The antioxidant N-acetyl cysteine suppresses lidocaine-induced intracellular reactive oxygen species production and cell death in neuronal SH-SY5Y cells

Abstract

BackgroundThe local anesthetic lidocaine can affect intra- and extra-cellular signaling pathways in both neuronal and non-neuronal cells, resulting in long-term modulation of biological functions, including cell growth and death. Indeed, lidocaine was shown to induce necrosis and apoptosis in vitro. While several studies have suggested that lidocaine-induced apoptosis is mitochondrial pathway-dependent, it remains unclear whether reactive oxygen species (ROS) are involved in this process and whether the observed cell death can be prevented by antioxidant treatment.MethodsThe effects of lidocaine and antioxidants on cell viability and death were evaluated using SH-SY5Y cells, HeLa cells, and HeLa cell derivatives. Cell viability was examined via MTS/PES ([3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt]/phenazine ethosulfate) assay. Meanwhile, cell apoptosis and necrosis were evaluated using a cell death detection assay with Annexin V-FITC and PI staining, as well as by assaying for caspase-3/7 and caspase-9 activity, and by measuring the release of lactate dehydrogenase, respectively. Mitochondrial transmembrane potential (ΔΨm) was assessed using the fluorescent probe tetramethylrhodamine ethyl ester.ResultsLidocaine treatment resulted in suppression of the mitochondrial electron transport chain and subsequent attenuation of mitochondrial membrane potential, as well as enhanced ROS production, activation of caspase-3/7 and caspase-9, and induction of apoptosis and necrosis in SH-SY5Y cells in a dose- and time-dependent manner. Likewise, the anesthetics mepivacaine and bupivacaine also induced apoptosis in SH-SY5Y cells. Notably, the antioxidants N-acetyl cysteine (NAC) and Trolox successfully scavenged the mitochondria-derived ROS and suppressed local lidocaine-induced cell death.ConclusionsOur findings demonstrate that the local anesthetics lidocaine, mepivacaine, and bupivacaine inhibited the activity of mitochondria and induced apoptosis and necrosis in a dose-dependent manner. Furthermore, they demonstrate that treatment with the antioxidants NAC, Trolox, and GGA resulted in preservation of mitochondrial voltage and inhibition of apoptosis via suppression of caspase activation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12871-016-0273-3) contains supplementary material, which is available to authorized users.

HIF-1-mediated suppression of mitochondria electron transport chain function confers resistance to lidocaine-induced cell death

“…We recently demonstrated that lidocaine treatment induced ROS generation 4 . As such, these data imply that lidocaine-mediated cell death is dependent on mitochondrial function.…”

“…Human neuroblastoma SH-SY5Y cells were maintained in RPMI 1640 medium that contained 10% FBS, 100 U/ml penicillin, and 0.1 mg/ml streptomycin 4 . n-propyl gallate (nPG), desferoxamine (DFX), and the anti-β-actin antibody (Ab) were obtained from Sigma (St. Louis, MO, USA).…”

Propofol induces a metabolic switch to glycolysis and cell death in a mitochondrial electron transport chain-dependent manner

“…Cell growth was assessed using a CellTiter 96 AQueous One Solution Cell Proliferation Assay™ with MTS (Promega, Madison, WI, USA) 47,49 . Briefly, cells were seeded into 96-well plates (2 × 10 4 cells/well) and cultured overnight.…”

“…Activity levels of caspase-3/7 and caspase-9 were assessed using an Apo-ONE™ homogeneous caspase-3/7 assay kit (Promega) and a Caspase-Glo™ 9 assay kit (Promega), respectively, according to the manufacturer's protocols 47,49 . Briefly, cells were seeded into 96-well plates (2 × 10 4 cells/well) and incubated overnight.…”

“…Cytotoxicity was measured using a CytoTox-ONE™ kit (Promega) as described previously 47,49 . Briefly, cells were cultured overnight in 96-well plates (2 × 10 4 cells/well) and treated with the indicated drug(s) for varying lengths of time.…”