ABSTRACT

Endothelial cell dysfunction and inflammatory responses play critical roles in the development of atherosclerosis. Recent data on the processes underlying atherogenesis indicate the substantial role of endotoxins (lipopolysaccharides; LPS) of the intestinal microflora in the initiation and progression of atherosclerosis. Mitogen-activated protein (MAP) kinase phosphatase-3 (MKP-3) is a cytoplasmic dual-specificity protein phosphatase that specifically binds to and inactivates MAP kinases in mammalian cells, but its biological function in endothelial cell dysfunction and inflammatory responses remains largely unknown. The aim of the present study was to investigate the role of MKP-3 in endotoxin-induced endothelial inflammation by western blotting, quantitative polymerase chain reaction, and immunofluorescence. The results of our study demonstrated that MKP-3 overexpression markedly inhibited the adhesion of human monocytic THP-1 cells to human umbilical vein endothelial cells (HUVECs) by downregulating the expression of vascular cell adhesion protein 1 (VCAM-1) and pro-inflammatory cytokines. In contrast, MKP-3-encoding gene knockdown by small interfering RNA (siRNA) exacerbated LPS-induced endothelial dysfunction. Additionally, we found that MKP-3 overexpression inhibited LPS-induced p38 MAPK phosphorylation and decreased the nuclear translocation of nuclear factor kappa B (NF-κB) after LPS treatment, suggesting its implication in the LPS/Toll-like receptor 4 (TLR4)/p38/NF-κB pathway. Overall, these observations suggest that MKP-3 plays a protective role in endothelial dysfunction and may be a therapeutic target.

摘要

内皮细胞功能障碍和炎症反应在动脉粥样硬化的发展中起着关键作用。最近关于动脉粥样硬化过程的数据表明，肠道菌群的内毒素（脂多糖；LPS）在动脉粥样硬化的发生和发展中起重要作用。丝裂原活化蛋白 (MAP) 激酶磷酸酶-3 (MKP-3) 是一种细胞质双特异性蛋白磷酸酶，可特异性结合哺乳动物细胞中的 MAP 激酶并使其失活，但其在内皮细胞功能障碍和炎症反应中的生物学功能仍然未知. 本研究的目的是通过蛋白质印迹、定量聚合酶链反应和免疫荧光研究 MKP-3 在内毒素诱导的内皮炎症中的作用。我们的研究结果表明，MKP-3 过表达通过下调血管细胞粘附蛋白 1 (VCAM-1) 的表达和促炎症反应，显着抑制人单核细胞 THP-1 细胞与人脐静脉内皮细胞 (HUVECs) 的粘附。细胞因子。相比之下，小干扰 RNA (siRNA) 对 MKP-3 编码基因的敲低加剧了 LPS 诱导的内皮功能障碍。此外，我们发现 MKP-3 过表达抑制了 LPS 诱导的 p38 MAPK 磷酸化并减少了 LPS 处理后核因子 kappa B (NF-κB) 的核转位，表明其对 LPS/Toll 样受体 4 (TLR4) 的影响/p38/NF-κB 通路。总体而言，这些观察结果表明 MKP-3 在内皮功能障碍中发挥保护作用，可能是治疗靶点。