Acrylonitrile-Induced Neurotoxicity in Normal Human Astrocytes: Oxidative Stress and 8-Hydroxy-2′-Deoxyguanosine Formation

Abstract

Studies in our laboratory and others have indicated that acrylonitrile (VCN) induces acute and chronic neurotoxicity and brain tumors in animal models. Reduced glutathione (GSH) depletion was suggested as the initiator of oxidative stress in VCN-induced neurotoxicity. Astrocytes possess the majority of total brain GSH and express various immunological functions that are characteristic of the brain, including the secretion of cytokines. We hypothesized that astrocytes could be the primary target of VCN's adverse activities in the brain. Therefore, VCN-induced neurotoxicity was studied by exposing proliferating normal human astrocytes (NHAs) to various concentrations of VCN (25-400 μM). We assessed cell viability; levels of endogenous antioxidants, GSH, and catalase; levels of reactive oxygen species; and secretion of TNF- α, a cellular marker for oxidative stress and oxidative damage to nuclear DNA, after treatment with VCN. At VCN concentrations of 25 and 50 μM, the oxidative stress markers were unaffected and at least 85% of the cells were viable. Cell viability was significantly affected at 200 and 400 μM VCN (22-42% less than control, p <. 05). The results also indicated VCN-induced depletion of GSH and a concomitant increase in levels of oxidized GSH (GSSG). The levels of total GSH and GSSG in control and treated (400 μM VCN) cells were 37 and 2, respectively. There was a significant upregulation of catalase activity (21% more than control, p <. 05) at 100 μM of VCN and a downregulation at 400 μM (40% lower than control, p <. 05). A dose-dependent, significant increase in the formation of reactive oxygen species was observed at 200 to 400 μM of VCN. Also, an elevation (two- to three fold as compared to control, p <. 05) in oxidative damage to DNA was observed at these concentrations of VCN. Increase in TNF- αsecretion (28% higher than control, p <. 05) was observed at 400 μM VCN. Therefore, redox imbalance in astrocytes may play a major role in VCN-induced neurotoxicity, which is indicated by compromised antioxidant defense mechanisms, such as depletion of GSH, increase in GSSG, inhibition of catalase, and increase in the formation of reactive oxygen species and TNF- αsecretion, resulting in DNA oxidation.

Keywords: :

• Acrylonitrile
• Astrocytes
• Catalase
• Glutathione
• 8-Hydroxy-2′-deoxyguanosine
• Oxidative Stress