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Modulating Macrophage Polarization through CCR2 Inhibition and Multivalent Engagement
Molecular Pharmaceutics ( IF 4.9 ) Pub Date : 2018-05-23 00:00:00 , DOI: 10.1021/acs.molpharmaceut.8b00237
Michael B. Deci 1 , Scott W. Ferguson 1 , Sydney L. Scatigno 1 , Juliane Nguyen 1
Affiliation  

Excessive or prolonged recruitment of inflammatory monocytes to damaged tissue can significantly worsen patient outcomes. Monocytes migrate to sites of tissue inflammation in response to high local concentrations of CCL2, a chemokine that binds to and signals through the CCR2 receptor. While the role of CCR2 in cellular migration is well studied, it is unclear how CCR2 inhibition affects macrophage polarization and if multivalency can increase downstream signaling effects. Using affinity selection with a phage library, we identified a novel single-chain variable fragment (scFv) (58C) that binds specifically and with high affinity to the N-terminal domain of CCR2 (KD = 59.8 nM). The newly identified 58C-scFv bound to native CCR2 expressed on macrophages and MDA-MB-231 cells, inhibited migration, and induced a pro-inflammatory M1-phenotype in macrophages. The M1/M2 macrophage phenotype ratio for monomeric 58C-scFv was significantly increased over the negative control by 1.0 × 104-fold (iNOS/Arg-1), 5.1 × 104-fold (iNOS/Mgl2), 3.4 × 105-fold (IL-6/Arg-1), and 1.7 × 106-fold (IL-6/Mgl2). The multivalent display of 58C-scFv on liposomes further reduced migration of both cell types by 25–40% and enhanced M1 polarization by 200% over monomeric 58C-scFv. These studies demonstrate that CCR2 inhibition polarizes macrophages toward an inflammatory M1 phenotype, and that multivalent engagement of CCR2 increases the effects of 58C-scFv on polarization and migration. These data provide important insights into the role of multivalency in modulating binding, downstream signaling, and cellular fate.

中文翻译:

通过抑制CCR2和多价参与来调节巨噬细胞极化。

炎性单核细胞过多或长时间募集到受损组织会显着恶化患者预后。单核细胞响应于局部高浓度的CCL2迁移到组织炎症部位,CCL2是一种结合CCR2受体并通过其发出信号的趋化因子。尽管对CCR2在细胞迁移中的作用进行了深入研究,但尚不清楚CCR2抑制作用如何影响巨噬细胞极化以及多价是否可以增加下游信号传导作用。使用与噬菌体库的亲和力选择,我们确定了一种新颖的单链可变片段(scFv)(58C),该片段特异性结合并以高亲和力与CCR2的N端结构域(K D= 59.8 nM)。新近鉴定的58C-scFv与巨噬细胞和MDA-MB-231细胞上表达的天然CCR2结合,抑制迁移并诱导巨噬细胞中的促炎性M1-表型。单体58C-scFv的M1 / M2巨噬细胞表型比率比阴性对照显着增加1.0×10 4倍(iNOS / Arg-1),5.1×10 4倍(iNOS / Mgl2),3.4×10 5倍(IL-6 / Arg-1)和1.7×10 6倍(IL-6 / Mgl2)。与单体58C-scFv相比,脂质体上58C-scFv的多价展示进一步将两种细胞类型的迁移减少了25-40%,将M1极化增强了200%。这些研究表明,CCR2抑制作用使巨噬细胞偏向炎症性M1表型,而CCR2的多价结合增加了58C-scFv对极化和迁移的影响。这些数据提供了重要的见解,了解多价在调节结合,下游信号传导和细胞命运中的作用。
更新日期:2018-05-23
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