Vaccine-Mediated Mechanisms Controlling Replication of Francisella tularensis in Human Peripheral Blood Mononuclear Cells Using a Co-culture System
Vaccine-Mediated Mechanisms Controlling Replication of Francisella tularensis in Human Peripheral Blood Mononuclear Cells Using a Co-culture System
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DOI：
10.3389/fcimb.2018.00027
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发表时间：
2018-02-07
影响因子：
5.7
通讯作者：
Sjostedt, Anders
中科院分区：
文献类型：
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作者：
Eneslatt, Kjell;Golovliov, Igor;Sjostedt, Anders
细胞介导的免疫（CMI）通常是有效保护免受细胞内感染所需的，然而，相关物的鉴定是具有挑战性的，并且它们通常缺乏。土拉热弗朗西丝菌是一种高毒性的兼性细胞内细菌，CMI是抵御病原体的关键，但这在人类中是如何实现的知之甚少。为了了解保护机制，我们建立了一个体外共培养试验，以确定如何控制感染F。土拉热是由人细胞完成的，并假设该模型将模拟体内免疫机制。用抗原扩增非贴壁外周血单核细胞（PBMC），并将其加入到用人疫苗株LVS或高毒力SCHU S4株感染的贴壁PBMC的培养物中。胞内F.跟踪土拉热72小时，分析分泌的和细胞内的细胞因子。添加从幼稚个体扩增的PBMC，即，无F.土拉热杆菌的PBMC的加入通常导致很少或没有细胞内细菌生长的控制，而来自大多数F.土拉热-免疫个体对细胞内细菌执行静态和有时杀死作用。无论感染何种菌株，两组间差异均有统计学意义，P < 0.05。感染72小时后分析了11种细胞因子的分泌，在免疫个体和未感染LV的个体之间观察到IFN-γ、TNF和MIP-1 β分泌的显著差异。此外，在LV感染的培养物中，来自疫苗接种者的CD 4 T细胞，而不是CD 8 T细胞，显示出IFN-γ、MIP-1 β、TNF和CD 107 a的表达显著高于来自初始个体的细胞。共培养系统似乎可以识别与理解宿主对F.土拉热。
Cell-mediated immunity (CMI) is normally required for efficient protection against intracellular infections, however, identification of correlates is challenging and they are generally lacking. Francisella tularensis is a highly virulent, facultative intracellular bacterium and CMI is critically required for protection against the pathogen, but how this is effectuated in humans is poorly understood. To understand the protective mechanisms, we established an in vitro co-culture assay to identify how control of infection of F. tularensis is accomplished by human cells and hypothesized that the model will mimic in vivo immune mechanisms. Non-adherent peripheral blood mononuclear cells (PBMCs) were expanded with antigen and added to cultures with adherent PBMC infected with the human vaccine strain, LVS, or the highly virulent SCHU S4 strain. Intracellular numbers of F. tularensis was followed for 72 h and secreted and intracellular cytokines were analyzed. Addition of PBMC expanded from naive individuals, i.e., those with no record of immunization to F. tularensis, generally resulted in little or no control of intracellular bacterial growth, whereas addition of PBMC from a majority of F. tularensis-immune individuals executed static and sometimes cidal effects on intracellular bacteria. Regardless of infecting strain, statistical differences between the two groups were significant, P < 0.05. Secretion of 11 cytokines was analyzed after 72 h of infection and significant differences with regard to secretion of IFN-gamma, TNF, and MIP-1 beta was observed between immune and naive individuals for LVS-infected cultures. Also, in LVS-infected cultures, CD4 T cells from vaccinees, but not CD8 T cells, showed significantly higher expression of IFN-gamma, MIP-1 beta, TNF, and CD107a than cells from naive individuals. The co-culture system appears to identify correlates of immunity that are relevant for the understanding of mechanisms of the protective host immunity to F. tularensis.