Chen Ku-Chung, Chang Long-Sen
Institute of Biomedical Sciences, National Sun Yat-Sen University-Kaohsiung Medical University Joint Research Center, National Sun Yat-Sen University, Kaohsiung 804, Taiwan.
Toxicology. 2009 Aug 21;262(3):199-206. doi: 10.1016/j.tox.2009.06.009. Epub 2009 Jun 21.
Arachidonic acid (AA)-induced apoptosis of human neuroblastoma SK-N-SH cells was characteristic of elevation of intracellular Ca(2+) concentration ([Ca(2+)]i), ROS generation, activation of 38 MAPK and JNK and loss of mitochondrial membrane potential (DeltaPsim). Subsequent modulation of Bcl-2 family members and cytochrome c release accompanied with activation of caspase-9 and -3 were involved in the death of SK-N-SH cells. BAPTA-AM (Ca(2+) chelator) pretreatment rescued viability of AA-treated cells through abolishing phosphorylation of p38 MAPK and JNK, DeltaPsim loss and ROS generation. N-Acetylcysteine (ROS scavenger) pretreatment reduced the dissipation of DeltaPsim, but insignificantly affected AA-induced p38 MAPK and JNK activation. SB202190 (p38 MAPK inhibitor) and SP600125 (JNK inhibitor) attenuated mitochondrial depolarization, degradation of Bcl-2/Bcl-xL, and mitochondrial translocation of Bax. Transfection of specific siRNA proved that p38alpha MAPK and JNK1 were involved in modulating Bcl-2 family proteins. Taken together, our data suggest that the cytotoxicity of AA toward SK-N-SH cells is mediated through mitochondria-dependent death pathway, eliciting by AA-induced ROS generation and Ca(2+)-evoked activation of p38alpha MAPK and JNK1.
花生四烯酸（AA）诱导的人神经母细胞瘤SK-N-SH细胞凋亡的特征是细胞内Ca(2+)浓度（[Ca(2+)]i）升高、活性氧（ROS）生成、38丝裂原活化蛋白激酶（MAPK）和应激活化蛋白激酶（JNK）激活以及线粒体膜电位（ΔΨm）丧失。随后Bcl-2家族成员的调节和细胞色素c释放，伴随着caspase-9和-3的激活，参与了SK-N-SH细胞的死亡。1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸乙酰甲酯（BAPTA-AM，Ca(2+)螯合剂）预处理通过消除p38 MAPK和JNK的磷酸化、ΔΨm丧失和ROS生成，挽救了AA处理细胞的活力。N-乙酰半胱氨酸（ROS清除剂）预处理减少了ΔΨm的消散，但对AA诱导的p38 MAPK和JNK激活影响不显著。SB202190（p38 MAPK抑制剂）和SP600125（JNK抑制剂）减弱了线粒体去极化、Bcl-2/Bcl-xL的降解以及Bax的线粒体转位。特异性小干扰RNA（siRNA）转染证明p38α MAPK和JNK1参与调节Bcl-2家族蛋白。综上所述，我们的数据表明，AA对SK-N-SH细胞的细胞毒性是通过线粒体依赖性死亡途径介导的，由AA诱导的ROS生成和Ca(2+)诱发的p38α MAPK和JNK1激活引起。
