Microglia plays crucial roles in Alzheimer’s disease (AD) development. Triggering receptor expressed on myeloid cells 2 (TREM2) in association with DAP12 mediates signaling affecting microglia function. Here we study the negative regulation of TREM2 functions by leukocyte immunoglobulin-like receptor subfamily B member 2 (LILRB2), an inhibitory receptor bearing ITIM motifs. To specifically interrogate LILRB2-ligand (oAβ and PS) interactions and microglia functions, we generated potent antagonistic LILRB2 antibodies with sub-nanomolar level activities. The biological effects of LILRB2 antagonist antibody (Ab29) were studied in human induced pluripotent stem cell (iPSC)–derived microglia (hMGLs) for migration, oAβ phagocytosis, and upregulation of inflammatory cytokines. Effects of the LILRB2 antagonist antibody on microglial responses to amyloid plaques were further studied in vivo using stereotaxic grafted microglia in 5XFAD mice. We confirmed the expression of both LILRB2 and TREM2 in human brain microglia using immunofluorescence. Upon co-ligation of the LILRB2 and TREM2 by shared ligands oAβ or PS, TREM2 signaling was significantly inhibited. We identified a monoclonal antibody (Ab29) that blocks LILRB2/ligand interactions and prevents TREM2 signaling inhibition mediated by LILRB2. Further, Ab29 enhanced microglia phagocytosis, TREM2 signaling, migration, and cytokine responses to the oAβ-lipoprotein complex in hMGL and microglia cell line HMC3. In vivo studies showed significantly enhanced clustering of microglia around plaques with a prominent increase in microglial amyloid plaque phagocytosis when 5XFAD mice were treated with Ab29. This study revealed for the first time the molecular mechanisms of LILRB2-mediated inhibition of TREM2 signaling in microglia and demonstrated a novel approach of enhancing TREM2-mediated microglia functions by blocking LILRB2-ligand interactions. Translationally, a LILRB2 antagonist antibody completely rescued the inhibition of TREM2 signaling by LILRB2, suggesting a novel therapeutic strategy for improving microglial functions.

中文翻译：

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LILRB2 介导的 TREM2 信号传导抑制抑制小胶质细胞功能

小胶质细胞在阿尔茨海默病 (AD) 的发展中起着至关重要的作用。与 DAP12 相关的骨髓细胞 2 (TREM2) 上表达的触发受体介导影响小胶质细胞功能的信号传导。在这里，我们研究了白细胞免疫球蛋白样受体亚家族 B 成员 2 (LILRB2) 对 TREM2 功能的负调控，这是一种带有 ITIM 基序的抑制性受体。为了专门询问 LILRB2-配体（oAβ 和 PS）相互作用和小胶质细胞功能，我们产生了具有亚纳摩尔水平活性的强效拮抗 LILRB2 抗体。LILRB2 拮抗剂抗体 (Ab29) 在人类诱导多能干细胞 (iPSC) 衍生的小胶质细胞 (hMGLs) 中对迁移、oAβ 吞噬作用和炎性细胞因子上调的生物学效应进行了研究。在 5XFAD 小鼠中使用立体定向移植的小胶质细胞进一步研究了 LILRB2 拮抗剂抗体对小胶质细胞对淀粉样斑块反应的影响。我们使用免疫荧光证实了 LILRB2 和 TREM2 在人脑小胶质细胞中的表达。在 LILRB2 和 TREM2 通过共享配体 oAβ 或 PS 共连接后，TREM2 信号传导被显着抑制。我们鉴定了一种单克隆抗体 (Ab29)，可阻断 LILRB2/配体相互作用并阻止 LILRB2 介导的 TREM2 信号传导抑制。此外，Ab29 增强了小胶质细胞的吞噬作用、TREM2 信号传导、迁移和细胞因子对 hMGL 和小胶质细胞系 HMC3 中 oAβ-脂蛋白复合物的反应。体内研究表明，当用 Ab29 处理 5XFAD 小鼠时，斑块周围小胶质细胞的聚集显着增强，小胶质细胞淀粉样蛋白斑块的吞噬作用显着增加。这项研究首次揭示了 LILRB2 介导的小胶质细胞中 TREM2 信号传导抑制的分子机制，并展示了一种通过阻断 LILRB2-配体相互作用来增强 TREM2 介导的小胶质细胞功能的新方法。从转化上讲，LILRB2 拮抗剂抗体完全挽救了 LILRB2 对 TREM2 信号传导的抑制，这表明了一种改善小胶质细胞功能的新治疗策略。这项研究首次揭示了 LILRB2 介导的小胶质细胞中 TREM2 信号传导抑制的分子机制，并展示了一种通过阻断 LILRB2-配体相互作用来增强 TREM2 介导的小胶质细胞功能的新方法。从转化上讲，LILRB2 拮抗剂抗体完全挽救了 LILRB2 对 TREM2 信号传导的抑制，这表明了一种改善小胶质细胞功能的新治疗策略。这项研究首次揭示了 LILRB2 介导的小胶质细胞中 TREM2 信号传导抑制的分子机制，并展示了一种通过阻断 LILRB2-配体相互作用来增强 TREM2 介导的小胶质细胞功能的新方法。从转化上讲，LILRB2 拮抗剂抗体完全挽救了 LILRB2 对 TREM2 信号传导的抑制，这表明了一种改善小胶质细胞功能的新治疗策略。