卡介苗感染人树突状细胞对miRNA-99b表达和Th17/Treg分化的影响
|
摘要
目的 探讨牛型分枝杆菌减毒活菌—卡介苗(BCG)感染对人树突状细胞(DCs)microRNA-99b(miRNA-99b)表达的影响及对新生儿脐血CD_4~+初始T淋巴细胞分化为Th17/Treg的作用。方法 BCG感染体外培养的人树突状细胞,然后与新生儿脐血CD_4~+初始T淋巴细胞混合培养,qRT-PCR观察人树突状细胞miRNA-99b的表达变化,qRT-PCR检测CD_4~+T淋巴细胞Foxp3、ROR-γt、γ干扰素(IFN-γ)、白细胞介素-10(IL-10)基因转录水平,观察CD_4~+T淋巴细胞分化亚型的改变;ELISA法检测培养上清INF-γ、IL-17和IL-23表达水平。结果 BCG感染后,人树突状细胞miRNA-99b基因表达增加(P<0.001);CD_4~+T淋巴细胞IFN-γ、IL-10基因表达增加(P<0.05),ROR-γt基因表达降低(P<0.05),Foxp3基因转录增加,但差异无统计学意义;感染组培养上清中INF-γ表达水平增加(P<0.001),IL-17和IL-23表达水平均有一定的增加,但差异无统计学意义。结论 结核感染可能通过影响树突状细胞miRNA-99b的表达进而诱导初始CD_4~+T淋巴细胞分化为Th17/Treg的失衡参与结核病的发病过程。
OBJECTIVE To investigate the effects of Bacillus Calmette Guerin vaccine(BCG) infection on microRNA-99 b(miRNA-99 b) expression in human dendritic cells and the differentiation of CD_4~+ naive T cells in neonate cord blood. METHODS After infected by BCG, human dendritic cells cultured in vitro were mixed cultured with CD_4~+ naive T cells of neonate cord blood. The change in expression of miRNA-99 b in human dendritic cells was observed by qRT-PCR. The transcriptional levels of Foxp3, ROR-γt, IFN-γ and IL-10 genes of CD_4~+ T cells were determined by qRT-PCR to observe the change in differentiation subtypes of T lymphocytes. The INF-γ, IL-17 and IL-23 levels in culture supernatant were analyzed by ELISA. RESULTS After infected by BCG, the miRNA-99 b expression was dramatically improved in human dendritic cells(P<0.001), the IFN-γ and IL-10 gene expression levels of CD_4~+ T cells significantly increased(P<0.05), ROR-γt gene expression of CD_4~+ T cells significantly decreased( P<0.05). INF-γ level increased significantly in culture supernatant(P<0.001). IL-17 and IL-23 expression levels also increased in culture supernatant, but the differences were not significant. CONCLUSION Tuberculosis infection may be involved in the pathogenesis of tuberculosis through controlling miRNA-99 b expression in human dendritic cells and inducing Th17/Treg differentiation imbalance.
OBJECTIVE To investigate the effects of Bacillus Calmette Guerin vaccine(BCG) infection on microRNA-99 b(miRNA-99 b) expression in human dendritic cells and the differentiation of CD_4~+ naive T cells in neonate cord blood. METHODS After infected by BCG, human dendritic cells cultured in vitro were mixed cultured with CD_4~+ naive T cells of neonate cord blood. The change in expression of miRNA-99 b in human dendritic cells was observed by qRT-PCR. The transcriptional levels of Foxp3, ROR-γt, IFN-γ and IL-10 genes of CD_4~+ T cells were determined by qRT-PCR to observe the change in differentiation subtypes of T lymphocytes. The INF-γ, IL-17 and IL-23 levels in culture supernatant were analyzed by ELISA. RESULTS After infected by BCG, the miRNA-99 b expression was dramatically improved in human dendritic cells(P<0.001), the IFN-γ and IL-10 gene expression levels of CD_4~+ T cells significantly increased(P<0.05), ROR-γt gene expression of CD_4~+ T cells significantly decreased( P<0.05). INF-γ level increased significantly in culture supernatant(P<0.001). IL-17 and IL-23 expression levels also increased in culture supernatant, but the differences were not significant. CONCLUSION Tuberculosis infection may be involved in the pathogenesis of tuberculosis through controlling miRNA-99 b expression in human dendritic cells and inducing Th17/Treg differentiation imbalance.
引文
[1] Walker NF,Stek C,Wasserman S,et al.The tuberculosis-associated immune reconstitution inflammatory syndrome:recent advances in clinical and pathogenesis research[J].Curr Opin HIV AIDS,2018,13(6):512-521.
[2] Li Y,Sun W.Effects of Th17/Treg cell imbalance on HIV replication in patients with AIDS complicated with tuberculosis[J].Exp Ther Med,2018,15(3):2879-2883.
[3] Luo J,Zhang M,Yan B,et al.Imbalance of Th17 and Treg in peripheral blood mononuclear cells of active tuberculosis patients[J].Braz J Infect Dis,2017,21(2):155-161.
[4] Ellwanger JH,Zambra F,Guimaraes RL,et al.MicroRNA-related polymorphisms in infectious diseases-tiny changes with a huge impact on viral infections and potential clinical applications[J].Front Immunol,2018,9:1316.
[5] von Both U,Berk M,Agapow PM,et al.Mycobacterium tuberculosis exploits a molecular off switch of the immune system for intracellular survival[J].Sci Rep,2018,8(1):661.
[6] Picchi H,Mateus C,Chouaid C,et al.Infectious complications associated with the use of immune checkpoint inhibitors in oncology:reactivation of tuberculosis after anti PD-1 treatment[J].Clin Microbiol Infect,2018,24(3):216-218.
[7] Yu S,Liu C,Li L,et al.Inactivation of Notch signaling reverses the Th17|[sol]|Treg imbalance in cells from patients with immune thrombocytopenia[J].Lab Invest,2015,95(2):157-167.
[8] Burel JG,Lindestam Arlehamn CS,Khan N,et al.Transcriptomic analysis of CD4+ T cells reveals novel immune signatures of latent tuberculosis[J].J Immunol,2018,200(9):3283-3290.
[9] Zhang Y,Zhang Y,Gu W,et al.Th1/Th2 cell differentiation and molecular signals[J].Adv Exp Med Biol,2014,841:15-44.
[10] Kononova TE,Urazova OI,Novitskii VV,et al.Factors of Th17 and Treg lymphocyte differentiation in pulmonary tuberculosis[J].Bull Exp Biol Med,2015,159(2):201-204.
[11] Mendez-Samperio P.Commentary:the role of neutrophils in the induction of specific Th1 and Th17 during vaccination against tuberculosis[J].Front Cell Infect Microbiol,2017,7:179.
[12] Mathew JM,Voss JH,Mcewen ST,et al.Generation and characterization of alloantigen-specific regulatory T cells for clinical transplant tolerance[J].Sci Rep,2018,8(1):1136.
[13] Taams LS,Vukmanovic-Stejic M,Smith J,et al.Antigen-specific T cell suppression by human CD4+CD25+ regulatory T cells[J].Eur J Immunol,2002,32(6):1621-1630.
[14] Vuddamalay Y,van Meerwijk JP.CD28- and CD28lowCD8+ Regulatory T Cells:Of Mice and Men[J].Front Immunol,2017,8:31.
[15] Liu B,Gao W,Zhang L,et al.Th17/Treg imbalance and increased interleukin-21 are associated with liver injury in patients with chronic severe hepatitis B[J].Int Immunopharmacol,2017,46:48-55.
[16] Boer MC,Joosten SA,Ottenhoff TH.Regulatory T-cells at the interface between human host and pathogens in infectious diseases and vaccination[J].Front Immunol,2015,6:217.
[17] Narayana JL,Huang HN,Wu CJ,et al.Efficacy of the antimicrobial peptide TP4 against Helicobacter pylori infection:in vitro membrane perturbation via micellization and in vivo suppression of host immune responses in a mousemodel[J].Oncotarget,2015,6(15):12936-12954.
[18] Boudewijns S,Westdorp H,Koornstra RH,et al.Immune-related adverse events of dendritic cell vaccination correlate with immunologic and clinical outcome in stage III and IV melanoma patients[J].J Immunother,2016,39(6):241-248.
[19] Siddle KJ,Tailleux L,Deschamps M,et al.Bacterial infection drives the expression dynamics of microRNAs and their isomiRs[J].PLoS Genet,2015,11(3):e1005064.
[20] Wang J,Zhu X,Xiong X,et al.Identification of potential urine proteins and microRNA biomarkers for the diagnosis of pulmonary tuberculosis patients[J].Emerg Microbes Infect,2018,7(1):63.
[21] Bettencourt P,Pires D,Anes E.Immunomodulating microRNAs of mycobacterial infections[J].Tuberculosis (Edinb),2016,97:1-7.
[22] Ahmed W,Zheng K,Liu ZF.Small non-coding RNAs:new insights in modulation of host immune response by intracellular bacterial pathogens[J].Front Immunol,2016,7:431.
[23] Das K,Saikolappan S,Dhandayuthapani S.Differential expression of miRNAs by macrophages infected with virulent and avirulent Mycobacterium tuberculosis[J].Tuberculosis (Edinb),2013,93(Suppl):S47-50.
[24] Coulter F,Parrish A,Manning D,et al.IL-17 production from T helper 17,mucosal-associated invariant T,and γδ cells in tuberculosis infection and disease[J].Front Immunol,2017,8:1252.
[25] 陈琳,钟山亮,张磊,等.微RNA-99b靶向调节mTOR基因表达可逆转人乳腺癌细胞对多柔比星的耐药性[J].肿瘤,2016,36(1):3-11.
[2] Li Y,Sun W.Effects of Th17/Treg cell imbalance on HIV replication in patients with AIDS complicated with tuberculosis[J].Exp Ther Med,2018,15(3):2879-2883.
[3] Luo J,Zhang M,Yan B,et al.Imbalance of Th17 and Treg in peripheral blood mononuclear cells of active tuberculosis patients[J].Braz J Infect Dis,2017,21(2):155-161.
[4] Ellwanger JH,Zambra F,Guimaraes RL,et al.MicroRNA-related polymorphisms in infectious diseases-tiny changes with a huge impact on viral infections and potential clinical applications[J].Front Immunol,2018,9:1316.
[5] von Both U,Berk M,Agapow PM,et al.Mycobacterium tuberculosis exploits a molecular off switch of the immune system for intracellular survival[J].Sci Rep,2018,8(1):661.
[6] Picchi H,Mateus C,Chouaid C,et al.Infectious complications associated with the use of immune checkpoint inhibitors in oncology:reactivation of tuberculosis after anti PD-1 treatment[J].Clin Microbiol Infect,2018,24(3):216-218.
[7] Yu S,Liu C,Li L,et al.Inactivation of Notch signaling reverses the Th17|[sol]|Treg imbalance in cells from patients with immune thrombocytopenia[J].Lab Invest,2015,95(2):157-167.
[8] Burel JG,Lindestam Arlehamn CS,Khan N,et al.Transcriptomic analysis of CD4+ T cells reveals novel immune signatures of latent tuberculosis[J].J Immunol,2018,200(9):3283-3290.
[9] Zhang Y,Zhang Y,Gu W,et al.Th1/Th2 cell differentiation and molecular signals[J].Adv Exp Med Biol,2014,841:15-44.
[10] Kononova TE,Urazova OI,Novitskii VV,et al.Factors of Th17 and Treg lymphocyte differentiation in pulmonary tuberculosis[J].Bull Exp Biol Med,2015,159(2):201-204.
[11] Mendez-Samperio P.Commentary:the role of neutrophils in the induction of specific Th1 and Th17 during vaccination against tuberculosis[J].Front Cell Infect Microbiol,2017,7:179.
[12] Mathew JM,Voss JH,Mcewen ST,et al.Generation and characterization of alloantigen-specific regulatory T cells for clinical transplant tolerance[J].Sci Rep,2018,8(1):1136.
[13] Taams LS,Vukmanovic-Stejic M,Smith J,et al.Antigen-specific T cell suppression by human CD4+CD25+ regulatory T cells[J].Eur J Immunol,2002,32(6):1621-1630.
[14] Vuddamalay Y,van Meerwijk JP.CD28- and CD28lowCD8+ Regulatory T Cells:Of Mice and Men[J].Front Immunol,2017,8:31.
[15] Liu B,Gao W,Zhang L,et al.Th17/Treg imbalance and increased interleukin-21 are associated with liver injury in patients with chronic severe hepatitis B[J].Int Immunopharmacol,2017,46:48-55.
[16] Boer MC,Joosten SA,Ottenhoff TH.Regulatory T-cells at the interface between human host and pathogens in infectious diseases and vaccination[J].Front Immunol,2015,6:217.
[17] Narayana JL,Huang HN,Wu CJ,et al.Efficacy of the antimicrobial peptide TP4 against Helicobacter pylori infection:in vitro membrane perturbation via micellization and in vivo suppression of host immune responses in a mousemodel[J].Oncotarget,2015,6(15):12936-12954.
[18] Boudewijns S,Westdorp H,Koornstra RH,et al.Immune-related adverse events of dendritic cell vaccination correlate with immunologic and clinical outcome in stage III and IV melanoma patients[J].J Immunother,2016,39(6):241-248.
[19] Siddle KJ,Tailleux L,Deschamps M,et al.Bacterial infection drives the expression dynamics of microRNAs and their isomiRs[J].PLoS Genet,2015,11(3):e1005064.
[20] Wang J,Zhu X,Xiong X,et al.Identification of potential urine proteins and microRNA biomarkers for the diagnosis of pulmonary tuberculosis patients[J].Emerg Microbes Infect,2018,7(1):63.
[21] Bettencourt P,Pires D,Anes E.Immunomodulating microRNAs of mycobacterial infections[J].Tuberculosis (Edinb),2016,97:1-7.
[22] Ahmed W,Zheng K,Liu ZF.Small non-coding RNAs:new insights in modulation of host immune response by intracellular bacterial pathogens[J].Front Immunol,2016,7:431.
[23] Das K,Saikolappan S,Dhandayuthapani S.Differential expression of miRNAs by macrophages infected with virulent and avirulent Mycobacterium tuberculosis[J].Tuberculosis (Edinb),2013,93(Suppl):S47-50.
[24] Coulter F,Parrish A,Manning D,et al.IL-17 production from T helper 17,mucosal-associated invariant T,and γδ cells in tuberculosis infection and disease[J].Front Immunol,2017,8:1252.
[25] 陈琳,钟山亮,张磊,等.微RNA-99b靶向调节mTOR基因表达可逆转人乳腺癌细胞对多柔比星的耐药性[J].肿瘤,2016,36(1):3-11.