In situ real-time chemiluminescence imaging of reactive oxygen species formation from cardiomyocytes.
In situ real-time chemiluminescence imaging of reactive oxygen species formation from cardiomyocytes.
复制标题
心肌细胞活性氧形成的原位实时化学发光成像。
DOI：
10.1155/2008/941729
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发表时间：
2008
影响因子：
7.6
通讯作者：
Wang,Ge
中科院分区：
文献类型：
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作者：
Li,Yunbo;Shen,Haiou;Zhu,Hong;Trush,MichaelA;Jiang,Ming;Wang,Ge
应用高灵敏度化学发光（CL）成像技术对培养的大鼠H9c2心肌细胞单层中ROS的原位形成进行了研究。H9c2细胞与鲁米诺和辣根过氧化物酶（HRP）培养后，通过一种创新的成像系统检测光子发射，表明心肌细胞形成了ROS。在培养的H9c2细胞中添加苯并(a)芘‐1,6‐醌（BPQ），通过CL成像检测到ROS的形成增加了4‐5倍。在培养的H9c2细胞中，组成型和BPQ刺激的CL反应均持续长达1小时。在超氧化物歧化酶和过氧化氢酶的存在下，CL反应被完全消除，表明超氧化物和过氧化氢（H2O2）的主要参与。与BPQ介导的氧化还原循环相反，抗霉素A或鱼藤酮阻断线粒体电子传递链对培养的H9c2细胞ROS形成的影响微乎其微。在培养的H9c2细胞中，3H‐1,2‐二噻吩‐3‐硫酮上调细胞抗氧化剂对超氧化物和h2o2的解毒作用，显著抑制了组成型和BPQ‐增强型ROS的形成。综上所述，我们证明了CL成像在培养的心肌细胞中对ROS的敏感检测。
We have applied the highly sensitive chemiluminescence (CL) imaging technique to investigate the in situ ROS formation in cultured monolayers of rat H9c2 cardiomyocytes. Photon emission was detected via an innovative imaging system after incubation of H9c2 cells in culture with luminol and horseradish peroxidase (HRP), suggesting constitutive formation of ROS by the cardiomyocytes. Addition of benzo(a)pyrene‐1,6‐quinone (BPQ) to cultured H9c2 cells resulted in a 4‐5‐fold increase in the formation of ROS, as detected by the CL imaging. Both constitutive and BPQ‐stimulated CL responses in cultured H9c2 cells were sustained for up to 1 hour. The CL responses were completely abolished in the presence of superoxide dismutase and catalase, suggesting the primary involvement of superoxide and hydrogen peroxide (H2O2). In contrast to BPQ‐mediated redox cycling, blockage of mitochondrial electron transport chain by either antimycin A or rotenone exerted marginal effects on the ROS formation by cultured H9c2 cells. Upregulation of cellular antioxidants for detoxifying both superoxide and H2O2by 3H‐1,2‐dithiole‐3‐thione resulted in marked inhibition of both constitutive and BPQ‐augmented ROS formation in cultured H9c2 cells. Taken together, we demonstrate the sensitive detection of ROS by CL imaging in cultured cardiomyocytes.