血管内皮细胞中三磷酸腺苷-结合盒转运子A1对炎性细胞因子表达的影响

The effects of ATP-binding cassette transporter A1 in vascular endothelial cells on expression of inflammatory cytokines

目的研究在公认的ATP-结合盒转运子A1(ABCA1)诱导因素8-溴-环磷酸腺苷(8-Br-cAMP)刺激下,人血管内皮细胞ECV304中ABCA1基因对炎性细胞因子细胞间黏附分子-1(ICAM-1)、单核细胞化学趋向蛋白-1(MCP-1)及白介素-1β(IL-1β)的调节作用。以阐明ABCA1基因在动脉粥样硬化(AS)发生中的作用机制。方法培养人血管内皮细胞株ECV304,加入8-Br-cAMP(0.5 mmol/L)刺激3、6、12及24 h,以荧光定量逆转录聚合酶链式反应检测ABCA1、ICAM-1及MCP-1 mRNA表达量,Western blot蛋白印迹法和酶联免疫吸附测定法检测ABCA1I、CAM-1、MCP-1及IL-1β蛋白质表达量;ABCA1的反义寡核苷酸(100 nmol/L)转染人血管内皮细胞,给予上述的8-Br-cAMP,同样的方法测定上述指标的改变。结果人血管内皮细胞ECV304在给予8-Br-cAMP刺激6、12 h后,ABCA1I、CAM-1、MCP-1的mR-NA和蛋白质水平及IL-1β蛋白质水平均增高;给予反义寡核苷酸转染后,8-Br-cAMP刺激后36、h ABCA1I、CAM-1、MCP-1mRNA的表达降低,122、4 h ABCA1I、CAM-1、MCP-1及IL-1β蛋白质的表达水平降低。结论在8-Br-cAMP作用下,血管内皮细胞中ABCA1可增加炎性因子的表达,从而在AS早期的发生中发挥作用。

Objective To investigate the effects of ATP-binding cassette transporter A1 （ABCA1） on ICAM-1, MCP-1 and IL-1β in vascular endothelial cells ECV304 after the treatment with 8-Br-cAMP in order to find the new possible mechanisms that ABCA1 contributes to atherosclerogenesis. Method 8-Br-cAMP （0.5 mmol/L） was added to the culture media of ECV304 cells which were harvested after 3, 6, 12 and 24 hours. The mRNA levels of ABCAI,ICAM-1 and MCP-1 were measured by real-time fluorescent quantitative RT-PCR, and protein levels of ABCA1, ICAM-1 ,MCP-1 and IL-1β were determined by Western blot and ELISA methods. After phosphorothioate antisense oligonucleotides of ABCA1 mixture were added to the culture media at a final concentration of 100 nmol/L, the same experiments were repeated. Results The mRNA and protein amounts of ABCA1, ICAM-1 and MCP-1 and also protein amount of IL-1β were increased after incubation with 8-Br-cAMP for 6 and 12 hours. After transfection with antisense oligonucleotide of ABCA1, the expression levels of ABCA1, ICAM-1 and MCP-1 mRNA were decreased at 3 and 6 hours, and ABCA1, ICAM-1, MCP-1 and IL- 1β protein levels at 12 and 24 hours. Conclusion ABCA1 could increase expression of inflammatory cytokines in hmnan vascular endothelial cells after 8-Br-cAMP stimulation and so take effects on atherosclerogenesis.