Effects Of Short-term Hyperoxia On Mitochondrial Reactive Oxygen Generation And Related Pathways Of Alveolar Type Ⅱ Cells

Objective:To explore the effect of short-term hyperoxia on the mitochondrial Ca²⁺/nicotinamide adenine dinucleotide（NAD⁺）/silencing information regulatory factor 3（SIRT3）/superoxide dismutase 2（SOD2）pathway and reactive oxygen species（ROS）production in alveolar typeⅡepithelial cells（AECⅡ）.Methods:The same generation cells of rat alveolar typeⅡepithelial cell line（RLE-6TN）were randomly divided into a control group,a hyperoxia group and a mitochondrial calcium channel antagonist group（antagonist group）.Cells in group H were placed in a hyperoxia box with an oxygen concentration of 90%for 4 hours,cells in group C were placed in a conventional cell incubator for 4 hours,cells in group HR were placed in an oxygen concentration of 90%after adding ruthenium red（2umol/L）4 hours in the high-oxygen box.Cell messenger RNA（messenger RNA,m RNA）was then extracted,and the levels of silencing information regulator（SIRT3）and superoxide dismutase 2（SOD2）m RNA were analyzed by real-time fluorescent quantitative PCR;the mitochondrial calcium ion specific fluorescent probe Rhod--2-AM detects cell mitochondrial Ca²⁺content;uses Coenzyme I amide adenine dinucleotide/reduced nicotinamide adenine dinucleotide（NAD⁺/NADH）content kit to measure the content and ratio of NAD⁺and NADH;use activity The oxygen detection kit detects the level of ROS.Results:Compared with the control group and the antagonist group,in the hyperoxia group,the cell NAD⁺content,NAD⁺/NADH ratio,SIRT3 and SOD2 m RNA expression levels were significantly reduced,and the cell mitochondrial Ca²⁺content,NADH content and ROS production were significantly increased,and the difference was statistically significant（P<0.05）.Compared with the control group,the cell Ca²⁺content,NAD⁺/NADH ratio and ROS content of the antagonist group were significantly reduced,and the difference was statistically significant（P<0.05）.There was no statistical difference in cell NAD⁺content,NADH content,SIRT3 and SOD2m RNA expression levels.Conclusions:Short-term hyperoxia can increase the production of ROS by the Ca²⁺/NAD⁺/SIRT3/SOD2 pathway in AECⅡ.