NRP1 在蜕膜基质和免疫细胞中的矛盾表达揭示了早孕期间一种新的炎症平衡机制。

Paradoxical expression of NRP1 in decidual stromal and immune cells reveals a novel inflammation balancing mechanism during early pregnancy.

机构信息

Laboratory for Reproductive Immunology, NHC Key Lab of Reproduction Regulation, Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Hospital of Obstetrics and Gynecology, Shanghai Institute of Planned Parenthood Research), Fudan University Shanghai Medical College, Zhao Zhou Road 413, Shanghai, 200032, China.

Department of Obstetrics and Gynecology, School of Medicine, Shanghai Fourth People's Hospital, Tongji University, Shanghai, 200434, China.

出版信息

Inflamm Res. 2023 Jul;72(7):1341-1357. doi: 10.1007/s00011-023-01734-y. Epub 2023 Jun 16.

DOI:10.1007/s00011-023-01734-y

PMID:37328599

Abstract

OBJECTIVE AND DESIGN

To investigate the balancing mechanisms between decidualization-associated inflammation and pregnancy-related immunotolerance.

MATERIAL OR SUBJECTS

Decidual samples from women with normal pregnancy (n = 58) or unexplained spontaneous miscarriage (n = 13), peripheral blood from normal pregnancy and endometria from non-pregnancy (n = 10) were collected. Primary endometrial stromal cells (ESCs), decidual stromal cells (DSCs), decidual immune cells (DICs) and peripheral blood mononuclear cells (PBMCs) were isolated.

TREATMENT

The plasmid carrying neuropilin-1 (NRP1) gene was transfected into ESC for overexpression. To induce decidualization in vitro, ESCs were treated with a combination of 10 nM estradiol, 100 nM progesterone and 0.5 mM cAMP. Anti-Sema3a and anti-NRP1 neutralizing antibodies were applied to block the ligand-receptor interactions.

METHODS

RNA-seq analysis was performed to identify differentially expressed genes in DSCs and DICs, and NRP1 expression was verified by Western blotting and flow cytometry. The secretion of inflammatory mediators was measured using a multifactor cytometric bead array. The effects of Sema3a-NRP1 pathway on DICs were determined by flow cytometry. Statistical differences between groups were compared using the T test and one way or two-way ANOVA.

RESULTS

Combined with five RNA-seq datasets, NRP1 was the only immune checkpoint changing oppositely between DSCs and DICs. The decreased expression of NRP1 in DSCs allowed intrinsic inflammatory responses required for decidualization, while its increased expression in DICs enhanced tolerant phenotypes beneficial to pregnancy maintenance. DSC-secreted Sema3a promoted immunosuppression in DICs via NRP1 binding. In women with miscarriage, NRP1 was abnormally elevated in DSCs but diminished in decidual macrophages and NK cells.

CONCLUSION

NRP1 is a multifunctional controller that balances the inflammatory states of DSCs and DICs in gravid uterus. Abnormal expression of NRP1 is implicated in miscarriage.

摘要

目的和设计

研究蜕膜化相关炎症与妊娠相关免疫耐受之间的平衡机制。

材料或对象

收集了正常妊娠（n=58）和不明原因自然流产（n=13）女性的蜕膜样本、正常妊娠外周血和非妊娠子宫内膜。分离原代子宫内膜基质细胞（ESCs）、蜕膜基质细胞（DSCs）、蜕膜免疫细胞（DICs）和外周血单核细胞（PBMCs）。

治疗

将携带神经纤毛蛋白-1（NRP1）基因的质粒转染入 ESC 以过表达。为了在体外诱导蜕膜化，用 10 nM 雌二醇、100 nM 孕酮和 0.5 mM cAMP 联合处理 ESCs。应用抗 Sema3a 和抗 NRP1 中和抗体阻断配体-受体相互作用。

方法

通过 RNA-seq 分析鉴定 DSCs 和 DICs 中差异表达的基因，并通过 Western blot 和流式细胞术验证 NRP1 的表达。使用多因子细胞因子 bead 阵列测定炎症介质的分泌。通过流式细胞术确定 Sema3a-NRP1 通路对 DICs 的影响。使用 T 检验和单向或双向 ANOVA 比较组间的差异。

结果

结合五个 RNA-seq 数据集，NRP1 是唯一在 DSCs 和 DICs 之间表达相反的免疫检查点。DSC 中 NRP1 表达下调允许蜕膜化所需的内在炎症反应，而 DIC 中 NRP1 表达上调增强了有利于妊娠维持的耐受表型。DSC 分泌的 Sema3a 通过与 NRP1 结合促进 DIC 中的免疫抑制。在流产的女性中，NRP1 在 DSCs 中异常升高，但在蜕膜巨噬细胞和 NK 细胞中降低。