摘要
目的探讨短时间高氧对肺泡Ⅱ型上皮细胞(AECⅡ)线粒体Ca2+/烟酰胺腺嘌呤二核苷酸(NAD+)/沉默信息调节因子3(SIRT3)/超氧化物歧化酶2(SOD2)通路及活性氧的影响。方法将RLE-6TN细胞株细胞分为对照组、高氧组及线粒体钙通道拮抗剂组(拮抗剂组)。对照组细胞置于常规细胞培养箱中,高氧组细胞置于氧浓度为90%的培养箱中,拮抗剂组细胞加入钌红(2μmol/L)后置于氧浓度为90%的培养箱中,各组均持续培养4 h。随后,对各组细胞线粒体内Ca2+、活性氧、NAD+、还原型烟酰胺腺嘌呤二核苷酸(NADH)含量进行检测,并计算NAD+/NADH比值;同时,采用实时荧光定量PCR检测SIRT3和SOD2信使RNA(m RNA)水平。结果各组间细胞线粒体内Ca2+、活性氧、NAD+、NADH、NAD+/NADH比值及SIRT3 m RNA、SOD2 m RNA表达水平的比较,差异均有统计学意义(F=183.500、135.900、32.140、51.520、128.300、59.970、45.020,P均<0.001)。且与对照组及拮抗剂组比较,高氧组细胞线粒体内Ca2+[(19.5±0.8)、(17.2±0.7)、(24.3±0.3)nmol/L]、活性氧[(491±9)、(480±5)、(530±6)相对荧光单位]及NADH[(0.85±0.03)、(0.87±0.04)、(1.06±0.06)nmol/104cells]含量均明显升高,而NAD+含量[(3.30±0.12)、(3.24±0.14)、(2.58±0.29)nmol/104cells]、NAD+/NADH比值[(3.89±0.15)、(3.71±0.15)、(2.44±0.27)]、SIRT3 mRNA[(1.01±0.11)、(0.96±0.08)、(0.45±0.09)]及SOD2 m RNA[(1.01±0.14)、(1.05±0.11)、(0.48±0.10)]表达水平均显著降低(P均<0.05)。结论短时间高氧可通过AECⅡ线粒体内Ca2+/NAD+/SIRT3/SOD2通路导致活性氧蓄积。
Objective To explore the effect of short-term hyperoxia on the mitochondrial Ca2+/nicotinamide adenine dinucleotide(NAD+)/silence information regulator 3(SIRT3)/superoxide dismutase 2(SOD2)pathway and reactive oxygen species(ROS)in alveolar epithelial typeⅡcells(AECⅡ).Methods The RLE-6 TN cells were randomly divided into a control group,a hyperoxia group and a mitochondrial calcium channel antagonist group(antagonist group).Cells in the control group were placed in a conventional cell culture box,cells in the hyperoxia group were placed in a box with an oxygen concentration of 90%,and cells in the antagonist group were given ruthenium red(2μmol/L)and then placed in a box with an oxygen concentration of90%.The cells in all groups were cultured continuously for 4 h.Subsequently,the mitochondrial Ca2+,ROS,NAD+and reduced nicotinamide adenine dinucleotide(NADH)contents were measured,and the NAD+/NADH level was calculated.The messenger RNA(mRNA)levels of SIRT3 and SOD2 were detected by real-time fluorescent quantitative PCR.Results The mitochondrial Ca2+,ROS,NAD+,NADH,NAD+/NADH,SIRT3 mRNA and SOD2 mRNA all showed significant differences among the three groups(F=183.500,135.900,32.140,51.520,128.300,59.970,45.020;all P<0.001).Compared with the control group and antagonist group,the contents of mitochondrial Ca2+[(19.5±0.8),(17.2±0.7),(24.3±0.3)nmol/L],ROS[(491±9),(480±5),(530±6)relative fluorescence units]and NADH[(0.85±0.03),(0.87±0.04),(1.06±0.06)nmol/104 cells]increased obviously,and the NAD+content[(3.30±0.12),(3.24±0.14),(2.58±0.29)nmol/104 cells],NAD+/NADH[(3.89±0.15),(3.71±0.15),(2.44±0.27)],SIRT3 mRNA[(1.01±0.11),(0.96±0.08),(0.45±0.09)]and SOD2 mRNA[(1.01±0.14),(1.05±0.11),(0.48±0.10)]decreased markedly in the hyperoxia group(all P<0.05).Conclusion Short-term hyperoxia can increase the production of ROS by the Ca2+/NAD+/SIRT3/SOD2 pathway in AECⅡ.
作者
赵彦琴
李玉兰
程晓彤
李春兰
殷玉江
Zhao Yanqin;Li Yulan;Cheng Xiaotong;Li Chunlan;Yin Yujiang(The First School of Clinical Medicine of Lanzhou University,Lanzhou 730000,China;Department of Anesthesiology,the First Hospital of Lanzhou University,Lanzhou 730000,China)
出处
《中华危重症医学杂志（电子版）》
CAS
CSCD
2020年第6期427-431,共5页
Chinese Journal of Critical Care Medicine:Electronic Edition
基金
甘肃省自然科学基金(17JR5RA262)。
关键词
高氧
线粒体
肺泡Ⅱ型上皮细胞
活性氧
Hyperoxia
Mitochondria
Alveolar epithelial typeⅡcells
Reactive oxygen species
