Differential Effect of apoE isoform on cholesterol-loaded macrophage

载脂蛋白E（apoE）在脂蛋白代谢和动脉粥样硬化性疾病中起重要作用。在人apoE中存在三种主要的常见同种型，即，E2、E3和E4。已知这些异构体对脂蛋白代谢和动脉粥样硬化具有不同的作用。在本研究中，为了研究这些异构体在1）巨噬细胞胆固醇逆向转运和2）VLDL-甘油三酯脂解中的作用，我们利用腺病毒载体表达这些异构体。加载胆固醇后，用腺病毒载体感染细胞以表达apoE同种型。与LacZ腺病毒感染的对照细胞相比，apoE 2的表达显著降低了细胞内胆固醇水平。ApoE 3和E4也能降低细胞胆固醇含量，但其作用不如apoE 2。在n ...更多信息 在第二个实验中，将负载胆固醇的RAW 264.7细胞与从预先用apoE腺病毒感染的HeLa细胞收获的培养基一起孵育，以阐明外源apoE对胆固醇反向转运的作用。本实验表明，apoE 3在胆固醇逆向转运中是有效的，而apoE 2和E4的作用很小。最后，为了阐明apoE对VLDL-甘油三酯脂解的作用，将腺病毒载体注射到apoE/LDL-受体双缺陷小鼠中，获得含有apoE的VLDL。改变apoE/TG的比值，用牛脂蛋白脂酶对这些VLDL颗粒进行体外脂解试验。本实验结果表明，载脂蛋白E无论在何种异构体上均能抑制VLDL颗粒的脂解，且载脂蛋白E 2/TG比值的增加对脂解的抑制作用大于E3和E4。总之，无论是内源性表达还是外源性表达，载脂蛋白E异构体对巨噬细胞胆固醇逆向转运的影响均不同。apoE对VLDL-脂解均有抑制作用，其中apoE 2的抑制作用较其他两种亚型更为显著。少

Apolipoprotein E (apoE) plays a key role in the lipoprotein metabolism and atherosclerotic diseases. There exist three major common isoforms in human apoE, i.e., E2, E3 and E4. These isoforms have been known to have differential effect on lipoprotein metabolism and atherosclerosis. In the present study, in order to investigate the roles of these isoforms on 1) the reverse cholesterol transport from the macrophages and 2) VLDL-triglycerides lipolysis, we utilized adenoviral vector for the expression of these isoforms.The RAW264.7 mouse macrophage cell line, which does not express apoE endogenously, was cholesterol-loaded. After loading cholesterol, the cells were infected with adenoviral vectors to express apoE isoforms. The expression of apoE2 reduced cellular esterified-cholesterol levels significantly compared with control cells infected with LacZ adenovirus. ApoE3 and E4 also reduced the cellular cholesterol content, however, their effect was not so much effective as apoE2. In the n … More ext experiment, cholesterol-loaded RAW264.7 cells were incubated with the medium harvested from the HeLa cells previously infected with apoE adenovirus, in order to elucidate the role of exogenous apoE on reverse cholesterol transport. This experiment revealed that apoE3 is effective in the reverse cholesterol transport, however, apoE2 and E4 have little effect. Finally, to elucidate the role of apoE on VLDL-triglycerides lipolysis, adenoviral vectors were injected to apoE/LDL-receptor double deficient mice and apoE-containing VLDL was obtained. These VLDL particles were subjected to in vitro lipolysis assay with bovine lipoprotein lipase, with changing the ratio of apoE/TG. This experiment revealed that the existence of apoE on VLDL particles inhibits lipolysis regardless of its isoform, and increasing ratio of apoE2/TG had more inhibitory effect on the lipolysis compared with E3 and E4.In summary, apoE isoforms have differential effect on reverse cholesterol transport from macrophages not only when they were expressed endogenously but also when they were added exogenously. The VLDL-lipolysis is inhibited by apoE regardless of its isoform, and apoE2 has more substantial inhibitory effect compared with the other two isoforms. Less