Post-stroke DHA Treatment Protects Against Acute Ischemic Brain Injury by Skewing Macrophage Polarity Toward the M2 Phenotype
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Systemic docosahexaenoic acid (DHA) has been explored as a clinically feasible protectant in stroke models. However, the mechanism for DHA-afforded neuroprotection remains elusive. Transient middle cerebral artery occlusion (tMCAO) was induced for 1 h. DHA (i.p., 10 mg/kg) was administered immediately after reperfusion and repeated daily for 3 days. Stroke outcomes, systemic inflammatory status, and microglia/macrophage phenotypic alterations were assessed 3 days after stroke. Macrophage depletion was induced by clodronate liposomes injection. Primary macrophage cultures were used to evaluate the direct effect of DHA on macrophages. We demonstrated that post-stroke DHA injection efficiently reduced brain infarct and ameliorated neurological deficits 3 days after tMCAO. Systemic DHA treatment significantly inhibited immune cell infiltration (macrophages, neutrophils, T lymphocytes, and B lymphocytes) and promoted macrophage polarization toward an anti-inflammatory M2 phenotype in the ischemic brain. Meanwhile, systemic DHA administration inhibited the otherwise elevated pro-inflammatory factors in blood and shifted circulating macrophage polarity toward M2 phenotype after ischemic stroke. The numbers of circulating immune cells in blood and spleen, however, were equivalent between DHA- and vehicle-treated groups. The protective effects of DHA were macrophage-dependent, as macrophage depletion abolished DHA-afforded neuroprotection. In vitro studies confirmed that DHA suppressed production of chemokines and pro-inflammatory cytokines from macrophages under inflammatory stimulation. These data indicate that post-stroke DHA treatment ameliorated acute ischemic brain injury in a macrophage-dependent manner and DHA enhanced macrophage phenotypic shift toward an anti-inflammatory phenotype to reduced central and peripheral inflammation after stroke.
KeywordsCerebral ischemia Macrophage Chemokine Cytokine DHA Neuroprotection
The authors thank Dr. Lisheng Peng for technical support with the tMCAO stroke model. We thank Michael Hezel for editorial assistance.
WC designed and performed the experiments, collected and analyzed data, and drafted the manuscript. SL carried out immunostaining, imaging, and quantification and drafted the manuscript. MH and XS performed animal experiments and collected data. WQ contributed to the experimental design and the manuscript. SZ contributed to the experimental design and revised the manuscript. XH designed the experiment and critically revised the manuscript. ZL designed and supervised the study and critically revised the manuscript. All authors read and approved the final manuscript.
This work was supported by grant 81671178 from the National Natural Science Foundation of China and funding from the Natural Science Foundation of Guangdong Province (2017A030311013).
Compliance with Ethical Standards
Conflict of Interest
The authors declare that they have no conflicts of interest.
All animal experiments were approved by the Third Affiliated Hospital of Sun Yat-sen University and performed following the Guide for the Care and Use of Laboratory Animals (8th edition, 2011) .
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