Caspase inhibition augments Dichlorvos-induced dopaminergic neuronal cell death by increasing ROS production and PARP1 activation.
Numerous epidemiological studies have shown an association between pesticide exposure and the increased risk of developing Parkinson's disease. Previously we have reported that Dichlorvos exposure can induce oxidative stress, resulting in over-expression of pro-apoptotic genes and finally caspase-dependent nigrostriatal dopaminergic neuronal cell death in rat brain. Here, we examined the effect of caspase inhibition on PC12 cell death induced by Dichlorvos (30 μM). Reactive oxygen species (ROS) generation followed by protein carbonylation, lipid peroxidation, decreased antioxidant defenses (decreased Mn-superoxide dismutase (MnSOD) activity and decreased glutathione levels) and subsequent caspase activation mediated the apoptosis. Inhibition of caspase cascade with Boc-aspartyl(OMe)-fluoromethylketone (BAF) enhanced the Dichlorvos-induced PC12 cell death, as assessed by the increased cellular efflux of lactate dehydrogenase (LDH). This increase in cell death was accompanied by a marked increase in poly(ADP-ribose) polymerase-1 (PARP1) activity, increased oxidative stress, a reduction in the mitochondrial membrane potential and reduced cellular NAD and ATP levels. Pretreatment of cells with PJ34, a PARP1 inhibitor prevented the cells from undergoing cell death and preserved intracellular NAD and ATP levels. Subsequent release of the apoptosis-inducing factor (AIF) from mitochondria and its translocation into the nucleus was also prevented by PJ34 pretreatment. In conclusion, the results of the present study show that caspase inhibition without concurrent inhibition of PARP1 is unlikely to be effective in preventing cell death because in the presence of the caspase inhibitor, caspase-independent cell death predominates due to PARP activation. These results suggest that combined therapeutic strategies directed at multiple cell death pathways may provide superior neuroprotection than those directed at a single mechanism.
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
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Wani
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
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Sunkaria
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
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Sharma
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
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Kandimalla
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India.
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Kaushal
Department of Preclinical and Clinical Pharmacology, University of Florence, Florence, Italy.
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Gerace
Department of Preclinical and Clinical Pharmacology, University of Florence, Florence, Italy.
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Chiarugi
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh 160012, India. Electronic address: kdgill2002@yahoo.co.in.
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Gill
D000255:Adenosine Triphosphate / Q000378:metabolism; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology*; D000818:Animals; D051033:Apoptosis Inducing Factor / Q000378:metabolism; D061945:Caspase Inhibitors / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D004006:Dichlorvos / Q000633:toxicity*; D004298:Dopamine / Q000378:metabolism; D007306:Insecticides / Q000633:toxicity*; D009243:NAD / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D016716:PC12 Cells; D010616:Phenanthrenes / Q000494:pharmacology; D000071137:Poly (ADP-Ribose) Polymerase-1; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism
C494831:Aifm1 protein, rat; D000590:Amino Acid Chloromethyl Ketones; D051033:Apoptosis Inducing Factor; D061945:Caspase Inhibitors; D007306:Insecticides; C434926:N-(oxo-5,6-dihydrophenanthridin-2-yl)-N,N-dimethylacetamide hydrochloride; D010616:Phenanthrenes; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D017382:Reactive Oxygen Species; C103192:butyloxycarbonyl-O-methyl-aspartyl-fluoromethyl ketone; D009243:NAD; D004006:Dichlorvos; D000255:Adenosine Triphosphate; C499322:Parp1 protein, rat; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D004298:Dopamine
2,4-dinitrophenyl hydrazine; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid; 4′,6-diamidino-2-phenylindole; AChE; AIF; BAF; Boc-aspartyl(OMe)-fluoromethylketone; DAPI; DCFH-DA; DNPH; Dichlorvos; EDTA; EGTA; ELISA; H2DCF-DA; HEPES; IC50; LDH; MDA; MMP; MPT; MTT; Mn-superoxide dismutase; MnSOD; OP; PARP1; PARP1 activation; PBS; ROS; RT; acetylcholinesterase; apoptosis-inducing factor; caspase inhibition; dichloro-dihydro-fluorescein diacetate; dihydrodichlorofluorescein diacetate; enzyme-linked immunosorbent assay; ethylene glycol tetraacetic acid; ethylenediaminetetraacetic acid; inhibitory concentration 50; lactate dehydrogenase; malondialdehyde; mitochondrial membrane potential; mitochondrial permeability transition; organophosphate; oxidative stress; phosphate-buffered saline; poly(ADP-ribose) polymerase-1; reactive oxygen species; room temperature