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Salidroside-Mediated Neuroprotection is Associated with Induction of Early Growth Response Genes (Egrs) Across a Wide Therapeutic Window

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Abstract

Salidroside exhibits anti-inflammatory, anti-oxidative, and anti-apoptotic properties. To identify whether salidroside might be a candidate for treating ischemic stroke, we investigated the effects of salidroside or vehicle, given daily for 6 days, after middle cerebral artery occlusion (MCAO) for 2 h and reperfusion for either 1 or 48 h in rats. Salidroside reduced cerebral infarct volume and significantly improved neurological scores whether started after 1 or 48 h of reperfusion. Microarray analysis showed that 20 % (133/678) of the genes down-regulated by ischemia and 1 h of reperfusion were up-regulated by salidroside, whereas 13 % (105/829) of the genes induced by ischemia–reperfusion were inhibited by salidroside, suggesting that salidroside can reverse effects of ischemia–reperfusion on gene expression. The main enriched functional categories induced by salidroside were genes related to synaptic plasticity, whereas salidroside inhibited genes related to inflammation. Induction of Egr1, Egr2, Egr4, and Arc by salidroside was confirmed by qRT-PCR and western blotting in ischemic brains treated after either 1 or 48 h of reperfusion. The potential protective role of Egr4 in salidroside-mediated neuroprotection was subsequently investigated in CoCl2-treated PC12 cells. Egr4 was dose-dependently induced by salidroside in PC12 cells, and depleting Egr4 with target-specific siRNA increased caspase-3 activity and Bax, but decreased Bcl-xl, which were reversed by salidroside. Finally, we confirmed that salidroside inhibited the Bax/Bcl-xl-related apoptosis after MCAO with reperfusion. In conclusion, salidroside is highly neuroprotective with a wide therapeutic time window after ischemia–reperfusion injury in the rat, and this partially involves induction of Egrs, leading to inhibition of Bax/Bcl-xl-related apoptosis.

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (No. 81473382), the Department of Science & Technology of Fujian Province (No. 2014Y4004), the Collaborative Innovation Center for Rehabilitation Technology of Fujian University of TCM and the TCM Rehabilitation Research Center of SATCM. The authors would like to thank Mr. Bin Chen for his support in the animal surgery.

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No competing financial interests are associated with this paper.

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    Authors and Affiliations

    1. Centre of Biomedical Research & Development, Fujian University of Traditional Chinese Medicine, No. 1 Huatou Road, Minhou Shangjie, Fuzhou, China

      Wenfang Lai, Zhenwei Zheng, Xiaoqin Zhang, Yicong Wei, Kedan Chu, John Brown, Guizhu Hong & Lidian Chen

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    1. Wenfang Lai
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    2. Zhenwei Zheng
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    Correspondence to Lidian Chen.

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    Wenfang Lai and Zhenwei Zheng have contributed equally to this work.

    Electronic supplementary material

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    12640_2015_9529_MOESM1_ESM.tif

    Online Resource 1: Representative Nissl-staining coronal sections show no infarcts in the brains of the sham-operated animals treated with placebo or with salidroside, starting 1 h after operation and continued for 6 d (n = 3 per group). Supplementary material 1 (TIFF 2178 kb)

    12640_2015_9529_MOESM2_ESM.xls

    Microarray analysis of gene expression in ischemic brain of sham-operated, MCAO received vehicle after 1 h of reperfusion (MCAO) and MCAO received salidroside after 1 h of reperfusion (MCAO + Sal). Differential gene expression was identified based on 1.5-fold change, p < 0.05 difference threshold, Student’s t test. Supplementary material 2 (XLS 310 kb)

    12640_2015_9529_MOESM3_ESM.tif

    No effect of salidroside on expression of Egr1, 2, 4 and Arc in sham-operated brains. Sham-operated animals were treated with placebo or with salidroside, starting 1 h after operation and continued for 6 d. mRNA levels were measured by qRT-PCR, and expressed as fold change relative to sham control ± SEM (n = 3 per group), after normalization to GAPDH. Supplementary material 3 (TIFF 4426 kb)

    12640_2015_9529_MOESM4_ESM.tif

    Representative images show no TUNEL-positive cells in the region of sham-operated animals that corresponds to the infarcted region in MCAO rats. Sham-operated animals were treated with placebo or with salidroside, starting 1 h after operation and continued for 6d. The images were captured using laser confocal microscope. Merged images are an overlay of TUNEL (green) and DAPI (blue) staining (magnification, 400 × ; bar = 50 μm). Supplementary material 4 (TIFF 4137 kb)

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    Lai, W., Zheng, Z., Zhang, X. et al. Salidroside-Mediated Neuroprotection is Associated with Induction of Early Growth Response Genes (Egrs) Across a Wide Therapeutic Window. Neurotox Res 28, 108–121 (2015). https://doi.org/10.1007/s12640-015-9529-9

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