巨噬细胞中ABCA1对炎症因子的调节及其意义

Modulation of inflammatory cytokines by ATP-binding cassette A1 in THP-1 macrophages

目的研究在8-溴-环磷酸腺苷(8-Br-cAMP)刺激下,巨噬细胞中三磷酸腺苷结合盒转运子(ABCA1)对细胞间黏附分子-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,在细胞水平证实ABCA1在动脉粥样硬化发生中的作用机制。方法用氟波酯(PMA)刺激THP-1细胞使之转变为巨噬细胞,8-Br-cAMP(0.5mmol/L)刺激3、6、12、24h后,以荧光定量RT-PCR和Western蛋白印迹法及ELISA法检测ABCA1、ICAM-1、MCP-1及IL-1βmRNA和蛋白质表达量;用反义寡核苷酸(100nmol/L)抑制ABCA1的表达,观察Ox-LDL刺激下上述指标的改变。结果予8-Br-cAMP刺激6、12h后,巨噬细胞ABCA1、ICAM-1、MCP-1mRNA和蛋白质水平及IL-1β蛋白质水平均增高;反义寡核苷酸转染巨噬细胞,8-Br-cAMP刺激后3、6h,ABCA1、ICAM-1、MCP-1mRNA的表达降低,刺激后12、24h,ABCA1、ICAM-1、MCP-1及IL-1β蛋白质的表达水平降低。结论在8-Br-cAMP刺激下,巨噬细胞ABCA1可增加炎症因子表达,参与动脉粥样硬化的发生。

Objective To observe the effects of ATP-binding cassette A1 （ABCA1） on intercellular cell adhesion molecule type 1 （ICAM- 1 ）, monocyte chemoattractant protein- 1 （MCP- 1 ） and interleukin- 1 β （IL- 1 β） in THP- 1 macrophages stimulated with 8-Br-cAMP to identify a possible new mechanism that ABCA1 contributes to atherosclerogenesis （AS）. Methods Monocytic THP-1 cells were cultured in the presence of 100 nmol/L phorbol myristate acetate （PMA） for 72 h to transform the cells into THP-1 macrophages. After the macrophages were stimulated with 8-Br-cAMP （final concentration 0.5 mmol/L） for 3, 6, 12 and 24 h respectively, the amounts ofABCA1, ICAM-1 and MCP-1 mRNA were examined by real-time fluorescent quantitative RT-PCR, and the protein amounts of ABCA1, ICAM-1, MCP-1 and IL-113 were determined by Western blotting and enzyme-linked immunosorbent assay （ELISA）. Phosphorothioate antisense oligonucleotides of ABCA1 were add into the culture media at a final concentration of 100 nmolFL and the experiments were repeated. Results ABCA1, 1CAM-1 and MCP-1 mRNA and protein and IL-1β protein were increased in the macrophages after stimulation with 8-Br-cAMP for 6 and 12 h. The mRNA expressions of ABCA1, ICAM-1 and MCP-1 were decreased significantly at 3 and 6 h （P〈0.01）, and the protein expressions of ABCA1, ICAM-1, MCP-1 and IL-1β declined significantly at 12 and 24 h （P〈0.01） after transfection of the macrophages with antisense oligonucleotides of ABCA1. Conclusion ABCA1 can increase the expressions of the inflammatory cytokines in THP-1 macrophages stimulated by 8-Br-cAMP and plays a role in the pathogenesis of AS.