Characteristics and Functional Analysis of CD4+ T Lymphocyte Subsets in Mice Infected with Streptococcus pneumoniae
-
摘要:
目的 分析肺炎链球菌（Streptococcus pneumoniae，S.P.）感染后小鼠脾和肺组织中CD4+ T细胞亚群水平和功能，探索S.P.感染后的免疫调控机制。 方法 通过流式细胞术检测正常组（n = 4）、S.P.感染12 h（n = 4）和24 h（n = 4）后小鼠脾组织中CD4+ T细胞亚群Th1、Th2、Th17和Treg细胞比例并通过H&E染色检测各组小鼠肺组织病理特征，分析S.P.感染2 d和5 d后小鼠肺组织差异基因集及功能变化并预测免疫细胞丰度。 结果 小鼠感染S.P.后肺组织出现明显的炎症性病理特征。S.P.感染后小鼠脾脏组织Th1和Treg细胞比例逐渐升高，感染24 h后Th1和Treg细胞比例明显较对照组升高（P < 0.05），而S.P.感染5 d后仅Treg细胞比例明显高于对照组（P < 0.05）。功能分析发现S.P.感染2 d后肺组织中产生IL6、IL10和IL4/IL13的信号通路异常激活，而感染5 d后IL-2和IL-6/TGF-β通路异常富集。 结论 Treg和Th1细胞是小鼠S.P.感染后重要的免疫调节细胞，干预IL-10等介导的Treg细胞功能和IL-2介导的Th1细胞功能可改善S.P.感染后的免疫功能。 Abstract:Objective To analyze the levels and functions of CD4+ T cell subsets in mouse spleen and lung tissues after Streptococcus pneumoniae （S.P.） infection, and to explore the immune regulatory mechanisms S.P. infection. Methods Flow cytometry was used to detect the proportions of CD4+ T cell subsets （Th1, Th2, Th17, and Treg cells） in mouse spleen tissues from control group （n = 4） , S.P. infection at 12 h （n = 4）, and 24 h （n = 4）. H&E staining was used to examine lung tissue pathological characteristics. Differential gene sets and functional changes in lung tissues were analyzed after S.P. infection for 2 and 5 days, and immune cell abundance was predicted. Results Significant inflammatory pathological features were observed in the lung tissues of mice after S.P. infection. The proportions of Th1 and Treg cells in the spleen tissues gradually increased after S.P. infection, with Th1 and Treg cell proportions significantly higher than the control group at 24 h post-infection （P < 0.05）. At 5 d post-infection, only Treg cell proportion was significantly higher than the control group （P < 0.05）. Functional analysis revealed abnormal activation of IL6, IL10, and IL4/IL13 signaling pathways 2 days after infection, and abnormal enrichment of IL-2 and IL-6/TGF-β pathways 5 days after infection. Conclusion Treg and Th1 cells are key immune regulatory cells in mice following S.P. infection. Modulating Treg cell function mediated by IL-10 and Th1 cell function mediated by IL-2 can improve immune responses after S.P. infection. -
Key words:
- Streptococcus pneumoniae /
- Pneumonia /
- CD4+ T cell /
- Treg cell /
- Th1 cell /
- Immunoregulation
-
图 1 S.P.感染后小鼠肺组织病理变化（左10×，右40×）
A：正常小鼠肺组织；B：S.P.感染12 h后小鼠肺组织；C：S.P.感染24 h后小鼠肺组织。橙色箭头为肺泡壁断裂；黑色箭头为淋巴细胞；绿色箭头为嗜酸性粒细胞；蓝色箭头为中性粒细胞；I～IV为肺泡壁厚度。
Figure 1. Pathological changes in lung tissues of mice after S.P. infection （left 10×，right 40×）
图 2 S.P.感染后小鼠脾脏CD4+ T细胞亚型分析（$\bar x \pm s $，n = 4）
A：流式检测CD4+ T细胞亚型的流程；B：流式检测Th1、Th2、Th17和Treg细胞亚群的结果示例；C：各组Th1、Th2、Th17和Treg细胞比例统计分析；*P < 0.05。
Figure 2. Analysis of CD4+ T cell subsets in the spleen of mice after S.P. infection （$\bar x \pm s $，n = 4）
图 3 S.P.感染2 d后小鼠肺组织细胞差异基因及功能分析
A：对照组与S.P.感染2 d后小鼠肺组织中差异性表达基因的火山图；B：差异性表达基因的蛋白质互作网络，红色为SP组上调表达，蓝色为SP组下调表达，颜色越深标识差异越显著；圆点大小表示相互作用节点，点越大表示相互作用节点越多；线段灰度值表示相互作用可能性，颜色越深表示相互作用可能性越大；C：差异性表达基因功能集，圆点代表信号通路，同种颜色表示功能相似/相关的信号通路；D：差异性表达基因相关信号通路的富集气泡图，圆点大小代表富集到该信号通路中的差异基因数。
Figure 3. Differential gene expression and functional analysis of lung tissue cells in mice two days after S.P. infection
图 4 S.P.感染5 d后小鼠肺组织细胞功能及免疫细胞浸润分析
A：GAEA分析小鼠肺组织细胞功能改变；NES：校正后的归一化的ES值（富集评分）；|NES| > 1为功能富集标准；NOM P：即P-value，是对富集得分ES的统计学分析；Exp.为基因表达水平，红色为上调，蓝色为下调表达，颜色越深表示差异变化越大；B：小鼠肺组织中免疫细胞比例分析；*P < 0.05。
Figure 4. Functional analysis and immune cell infiltration in lung tissue of mice 5 days after S.P. infection
-
[1] Li L,Ma J,Yu Z,et al. Epidemiological characteristics and antibiotic resistance mechanisms of streptococcus pneumoniae: An updated review[J]. Microbiol Res,2023,266(1):127221. [2] Marquart M E. Pathogenicity and virulence of streptococcus pneumoniae: Cutting to the chase on proteases[J]. Virulence,2021,12(1):766-787. doi: 10.1080/21505594.2021.1889812 [3] Ramos-Sevillano E,Ercoli G,Brown J S. Mechanisms of naturally acquired immunity to streptococcus pneumoniae[J]. Front Immunol,2019,10(1):358. [4] Silva M J A,Ribeiro L R,Lima K V B,et al. Adaptive immunity to SARS-CoV-2 infection: A systematic review[J]. Front Immunol,2022,13(1):1001198. [5] Chatzileontiadou D S M,Sloane H,Nguyen A T,et al. The many maces of CD4(+) T cells: Immunological and structural characteristics[J]. Int J Mol Sci,2020,22(1):73. doi: 10.3390/ijms22010073 [6] Drescher H K,Bartsch L M,Weiskirchen S,et al. Intrahepatic T(H)17/T(Reg) cells in homeostasis and disease-It's all about the balance[J]. Front Pharmacol,2020,11(1):588436. [7] Pissas G,Tziastoudi M,Poulianiti C,et al. In human CD4+ T-Cells,omeprazole suppresses proliferation,downregulates V-ATPase,and promotes differentiation toward an autoimmunity-favoring phenotype[J]. Int Immunopharmacol,2025,144(1):113728. [8] 田琴琴,刘茹,陈云秀,等. 新生期肺炎链球菌肺炎对成年期肺组织免疫状态及气道高反应的影响[J]. 免疫学杂志,2015,31(12):1051-1055. [9] Larche M,Robinson D S,Kay A B. The role of T lymphocytes in the pathogenesis of asthma[J]. J Allergy Clin Immunol,2003,111(3):450-463. doi: 10.1067/mai.2003.169 [10] Zhang Y,Zhang Y,Gu W,et al. Th1/Th2 cell's function in immune system[J]. Adv Exp Med Biol,2014,841(1):45-65. [11] Ercoli G,Fernandes V E,Chung W Y,et al. Intracellular replication of streptococcus pneumoniae inside splenic macrophages serves as a reservoir for septicaemia[J]. Nat Microbiol,2018,3(5):600-610. doi: 10.1038/s41564-018-0147-1 [12] Van Breedam W,Verbeeck M,Christiaens I,et al. Porcine,murine and human sialoadhesin (Sn/Siglec-1/CD169): Portals for porcine reproductive and respiratory syndrome virus entry into target cells [J]. J Gen Virol,2013,94(Pt 9): 1955-1960. [13] Miao Y R,Xia M,Luo M,et al. ImmuCellAI-mouse: A tool for comprehensive prediction of mouse immune cell abundance and immune microenvironment depiction[J]. Bioinformatics,2022,38(3):785-791. doi: 10.1093/bioinformatics/btab711 [14] 马建芳,黄晓玲,李艳华,等. 社区获得性肺炎合并糖尿病患者的临床特征和死亡危险因素分析[J]. 昆明医科大学学报,2024,45(2):123-130. doi: 10.12259/j.issn.2095-610X.S20240217 [15] Wei H,Zhao H,Li R,et al. Rhinovirus impairs the immune response of alveolar macrophages to facilitate streptococcus pneumonia infection[J]. Pathog Dis,2020,78(3):ftaa020. doi: 10.1093/femspd/ftaa020 [16] Ip W K E,Hoshi N,Shouval D S,et al. Anti-inflammatory effect of IL-10 mediated by metabolic reprogramming of macrophages[J]. Science,2017,356(6337):513-519. doi: 10.1126/science.aal3535 [17] 张群威,任志宏,程力平,等. 肺炎支原体肺炎患儿外周血IL-10/IL-17表达与肺功能变化的相关性 [J] 安徽医药,2016,20(4): 762-763. [18] Xiong Y,Zhong Q,Palmer T,et al. Estradiol resolves pneumonia via ERβ in regulatory T cells[J]. JCI Insight,2021,6(3):e133251. doi: 10.1172/jci.insight.133251 [19] McKinley L,Logar A J,McAllister F,et al. Regulatory T cells dampen pulmonary inflammation and lung injury in an animal model of pneumocystis pneumonia[J]. J Immunol,2006,177(9):6215-6226. doi: 10.4049/jimmunol.177.9.6215 [20] D'Alessio F R,Kurzhagen J T,Rabb H. Reparative T lymphocytes in organ injury[J]. J Clin Invest,2019,129(7):2608-2618. doi: 10.1172/JCI124614 [21] Chun Y Y,Tan K S,Yu L,et al. Influence of glycan structure on the colonization of streptococcus pneumoniae on human respiratory epithelial cells[J]. Proc Natl Acad Sci U S A,2023,120(13):e2213584120. doi: 10.1073/pnas.2213584120 [22] Sheng K,Sang L,Ge H. TRPM8 knockdown relieved inflammatory response and cell apoptosis in pneumonia model induced by streptococcus pneumoniae in vitro[J]. Cell Mol Biol (Noisy-le-grand),2023,69(15):198-203. doi: 10.14715/cmb/2023.69.15.34 [23] Han L,Zhang F,Liu Y,et al. Uterus globulin associated protein 1 (UGRP1) binds podoplanin (PDPN) to promote a novel inflammation pathway during streptococcus pneumoniae infection[J]. Clin Transl Med,2022,12(6):e850. doi: 10.1002/ctm2.850 [24] Cui A H,Zhao J,Liu S X,et al. Associations of IL-4,IL-6,and IL-12 levels in peripheral blood with lung function,cellular immune function,and quality of life in children with moderate-to-severe asthma[J]. Medicine (Baltimore),2017,96(12):e6265. [25] Yang J,Hooper W C,Phillips D J,et al. Cytokines in mycoplasma pneumoniae infections[J]. Cytokine Growth Factor Rev,2004,15(2-3):157-168. doi: 10.1016/j.cytogfr.2004.01.001 [26] Calum H P,Moser C,Jensen P O,et al. Early IL-2 treatment of mice with pseudomonas aeruginosa pneumonia induced PMN-dominating response and reduced lung pathology[J]. APMIS,2020,128(12):647-653. doi: 10.1111/apm.13072 [27] Nakashima M,Kinoshita M,Nakashima H,et al. Mouse liver B cells phagocytose streptococcus pneumoniae and initiate immune responses against their antigens[J]. J Immunol,2022,209(1):26-37. doi: 10.4049/jimmunol.2100520 [28] 刘洋,崔玉霞. 儿童肺炎支原体肺炎Th1/Th2细胞因子变化特征及其相关免疫机制的研究进展[J]. 贵州医药,2023,47(4):520-521. doi: 10.3969/j.issn.1000-744X.2023.04.006 [29] Chen K,Kolls J K. T cell-mediated host immune defenses in the lung[J]. Annu Rev Immunol,2013,31(1):605-633. doi: 10.1146/annurev-immunol-032712-100019 [30] Chowdhury D,Lieberman J. Death by a thousand cuts: Granzyme pathways of programmed cell death[J]. Annu Rev Immunol,2008,26(1):389-420. doi: 10.1146/annurev.immunol.26.021607.090404 [31] Zhang J,Xiao Z,Qu C,et al. CD8 T cells are involved in skeletal muscle regeneration through facilitating MCP-1 secretion and Gr1(high) macrophage infiltration[J]. J Immunol,2014,193(10):5149-5160. doi: 10.4049/jimmunol.1303486 [32] Zhang X,Ali M,Pantuck M A,et al. CD8 T cell response and its released cytokine IFN-gamma are necessary for lung alveolar epithelial repair during bacterial pneumonia[J]. Front Immunol,2023,14(1):1268078. -
下载:
点击查看大图
图(4)
计量
- 文章访问数: 387
- HTML全文浏览量: 207
- PDF下载量: 15
- 被引次数: 0
