角鲨烯通过调节细胞铁死亡减轻脑缺血再灌注损伤  

Squalene attenuates cerebral ischemia-reperfusion induced brain tissue injury by regulating ferroptosis

作　　者：蒋霞 袁钰萍 陈舒怀 段名扬 邓祖跃 JIANG Xia;YUAN Yu-ping;CHEN Shu-huai;DUAN Ming-yang;DENG Zu-yue(Green Pharmaceutical Collaborative Innovation Center,Zhejiang University of Technology,Hangzhou 310014,China;Zhejiang Institute for Food and Drug Control,Hangzhou 310052,China)

机构地区：[1]浙江工业大学绿色制药协同创新中心,浙江杭州310014 [2]浙江省食品药品检验研究院,浙江杭州310052

出　　处：《中国病理生理杂志》2021年第10期1784-1793,共10页Chinese Journal of Pathophysiology

基　　金：浙江省省级公益性技术应用研究计划项目(No.2014C37020);浙江省中医药科技计划项目(No.2021ZB178)。

摘　　要：目的:研究角鲨烯(SQU)对脑缺血再灌注损伤大鼠脑组织的保护作用及其抗细胞铁死亡的机制。方法:采用大脑中动脉栓塞(MCAO)2 h后复灌建立大鼠脑缺血再灌注损伤模型,术后12 h和72 h采用Longa神经行为学评分标准和TTC染色评价脑损伤。用FeCl3(200μmol/L)诱导PC12细胞建立铁死亡细胞模型,比较SQU和铁死亡抑制剂Fer-1或诱导剂erastin对细胞铁死亡的影响。MTT法测定细胞存活率,荧光显微镜检测线粒体膜电位,发色底物法测定细胞和大鼠脑组织中的铁离子含量、ROS荧光强度、谷胱甘肽(GSH)和丙二醛(MDA)水平,Western blot检测各组脑组织和PC12细胞转铁蛋白受体(TFR1)、脂酰辅酶A合成酶长链家族成员4(ACSL4)、谷胱甘肽过氧化物酶4(GPX4)、膜铁转运蛋白1(FPN1)和胱氨酸/半胱氨酸-谷氨酸的转运蛋白轻链亚基SLC7A11的表达。结果:与MCAO模型组比较,各剂量(20、40和80 mg·kg^(-1)·d^(-1))SQU治疗组大鼠的神经行为学评分下降,脑梗死体积减小(P<0.05);在铁死亡细胞模型中,SQU可增加PC12细胞存活率(P<0.05);在相关指标检查中,SQU可下调大鼠脑组织和PC12细胞中TFR1、ACSL4蛋白表达,上调FPN1、SLC7A11和GPX4蛋白的表达,降低细胞铁离子浓度、ROS的荧光强度和MDA水平,增加细胞GSH的含量(P<0.05);SQU能拮抗erastin诱导的PC12细胞铁死亡,并且能增强阻断剂Fer-1抗PC12细胞铁死亡的作用。结论:角鲨烯对脑缺血再灌注损伤大鼠的脑组织有保护作用,机制可能与其降低细胞铁积累、提高细胞抗氧化能力、降低ROS和脂质过氧化水平从而降低细胞铁死亡的发生有关。AIM:To study the protective effect of squalene(SQU)on brain tissue of rats with cerebral ischemia-reperfusion injury and its mechanism of anti-ferroptosis.METHODS:Cerebral ischemia-reperfusion injury was established by middle cerebral artery occlusion(MCAO)for 2 h followed by reperfusion in rats. The Longa neurobehavioral score standard and TTC staining were used to assess brain damage at 12 h and 72 h after surgery. The PC12 cells were treated with FeCl3(200 μmol/L)to establish a cell model of ferroptosis,and then the effects of SQU and ferroptosis inhibitor Fer-1 or inducer erastin on ferroptosis were compared. The viability was measured by MTT assay. The mitochondrial membrane potential was analyzed by fluorescence microscopy. The iron concentration,reactive oxygen species(ROS)fluorescence intensity,glutathione(GSH)and malondialdehyde(MDA)in cells and brain tissue were evaluated by chromogenic substrate method. The expression of transferrin receptor 1(TFR1),acyl-CoA synthetase long-chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),ferroportin 1(FPN1)and SLC7 A11 in brain tissues and PC12 cells of each group was investigated by Western blot.RESULTS:Compared with MCAO group,the neurobehavioral scores and the cerebral infarction volume of each dose(20,40 and 80 mg·kg^(-1)·d^(-1))of SQU treated group were decreased(P<0. 05). SQU increased the cell viability of FeCl3-induced PC12 cells(P<0. 05). SQU inhibited the expression of TFR1 and promoted the expression of FPN1 in rat brain tissues and PC12 cells and then reduced the iron concentration(P<0. 05). At the same time,SQU reduced the expression of ACSL4 and increased the expression of SLC7 A11 and GPX4,and then reduced the fluorescence intensity of ROS and MDA levels,and increased the content of GSH(P<0. 05). In addition,SQU antagonized erastin-induced ferroptosis and increased the effect of Fer-1 to against the ferroptosis in PC12 cells.CONCLUSION:Squalene has a protective effect on brain tissue in rats with cerebral ischemia-reperfusion injury,the un

关 键 词：角鲨烯 脑缺血再灌注损伤 铁死亡 脂质代谢 脂质过氧化 

分 类 号：R743.3[医药卫生—神经病学与精神病学] R363.2[医药卫生—临床医学]