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Neuroprotective Effect of the LRRK2 Kinase Inhibitor PF-06447475 in Human Nerve-Like Differentiated Cells Exposed to Oxidative Stress Stimuli: Implications for Parkinson’s Disease

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Abstract

Leucine-rich repeat kinase 2 (LRRK2) has been implicated in oxidative stress (OS) and neurodegeneration in Parkinson’s disease (PD). However, the pathophysiological mechanism of the LRRK2 kinase in neurons under stress stimuli is not yet understood. We demonstrate that rotenone (ROT), a mitochondria complex I inhibitor frequently used to generate in vitro and in vivo experimental models of PD, induces LRRK2 phosphorylation at serine 935 p-(S935) concomitant with cell death in nerve-like differentiated cells (NLCs). Indeed, ROT (50 µM) at 6 h exposure significantly increased reactive oxygen species (ROS) (~100 %), p-(S935)-LRRK2 kinase [~2 f(old)-(i)ncrease] level, induced nuclei condensation/fragmentation (16 %), increased the expression of NF-κB (5.6 f-i), p53 (5.3 f-i), c-Jun (5.4 f-i) transcription factors, activated caspase-3 (8.0 f-i) and AIF (6.8 f-i) proteins; but significantly decreased mitochondrial membrane potential (∆Ψm, ~21 %), indicative of apoptosis -a type of regulated cell death process- compared to untreated cells. Strikingly, the LRRK2 kinase inhibitor PF-06447475 (PF-475, 1 µM) protects NLCs against ROT induced noxious effect. The inhibitor not only blocked the p-(S935)-LRRK2 kinase phosphorylation but also completely abolished ROS, and significantly reversed all ROT-induced apoptosis signaling and OS associated markers to comparable control values. We conclude that wild-type LRRK2 may act as a pro-apoptotic factor under OS stimuli. Our findings suggest an association between OS and LRRK2 phosphorylation in the NLCs death process, as PD model. Therefore, the pharmacological inhibition of LRRK2 might help to understand the OS-mediated kinase activation in PD neurodegenerative disorder.

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Acknowledgments

This study was supported by Colciencias grants #1115-545-31420 (contract 391) to MJ-Del-Rio and CV-P. MM-P is a Doctoral student at the “Corporacion Ciencias Basicas Biomedicas” program (CCBB)-University of Antioquia-UdeA. MM-P is funded by Colciencias Grants 567–2012. The use and technical assistance of the Flow Cytometry Unit of GICIG-SIU-UdeA and use of Odyssey Infrared Imaging System (Neuroscience Research Group, UdeA) are acknowledged.

Author Contributions

MM-P designed and performed experiments, analyzed data; CV-P and MJ-R obtained funding, designed experiments, provided reagents, analyzed data, critical reading of the manuscript and wrote the manuscript.

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  1. Neuroscience Research Group, Medical Research Institute, Faculty of Medicine, University of Antioquia (UdeA), Calle 70 No. 52-21, and Calle 62 # 52-59, Building 1, Room 412, SIU Medellin, Colombia

    Miguel Mendivil-Perez, Carlos Velez-Pardo & Marlene Jimenez-Del-Rio

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  1. Miguel Mendivil-Perez
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Correspondence to Marlene Jimenez-Del-Rio.

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Mendivil-Perez, M., Velez-Pardo, C. & Jimenez-Del-Rio, M. Neuroprotective Effect of the LRRK2 Kinase Inhibitor PF-06447475 in Human Nerve-Like Differentiated Cells Exposed to Oxidative Stress Stimuli: Implications for Parkinson’s Disease. Neurochem Res 41, 2675–2692 (2016). https://doi.org/10.1007/s11064-016-1982-1

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