neuronal differentiation in PC12 cells. NeuroScreen1, a clone of PC12 (rat adrenal pheochromocytoma) cell line is used. NS-1 cells were treated with Vorinostat for 72 hrs. Cytotoxicity was measured and cell cultures were analyzed for neuritic outgrowth by digital microscope imaging and NIS element software for measurement of neurite numbers and length. Vorinostat produced significant neuronal differentiation, similar to NGF in PC12 cells. However, it did not produce further induction of NGF-stimulated neuritic outgrowth in PC12 cells. Further, to investigate the mechanism for Vorionstat-induced differentiation of PC12 cells the MEK1/2 inhibitors (PD98059 and U0126) and PI3K inhibitor (LY294002) were also tested. The cell cultures treated with Vorinostat were analyzed for acetylation patterns of proteins by Western blots probed with antibodies for acetylated-lysine. The neurotrophic actions of both NGF and Vorinostat were almost completely abolished by co-treatment of the PC12 cell cultures with the inhibitors of MEK1/2 & PI3K. Vorinostat produced selective time-dependent hyperacetylation of histones as well as non-histone proteins in PC12 cells. In conclusion, Vorinostat exerts neurotropic action via activation of MEK1/2 & PI3K pathways similar to that by NGF.

Fig. 1: In vitro neurotrophic activity of Vorinostat (SAHA) and NGF on PC 12 cells

Acknowledgements: Financial support from the USDA Agricultural Research Service Specific Cooperative Agreement No. 58 – 6408 – 02 – 1-612.