Centre Hospitalier Universitaire de Rennes, Laboratoire de Parasitologie, Rennes, France.
PLoS One. 2012;7(3):e33413. doi: 10.1371/journal.pone.0033413. Epub 2012 Mar 22.
The development of inflammatory granulomas around infected Kupffer cells is necessary for hepatic parasite clearance during visceral leishmaniasis. Invariant NKT (iNKT) cells are predominant T cells in the mouse liver and can synthesize large quantities of IL-4 and IFN-γ, two cytokines involved in granuloma formation. This study analyzed the role of iNKT cells in the hepatic immune response during Leishmania donovani infection, using a murine model of wild-type (WT) and iNKT cell-deficient (Jα18⁻/⁻) C57BL/6 mice sacrificed 15, 30 or 60 days post-infection. We recorded hepatic parasite loads, cytokine expression, and analyzed granulomatous response by immunohistochemistry and hepatic immune cell infiltration by flow cytometry. Whereas WT animals rapidly controlled the infection and developed an inflammatory response associated with a massive influx of iNKT cells observed by flow cytometry, Jα18⁻/⁻ mice had significantly higher parasitic loads on all time points. This lack of control of parasite burden was associated with a delay in granuloma maturation (28.1% of large granulomas at day 60 versus 50.7% in WT). Cytokine transcriptome analysis showed that mRNA of 90/101 genes encoding chemokines, cytokines and their receptors, was underexpressed in Jα18⁻/⁻ mice. Detection of IL-4 and TNF-α by ELISA in liver extracts was also significantly lower in Jα18⁻/⁻ mice. Consistent with flow cytometry analysis, cytokinome profile in WT mice showed a bias of expression towards T cell-chemoattractant chemokines on D15, and displayed a switch towards expression of granulocytes and/or monocytes -chemoattractant chemokines on D60. In Jα18⁻/⁻ mice, the significantly lower expression of CXCL5, MIP-2 and CCL2 mRNA was correlated with a defect in myeloperoxidase positive-cell attraction observed by immunohistochemistry and with a lower granulocyte and monocyte infiltration in the liver, as shown by flow cytometry. These data indicate that iNKT cells play a role in early and sustained pro-inflammatory cytokine response warranting efficient organization of hepatic granulomas and parasite clearance.
在内脏利什曼病期间，感染的枯否细胞周围炎症性肉芽肿的发展对于肝寄生虫的清除是必要的。不变自然杀伤 T(iNKT)细胞是小鼠肝脏中的主要 T 细胞，能够合成大量的 IL-4 和 IFN-γ，这两种细胞因子参与肉芽肿的形成。本研究使用野生型(WT)和 iNKT 细胞缺陷型(Jα18⁻/⁻) C57BL/6 小鼠的感染模型，分析了 iNKT 细胞在利什曼原虫感染期间肝免疫反应中的作用，这些小鼠在感染后 15、30 或 60 天被处死。我们记录了肝寄生虫负荷、细胞因子表达，并通过免疫组织化学分析了肉芽肿反应，通过流式细胞术分析了肝免疫细胞浸润。尽管 WT 动物迅速控制了感染并产生了与大量 iNKT 细胞流入相关的炎症反应，但 Jα18⁻/⁻小鼠在所有时间点的寄生虫负荷都明显更高。这种对寄生虫负担的控制缺失与肉芽肿成熟的延迟有关(第 60 天的大肉芽肿中只有 28.1%，而 WT 中的肉芽肿为 50.7%)。细胞因子转录组分析显示，编码趋化因子、细胞因子及其受体的 90/101 个基因的 mRNA 在 Jα18⁻/⁻小鼠中表达下调。通过 ELISA 在肝提取物中检测到的 IL-4 和 TNF-α也明显较低。与流式细胞术分析一致，WT 小鼠的细胞因子组显示出在 D15 时偏向于 T 细胞趋化因子的表达谱，并在 D60 时显示出向粒细胞和/或单核细胞趋化因子表达的转变。在 Jα18⁻/⁻小鼠中，CXCL5、MIP-2 和 CCL2 mRNA 的表达显著下调与免疫组织化学观察到的髓过氧化物酶阳性细胞吸引缺陷相关，并与通过流式细胞术观察到的肝粒细胞和单核细胞浸润减少相关。这些数据表明，iNKT 细胞在早期和持续的促炎细胞因子反应中发挥作用，这保证了肝肉芽肿的有效组织和寄生虫的清除。
