Chemical hypoxia in human pluripotent NT2 stem cell-derived neurons: Effect of hydroxamic acid and benzamide-based epigenetic drugs

  • Rushita A Bagchi Department of Medicine, University of Colorado Anschutz Medical Campus, Aurora, Colorado, USA.
  • Ashim K Bagchi Institute of Cardiovascular Sciences, St. Boniface Albrechtsen Research Centre, Winnipeg, Manitoba, Canada.
  • Ankita Salunke Department of Zoology, The Maharaja Sayajirao University of Baroda, Vadodara, Gujarat, India.
  • Dipak K Hens Sonamukhi College, The University of Burdwan, West Bengal, India.
  • Pragna H Parikh Department of Zoology, The Maharaja Sayajirao University of Baroda, Vadodara, Gujarat, India.
Keywords: pluripotent, neuron, hypoxia, histone deacetylase, gene expression


Hypoxia-induced oxidative stress contributes to neuronal damage leading to many neurodegenerative disorders. Hypoxia promotes many downstream effectors such as hypoxia-inducible factor-1α (HIF-1α) in order to restore respiratory homeostasis due to low oxygen availability and increased ROS. Use of histone deacetylase (HDAC) inhibitors may modulate hypoxia-induced neuronal cell damage.  In this study, we used two chemically diverse HDAC inhibitors to investigate their effect on hypoxia-exposed neuronal cells. Human pluripotent NT-2 stem cell-derived neuronal differentiated cells were exposed to CoCl2 pre-treatment for 6h to induce hypoxia, prior to supplementation of HDAC inhibitor (SAHA or MGCD0103). Treatment with HDAC inhibitor improved cell viability in hypoxia-induced neuronal cells. The increased HIF1α expression in hypoxia-induced neuronal cells was blunted by these HDAC inhibitors with a concomitant decrease in ROS production. CoCl2 treatment caused an increase in IL-1β, which was significantly inhibited by these HDAC inhibitors. Furthermore, apoptosis induced in these CoCl2 treated neuronal cells was mitigated by SAHA as well MGCD0103 suggesting that these HDAC inhibitors are capable of reducing cellular toxicity, inflammation and apoptosis, and thus, could be beneficial as therapeutic molecules for many neuropathological conditions.


Adam-Vizi V. Production of reactive oxygen species in brain mitochondria: contribution by electron transport chain and non-electron transport chain sources. Antioxid Redox Signal. 2005;7(9-10):1140-1149.

Alarifi S, Ali D, Omar Suliman Y Al, Ahamed M, Siddiqui MA, Al-Khedhairy AA. Oxidative stress contributes to cobalt oxide nanoparticles-induced cytotoxicity and DNA damage in human hepatocarcinoma cells. Int J Nanomedicine. 2013;8(10):189-199.

Butler KV, Kalin J, Brochier C, Vistoli G, Langley B, Kozikowski AP. Rational design and simple chemistry yield a superior, neuroprotective HDAC6 inhibitor, tubastatin A. J Am Chem Soc. 2010;132(31):10842-10846.

DeGracia DJ, Kumar R, Owen CR, Krause GS, White BC. Molecular pathways of protein synthesis inhibition during brain reperfusion: implications for neuronal survival or death. J Cereb Blood Flow Metab. 2002;22(2):127-141.

Fournel M, Bonfils C, Hou Y, Yan PT, Trachy-Bourget M-C, Kalita A, et al. MGCD0103, a novel isotype-selective histone deacetylase inhibitor, has broad spectrum antitumor activity in vitro and in vivo. Mol Cancer Ther. 2008;7(4):759-768.

Haile Y, Fu W, Shi B, Westaway D, Baker G, Jhamandas J, et al. Characterization of the NT2-derived neuronal and astrocytic cell lines as alternative in vitro models for primary human neurons and astrocytes. J Neurosci Res. 2014;92(9):1187-1198.

Hardy M, Younkin D, Tang CM, Pleasure J, Shi QY, Williams M, et al. Expression of non-NMDA glutamate receptor channel genes by clonal human neurons. J Neurochem. 1994;63(2):482-489.

Hull EE, Montgomery MR, Leyva KJ. HDAC Inhibitors as Epigenetic Regulators of the Immune System: Impacts on Cancer Therapy and Inflammatory Diseases. BioMed Research International. 2016;2016:8797206.

Langlois A, Duval D. Differentiation of the human NT2 cells into neurons and glia. Methods Cell Sci. 1997;19:213-219.

Lee PJ, Jiang BH, Chin BY, Iyer NV, Alam J, Semenza GL, et al. Hypoxia-inducible factor-1 mediates transcriptional activation of the heme oxygenase-1 gene in response to hypoxia. J Biol Chem. 1997;272(9):5375-5381.

Morrison BE, Majdzadeh N, Zhang X, Lyles A, Bassel-Duby R, Olson EN, et al. Neuroprotection by histone deacetylase-related protein. Molecular and Cellular Biology. 2006;26(9):3550-3564.

Movafagh S, Crook S, Vo K. Regulation of hypoxia-inducible factor-1a by reactive oxygen species: new developments in an old debate. J Cell Biochem. 2014;116(5):696-703.

Pistritto G, Papaleo V, Sanchez P, Ceci C, Barbaccia ML. Divergent modulation of neuronal differentiation by caspase-2 and -9. PLoS ONE. 2012;7(5):e36002.

Poss KD, Tonegawa S. Reduced stress defense in heme oxygenase 1-deficient cells. Proc Natl Acad Sci USA. 1997;94(20):10925-10930.

Rangwala S, Zhang C, Duvic M. HDAC inhibitors for the treatment of cutaneous T-cell lymphomas. Future Med Chem. 2012;4(4):471-486.

Sagulenko V, Vitak N, Vajjhala PR, Vince JE, Stacey KJ. Caspase-1 Is an Apical Caspase Leading to Caspase-3 Cleavage in the AIM2 Inflammasome Response, Independent of Caspase-8. J Mol Biol. 2017;430(2):238-247.

Samanta D, Prabhakar NR, Semenza GL. Systems biology of oxygen homeostasis. Wiley Interdiscip Rev Syst Biol Med. 2017;9(4).

Schweizer S, Meisel A, Märschenz S. Epigenetic mechanisms in cerebral ischemia. J Cereb Blood Flow Metab. 2013;33(9):1335-1346.

Sebastián VP, Salazar GA, Coronado-Arrázola I, Schultz BM, Vallejos OP, Berkowitz L, et al. Heme Oxygenase-1 as a Modulator of Intestinal Inflammation Development and Progression. Front Immunol. 2018;9:1956.

Semenza GL. Hypoxia-inducible factor 1 and cardiovascular disease. Annu Rev Physiol. 2013;76:39-56.

Semenza GL. Transcriptional regulation by hypoxia-inducible factor 1 molecular mechanisms of oxygen homeostasis. Trends Cardiovasc Med. 1996;6(5):151-157.

Shimazu T, Hirschey MD, Newman J, He W, Shirakawa K, Le Moan N, et al. Suppression of oxidative stress by β-hydroxybutyrate, an endogenous histone deacetylase inhibitor. Science. 2012;339(6116):211-214.

Turrens JF. Mitochondrial formation of reactive oxygen species. J Physiol (Lond). 2003;552(Pt 2):335-344.

Unal-Cevik I, Kilinç M, Can A, Gürsoy-Ozdemir Y, Dalkara T. Apoptotic and necrotic death mechanisms are concomitantly activated in the same cell after cerebral ischemia. Stroke. 2004;35(9):2189-2194.

Uttara B, Singh AV, Zamboni P, Mahajan RT. Oxidative stress and neurodegenerative diseases: a review of upstream and downstream antioxidant therapeutic options. Curr Neuropharmacol. 2009;7(1):65-74.

Wang GL, Jiang BH, Rue EA, Semenza GL. Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension. Proc Natl Acad Sci USA. 1995;92(12):5510-5514.

Younkin DP, Tang CM, Hardy M, Reddy UR, Shi QY, Pleasure SJ, et al. Inducible expression of neuronal glutamate receptor channels in the NT2 human cell line. Proc Natl Acad Sci USA. 1993;90(6):2174-2178.

Zhang X, Yuan Z, Zhang Y, Yong S, Salas-Burgos A, Koomen J, et al. HDAC6 modulates cell motility by altering the acetylation level of cortactin. Neuroimage. 2007;27(2):197-213.

Zhao R, Feng J, He G. Hypoxia increases Nrf2-induced HO-1 expression via the PI3K/Akt pathway. Front Biosci (Landmark Ed). 2016;21:385-396.

Ziemka-Nalecz M, Zalewska T. Neuroprotective effects of histone deacetylase inhibitors in brain ischemia. Acta Neurobiol Exp (Wars). 2015;74(4):383-395.

How to Cite
Bagchi, R. A., Bagchi, A. K., Salunke, A., Hens, D. K. and Parikh, P. H. (2019) “Chemical hypoxia in human pluripotent NT2 stem cell-derived neurons: Effect of hydroxamic acid and benzamide-based epigenetic drugs”, Neuroscience Research Notes, 2(3), pp. 12-19. doi: 10.31117/neuroscirn.v2i3.30.
Research Notes