Natural killer cells regulate pulmonary macrophages polarization in host defense chlamydial lung infection. Natural killer cells regulate pulmonary macrophages polarization in host defense chlamydial lung infection

NK cells and pulmonary macrophages both are important components of innate immunity. The interaction between NK cells and pulmonary macrophages during Chlamydia muridarum（C. muridarum）respiratory infections is poorly understood. In this study, we explored the effect of NK cells on regulation of pulmonary macrophage function during chlamydial lung infection. We found that NK depletion led to polarization of pulmonary macrophages from M1 to M2 phenotype, and this related to significantly reduced miR-155 expression in pulmonary macrophage. Using adoptive transfer approach, we found that the recipient mice receiving lung macrophages isolated from C. muridarum-infected NK-cell-depleted mice exhibited an increased bacterial load and severe inflammation in the lung upon chlamydial challenge when compared with the recipients of lung macrophages from infected IgG -treated mice. Herein, the effects of NK cells on macrophage polarization were examined in vitro. We found that NK cells from chlamydial-infected mice (iNK) significantly induced M1 polarization compared to that from sham-infected mice (uNK). Inhibition of miR-155 expression in macrophages attenuated M1 polarization induced by iNK, while miR-155 over-expression enhanced it. Furthermore, neutralization of IFN-γ in the coculture system decreased the expression of miR-155 by macrophages, and resulted in diminished M1 polarization induced by iNK cells. The data indicates that NK cells direct M1 polarization through up-regulation of miR-155 by IFN-γ production, and NK-regulated macrophage polarization is functionally relevant to host defense against chlamydial infection. Overall design: Male BALB/c mice were maintained in a specific pathogen-free animal care facility of Shandong University. All mice used in this study were males between 6 and 8 weeks old. The experiment was divided into NK+macrophage and NK-macrophage group, N=3 per group.

自然杀伤细胞（NK cells）与肺巨噬细胞均为固有免疫的重要组成成分。鼠衣原体（Chlamydia muridarum，C. muridarum）呼吸道感染过程中，NK细胞与肺巨噬细胞之间的互作机制尚未明确。本研究探讨了衣原体肺部感染时，NK细胞对肺巨噬细胞功能的调控作用。

研究发现，NK细胞耗竭会导致肺巨噬细胞从M1型极化为M2型，该现象与肺巨噬细胞内微小RNA-155（miR-155）的表达水平显著下调相关。通过过继转移实验，我们观察到：相较于接受经IgG处理的感染小鼠肺巨噬细胞的受体小鼠，接受来自NK细胞耗竭的衣原体感染小鼠肺巨噬细胞的受体小鼠，在衣原体攻毒后肺部细菌载量更高、肺部炎症更为严重。

本研究进一步在体外验证了NK细胞对巨噬细胞极化的调控作用。结果显示，相较于假感染小鼠来源的NK细胞（uNK），衣原体感染小鼠来源的NK细胞（iNK）可显著诱导巨噬细胞向M1型极化。抑制巨噬细胞内miR-155的表达可削弱iNK诱导的M1极化，而过表达miR-155则可增强该极化效应。进一步研究发现，在共培养体系中中和干扰素-γ（IFN-γ）可降低巨噬细胞的miR-155表达水平，进而减弱iNK诱导的M1极化。

上述实验数据表明，NK细胞可通过分泌IFN-γ上调巨噬细胞内miR-155的表达，从而诱导巨噬细胞向M1型极化；且NK细胞调控的巨噬细胞极化与宿主抗衣原体感染的免疫防御功能密切相关。

总体实验设计：将6~8周龄的雄性BALB/c小鼠饲养于山东大学无特定病原体（SPF）级动物实验中心。本研究所有实验小鼠均为雄性，分为NK+巨噬细胞组与NK-巨噬细胞组，每组n=3。