Toll-like receptor 2-mediated signaling requirements for Francisella tularensis live vaccine strain infection of murine macrophages
Toll-like receptor 2-mediated signaling requirements for Francisella tularensis live vaccine strain infection of murine macrophages
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DOI：
10.1128/iai.01868-06
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发表时间：
2007-08-01
影响因子：
3.1
通讯作者：
Vogel, Stefanie N.
中科院分区：
文献类型：
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作者：
Cole, Leah E.;Shirey, Kari Ann;Vogel, Stefanie N.
图拉氏弗朗西斯杆菌是一种需氧、无芽胞、革兰氏阴性的球菌，是引起图拉热病的病原体。我们以前报道过图拉氏丝虫活疫苗株(LVS)在表达Toll样受体2(TLR2)的HEK293T细胞中诱导出强的、剂量依赖的NF-kappa B报告活性，并在原代小鼠巨噬细胞中表达促炎基因。在此，我们报道了图拉氏丝虫诱导的小鼠巨噬细胞促炎细胞因子基因和蛋白的表达是压倒性的TLR2依赖，TLR2(-/-)巨噬细胞的抑制反应证明了这一点。在体外，无论是在小鼠巨噬细胞中，还是在体内，感染图拉氏丝线虫的小鼠肝脏中TLR2的表达都增加了。通过共聚焦显微镜观察细胞内TLR2和MyD88的共定位。如果加热或福尔马林灭活或氯霉素处理，TLR2激动剂的活性依赖于新的细菌蛋白质合成，则信号被取消。在巨噬细胞中进行复制；然而，细菌复制对于TLR2信号不是必需的，因为LVS Delta guaA，一种在没有外源鸟嘌呤的情况下无法复制的LVS鸟嘌呤营养缺陷型，激活了TLR2转基因HEK293T细胞中的NF-kappa B，并诱导了野生型巨噬细胞中细胞因子的表达，与野生型图拉氏福尔马氏菌LVS相似。综上所述，这些数据表明，原始巨噬细胞对图拉氏杆菌LVS的反应绝大多数依赖于TLR2，需要从头开始的细菌蛋白质合成，并且不依赖于图拉氏杆菌细胞内的复制。
Francisella tularensis, an aerobic, non-spore-forming, gram-negative coccobacillus, is the causative agent of tularemia. We reported previously that F. tularensis live vaccine strain (LVS) elicited strong, dose-dependent NF-kappa B reporter activity in Toll-like receptor 2 (TLR2) -expressing HEK293T cells and proinflammatory gene expression in primary murine macrophages. Herein, we report that F. tularensis LVS-induced murine macrophage proinflammatory cytokine gene and protein expression are overwhelmingly TLR2 dependent, as evidenced by the abrogated responses of TLR2(-/-) macrophages. F. tularensis LVS infection also increased expression of TLR2 both in vitro, in mouse macrophages, and in vivo, in livers from F. tularensis LVS-infected mice. Colocalization of intracellular F. tularensis LVS, TLR2, and MyD88 was visualized by confocal microscopy. Signaling was abrogated if the F. tularensis LVS organisms were heat or formalin killed or treated with chloramphenicol, indicating that the TLR2 agonist activity is dependent on new bacterial protein synthesis. F. tularensis LVS replicates in macrophages; however, bacterial replication was not required for TLR2 signaling because LVS Delta guaA, an F. tularensis LVS guanine auxotroph that fails to replicate in the absence of exogenous guanine, activated NF-kappa B in TLR2-transfected HEK293T cells and induced cytokine expression in wild-type macrophages comparably to wild-type F. tularensis LVS. Collectively, these data indicate that the primary macrophage response to F. tularensis LVS is overwhelmingly TLR2 dependent, requires de novo bacterial protein synthesis, and is independent of intracellular F. tularensis replication.