针对脑缺血再灌注损伤设计的双功能蛋白的脑保护作用优于单功能蛋白

doi:10.5246/jcps.2019.11.072

中国药学(英文版) ›› 2019, Vol. 28 ›› Issue (11): 760-769.DOI: 10.5246/jcps.2019.11.072

针对脑缺血再灌注损伤设计的双功能蛋白的脑保护作用优于单功能蛋白

于快, 李茹一, 朱元军^*, 刘晓岩, 王银叶^*

北京大学医学部药学院分子与细胞药理学系, 北京 100191

收稿日期:2019-08-23 修回日期:2019-09-15 出版日期:2019-12-01 发布日期:2019-10-25
通讯作者: Tel.: +86-10-82802652, E-mail: zhuyuanjun@bjmu.edu.cn; wangyinye@bjmu.edu.cn
基金资助:
National Natural Science Foundation of China (Grant No. 81573333, 81503060).

Engineering the protein targeting two pathways of cerebral ischemia reperfusion injury provides better neuroprotective effect than targeting one pathway

Kuai Yu, Ruyi Li, Yuanjun Zhu^*, Xiaoyan Liu, Yinye Wang^*

Department of Molecular and Cellular Pharmacology, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing, 100191, China

Received:2019-08-23 Revised:2019-09-15 Online:2019-12-01 Published:2019-10-25
Contact: Tel.: +86-10-82802652, E-mail: zhuyuanjun@bjmu.edu.cn; wangyinye@bjmu.edu.cn
Supported by:
National Natural Science Foundation of China (Grant No. 81573333, 81503060).

摘要/Abstract

摘要：

为了验证针对缺血再灌注损伤过程所涉及的多种途径的神经保护剂优于只针对单途径的神经保护剂的设想, 本研究选择了抗凋亡蛋白Bcl-xL (B)和抗炎蛋白IL-10 (I)作为两个功能单元, 设计了双功能融合蛋白Bcl-xL-IL-10 (B-I), 并在大肠杆菌中高水平表达。通过亲和层析对三种重组蛋白的包涵体进行纯化并复性, 获得B-I、B和I纯化蛋白。得到的B-I可以在其N端的TAT肽段的引导下跨过血脑屏障, 并被caspase-1切割为两部分。B-I处理可以显著降低脑梗死体积, 优于单纯B或I处理, 与两种单功能蛋白联合处理效果相当。用B或B-I治疗可显著减少缺血再灌注诱导的神经元凋亡, 表现为凋亡率的降低和caspase-3活性的抑制。另外, 所有重组蛋白, 尤其是B-I, 显著下调脑皮层TNF-α的含量。这些结果表明:融合蛋白B-I抑制炎症和凋亡比单独抑制任何一种都具有更好的神经保护作用。本研究证明双功能融合蛋白的脑缺血保护作用优于单功能蛋白, 该结果为缺血性脑卒中的神经保护治疗药物研究提供了新的思路。

关键词: 脑缺血再灌注, 双功能, 凋亡, 炎症, 神经保护

Abstract:

We hypothesized that neuroprotective agents targeting various pathways involved in cerebral ischemia/reperfusion (I/R) injury might be superior to that targeting single pathway. Here, we prepared a fusion protein (B-I) by combining anti-apoptotic Bcl-xL (B) and anti-inflammatory IL-10 (I). B-I could cross blood brain barrier by its N-terminal TAT domain, and be cleaved into separate B and I by Caspase-1. B-I treatment significantly reduced the cerebral infarct volume, better than B or I treatment alone, and equivalent to B and I treatment (B+I). Treatment with B or B-I significantly attenuated I/R-induced neuronal apoptosis as shown by the decrease in apoptotic rate and the inhibition of caspase-3 activity. Moreover, all recombinant proteins, especially B-I, remarkably attenuated I/R-induced up-regulation of TNF-α. These results suggested that fusion protein B-I inhibiting both inflammation and apoptosis provided better neuroprotective effects than inhibiting either one alone. Our study suggested that multiple pathways targeting brain I-R injury could enhance the neuroprotective effect, and it provided a new idea for the study of neuroprotective drugs for ischemic stroke.

Key words: Cerebral ischemia reperfusion, Bifunction, Apoptosis, Inflammation, Neuroprotection

中图分类号:

R965

Supporting:

于快, 李茹一, 朱元军, 刘晓岩, 王银叶. 针对脑缺血再灌注损伤设计的双功能蛋白的脑保护作用优于单功能蛋白[J]. 中国药学(英文版), 2019, 28(11): 760-769.

Kuai Yu, Ruyi Li, Yuanjun Zhu, Xiaoyan Liu, Yinye Wang. Engineering the protein targeting two pathways of cerebral ischemia reperfusion injury provides better neuroprotective effect than targeting one pathway[J]. Journal of Chinese Pharmaceutical Sciences, 2019, 28(11): 760-769.

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针对脑缺血再灌注损伤设计的双功能蛋白的脑保护作用优于单功能蛋白

Engineering the protein targeting two pathways of cerebral ischemia reperfusion injury provides better neuroprotective effect than targeting one pathway

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