Departments of Central Laboratory, Jinhua Municipal Central Hospital, Jinhua, Zhejiang, 321000, PR China.
Placenta. 2024 Sep 2;154:1-8. doi: 10.1016/j.placenta.2024.05.137. Epub 2024 May 29.
Toxoplasma gondii is an opportunistic intracellular parasite that is a major pathogenic factor in miscarriage, especially when it occurs early in pregnancy. We have previously demonstrated that the regulation of forkhead box transcription factor (Foxp3) is associated with abortion in early pregnancy caused by excretory-secretory antigen (ESA) of strain China 1. We aimed to reveal the underlying mechanism of miscarriage caused by ESA.
A TLR4 pregnant mouse model was successfully constructed. Pregnant mice at gestational day 5 (G5) were injected with ESA. All animals were sacrificed on G13, pregnancy outcomes were observed, and abortion rates were calculated. Placental status observed by Hematoxylin-eosin staining; gene expression was measured by IHC; flow cytometry analysis was used to determine the number and function of regulatory T cells. In EL4 cells, real-time PCR and Western blot were used to evaluate gene expression and cytokines assay.
In vivo studies revealed that ESA injection caused 83% abortion in pregnant mice but only 35% abortion in TLR4-/- pregnant mice. In addition, ESA attenuated the number and function of regulatory T cells, further suppressed Foxp3, FOXO1 levels, and upregulated CD127 expression. TLR4 mice partially reversed this inhibitory effect on regulatory T cells. Furthermore, in vitro studies revealed that ESA inhibited TLR4/NF-κB signaling pathway expression and that TLR4 agonists significantly restored the ESA-induced decrease in Foxp3.
These findings suggest that ESA suppresses Foxp3 expression by blocking TLR4/NF-κB signaling, resulting in miscarriage. More importantly, the results indicated that miscarriage caused by ESA is TLR4 dependent.
刚地弓形虫是一种机会性细胞内寄生虫，是导致流产的主要致病因素，尤其是在妊娠早期。我们之前的研究表明，叉头框转录因子（Foxp3）的调节与由中国 1 株排泄分泌抗原（ESA）引起的早期妊娠流产有关。我们旨在揭示 ESA 引起流产的潜在机制。
成功构建了 TLR4 妊娠小鼠模型。在妊娠第 5 天（G5）给妊娠小鼠注射 ESA。所有动物均在 G13 处死，观察妊娠结局，计算流产率。通过苏木精-伊红染色观察胎盘状态；通过免疫组化测量基因表达；通过流式细胞术分析确定调节性 T 细胞的数量和功能。在 EL4 细胞中，使用实时 PCR 和 Western blot 评估基因表达和细胞因子测定。
体内研究表明，ESA 注射导致妊娠小鼠 83%流产，但 TLR4-/-妊娠小鼠仅 35%流产。此外，ESA 减弱了调节性 T 细胞的数量和功能，进一步抑制了 Foxp3、FOXO1 水平，并上调了 CD127 的表达。TLR4 小鼠部分逆转了对调节性 T 细胞的这种抑制作用。此外，体外研究表明，ESA 抑制 TLR4/NF-κB 信号通路表达，TLR4 激动剂显著恢复了 ESA 诱导的 Foxp3 减少。
这些发现表明，ESA 通过阻断 TLR4/NF-κB 信号通路抑制 Foxp3 表达，导致流产。更重要的是，结果表明 ESA 引起的流产依赖于 TLR4。
