Purpose: Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid, play a crucial role in cytoprotection by attenuating oxidative stress, inﬂammation and apoptosis. EETs are rapidly metabolised in vivo by the soluble epoxide hydrolase (sEH). Increasing the half life of EETs by inhibiting the sEH enzyme is a novel strategy for neuroprotection. In the present study, eight sEH inhibitors (t-TUCB, t-AUCB, t-CUPM, AUDA, TPPU, APAU, AEPU, and t-MTUCB) were screened in silico and further evaluated for their antiparkinson activity against rotenone (ROT) induced neurodegeneration in N27 dopaminergic cell line and Drosophila melanogaster model of Parkinson disease (PD).
Methods: In the in vitro study cell viability (MTT and LDH release assay), oxidative stress parameters (total intracellular ROS, hydroperoxides, protein oxidation, lipid peroxidation, superoxide dismutase, catalase, glutathione peroxidise, glutathione reductase, glutathione, total antioxidant status, mitochondrial complex-1activity and mitochondrial membrane potential), inflammatory markers (IL-6, COX-1 and COX-2), and apoptotic markers (JNK, phospho-JNK, c-jun, phospho-c-jun, pro and active caspase-3) were assessed to study the neuroprotective effects. In vivo activity of APAU was assessed in Drosophila melanogaster by measuring survival rate, negative geotaxis, oxidative stress parameters (total intracellular ROS, hydroperoxides, glutathione levels) were measured. Dopamine and its metabolites were estimated by LC-MS/MS analysis. In the in silico study the molecules showed good binding interaction of inhibitors at the active site of sEH (PDB: 1VJ5).
Results: In the in silico study the molecules showed good binding interaction of inhibitors at the active site of sEH (PDB: 1VJ5). In the in vitro study, among the tested molecules, APAU significantly attenuated ROT induced changes in oxidative, pro-inflammatory and apoptotic parameters. In the in vivo study, APAU significantly attenuates ROT induced changes in survival rate, negative geotaxis, oxidative stress, dopamine and its metabolites levels (p<0.05).
Conclusion: Our study, therefore, concludes that the molecule APAU, has significant neuroprotection benefits against ROT induced Parkinsonism.
Praveen T. Krishnamurthy– professor, JSS college of Pharmacy, ooty, Tamil Nadu
Pandareesh M. d– Post Doc, New York State Institute for Basic Research in Developmental Disabilities, New York
Bruce D Hammock– professor, Department of Entomology and Nematology, and Comprehensive Cancer Research Center, University of California, Davis, United States