Paraquat induces apoptosis in human lymphocytes: protective and rescue effects of glucose, cannabinoids and insulin-like growth factor-1

Abstract

In order to establish causal or protective treatments for Parkinson's disease (PD), it is necessary to identify the cascade of deleterious events that lead to the dysfunction and death of dopaminergic neurons. Paraquat (PQ) is a pesticide used as xenobiotic compound to model PD. However, the mechanism(s) of PQ-induced cell death and the mechanism(s) of cytoprotection in a single cell model are still unknown. In this study, lymphocytes were treated with (0.1-1 mM) PQ. Apoptotic morphology was assessed with acridine orange/ethidium bromide staining. Further evaluation included (i) superoxide radicals, reflected by nitroblue tetrazolium reduction to formazan, (ii) the production of hydrogen peroxide, reflected by rhodamine-positive fluorescent cells, (iii) the generation of hydroxyl radicals, reflected by dimethylsulfoxide and melatonin ( radical)OH scavengers, (iv) activation and/or translocation of NF-kappaB, p53 and c-Jun transcription factors showed by immunocytochemical staining, and by ammonium pyrrolidinedithiocarbamate, pifithrin-alpha and SP600125 inhibition and (V) caspase-3 activation, reflected by caspase Ac-DEVD-cho inhibition. To elucidate the mechanism of cytoprotection, lymphocytes were treated with PQ in the presence of cannabinoids, insulin-like growth factor-1 and glucose. We provide evidence that PQ induces apoptosis in lymphocytes in a concentration- and time-dependent fashion by an oxidative stress mechanism involving O(2)( radical - ), H(2)O(2)/(( radical)OH) generation, simultaneous activation of NF-kappaB/p53/c-Jun transcription factors, mitochondrial depolarization and caspase-3 activation leading to morphological apoptosis. Moreover, dying lymphocytes are protected and rescued from PQ noxious stimuli by direct antioxidant effect by cannabinoids, receptor mediated signaling by IGF-1, and/or energetic protection by glucose. It is concluded that PQ-induced apoptosis in lymphocytes by a mechanism involving reactive oxygen species generation, mitochondrial dysfunction, transcriptional factors and caspase-3 activation. However, this cell death routine can be reversed by the action of cannabinoids, IGF-1 and glucose. These data may provide innovating therapeutic strategies to intervene environmentally or genetically susceptible PD population to oxidative stress.