炎性环境下内皮微粒介导单核细胞促内皮细胞增殖的作用

目的 探讨炎性环境中内皮微粒(EMPs)在单核细胞参与的促内皮细胞增殖中的作用,并探讨EMPs对单核细胞表达内皮细胞标志物的影响.方法 体外培养人脐静脉内皮细胞(HUVECs),炎性因子肿瘤坏死因子α(TNF-α)刺激内皮细胞,超速离心法获取EMPs；用透射电镜观察其形态和结构；运用酶联免疫吸附法(ELISA)检测经EMPs激活的单核细胞株THP-1对血管内皮生长因子(VEGF)中VEGF-A、VEGF-C的表达情况；将HUVECs和THP-1细胞或者经EMPs诱导的THP-1细胞共培养,检测HUVECs的增殖情况；运用RT-PCR和免疫荧光细胞化学技术,观察经EMPs激活的THP-1细胞对内皮细胞标志物vWF和血管内皮生长因子受体2(VEGFR-2)的表达情况.结果 内皮细胞经炎性因子刺激后释放EMPs,EMPs刺激的THP-1细胞上清中VEGF-A和VEGF-C蛋白水平明显升高；HUVECs细胞增殖实验中,与THP-1细胞或者经EMPs诱导的THP-1共培养的两种情况下,HUVECs均显著增多；与EMPs共培养3d后,THP-1中vWF和VEGFR-2蛋白表达为阳性,基因表达为阴性,共培养8d后,vWF和VEGFR-2的蛋白和基因表达均为阴性.结论 炎性环境中内皮细胞释放EMPs可以促使单核细胞分泌VEGF-A和VEGF-C,后两者可使HUVECs增殖,EMPs仅能使单核细胞一过性呈现内皮表型,而不能将其转分化为内皮细胞.

Effect of endothelial microparticles on monocytes to promote endothelial cell proliferation in inflammation

Objective To investigate the effect of endothelial microparticles (EMPs) on monocytes in promoting inflammatory angiogenesis and in expressing endothelial markers. Methods EMPs were isolated from the supernatants of cultured human umbilical vein endothelial cells (HUVECs) treated with TNF-αby ultracentrifugation, identified under an electron microscope and applied to cultured THP-1 cells. The level of VEGF-A and VEGF-C expressed by THP-1 cells was examined with ELISA. The expression of von Willebrand factor (vWF) and vascular endothelial growth factor receptor 2 (VEGFR-2) in THP-1 cells were studied with RT-PCR and immunofluorescence. HUVECs were co-cultured with THP-1 cells either treated with EMPs not. The proliferation of HUVECs was assessed. Results TNF-α enhanced HUVECs to release EMPs, which promoted THP-1 cells to secrete vascular endothelial growth factor A (VEGF-A) and vascular endothelial growth factor C (VEGF-C). THP-1 cells did not express vWF or VEGFR-2 genetically after either 3 days or 8 days of culture. When cultured with THP-1 cells, HUVECs were more active in proliferation. Conclusion EMPs are able to activate monocytes which in turn enhanced endothelial cells growth by secreting