ApoAⅠ促进THP-1源性泡沫细胞ApoE分泌的研究

摘要

Objective We hypothesized that interaction between ApoAⅠ and ApoE contributes to the antiatherosclerosis. To this end,we set up the cell models of atherosclerosis to investigate the effects of ApoAⅠ on secretion of ApoE in human THP-1 macrophages. Methods Human plasma HDL was first isolated and purified by PEG-6000 of different density,and ApoAⅠ was then separated with gel-filtration chromatography from the HDL. Human monocyte cell line THP-1 was stimulated and transformed to macrophages by PMA,then incubated with ox-LDL. To set up the cell models of atherosclerosis,THP-1 macrophages were incubated with ApoAⅠ at dose range of 0,5,10,15,and 25 g/ml. In the time course experiment,THP-1 macrophages were incubated with 25 g/ml ApoAⅠ for 0,3,6,12,and 24 hours,respectively. Secretion of ApoE was detected with ELISA. The mRNA level of ApoE was determined by reverse transcription polymerase chain reaction(RT-PCR).Results ApoAⅠ increased secretion of ApoE out of THP-1 derived foam macrophages. Secretion of ApoE was increase with the dose range and time course,reaching the highest amount correspondingly at the time point of 24 hour incubation with ApoAⅠ and the dose of 25 g/ml ApoAⅠ. The effect of ApoAⅠ on the mRNA of ApoE was not obvious.Conclusion ApoAⅠ stimulates ApoE secretion by THP-1 derived foam macrophages. The amount of ApoE secretion is time-course and dose dependent on ApoAⅠ. But ApoAⅠ has no effect on the mRNA level of ApoE.%目的 通过建立动脉粥样硬化损伤部位细胞模型,观察ApoAⅠ对THP-1源性泡沫细胞 ApoE分泌的影响,探讨ApoAⅠ与ApoE之间是否存在相互关系以及这种关系对动脉粥样硬化的影响.方法 采用聚乙二醇-6000(PEG-6000)沉淀法制备人血浆HDL,再利用凝胶过滤柱层析法分离ApoAⅠ;佛波酯(PMA)诱导THP-1单核细胞分化为巨噬细胞,氧化型低密度脂蛋白(oxidized low density lipoprotein,ox-LDL)使分化后THP-1细胞荷脂,建立动脉粥样硬化细胞模型;ELISA检测不同浓度ApoAⅠ(0,5,10,15,25 μg/ml)及不同孵育时间(0,3,6,12,24 h),THP-1源性泡沫细胞ApoE的分泌;RT-PCR检测细胞ApoE mRNA的表达情况.结果 ApoAⅠ增加THP-1源性泡沫细胞ApoE的分泌量;且ApoE蛋白质的分泌随着ApoAⅠ孵育时间增加而增加,在24 h ApoE的分泌量达最大;在剂量试验中,ApoE的分泌量随着ApoAⅠ剂量的增加有增加趋势,ApoAⅠ浓度为25 μmol/L时,ApoE分泌量最大;而ApoE基因的表达不受ApoAⅠ的影响.结论 一定浓度的ApoAⅠ促进THP-1源性泡沫细胞ApoE的分泌,且具有时间及剂量依赖性.而在基因水平上,ApoAⅠ对ApoE mRNA表达没有影响.