Supplementary Material for: Defective Inflammatory Monocyte Development in IRF8-Deficient Mice Abrogates Migration to the West Nile Virus-Infected Brain

IRF8 (interferon-regulatory factor-8) plays a critical role in regulating myeloid cell differentiation. However, the role of this transcription factor in the development of Ly6C+ inflammatory monocytes and their migration to the infected brain has not been examined. We have previously shown that West Nile virus (WNV) infection of wild-type (WT) mice triggers a significant increase in numbers of Ly6C+ monocytes in the bone marrow. These cells traffic via the blood to the infected brain, where they give rise to proinflammatory macrophages. Here, we show that WNV-infected IRF8-deficient (IRF8-/-) mice had significantly reduced numbers of Ly6C+ monocytes in the periphery, with few of these cells found in the blood. Furthermore, low numbers of inflammatory monocyte-derived macrophages were observed in the brains of IRF8-/- mice throughout infection. Adoptive transfer of IRF8-/- Ly6C+ monocytes demonstrated that these cells were intrinsically unable to traffic to the inflamed brain. Low expression of the chemokine receptor CCR2 and integrin VLA-4 by IRF8-/- monocytes likely contributed to this defect, as the interactions between these proteins and their ligands are critical for monocyte egress and migration to inflammatory foci. These data highlight a critical role for IRF8 in inflammatory monocyte differentiation and migration during WNV infection.

干扰素调节因子8（interferon-regulatory factor-8，IRF8）在调控髓系细胞分化过程中发挥关键作用。然而，该转录因子在Ly6C+炎性单核细胞（Ly6C+ inflammatory monocytes）的发育及其向感染脑组织迁移过程中的作用尚未得到研究。我们此前的研究表明，野生型（wild-type，WT）小鼠感染西尼罗河病毒（West Nile virus, WNV）后，骨髓内Ly6C+炎性单核细胞的数量会显著升高。此类细胞经血液循环迁移至受感染的脑组织，并分化为促炎性巨噬细胞。本研究显示，感染WNV的IRF8缺陷型（IRF8-/-）小鼠外周组织中Ly6C+炎性单核细胞数量显著减少，血液中此类细胞亦极为罕见。此外，在整个感染过程中，IRF8-/-小鼠脑组织内由炎性单核细胞分化而来的巨噬细胞数量均处于较低水平。对IRF8-/-小鼠的Ly6C+炎性单核细胞进行过继转移实验证实，此类细胞本身无法向炎性脑组织迁移。IRF8-/-单核细胞表面趋化因子受体CCR2（chemokine receptor CCR2）与整合素VLA-4（integrin VLA-4）的低表达可能是这一迁移缺陷的成因——上述蛋白与其配体的相互作用对于单核细胞迁出循环系统及向炎性病灶迁移至关重要。上述数据表明，IRF8在WNV感染过程中的炎性单核细胞分化与迁移过程中发挥关键调控作用。