Zhang Xiaolei, Xie Yachao, Zhou Hong, Xu Ya, Liu Jingjing, Xie Hongxiang, Yan Jinchuan
Department of Cardiology, Affiliated Hospital of Jiangsu University.
J Atheroscler Thromb. 2014;21(11):1140-51. doi: 10.5551/jat.24372. Epub 2014 Jul 5.
It has been reported that oxidized low-density lipoprotein (oxLDL) forms a stable and non-dissociable complex with β2-glycoprotein I (β2GPI) and that IgG anti-β2GPI autoantibodies are able to recognize this complex, thus facilitating macrophage-derived foam cell formation in patients with antiphospholipid syndrome (APS). However, the immunopathological mechanisms of oxLDL/β2GPI complexes in promoting foam cell formation are not fully understood. In this study, we examined the role of toll-like receptor 4 (TLR4) in the oxLDL/β2GPI/anti-β2GPI complex-induced transformation of mouse peritoneal macrophages to foam cells.
Oil red O staining and optical density (OD) measurements of intracellular stained oil red O solution were used to monitor the transformation of peritoneal macrophages to foam cells in TLR4-competent C3H/HeN and TLR4-mutant C3H/HeJ mice. During foam cell formation induced by the oxLDL/β2GPI/anti-β2GPI complex, the expression of TLR4 and activation of nuclear factor kappa B (NF-κB) were confirmed by analyzing the protein and mRNA levels of these compounds. Furthermore, the related active molecule expression during foam cell formation induced by the oxLDL/β2GPI/anti-β2GPI complex was examined in the presence or absence of TLR4.
The data showed that treatment with the oxLDL/β2GPI/anti-β2GPI complex markedly increased foam cell formation, the TLR4 expression, NF-κB activation, the tissue factor (TF) expression and tumor necrosis factor-α (TNF-α) and monocyte chemotactic protein-1 (MCP-1) secretion in the C3H/HeN mice. However, the transformation of macrophages to foam cells and the expression levels of phosphorylated NF-κB, TF, TNF-α and MCP-1 were significantly reduced in the C3H/HeJ mice treated with the oxLDL/β2GPI/anti-β2GPI complex. In addition, compared with that achieved by oxLDL alone, the oxLDL/β2GPI complex decreased foam cell formation and the related signaling molecule expression in the C3H/HeN mice.
Our results indicate that TLR4 plays an important role in the process of oxLDL/β2GPI/anti-β2GPI complex-induced transformation of macrophages to foam cells, which may accelerate the development of atherosclerosis in the setting of APS. However, β2GPI alone functions as an antiatherogenic protein by preventing the foam cell formation induced by oxLDL.
据报道，氧化型低密度脂蛋白（oxLDL）与β2糖蛋白I（β2GPI）形成稳定且不可解离的复合物，抗β2GPI自身抗体IgG能够识别该复合物，从而促进抗磷脂综合征（APS）患者巨噬细胞源性泡沫细胞的形成。然而，oxLDL/β2GPI复合物促进泡沫细胞形成的免疫病理机制尚未完全阐明。在本研究中，我们检测了Toll样受体4（TLR4）在oxLDL/β2GPI/抗β2GPI复合物诱导的小鼠腹腔巨噬细胞向泡沫细胞转化中的作用。
采用油红O染色及对细胞内油红O染色溶液进行光密度（OD）测定，以监测野生型TLR4的C3H/HeN小鼠和TLR4突变型的C3H/HeJ小鼠腹腔巨噬细胞向泡沫细胞的转化。在oxLDL/β2GPI/抗β2GPI复合物诱导泡沫细胞形成过程中，通过分析这些化合物的蛋白质和mRNA水平，确认TLR4的表达及核因子κB（NF-κB）的激活。此外，在有无TLR4的情况下，检测oxLDL/β2GPI/抗β2GPI复合物诱导泡沫细胞形成过程中相关活性分子的表达。
数据显示，用oxLDL/β2GPI/抗β2GPI复合物处理显著增加了C3H/HeN小鼠的泡沫细胞形成、TLR4表达、NF-κB激活、组织因子（TF）表达以及肿瘤坏死因子-α（TNF-α）和单核细胞趋化蛋白-1（MCP-1）的分泌。然而，用oxLDL/β2GPI/抗β2GPI复合物处理的C3H/HeJ小鼠中，巨噬细胞向泡沫细胞的转化以及磷酸化NF-κB、TF、TNF-α和MCP-1的表达水平显著降低。此外，与单独使用oxLDL相比，oxLDL/β2GPI复合物降低了C3H/HeN小鼠的泡沫细胞形成及相关信号分子的表达。
我们的结果表明，TLR4在oxLDL/β2GPI/抗βGPI复合物诱导巨噬细胞向泡沫细胞转化的过程中起重要作用，这可能会加速APS患者动脉粥样硬化的发展。然而，单独的β2GPI通过阻止oxLDL诱导的泡沫细胞形成而发挥抗动脉粥样硬化蛋白的作用。
