Bardoxolone methyl induces neuritogenesis in Neuro2a cells

Background: Bardoxolone methyl (RTA 402, CDDOMe) has been long known for its anti-inflammatory and exceptional cytotoxic activity. The biological responses to CDDOMe are truly dose dependent. And owing to the structural modifications introduced in its parent molecule oleanolic acid, CDDOMe is able to form reversible adducts with cellular proteins containing redox sensitive cysteine residues. This nature of CDDOMe makes it a multifunctional molecule targeting multiple signaling pathways. This study was initiated to study the response of Neuro2a, a mouse neuroblastoma cell line to CDDOMe. Methods: Neuro2a cells were treated with CDDOMe and all trans retinoic acid (ATRA) for 4 days and observed for neurite outgrowth. The neurite length was estimated using ImageJ software (Neuron growth plugin). Cell viability was investigated using MTT dye reduction and trypan blue dye exclusion method. Gene expression of differentiation markers was analyzed using quantitative PCR. Cellular localization of Tujl and synaptophysin in differentiated Neuro2a cells was observed using immunofluorescence. Results: CDDOMe ceased proliferation and induced dramatic neurite outgrowth in Neuro2a cells. These morphological changes were accompanied by time dependent increase in the mRNA levels of tyrosine hydroxylase, neurofilament 200 and synaptophysin. Besides, cytoskeleton protein Tujl and the synaptic vesicle protein synaptophysin were also observed to be localized in the neurites induced by CDDOMe. Conclusions: These early shreds of evidence suggest that CDDOMe induces differentiation in Neuro2a cells at concentrations ranging from 0.2 to 0.4 mu M and indeed contributes the existing knowledge on CDDOMe induced activities in cells. (C) 2018 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier B.V. All rights reserved.