[Show abstract][Hide abstract] ABSTRACT: Effect of 4-hydroxynonenal (HNE), a long-chain alpha, beta unsaturated aldehyde product, generated by the oxidation of omega-6 polyunsaturated fatty acids on the sensitivity of selected neurotransmitter receptors was studied in PC-12 cells. Cytotoxicity profiling was carried out at varying concentrations of HNE (0.1-50microM) for 30min to 24h. Trypan blue dye exclusion, MTT, LDH release and neutral red uptake (NRU) assays were carried out to assess the cytotoxicity of HNE. Cytotoxic response was found to be significant at 2h of exposure. Cytotoxicity of HNE at 50microM was exerted even at 90min. HNE 10-50microM was found to be cytotoxic, whereas, 2-5microM causes physiological stress only and 1-0.1microM non-cytotoxic. Effect on dopamine, cholinergic, serotonin and benzodiazepine receptors was studied at varying concentrations of HNE (1, 10, 25 and 50microM for 1-8h). A significant decrease in binding of 3H-QNB, 3H-Fluinitrazepam and 3H-Ketanserin, known to label cholinergic (muscarinic), benzodiazepine and serotonin (5HT(2A)) receptors respectively was observed at 1h exposure of PC-12 cells to HNE at 25 and 50microM concentrations. The decrease in the binding of (3)H-Spiperone, known to label dopamine (DA-D2) receptors was evident at 4h of exposure of PC-12 cells to HNE. The decrease in the binding with DA-D2 receptors continued till 8h. Effect on the binding of (3)H-Fluinitrazepam and 3H-Ketanserin appeared to be maximum at 25 and 50microM concentrations of HNE for 4h and 8h. The PC-12 cells appear to be vulnerable to cytotoxic concentrations of HNE. Experimental HNE exposure provides an intriguing model of toxicant-cell interactions involving neurotransmitter receptors in HNE neurotoxicity.
Toxicology in Vitro 08/2008; 22(7):1681-8. DOI:10.1016/j.tiv.2008.07.001 · 2.90 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Investigations were carried out to elucidate the involvement of oxidative stress in 4-hydroxynonenal (HNE) mediated cytotoxicity in NGF induced differentiated PC12 cells. Well differentiated PC12 cells at day 8 were exposed to various concentrations (0.1-50mM) of HNE for 30 minutes-24h. Non-differentiated PC12 cells were also exposed under identical conditions and served as control. Following exposure, cytotoxicity assessment was done using standard endpoints viz., MTT, LDH release and neutral red uptake (NRU) assays. Further levels of glutathione and intracellular calcium, immunocytochemical localization of DA-D 2 receptor and HNE-protein binding were carried out using concentrations of HNE (1, 10, 25, 50mM), exerting the effect on cell growth in cytotoxicity studies. Significant cytotoxic responses were observed earliest by 2h of exposure, except for the concentration 50mM, where cytotoxicity was exerted even at 90min. HNE concentrations 10-50mM were found to be cytotoxic, 2-5mM cytostatic and 1-0.1mM non-cytotoxic. The depletion in GSH levels was statistically significant (p<0.001) since 1h of exposure and became more intensive at 2 and 4h exposures, however the exposure of 8h could not lead further depletion in GSH level. A correlation between depletion of GHS levels and increase in the intracellular Ca ++ , binding of HNE with intracellular proteins and involvement of DA-D 2 receptor in HNE mediated cytotoxicity could be well established. Moreover, no significant differences in the responsiveness to HNE by NGF differentiated and non-differentiated PC12 cells could be observed. Results are suggestive the involvement of oxidative stress in HNE mediated cytotoxicity in PC12 cells.
Journal of Ecophysiology and Occupational Health 06/2007; 6(1-6(2007) 99-111):99-111.