组织激肽释放酶对酸敏离子通道介导的神经元缺血／再灌注损伤的影响  

Effects of tissue kallikrein on acid-sensing ion channels mediated neuronal damage during ischemia/reoxygen

作　　者：刘玲[1,2] 唐敏[1] 任惠民[1] 杨雪莲[1] 周厚广[1] 董强[1,3] 

机构地区：[1]复旦大学附属华山医院神经内科,上海200040 [2]南京大学附属南京军区南京总医院神经内科,南京200022 [3]复旦大学脑科学研究院,上海200032

出　　处：《神经解剖学杂志》2009年第1期47-54,共8页Chinese Journal of Neuroanatomy

基　　金：国家自然科学基金(No.30570632)资助项目

摘　　要：为了解缺血/再灌注过程中组织激肽释放酶(tissu kallikrein,TK)对神经元的保护作用与酸敏感离子通道(acid-sensing ion channels,ASICs)激活的关系以及其可能涉及的细胞内信号通路,本研究建立大鼠原代皮层神经元缺氧缺糖(oxygen-glucose deprivation,OGD)、缺氧缺糖-酸中毒(OGD-Acidosis)损伤模型以模拟在体缺血/再灌注损伤,应用细胞免疫化学方法观察损伤前后神经元形态学的变化,并检测ASIC1a的分布变化;用活细胞计数试剂盒CCK-8了解神经元存活情况并检测Caspase-3酶活性了解神经元凋亡情况。观察OGD和OGD-酸中毒对培养神经元的存活率和Caspase-3活性的影响及TK预处理、缓激肽(bradyki-nin,BK)预处理、ASIC1a特异性和非特异阻断剂预处理对OGD-酸中毒损伤神经元上述指标的影响,并检测细胞外信号调节激酶(ERK)、C-Jun-N末端激酶(JNK)和磷酸肌醇-3激酶/蛋白激酶B或Akt(PI3K/Akt)信号通路抑制剂对TK作用的影响。结果显示:OGD-酸中毒对神经元的损伤明显重于单纯OGD,阻断ASICs的激活能提高OGD-酸中毒神经的存活率、降低Caspase-3活性;TK和BK预处理能促进OGD-酸中毒损伤神经元存活、抑制Caspase-3激活;阻断ERK和PI3K/Akt信号通路能够抑制TK促OGD-酸中毒损伤神经元存活效应。结果表明,ASICs的激活参与了OGD-酸中毒神经元损伤过程;TK对OGD-酸中毒神经元的保护作用可能是拮抗了ASICs介导的谷氨酸非依赖的神经元毒性作用,并可能与ERK和PI3K/Akt信号通路的激活有关。In order to investigate the relationship between tissue kallikrein （TK） protection and acid - sensing ion channels （ASICs） activation and its signaling mechanisms during ischemia/reperfusion, we set up the oxygen-glucose deprivation （OGD） and OGD - Acidosis models to mimick ischema/reperfusion in vivo. ASIC1a distribution and morphological changes of neurons after OGD or OGD-Acidosis were determined by immunocytochemistry. Cell viability and apoptosis were examined by Cell Counting Kit-8 and Caspase-3 assay. Here we demonstrated that OGD-Acidosis, but not OGD induced more remarkable changes in neuronal morphology, neuron survival and Caspase-3 activity, which could be inhibited by ASICs blockers, suggesting that ASICs played a crucial role in OGD-Acidosis-induced neuronal injury. In addition, TK or BK （bradykinin, the agonist of bradykinin receptor 2, B2R） pre-treatment in cultured neurons also significantly promoted neuronal viability and reduced Caspase-3 activation induced by OGD-Acidosis. Furthermore, the effect of TK on neuronal survival was blocked by the inhibitor of extraeellular signal regulated kinase （ERK） , c-Jun-N terminal kinase （JNK） or PI3 K inhibitor. In conclusion, ASICs activation during OGD-Acidosis contributes to neuronal injury. TK attenuates OGD-Acidosis-induced neuronal damage, which may be associated with its inhibitory effects on ASICs by B2R activation, involving the activation of ERK and PI3K/Akt signaling pathways and the inhibition of Caspase-3 activity.

关 键 词：组织激肽释放 酶酸敏感离子通道 缺氧缺糖 酸中毒 神经元 

分 类 号：R363[医药卫生—病理学]