TNF-α对大鼠肺泡Ⅱ型上皮细胞增殖及凋亡的影响

Effects of TNF-αon Proliferation and Apoptosis of Alveolar TypeⅡEpithelial Cells in Rats

目的探究TNF-α对大鼠肺泡Ⅱ型上皮细胞(alveolar typeⅡepithelial cells,AEC-Ⅱ)增殖及凋亡的影响。方法体外培养大鼠AEC-Ⅱ,透射电镜鉴定大鼠肺泡Ⅱ型上皮细胞后,将细胞分为对照组和TNF-α组(5、10、20、40 ng/mL),分别干预24、48、72 h。MTT比色法检测各组细胞增殖抑制率,AnnexinⅤ/PI双染色联合流式细胞仪检测各组细胞的早期凋亡率,ELISA法检测各组细胞上清液中IL-6的含量。结果透射电镜下可见细胞内含有大量的特征性板层小体。MTT法示TNF-α组ACE-Ⅱ的增殖抑制率均高于对照组(均P<0.05),且随着剂量的增大,作用时间延长,细胞的增殖抑制率越高。流式细胞术检测结果可见TNF-α组细胞凋亡率明显高于对照组(均P<0.05),且呈剂量-时间依赖关系。ELISA结果示40 ng/mL TNF-α组细胞上清液中IL-6含量高于对照组及5 ng/mL TNF-α组(P<0.05),而各作用时点组间两两比较,差异均无统计学意义(均P>0.05)。结论TNF-α通过诱导AEC-Ⅱ的凋亡导致肺损伤,并通过抑制其增殖阻碍损伤肺组织的自我修复;同时,高浓度TNF-α可诱导AEC-Ⅱ合成分泌IL-6水平的上调,加重肺部的炎症反应,加重肺损伤。

Objective To investigate the effects of TNF-αon the proliferation and apoptosis of alveolar typeⅡepithelial cells in rats.Methods Rat AEC-Ⅱwere cultured in vitro.Transmission electron microscope was used to identify AEC-Ⅱ.Cells were divided into control group and TNF-αgroups(5,10,20,40 ng/mL)for 24,48 and 72 h.Cell proliferation inhibition was detected by MTT assay.The early apoptosis rate of each group was detected by flow cytometry.The content of IL-6 in supernatant was detected by ELISA.Results MTT showed that the inhibitory rate of ACE-Ⅱin TNF-αgroups was higher than that in control group(P<0.05),and it increased with the dosage and time of TNF-α.Flow cytometry methods showed that the apoptosis rate in TNF-αgroups was significantly higher than that of the control group in a dose-time dependent manner(P<0.05).ELISA method showed that the content of IL-6 in the cell supernatant of TNF-α40 ng/mL group was higher than that of the control group and the TNF-α5 ng/mL group(P<0.05),but there was no statistical difference between each two groups at different time points(P>0.05).Conclusion TNF-αinduces lung injury by inducing cell apoptosis,and inhibits the proliferation of AEC-Ⅱ,and high concentration of TNF-αincreases IL-6.It aggravates the inflammatory reaction of lung and lung injury.