Investigating the Synergistic Effects of Chlorpyrifos and Cadmium Neurotoxicity in AlphasynucleinOverexpressing Dopaminergic Cell Model of Parkinson's Disease

Science Center, Bent Corridor

Parkinson’s disease (PD) is a progressive neurological disorder characterized by selective loss of dopaminergic cells in the substantia nigra of the brain that results in motor, cognitive and psychiatric abnormalities. Alpha-synuclein (α-syn) is a protein that is predominantly expressed in the brain and comprises about 60% of cytoplasmic inclusions called Lewy bodies, which are often present in the substantia nigra of PD patients. Although α-syn has been implicated in PD, the native function remains unclear. Previous studies have implicated chlorpyrifos (CPF) and cadmium (Cd), two neurotoxins, in causing oxidative stress, protein aggregation and mitochondrial dysfunction in PD. Recognizing some of the shared similarities in the cellular mechanisms between PD, pesticides and heavy metal neurotoxicity, we hypothesized that CPF, Cd, and human native α-syn will exhibit a gene-environment interaction, wherein CPF and Cd co-exposure will synergistically interact and enhance neuronal loss in α-syn expressing cells compared to vector (VEC) controls. Furthermore, we examined the synergistic effect of CPF and Cd co-exposure on native human α-syn. Utilizing an established immortalized rat mesencephalic cell line that expresses human native α-syn or empty vector control, we examined the effect of CPF, Cd, and CPF-Cd neurotoxicity using a mitochondrial-dependent cell viability assay (MTT 3-4,5-Dimethylthiazol-2-yl-2,5-Diphenyltetrazolium Bromide Assay). We report that expression of human native α-syn enhances susceptibility to Cd and CPF-Cd interaction following 24 hours exposure. Furthermore, we investigated the contribution of oxidative stress pathways in the synergistic effect between α-syn and CPF-Cd with an oxyblot technique that detects carbonyl group formation on protein chains as well as real-time free radical production detection assay. We report differential rates of protein oxidation and production of free radicals following Cd, CPF, and CPF-Cd exposures. In summary, we have identified a selective response of human native α-syn to CPF, Cd, and CPF-Cd combination. Our data suggests that neurotypical individuals that express native α-syn and potentially exposed to Cd or CPF-Cd may experience neuronal dysfunction and loss regardless of the genetic mutation. This study further establishes the need for future research to elucidate potential adverse effects of multiple environmental pollutant exposures in neurodegenerative diseases.