Toxoplasma gondii-induced adverse pregnancy outcomes: insight into the inhibitory role of Trem2 on TLR4/TRAF6/JNK signaling pathway

Decidual macrophages (dMφs) are not only essential for maintaining normal pregnancy but also serve as crucial immune defenders against infections, including Toxoplasma gondii (T. gondii). Triggering receptor expressed on myeloid cells 2 (Trem2), as a critical immunoregulatory receptor on dMφs, can counteract inflammation and defend against pathogen infection. However, the mechanisms underlying the Trem2 downstream pathways during T. gondii infection—particularly their impact on adverse pregnancy outcomes (APOs)—remain elusive. Here, via establishing T. gondii-challenged wild type and Trem2-/- mouse pregnancy models, we observed that genetic ablation of Trem2 exacerbated T. gondii-triggered APOs. Importantly, the interaction of Trem2 with Toll-like receptor 4 (TLR4) was predicated by molecular docking models with a calculated binding energy (-15 kcal/mol) and confirmed by co-IP. Both animal and cellular models were employed, revealing that Trem2 downregulation in the response to T. gondii, concomitant with an increase in TLR4 and its downstream signaling molecules like TNF receptor-associated factor 6 (TRAF6) and c-Jun N-terminal kinase (JNK). More importantly, Trem2 deficiency in mice and macrophages further enhanced the activation of TLR4/TRAF6/JNK signaling axis. In contrast, Trem2 overexpression in macrophages inhibited the expression of the downstream signaling cascade and reversed the activation induced by T. gondii antigens. Additionally, the treatment with a TLR4-blocking antibody suppressed the activation of TRAF6 and P-JNK in both Raw 264.7 cells and bone marrow-derived macrophages stimulated with T. gondii antigens, whereas it failed to inhibit Trem2 expression. Taken together, Trem2 deficiency in mice promotes the TLR4/TRAF6/JNK signaling cascade, contributing to exacerbate APOs following T. gondii infection. This study provides novel mechanistic insights into T. gondii-induced pregnancy outcomes, highlighting the pregnancy-specific inhibitory function of Trem2 on TLR4/TRAF6/JNK signaling pathway.

蜕膜巨噬细胞（decidual macrophages，dMφs）不仅是维持正常妊娠的必需细胞，同时也是抵御包括刚地弓形虫（Toxoplasma gondii，T. gondii）在内的病原体感染的关键免疫防御细胞。髓系细胞触发受体2（Triggering receptor expressed on myeloid cells 2，Trem2）作为蜕膜巨噬细胞表面的重要免疫调节受体，能够拮抗炎症反应并抵御病原体感染。然而，刚地弓形虫感染过程中Trem2下游信号通路的调控机制——尤其是其对不良妊娠结局（adverse pregnancy outcomes，APOs）的影响——仍有待阐明。
本研究通过构建刚地弓形虫感染的野生型与Trem2基因敲除小鼠妊娠模型，发现Trem2基因缺失会加剧刚地弓形虫诱导的不良妊娠结局。值得注意的是，通过结合能为-15 kcal/mol的分子对接模型预测，并经免疫共沉淀（co-IP）验证，Trem2与Toll样受体4（Toll-like receptor 4，TLR4）存在特异性相互作用。
本研究同时采用动物与细胞两种模型，结果显示在刚地弓形虫刺激下，Trem2表达下调，同时伴随TLR4及其下游信号分子（如肿瘤坏死因子受体相关因子6（TNF receptor-associated factor 6，TRAF6）与c-Jun氨基末端激酶（c-Jun N-terminal kinase，JNK））的表达水平升高。更重要的是，小鼠与巨噬细胞中Trem2的缺失会进一步增强TLR4/TRAF6/JNK信号轴的激活。与之相反，在巨噬细胞中过表达Trem2则会抑制该下游信号级联的表达，并逆转刚地弓形虫抗原诱导的信号激活。
此外，使用TLR4阻断抗体处理经刚地弓形虫抗原刺激的RAW 264.7细胞与骨髓源巨噬细胞，可抑制TRAF6与磷酸化JNK（P-JNK）的激活，但无法抑制Trem2的表达。
综上，小鼠体内Trem2缺失会促进TLR4/TRAF6/JNK信号级联的激活，进而加剧刚地弓形虫感染后的不良妊娠结局。本研究为刚地弓形虫诱导的妊娠结局异常提供了全新的机制视角，揭示了Trem2通过抑制TLR4/TRAF6/JNK信号通路发挥的妊娠特异性调控功能。