Objective: Reactive oxygen species(ROS) are involved in a variety of biological phenomena and serve both deleterious and beneficial roles. ROS quantification and assessment of reaction networks are desirable but diffi...Objective: Reactive oxygen species(ROS) are involved in a variety of biological phenomena and serve both deleterious and beneficial roles. ROS quantification and assessment of reaction networks are desirable but difficult because of their short half-life and high reactivity. Here, we describe a pro-oxidative model in a single human lung carcinoma SPC-A-1 cell that was created by application of extracellular H2O2 stimuli. Methods: Modified microfluidics and imaging techniques were used to determine O2·- levels and construct an O2^·- reaction network. To elucidate the consequences of increased O2^·- input, the mitochondria were given a central role in the oxidative stress mode, by manipulating mitochondria-interrelated cytosolic Ca2+ levels, mitochondrial Ca^2+ uptake, auto-amplification of intracellular ROS and the intrinsic apoptotic pathway. Results and conclusions: Results from a modified microchip demonstrated that 1 mmol/L H·-2 O2 induced a rapid increase in cellular O2 levels(>27 vs.>406 amol in 20 min), leading to increased cellular oxidizing power(evaluated by ROS levels) and decreased reducing power(evaluated by glutathione(GSH) levels). In addition, we examined the dynamics of cytosolic Ca^2+ and mitochondrial Ca^2+ by confocal laser scanning microscopy and confirmed that Ca^2+ stores in the endoplasmic reticulum were the primary source of H2O2-induced cytosolic Ca^2+ bursts. It is clear that mitochondria have pivotal roles in determining how exogenous oxidative stress affects cell fate. The stress response involves the transfer of Ca^2+ signals between organelles,ROS auto-amplification, mitochondrial dysfunction, and a caspase-dependent apoptotic pathway.展开更多
基金Project supported by the Zhejiang Provincial Natural Science Foundation of China(No.LY18H300002)the Medical Health Science and Technology Project of Zhejiang Provincial Health Commission(No.2019RC061/2019312897)+1 种基金the Zhejiang Provincial Natural Science Foundation of China(Nos.Y4110212 and LY19H090001)partly by the National Natural Science Foundation of China(Nos.81372301 and 81301113)
文摘Objective: Reactive oxygen species(ROS) are involved in a variety of biological phenomena and serve both deleterious and beneficial roles. ROS quantification and assessment of reaction networks are desirable but difficult because of their short half-life and high reactivity. Here, we describe a pro-oxidative model in a single human lung carcinoma SPC-A-1 cell that was created by application of extracellular H2O2 stimuli. Methods: Modified microfluidics and imaging techniques were used to determine O2·- levels and construct an O2^·- reaction network. To elucidate the consequences of increased O2^·- input, the mitochondria were given a central role in the oxidative stress mode, by manipulating mitochondria-interrelated cytosolic Ca2+ levels, mitochondrial Ca^2+ uptake, auto-amplification of intracellular ROS and the intrinsic apoptotic pathway. Results and conclusions: Results from a modified microchip demonstrated that 1 mmol/L H·-2 O2 induced a rapid increase in cellular O2 levels(>27 vs.>406 amol in 20 min), leading to increased cellular oxidizing power(evaluated by ROS levels) and decreased reducing power(evaluated by glutathione(GSH) levels). In addition, we examined the dynamics of cytosolic Ca^2+ and mitochondrial Ca^2+ by confocal laser scanning microscopy and confirmed that Ca^2+ stores in the endoplasmic reticulum were the primary source of H2O2-induced cytosolic Ca^2+ bursts. It is clear that mitochondria have pivotal roles in determining how exogenous oxidative stress affects cell fate. The stress response involves the transfer of Ca^2+ signals between organelles,ROS auto-amplification, mitochondrial dysfunction, and a caspase-dependent apoptotic pathway.
