DosR抗原Rv2626c通过lncRNA MacORIS诱导巨噬细胞活化的结核潜伏感染机制研究  

Study of latent tuberculosis infection mechanism of DosR antigen Rv2626c inducing macrophage activation through lncRNA MacORIS

作　　者：汪兆慧 郑振鲁 李瑞娜 马媛媛 徐辉 魏振宏[4] 贾彦娟[4] 齐晓明[4] 高小玲 WANG Zhaohui;ZHENG Zhenlu;LI Ruina;MA Yuanyuan;XU Hui;WEI Zhenhong;JIA Yanjuan;QI Xiaoming;GAO Xiaoling(Department of Medical Genetics and Prenatal Diagnostics,Luoyang Maternal and Child Health Care Hospital,Luoyang 471099,Henan,China;Gansu Chinese Medical University,Lanzhou 730000,Gansu,China;Medical Laboratory Center,Hainan Provincial People's Hospital,Haikou 570311,Hainan,China;The Institute of Clinical Research and Translational Medicine,Gansu Provincial People's Hospital,Lanzhou 730000,Gansu,China)

机构地区：[1]洛阳市妇幼保健院医学遗传与产前诊断科,河南洛阳471099 [2]甘肃中医药大学,甘肃兰州730000 [3]海南省人民医院检验科,海南海口570311 [4]甘肃省人民医院临床研究与转化医学研究所,甘肃兰州730000

出　　处：《检验医学》2024年第10期923-932,共10页Laboratory Medicine

基　　金：国家自然科学基金项目(81830372)。

摘　　要：目的 探讨结核潜伏感染(LTBI)表达的DosR抗原Rv2626c在巨噬细胞活化中的作用和长链非编码RNA(lncRNA)MacORIS对其的调控作用。方法 将人急性单核细胞白血病细胞系THP-1诱导为巨噬细胞,根据转染物的不同分为4组:阴性对照(NC)组[转染NC-小干扰RNA(siRNA) 24 h]、MacORIS-siRNA组(转染MacORIS-siRNA 24 h);NC-siRNA+重组蛋白Rv2626c(rRv2626c)组(转染NC-siRNA 24 h,再用3μg·mL^(-1)rRv2626c刺激24h)、MacORIS-siRNA+rRv2626c组(转染MacORIS-siRNA24h,再用3μg·mL^(-1)rRv2626c刺激24h)。检测各组细胞lncRNAMacORIS、Toll样受体(TLR)2、TLR4、表面共刺激分子(CD80、CD86、CD163、CD206)、吲哚胺2,3-双加氧酶1(IDO1)、肿瘤坏死因子-α(TNF-α)、白细胞介素(IL)-6、IL-8、IL-1β的表达情况。采用生物信息学方法预测并构建lncRNA MacORIS的竞争内源RNA(ceRNA)网络,对lncRNA MacORIS潜在的靶微小RNA进行验证。结果 rRv2626c可有效促进巨噬细胞lncRNA MacORIS和IDO1上调(P<0.05)。rRv2626c可依赖于lncRNA MacORIS诱导THP-1细胞中TLR2的表达(P<0.05)。rRv2626c可依赖于lncRNA MacORIS上调巨噬细胞中CD80、CD86、TNF-α、IL-1β的表达(P<0.001)。与NC组比较,MacORIS-siRNA组miR-141-3p、miR-150-5p、miR-33-5p、miR-200-3p和miR-203-3p相对表达量均显著升高(P<0.05)。结论 rRv2626c可促进巨噬细胞极化到M1表型,并促进TLR2和炎症因子表达,这些变化均依赖于lncRNA MacORIS。lncRNA MacORIS可负调控miR-141-3p、miR-150-5p、miR-33-5p、miR-200-3p和miR-203-3p的表达。Objective To investigate the regulation role of dormancy survival regulator(DosR)antigen Rv2626c expressed in latent tuberculosis infection(LTBI)in the activation of macrophage and the regulation influence of long non-coding RNA(lncRNA)MacORIS.Methods Human acute monocytic leukemia cell line THP-1 cells were differentiated into macrophage and categorized into 4 groups based on different transfections:negative control(NC)group[transfected with NC-small interfering RNA(siRNA)for 24 h]and MacORIS-siRNA group[transfected with MacORIS-siRNA for 24 h],NC-siRNA+recombinant protein Rv2626c(rRv2626c)group[transfected with NC-siRNA for 24 h,then stimulated with 3μg·mL^(-1) rRv2626c for 24 h],MacORIS-siRNA+rRv2626c group[transfected with MacORIS-siRNA for 24 h,then stimulated with 3μg·mL^(-1) rRv2626c for 24 h].The expression of lncRNA MacORIS,Toll-like receptor(TLR)2,TLR4,surface co-stimulatory molecules(CD80,CD86,CD163,CD206),indoleamine 2,3-dioxygenase 1(IDO1),tumor necrosis factor-alpha(TNF-α),interleukin(IL)-6,IL-8 and IL-1βwere determined.The competitive endogenous RNA(ceRNA)networks of lncRNA MacORIS were predicted and constructed by bioinformatics methods,and the potential target microRNA of lncRNA MacORIS was verified.Results The rRv2626c effectively promoted the upregulation of lncRNA MacORIS and IDO1 in macrophage(P<0.05).The rRv2626c induced the expression of TLR2 in THP-1 cells,which was dependent on lncRNA MacORIS(P<0.05).Furthermore,rRv2626c depended on lncRNA MacORIS to upregulate the expressions of CD80,CD86,TNF-αand IL-1βin macrophage(P<0.001).Compared to the NC group,the relative expression levels of miR-141-3p,miR-150-5p,miR-33-5p,miR-200-3p and miR-203-3p in the MacORIS-siRNA group were increased(P<0.05).Conclusions The rRv2626c promotes the polarization of macrophage toward M1 phenotype,and it enhances the expression of TLR2 and inflammatory factors,which depends on lncRNA MacORIS.The lncRNA MacORIS could negatively regulate the expression of miR-141-3p,miR-150-5p,miR-33-5p,miR-200-3p and miR-2

关 键 词：长链非编码RNAMacORIS Rv2626c 巨噬细胞 结核潜伏感染 

分 类 号：R446.7[医药卫生—诊断学]