南京农业大学学报2018，Vol.41Issue(3)：481-488,8.DOI:10.7685/jnau.201801042
ASAP1调节巨噬细胞对结核分枝杆菌吞噬作用的研究
ASAP1 involvement of the phagocytosis of Mycobacterium tuberculosis in macrophages
摘要
Abstract
[Objectives]Tuberculosis(TB)is one of the most important zoonosis all over the world. ASAP1 is associated with the susceptibility to TB in humans. The purpose of this research is to investigate the relationship between ASAP1 expression and Mtb infection,prove whether ASAP1 involves the phagocytosis of Mycobacterium tuberculosis(Mtb)in macrophages,and understand how ASAP1 affects the phagocytosis of Mtb in macrophages. [Methods]By using cell model,a THP-1 derived macrophage model expressing different levels of ASAP1 was generated,then a serials experiments were performed for analysis of affection of ASAP1 on phagocytosis of Mtb after infection of Mtb or BCG in vitro. BCG and Mtb H37Ra were used to infect macrophages,and ASAP1 expression was tested by using Western blotting and Real-time qPCR. ASAP1 specific siRNA was used to generate ASAP1 knockdown macrophages,ASAP1 expression lentiviral system was constructed to transduce THP-1 cell,and then it was made to differentiate into ASAP1 overexpression macrophages. Mtb H37Ra was used to infect macrophages expressing three different ASAP1 expression levels including knockdown,overexpression and normal expression of macrophages which were directly derived from THP-1 cell line,and then the intracellular Mtb number post-infection was counted by using colony forming unit(CFU)counting assays and image acquisition technique. [Results]This research first demonstrated that Mtb H37Ra infection significantly provoked higher ASAP1 expression in macrophages derived from THP-1 compared with BCG infection and non-infection control. ASAP1 overexpression macrophages(P<0.01)and knockdown macrophages(P<0.01)were generated to establish a macrophage cellular model for investigation into the affection of ASAP1 on phagocytosis of Mtb in macrophages. The CFU assays showed that ASAP1 overexpression macrophages loaded more bacteria(P<0.001),and ASAP1 knockdown macrophages loaded less bacteria compared with macrophages directly derived from wild type THP1 as a control(P<0.001).[Conclusions]Mtb infection provokes higher ASAP1 expression in macrophages,and a macrophage cellular model has been established for investigation into the mechanisms underling ASAP1 expression affecting on phagocytosis of Mtb in macrophages. Results also have shown ASAP1 expression has positive correlation with phagocytosis of Mtb,and overexpression of ASAP1 may introduce redundant phagocytosis of Mtb in macrophages, which may affect humans' susceptibility to TB.关键词
ASAP1/巨噬细胞/结核分枝杆菌/吞噬Key words
ASAP1/macrophages/Mycobacterium tuberculosis(Mtb)/phagocytosis分类
医药卫生引用本文复制引用
李鹏,崔佳,王丽清,吴长新..ASAP1调节巨噬细胞对结核分枝杆菌吞噬作用的研究[J].南京农业大学学报,2018,41(3):481-488,8.基金项目
国家重点研发计划子课题(2017YFD0500303) （2017YFD0500303）
