Abstract
Receptor-interacting protein kinase 1/3 (RIPK1/3) and mixed lineage kinase domain-like (MLKL)-mediated necroptosis contributes to brain injury after ischemic stroke. Ligustroflavone is an ingredient of common privet with activities of anti-inflammation and complement inhibition. This study aims to explore the effect of ligustroflavone on ischemic brain injury in stroke rat and the underlying mechanisms. A rat model of ischemic stroke was established by middle cerebral artery occlusion (MCAO), which showed ischemic injury (increase in neurological deficit score and infarct volume) and upregulation of necroptosis-associated proteins (RIPK1, RIPK3 and MLKL/p-MLKL). Administration of ligustroflavone (30 mg/kg, i.g.) 15 min before ischemia evidently improved neurological function, reduced infarct volume, and decreased the levels of necroptosis-associated proteins except the RIPK1. Consistently, hypoxia-cultured PC12 cells (O2/N2/CO2, 1:94:5, 8 h) caused cellular injury (LDH release and necroposis) concomitant with up-regulation of necroptosis-associated proteins, and these phenomena were blocked in the presence of ligustroflavone (25 μM) except the elevated RIPK1 levels. Using the Molecular Operating Environment (MOE) program, we identified RIPK1, RIPK3, and MLKL as potential targets of ligustroflavone. Further studies showed that the interaction between RIPK3 and RIPK1 or MLKL was significantly enhanced, which was blocked in the presence of ligustroflavone. Based on these observations, we conclude that ligustroflavone protects rat brain from ischemic injury, and its beneficial effect is related to the prevention of necroptosis through a mechanism involving targeting RIPK1, RIPK3, and/or MLKL.
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Funding
This work was supported by the National Nature Science Foundation of China (No. 81573430 to Xiu-Ju Luo, No. 81872873 to Jun Peng), and Hunan Provincial Natural Science Foundation of China (No. 2015JJ2156 to Xiu-Ju Luo).
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LXJ, PJ, and YJ conceived and designed the research. ZYY, LWN, and LYQ conducted experiments. ZYY and ZXJ analyzed data. ZYY, LXJ, and PJ wrote the manuscript. All authors read and approved the manuscript.
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The study was done with compliance to the ethics standards and approval from the ethics committee of the Central South University, China.
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Zhang, YY., Liu, WN., Li, YQ. et al. Ligustroflavone reduces necroptosis in rat brain after ischemic stroke through targeting RIPK1/RIPK3/MLKL pathway. Naunyn-Schmiedeberg's Arch Pharmacol 392, 1085–1095 (2019). https://doi.org/10.1007/s00210-019-01656-9
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DOI: https://doi.org/10.1007/s00210-019-01656-9