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Combining Normobaric Oxygen with Ethanol or Hypothermia Prevents Brain Damage from Thromboembolic Stroke via PKC-Akt-NOX Modulation

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Abstract

In a thromboembolic stroke model after reperfusion by recombinant tissue plasminogen activator (rt-PA), we aimed to determine whether therapeutic hypothermia (TH) and ethanol (EtOH) in combination with low concentration (60 %) of normobaric oxygen (NBO) enhanced neuroprotection, as compared to using each of these agents alone. We further aimed to elucidate a potential role of the NADPH oxidase (NOX), phosphorylated protein kinase B (Akt), and protein kinase C-δ (PKC-δ) pathway in oxidative stress and neuroprotection. In Sprague–Dawley rats, a focal middle cerebral artery (MCA) occlusion was induced by an autologous embolus in the following experimental groups: rt-PA treatment alone, rt-PA + NBO treatment, rt-PA + TH at 33 °C, rt-PA + EtOH, rt-PA + NBO + EtOH, rt-PA + NBO + TH, rt-PA + NOX inhibitor, rt-PA + EtOH + NOX inhibitor, or rt-PA + EtOH + Akt inhibitor. Control groups included sham-operated without stroke or stroke without treatment. Infarct volume and neurological deficit were assessed at 24 h after rt-PA-induced reperfusion with or without treatments. ROS levels, NOX activity, and the protein expression of NOX subunits p22phox, p47phox, p67phox, gp91phox, as well as PKC-δ and phosphorylated Akt were measured at 3 and 24 h after rt-PA-induced reperfusion. Following rt-PA in thromboembolic stroke rats, NBO combined with TH or EtOH more effectively decreased infarct volume and neurological deficit, as well as reactive oxygen species (ROS) production than with any of the used monotherapies. NOX activity and subunit expressions were downregulated and temporally associated with reduced PKC-δ and increased p-Akt expression. The present study demonstrated that combining NBO with either TH or EtOH conferred similar neuroprotection via modulation of NOX activation. The results suggest a role of Akt in NOX activation and implicate an upstream PKC-δ pathway in the Akt regulation of NOX. It is possible to substitute EtOH for TH, thus circumventing the difficulties in clinical application of TH through the comparatively easier usage of EtOH as a potential stroke management.

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Acknowledgments

This work was partially supported by the Wayne State University Neurosurgery Fund, American Heart Association Grant-in-Aid (14GRNT20460246), and National Nature Science Foundation of China (no. 81501141).

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Authors and Affiliations

  1. China-America Institute of Neuroscience, Xuanwu Hospital, Capital Medical University, Beijing, 101149, China

    Lipeng Cai, Xunming Ji & Yuchuan Ding

  2. Department of Neurology, Luhe Hospital, Capital Medical University, Beijing, China

    Lipeng Cai, Xiaokun Geng, Xunming Ji & Yuchuan Ding

  3. Department of Neurological Surgery, Wayne State University School of Medicine, 550 E Canfield, Detroit, MI, 48201, USA

    Lipeng Cai, James Stevenson, Xiaokun Geng, Changya Peng, Ruiqiang Xin, Radhika Rastogi, Christopher Sy & Yuchuan Ding

  4. Department of Radiology, Luhe Hospital, Capital Medical University, Beijing, China

    Ruiqiang Xin

  5. Department of Anatomy and Cell Biology, Wayne State University School of Medicine, Detroit, MI, USA

    Jose A. Rafols

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  1. Lipeng Cai
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  2. James Stevenson
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  3. Xiaokun Geng
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Cai, L., Stevenson, J., Geng, X. et al. Combining Normobaric Oxygen with Ethanol or Hypothermia Prevents Brain Damage from Thromboembolic Stroke via PKC-Akt-NOX Modulation. Mol Neurobiol 54, 1263–1277 (2017). https://doi.org/10.1007/s12035-016-9695-7

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