Role of TLR-4 in anti-2-glycoprotein I-induced activation of peritoneal macrophages and vascular endothelial cells in mice
Role of TLR-4 in anti-2-glycoprotein I-induced activation of peritoneal macrophages and vascular endothelial cells in mice
复制标题
DOI：
10.3892/mmr.2019.10084
复制
发表时间：
2019-05-01
影响因子：
3.4
通讯作者：
Zhou, Hong
中科院分区：
文献类型：
--
作者：
Wang, Meiyun;Kong, Xiangmin;Zhou, Hong
抗磷脂综合征 (APS) 是一种系统性自身免疫性疾病，与抗磷脂抗体 (aPL) 的存在、复发性血栓形成和妊娠期胎儿发病有关。 Toll 样受体 4 (TLR-4) 是 TLR 家族的成员，已知在病原体识别和先天免疫激活中具有重要作用。 2-糖蛋白 I (2GPI) 是一种在血液中以高浓度循环的蛋白质，能够清除脂多糖 (LPS) 并清除循环中不需要的阴离子细胞残留物，例如微粒。我们前期的研究表明，TLR-4及其信号通路有助于体外抗2GPI诱导的单核细胞中促凝血因子和促炎细胞因子的上调。本研究旨在明确 TLR-4 在体内的作用。通过腹腔注射抗2GPI免疫球蛋白G（IgG）刺激C3H/HeN小鼠（TLR-4完整）和C3H/HeJ小鼠（TLR-4缺陷），然后提取处理小鼠的腹腔巨噬细胞和血管内皮细胞（VEC），并分析促炎细胞因子和粘附分子的表达谱。结果表明，C3H/HeN 小鼠（TLR-4 完整）腹腔巨噬细胞中促炎细胞因子（包括肿瘤坏死因子-（TNF-）、白细胞介素（IL）-1 和 IL-6）以及粘附分子（包括细胞间细胞粘附分子-1（ICAM-1）、血管细胞粘附分子-1（VCAM-1）和 E-选择素）在 VEC 中的表达显着高于对照组。 C3H/HeJ 小鼠（TLR-4 缺陷）。与 C3H/HeJ 小鼠（TLR-4 缺陷）相比，用抗 2GPI-IgG 刺激的 C3H/HeN 小鼠腹腔巨噬细胞和 VEC 中 p38 丝裂原激活蛋白激酶 (MAPK) 和核因子 B (NF-B) p65 的磷酸化水平显着升高。同型对照抗体 (NR-IgG) 对腹膜巨噬细胞和 VEC 没有这种影响。此外，TLR-4、p38 MAPK 和 NF-B 抑制剂可显着降低 TLR-4 完整小鼠腹膜巨噬细胞中抗 2GPI-IgG 诱导的 TNF、IL-1 和 IL-6 mRNA 表达。结果表明，TLR-4信号转导途径参与抗2GPI-IgG诱导的腹膜巨噬细胞和VEC的激活。该研究为后续研究阐明抗磷脂综合征的病理机制提供了基础。
Anti-phospholipid syndrome (APS) is a systematic autoimmune disease that is associated with presence of antiphospholipid antibodies (aPL), recurrent thrombosis, and fetal morbidity in pregnancy. Toll-like receptor-4 (TLR-4), a member of TLR family, is known to have a fundamental role in pathogen recognition and activation of innate immunity. The 2-glycoprotein I (2GPI), a protein circulating in the blood at a high concentration, is able of scavenging lipopolysaccharide (LPS) and clear unwanted anionic cellular remnants, such as microparticles, from the circulation. Our previous study demonstrated that TLR-4 and its signaling pathways contribute to the upregulation of pro-coagulant factors and pro-inflammatory cytokines in monocytes induced by anti-2GPI in vitro. The present study aimed to define the roles of TLR-4 in vivo. C3H/HeN mice (TLR-4 intact) and C3H/HeJ mice (TLR-4 defective) were stimulated with an intraperitoneal injection with anti-2GPI-immunoglobulin G(IgG), then peritoneal macrophages and vascular endothelial cells (VECs) were extracted from treated mice, and analyses were conducted on the expression profiles of pro-inflammatory cytokines and adhesion molecules. The results demonstrated that the expression of pro-inflammatory cytokines, including tumor necrosis factor- (TNF-), interleukin (IL)-1 and IL-6, in the peritoneal macrophages, and adhesion molecules, including intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin, in VECs of C3H/HeN mice (TLR-4 intact) were significantly higher than those of C3H/HeJ mice (TLR-4 defective). The phosphorylation levels of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-B (NF-B) p65 in peritoneal macrophages and VECs from C3H/HeN mice stimulated with anti-2GPI-IgG were significantly increased compared with those from C3H/HeJ mice (TLR-4 defective). The isotype control antibody (NR-IgG) had no such effects on peritoneal macrophages and VECs. Furthermore, the inhibitors of TLR-4, p38 MAPK and NF-B may significantly reduce the anti-2GPI-IgG-induced TNF-, IL-1 and IL-6 mRNAs expression in the peritoneal macrophages from TLR-4 intact mice. The results indicated that a TLR-4 signal transduction pathway is involved in anti-2GPI-IgG-induced activation of peritoneal macrophages and VECs. This study has provided a basis for subsequent investigations to elucidate the pathological mechanisms underlying anti-phospholipid syndrome.