结核杆菌耐热抗原对A549细胞分泌SP-B和凋亡的影响

摘要

Objective To investigate secreting pulmonary surfactant-associated protein B (SP-B) and apoptosis in human lung adenocarcinoma cell line (A549) cells,treated with heat-resistant antigen of Mycobacterium tuberculosis (Mtb-HAg).Methods Different concentrations of Mtb-HAg were used to culture A549 cells for 24 h and 48 h respectively,and the blank control groups(control group 1 and control group 2) and positive control groups [lipopolysaccharide (LPS) group and curcumin group] were set up.SP-B in the culture supernatant,was assessed by ELISA in control group 1,LPS group and experimental group 1 (A549 cells were respectively induced by 2 μg/ml,3 μg/ml Mtb-HAg to culture for 24 h and 48 h).For control group 1,LPS group and experimental group 1,the relative expression of SFTPB gene was quantified with quantitative real-time fluorescence quantitative PCR (qRT-PCR).The apoptosis rate of control group 2,curcumin group and experimental group 2(A549 cells were respectively induced by 2 μg/ml,3 μg/ml Mtb-HAg to culture for 24 h) in A549 cells were detected by flow cytometry.Results SP-B expression in the experimental group 1 was significantly lower than the control group 1,the difference was statistically significant (P ＜ 0.05).The difference of SP-B expression in the experimental group 1 was not obvious with the prolongation of the same concentration.At the same incubation time,the expression of SFTPB in the experimental group 1 decreased obviously with the increasing concentration of Mtb-HAg,the difference was statistically significant (P ＜ 0.05).The change with time was not significant.The apoptosis rate of curcumin group and experimental group 2 were significantly higher than that in control group 2 in A549 cells (P ＜ 0.05).Conclusion Mtb-HAg inhibited the expression of SP-B in A549 cells significantly,and induced apoptosis of A549 cells.%目的 探讨结核杆菌耐热抗原(Mtb-HAg)刺激诱导人肺腺癌细胞系(A549)细胞后对其肺泡表面活性物质相关蛋白B(SP-B)的分泌和凋亡的影响.方法 用不同浓度的Mtb-HAg分别刺激诱导A549细胞,相同条件分别培养24、48 h,同时设置空白对照组(对照组1和对照组2)和阳性对照组[脂多糖(LPS)组和姜黄素组].运用ELISA法检测对照组1、LPS组和实验组1(2、3μg/ml Mtb-HAg分别诱导A549细胞培养24 h和48 h)培养上清液中的SP-B表达量;使用实时荧光定量PCR法检测对照组1、LPS组和实验组1中基因SFTPB的相对表达量;采用流式细胞技术检测对照组2、姜黄素组和实验组2(2、3 μg/ml Mtb-HAg分别诱导A549细胞培养24 h)中A549细胞的凋亡率.结果 刺激诱导相同时间,实验组1的SP-B表达量低于对照组1,差异有统计学意义(P ＜0.05);相同浓度刺激诱导随时间的延长,实验组1中SP-B表达量降低的不显著.相同培养时间,随着Mtb-HAg浓度的增加实验组1表达SP-B的基因SFTPB的相对表达量显著低于对照组1,差异有统计学意义(P＜0.05);而在相同有效刺激浓度下,其相对表达量随作用时间变化不显著.姜黄素组和实验组2中的A549细胞凋亡率显著高于对照组2(P ＜0.05).结论 Mtb-HAg对A549细胞表达SP-B的抑制作用明显,并能诱导A549细胞发生凋亡.