致病菌金黄色葡萄球菌及肺炎链球菌代谢产物及菌体成分对人呼吸道上皮细胞结构与功能的影响  

Effects of pathogenic Staphylococcus aureus and Streptococcus pneumoniae metabolites and bacterial components on structure and function of human airway epithelial cell

作　　者：张鑫[1] 秦啸峰[1] 黄琼[1] 杨义然 杜晓楠[1] 李琴 吕喆[1] 陈彦[1] 王炜[1] 孙英[1] ZHANG Xin;QIN Xiao-feng;HUANG Qiong;YANG Yi-ran;DU Xiao-nan;LI Qin;LV Zhe;CHEN Yan;WANG Wei;SUN Ying(Department of Immunology,School of Basic Medical Sciences,Capital Medical University,Beijing 100069,China)

机构地区：[1]首都医科大学基础医学院免疫学系

出　　处：《中华临床免疫和变态反应杂志》2019年第6期492-500,共9页Chinese Journal of Allergy & Clinical Immunology

基　　金：国家自然科学基金面上项目(81770049);北京市自然科学基金(7192023)~~

摘　　要：目的探究呼吸系统常见致病菌金黄色葡萄球菌(Staphylococcus aureus,S. aureus)、肺炎链球菌(Streptococcus pneumonia,S. pneumonia)不同成分对人呼吸道肺泡Ⅱ型上皮细胞(A549细胞)结构与功能的影响。方法 A549细胞分别与S.aureus、S. pneumonia细菌培养上清、细菌裂解物及灭活菌体共培养,采用CCK8方法检测上述不同细菌成分对A549细胞增殖功能的影响,采用流式细胞术(flow cytometry)检测不同刺激物诱导细胞发生凋亡与坏死的作用,采用酶联免疫吸附试验(ELISA)测定其对炎症细胞因子白细胞介素6、白细胞介素8(IL-6、IL-8)分泌的影响,利用Western blot及细胞免疫荧光染色方法探究不同刺激成分对A549细胞紧密连接蛋白ZO-1、Claudin1、Occludin表达的影响。结果两种细菌培养上清及S. pneumonia裂解物可显著抑制A549细胞增殖并诱导细胞出现凋亡和坏死现象(P<0.05);两种细菌培养上清及细菌裂解物均可促进上皮细胞分泌IL-6,且两种细菌培养上清及S. pneumonia裂解物亦可促进上皮细胞分泌IL-8;两种细菌培养上清及各成分,仅S. aureus灭活菌体可降低紧密连接蛋白ZO-1的表达,但其对另外两种紧密连接蛋白Claudin1和Occludin的表达无显著影响。结论两种细菌感染过程中,不同细菌及细菌的不同成分对上皮细胞作用并不完全相同。细菌培养上清和细菌裂解物主要影响上皮细胞增殖、炎症因子分泌等功能,S. aureus灭活菌体可降低紧密连接蛋白ZO-1的表达进而影响上皮细胞间紧密连接的完整性。Objective To investigate the effects of different components of the common respiratory pathogenic bacteria Staphylococcus aureus(S. aureus) and Streptococcus pneumonia(S. pneumonia) on structure and function of human airway epithelial cell(A549). Methods A549 cells were cultured in the presences of S. aureus/S. pneumonia culture supernatants, bacterial lysates, and inactivated bacteria respectively. The effects of different components on A549 cells proliferation were detected by CCK8. Flow cytometry was used to explore the apoptosis and necrosis of cultured A549 cells. Enzyme-linked immunosorbent assay(ELISA) was used to measure the secretion of inflammatory cytokines interleukin-6(IL-6) and interleukin-8(IL-8). Expression of cell-cell tight junction associated proteins ZO-1, Claudin1 and Occludin by A549 cells was analyzed by using Western Blot and immunofluorescence staining after different stimuli mentioned above. Results A549 cells proliferation was significantly inhibited and apoptosis and necrosis of A549 cells was induced by supernatants of two bacterial cultures and S. pneumonia lysates(P<0.05). The supernatants and bacterial lysates enhanced IL-6 secretion of A549 epithelial cells, while IL-8 increased only after stimulation with the supernatants and S. pneumonia lysates. Inactivated S. aureus, but not S. aureus and S. pneumonia superatants or other bacterial compononts could reduce the expression of tight junction associated protein ZO-1, whereas it had no significant effect on the expression of the other two tight junction associated proteins Occludin and Claudin1.Conclusions The effects of cultured supernatants and different components on epithelial cell structure and function are variable between S. aureus and S. pneumonia. Bacterial culture supernatants, bacterial lysates mainly affect epithelial cell proliferation and secretion of inflammatory cytokines. Inactivated S. aureus bacteria can also reduce the expression of tight junction associated protein ZO-1, which may further affect the integrity of

关 键 词：金黄色葡萄球菌 肺炎链球菌 细菌培养上清 细菌裂解物 灭活菌体 上皮细胞 

分 类 号：R378.11[医药卫生—病原生物学]