日本血吸虫感染的小鼠肺脏组织CD69~+TRM细胞免疫学特性的变化
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摘要
目的探究日本血吸虫感染的小鼠肺脏CD69~+组织定居记忆T细胞(TRM)的表型和功能的变化。方法使用血吸虫感染C57BL/6小鼠,6周后分离感染组及正常对照组小鼠的肺脏组织,制备石蜡切片,使用HE染色,观察肺部病变情况;制备单细胞悬液,用流式细胞术检测CD4~+和CD8~+的CD69~+TRM的含量;使用细胞表面染色方法,观察CD69~+TRM细胞表面CD103、CD62L和CD107a分子的表达情况;使用细胞内细胞因子染色法,检测CD69~+TRM细胞白细胞介素-4(IL-4)、干扰素-γ(IFN-γ)、IL-9和IL-10的分泌情况。结果日本血吸虫感染小鼠肺脏组织出现明显肉芽肿病变;流式细胞术检测结果显示,在CD4~+T和CD8~+T细胞中,感染组CD69的表达比例均高于正常组[(19.02±2.87)%vs.(4.29±0.21)%,(7.61±1.99)%vs.(2.46±0.49)%],差异有统计学意义(P<0.05)。在CD4~+T细胞中,CD62L和CD107a分子在感染组小鼠CD69~+细胞的表达比例低于正常组[(6.41±0.47)%vs.(23.95±1.63)%,(30.19±2.70)%vs.(48.15±2.75)%],差异均有统计学意义(P<0.05)。在CD8~+T细胞中,感染组小鼠CD69~+细胞表面CD103和CD62L分子的表达比例远低于正常组小鼠[(19.12±0.41)%vs.(44.79±3.64)%,(38.89±4.59)%vs.(58.69±2.12)%],差异均有统计学意义(P<0.05)。在CD4~+T细胞中,感染组CD69~+T细胞相比正常组表达较高水平的IL-4[(27.25±0.88)%vs.(14.57±1.36)%],差异有统计学意义(P<0.05)。在CD8~+T细胞中,感染组CD69~+T细胞相比正常组表达较高水平的IFN-γ[(45.08±4.25)%vs.(2.75±0.07)%],差异有统计学意义(P<0.05)。结论小鼠肺脏组织中存在着一群表达CD69的TRM细胞,在日本血吸虫感染诱导的肺部肉芽肿病变过程中发挥了重要的作用。
Objective To explore the phenotypic and functional changes of CD69~+tissue-resident memory T cells(TRM)in the lung of Schistosoma japonicum infected mice.MethodsC57BL/6 mice were infected by Schistosoma japonicum.The lungs of both infected and normal control mice were isolated 6 weeks later.Paraffin sections were prepared,and HE staining was done to observe lung lesions.Single cell suspension solution was also prepared,and the contents of CD4~+CD69~+and CD8~+CD69~+TRM were detected by flow cytometry.The expressions of CD103,CD62L and CD107a on the surface of CD69~+TRM cells were observed by cell surface staining.The expressions of interleukin-4(IL-4),interferon-γ(IFN-γ),IL-9 and IL-10 on CD69~+TRM cells were detected by intracellular cytokine staining.ResultsObvious granuloma could be seen in the lung tissues of mice infected by Schistosoma japonicum.FACs results showed that the proportion of CD69~+cells in CD4~+and CD8~+T lymphocytes in the lung tissues of the infected group were significantly higher than that of control group[(19.02±2.87)%vs.(4.29±0.21)%,(7.61±1.99)%vs.(2.46±0.49)%,P<0.05].In the population of CD4~+T cells,the percentage of CD62L and CD107a cells in CD69~+cells of the infected group was lower than those in the normal group[(6.41±0.47)%vs.(23.95±1.63)%,(30.19±2.70)%vs.(48.15±2.75)%,P<0.05].In the CD8~+T cells,the percentage of CD103 and CD62L cells in CD69~+cells in the infected group was much lower than those in the normal group[(19.12±0.41)%vs.(44.79±3.64)%,(38.89±4.59)%vs.(58.69±2.12)%,P<0.05].In the population of CD4~+T cells,CD69~+T cells from the infected group expressed higher levels of IL-4 than the normal group[(27.25±0.88)%vs.(14.57±1.36)%,P<0.05].Whereas the CD8~+T,CD69~+T cells infected group expressed higher levels of IFN-γthan the normal group[(45.08±4.25)%vs.(2.75±0.07)%,P<0.05].ConclusionA group of T cells expressing CD69 existed in the lung tissue of mice,which played an important role in the process of pulmonary granuloma induced by Schistosoma japonicum infection.
Objective To explore the phenotypic and functional changes of CD69~+tissue-resident memory T cells(TRM)in the lung of Schistosoma japonicum infected mice.MethodsC57BL/6 mice were infected by Schistosoma japonicum.The lungs of both infected and normal control mice were isolated 6 weeks later.Paraffin sections were prepared,and HE staining was done to observe lung lesions.Single cell suspension solution was also prepared,and the contents of CD4~+CD69~+and CD8~+CD69~+TRM were detected by flow cytometry.The expressions of CD103,CD62L and CD107a on the surface of CD69~+TRM cells were observed by cell surface staining.The expressions of interleukin-4(IL-4),interferon-γ(IFN-γ),IL-9 and IL-10 on CD69~+TRM cells were detected by intracellular cytokine staining.ResultsObvious granuloma could be seen in the lung tissues of mice infected by Schistosoma japonicum.FACs results showed that the proportion of CD69~+cells in CD4~+and CD8~+T lymphocytes in the lung tissues of the infected group were significantly higher than that of control group[(19.02±2.87)%vs.(4.29±0.21)%,(7.61±1.99)%vs.(2.46±0.49)%,P<0.05].In the population of CD4~+T cells,the percentage of CD62L and CD107a cells in CD69~+cells of the infected group was lower than those in the normal group[(6.41±0.47)%vs.(23.95±1.63)%,(30.19±2.70)%vs.(48.15±2.75)%,P<0.05].In the CD8~+T cells,the percentage of CD103 and CD62L cells in CD69~+cells in the infected group was much lower than those in the normal group[(19.12±0.41)%vs.(44.79±3.64)%,(38.89±4.59)%vs.(58.69±2.12)%,P<0.05].In the population of CD4~+T cells,CD69~+T cells from the infected group expressed higher levels of IL-4 than the normal group[(27.25±0.88)%vs.(14.57±1.36)%,P<0.05].Whereas the CD8~+T,CD69~+T cells infected group expressed higher levels of IFN-γthan the normal group[(45.08±4.25)%vs.(2.75±0.07)%,P<0.05].ConclusionA group of T cells expressing CD69 existed in the lung tissue of mice,which played an important role in the process of pulmonary granuloma induced by Schistosoma japonicum infection.
引文
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[12]袁祥.组织原位记忆T细胞在系统性红斑狼疮的表达及与皮损的相关性研究.合肥:安徽医科大学,2017.
[13]Thome JJ,Farber DL.Emerging concepts in tissue-resident Tcells:lessons from humans[J].Trends Immunol,2015,36(7):428-435.
[14]Fan X,Rudensky AY.Hallmarks of tissue-resident lymphocytes[J].Cell,2016,164(6):1198-1211.
[15]Slütter B,Harty JT.Instructing the instructor:tissue-resident Tcells activate innate immunity[J].Cell Host Microbe,2014,16(4):421-423.
[16]Schenkel JM,Masopust D.Tissue-resident memory T cells[J].Immunity,2014,41(6):886-897.
[2]Olveda DU,Olveda RM,McManus DP,et al.The chronic enteropathogenic disease schistosomiasis[J].Int J Infect Dis,2014,28:193-203.
[3]曲佳乐,冯源发,金晨曦,等.日本血吸虫感染C57BL/6小鼠脾脏调节性B细胞的检测[J].热带医学杂志,2018,18(3):289-293.
[4]王静,王一任,王克强.一种CD3+T细胞细胞内CD69分子的检测方法[J].泰山医学院学报,2017,38(12):1337-1339.
[5]Mueller SN,Gebhardt T,Carbone FR,et al.Memory T cell subsets,migration patterns,and tissue residence[J].Annu Rev Immunol,2013,31:137-161.
[6]Gebhardt T,Mueller SN,Heath WR,et al.Peripheral tissue surveillance and residency by memory T cells[J].Trends Immunol,2013,34(1):27-32.
[7]Carbone FR,Mackay LK,Heath WR,et al.Distinct resident and recirculating memory T cell subsets in non-lymphoid tissues[J].Curr Opin Immunol,2013,25(3):329-333.
[8]Mueller SN,Mackay LK.Tissue-resident memory T cells:local specialists in immune defence[J].Nat Rev Immunol,2016,16(2):79-89.
[9]余思菲,吴长有.组织定居记忆性T细胞的免疫学特征研究进展[J].中国免疫学杂志,2017,33(7):1093-1100.
[10]康双朋,吴琼丽,吴长有.子宫组织定居记忆T细胞的免疫学特征[J].免疫学杂志,2017,33(11):937-942.
[11]陈虹虹,张仁利,朱兴全.日本血吸虫病免疫病理的研究进展[J].中国人兽共患病学报,2006,22(1):75-77,80.
[12]袁祥.组织原位记忆T细胞在系统性红斑狼疮的表达及与皮损的相关性研究.合肥:安徽医科大学,2017.
[13]Thome JJ,Farber DL.Emerging concepts in tissue-resident Tcells:lessons from humans[J].Trends Immunol,2015,36(7):428-435.
[14]Fan X,Rudensky AY.Hallmarks of tissue-resident lymphocytes[J].Cell,2016,164(6):1198-1211.
[15]Slütter B,Harty JT.Instructing the instructor:tissue-resident Tcells activate innate immunity[J].Cell Host Microbe,2014,16(4):421-423.
[16]Schenkel JM,Masopust D.Tissue-resident memory T cells[J].Immunity,2014,41(6):886-897.
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