isoflavones mixture from Trifolium pratense L. protects HCN 1-A neurons from oxidative stress

Oxidative stress-induced neuronal cell death has been implicated in different neurological disorders and neurodegenerative diseases such as Alzheimer's disease and Parkinson's. Using the Alzheimer's disease-associated hydrogen peroxide (H₂O₂), we investigated the neuroprotective efficacy of a natural mixture of phytoestrogenic isoflavones (genistein, daidzein, biochanin A and formononetin) from Trifolium pratense L. (Red clover) against oxidative stress-induced cell death in human cortical cell line HCN 1-A maintained in culture. Neuronal viability was determined by MTT or trypan blue test and neuronal integrity by morphological analysis.The results obtained indicate that exposure of HCN 1-A cell cultures to hydrogen peroxide resulted in a concentration-dependent decrease in neuron viability. Concentration of H₂O₂ ranging from 50 to 200 μg/ml were toxic to these cultures. A 24-hour pretreatment with 0.5, 1 and 2 μg/ml isoflavones extract significantly increased cell survival as evidenced by MTT or trypan blue test and significantly prevented the morphological disruption caused by H₂O₂ as shown by microscopical inspection, indicating that neurons treated with isoflavones were protected from the cell death induced by H₂O₂ exposure. These findings imply that the neuroprotective effect of isoflavones extract is partly associated with its antioxidant activity. Further, results of these investigations indicate that although isoflavones extract exert a neuroprotective effect, it do not promoted cortical neuron process outgrowth. Copyright © 2008 John Wiley & Sons, Ltd.