二氢辣椒素不依赖诱导亚低温对心脏骤停复苏后脑损伤的防治作用及分子机制  

Protective effect and molecular mechanism of dihydrocapsaicin independent of induced mild hypothermia on brain injury after the resuscitation in cardiac arrest patients

作　　者：钟晓芃[1] 郭义[2] 张诗武[3] Zhong Xiao-peng;Guo Yi;Zhang Shi-wu(Emergency Department,Tianjin People's Hospital,Tianjin 300122,China)

机构地区：[1]天津市人民医院急诊科,天津300122 [2]天津中医药大学中医学院 [3]天津市人民医院病理科

出　　处：《中国急救医学》2022年第10期850-857,共8页Chinese Journal of Critical Care Medicine

基　　金：国家自然科学基金重点项目(82030125);天津市卫健委课题(2019121);天津市人民医院院内课题(2017YJ019)。

摘　　要：目的研究二氢辣椒素不依赖诱导亚低温作用对心脏骤停复苏后脑损伤的保护作用机制。方法①细胞实验:体外培养神经瘤母细胞(SH-SY5Y)建立氧糖剥夺/复糖复氧模型,流式细胞学检测细胞的凋亡,Western blot检测TRPV1、UCP-4和PKA、caspase-3的表达变化;应用流式细胞学检测线粒体膜电位、线粒体内钙离子及细胞内ROS含量。②动物实验:使用窒息法诱导心脏骤停建立心脏骤停后脑损伤模型。纸带移除法测定大鼠神经功能结果,TUNEL法检测大鼠大脑皮层神经细胞凋亡,免疫组化法、Western blot检测大鼠大脑皮层神经细胞TRPV1、PKA、UCP-4蛋白含量。结果氧糖剥夺/复糖复氧SH-SY5Y及窒息法诱导大鼠心脏骤停复苏后中枢神经细胞发生凋亡,二氢辣椒素导致TRPV1过表达,cAMP-PKA信号被激活,UCP-4表达上升,线粒体膜电位恢复,线粒体内钙离子减少,细胞内ROS含量减少,caspase-3表达减少,细胞凋亡被抑制。抑制TRPV1表达,cAMP-PKA信号被抑制,且UCP-4表达减少,线粒体膜电位不能恢复,线粒体内钙离子增加,细胞内ROS含量增加,caspase-3表达增加,神经细胞凋亡增加。抑制PKA,UCP-4表达减少,线粒体膜电位不能恢复,线粒体内钙离子增加,细胞内ROS含量增加,caspase-3表达增加,神经细胞凋亡增加。结论在氧糖剥夺/复糖复氧SH-SY5Y细胞中经二氢辣椒素预处理可致TRPV1过表达,且通过PKA/UCP-4抑制SH-SY5Y细胞凋亡。在心脏骤停复苏后综合征SD大鼠中,二氢辣椒素同样可以通过TRPV1/PKA/UCP-4抑制大脑皮层神经细胞凋亡。Objective To study the protective mechanism of dihydrocapsaicin(DHC)independent of induced mild hypothermia on brain injury after the resuscitation in cardiac arrest patients.Methods①Cell experiment:the model of oxygen-glucose deprivation/reoxygenation(OGD/R)was established by culturing neuroblastoma cells(SH-SY5Y)in vitro,the apoptosis was detected by flow cytometry,the expressions of TRPV1,UCP-4,PKA and caspase-3 were detected by Western blot,and the mitochondrial membrane potential,intracellular calcium concentration and intracellular reactive oxygen species(ROS)concentration were detected by flow cytometry.②Animal experiment:the model of brain injury after cardiac arrest was induced by asphyxia.The results of neurological function of rats were measured by paper tape removal method,the apoptosis of cerebral cortical nerve cells was detected by TUNEL method,and the protein contents of TRPV1,PKA and UCP-4 in cerebral cortical nerve cells were detected by immunohistochemistry and Western blot.Results OGD/R and asphyxia induced cardiac arrest led to neuronal apoptosis,DHC led to the overexpression of TRPV1,cAMP-PKA signal was activated,UCP-4 expression rose,mitochondrial membrane potential was restored,and calcium ions in mitochondria decreased,the content of intracellular ROS decreased,the expression of caspase-3 decreased,and cell apoptosis was inhibited.Owing to the inhibition of TRPV1 expression,cAMP-PKA signal was inhibited,and UCP-4 expression reduced,mitochondrial membrane potential can not be restored,calcium ion concentration in mitochondria increased,intracellular ROS content increased,caspase-3 expression increased,and nerve cell apoptosis increased.After PKA inhibition,the expression of UCP-4 decreased,the mitochondrial membrane potential can not be restored,the calcium ion concentration in the mitochondria increased,the intracellular ROS content increased,the expression of caspase-3 increased,and the apoptosis of nerve cells increased.Conclusions DHC preconditioning can induce TRPV1 overexpression

关 键 词：复苏后综合征 氧糖剥夺/复糖复氧 TRPV1/PKA/UCP-4 凋亡 

分 类 号：R965[医药卫生—药理学]