神经肽P物质对高氧损伤肺泡Ⅱ型上皮细胞氧化/抗氧化状态的影响

Effect of neuropeptide substance P on oxidative / antioxidant status of typeⅡalveolar epithelial cells in rats with lung injury induced by hyperoxia

目的探讨神经肽P物质(Substance P,SP)对高氧损伤早产大鼠肺泡Ⅱ型上皮细胞(Alveolar epithelial celltypeⅡ,AECⅡ)氧化/抗氧化状态的影响。方法将分离培养的原代早产大鼠AECⅡ随机分为4组:空气组(将细胞置5%CO2、21%O2的孵箱中)、高氧组(将细胞置5%CO2、95%O2的密闭氧仓中)、高氧SP组(细胞培养液中预先加入5×10-8mol/L的SP,再置高氧中)和高氧SP受体拮抗剂组(细胞培养液中预先加入5×10-8mol/L的SP和5×10-7mol/L的L703.606,再置高氧中),各组细胞培养24 h后,镜下观察细胞形态的变化;MTT法检测细胞的增殖活力;流式细胞术检测细胞周期;JC-1探针法检测细胞凋亡早期线粒体膜电位的变化;化学比色法检测细胞丙二醛(Malondialdehyde,MDA)含量、总抗氧化能力(Total antioxidative capacity,TAOC)及超氧化物歧化酶(Superoxidedismutase,SOD)的活力。结果与空气组比较,高氧组AECⅡ的增殖活力、S+G2期细胞比例、线粒体膜电位、SOD活力和TAOC均明显下降(P<0.01),MDA含量明显升高(P<0.01);与高氧组比较,高氧SP组AECⅡ的增殖活力、S+G2期细胞比例、线粒体膜电位、SOD活力和TAOC均明显升高(P<0.01),而MDA含量明显下降(P<0.01),SP受体拮抗剂可阻断该效应。结论 SP可明显减轻高氧导致的早产大鼠AECⅡ的氧化损伤,增强细胞的抗氧化能力,进而起到促进细胞增殖的作用。

Objective To investigate the effect of neuropeptide substance P（SP） on oxidative / antioxidant status of hyperoxia typeⅡ alveolar epithelial cells（AECⅡ） in premature rats with lung injury induced by hyperoxia.Methods Primary AECⅡ of premature rats were randomly divided into four groups.The cells in air group were incubated in an incubator containing 5% carbon dioxide and 21% oxygen,while those in hyperoxia group in sealed chamber containing 5% carbon dioxide and 95% oxygen,those in hyperoxia ＋ SP group were cultured in medium pre-added with 5 × 10^-8 mol / L SP and incubated in hyperoxia,and those in hyperoxia ＋ SP antagonist group cultured in medium pre-added with 5 × 10^-8 mol / L SP and 5 × 10^-7 mol / L L703.606,then incubated in hyperoxia.The cells in various groups were observed for morphology 24 h after incubation by microscopy,while determined for proliferation activity by MTT colorimetric assay,for cell cycle by flow cytometry,for mitochondrial membrane potential by JC-1 probe method,and for malondialdehyde（MDA） content,total antioxidative capacity（TAOC） and SOD activity by chemical colorimetry.Results As compared with those in air group,the proliferation activity,percentage in S ＋ G2 phases,mitochondrial membrane potential,SOD activity and TAOC of AECⅡ in air group decreased significantly（P 0.01）,while MDA content increased significantly（P 0.01）.However,compared with those in hyperoxia group,the proliferation activity,percentage in S ＋ G2 phases,mitochondrial membrane potential,SOD activity and TAOC of AECⅡ in hyperoxia ＋ SP group increased significantly（P 0.01）,while the MDA content decreased significantly（P 0.01）,which were blocked by SP antagonist.Conclusion SP relived the injury induced by hyperoxia,enhanced the ability of oxidation resistance and promoted the proliferation of AECⅡ in premature rats.