摘要
前期设计合成了一种模拟人载脂蛋白E(apoE)结构域的小分子多肽EpK,体外实验证实该拟肽具有抑制巨噬细胞炎症及增强高密度脂蛋白(HDL)介导细胞胆固醇外流的作用.本文拟借助慢病毒体系分泌性表达EpK,研究EpK在体对apoE基因敲除(apoE-/-)小鼠动脉粥样硬化斑块的影响.将11月龄雌性apoE-/-小鼠18只,随机分两组,分别经眼球后静脉丛注射p WPI慢病毒(Lv-GFP对照组)和含EpK的重组慢病毒(Lv-EpK组).小鼠普食喂养,间隔采血监测血脂状态,检测血浆对氧磷酯酶(PON1)活性,病毒注射18周后小鼠安乐死,从主动脉根部连续冰冻切片及主动脉胸腹段纵剖面(en face)进行油红O染色分析斑块面积,采用实时荧光定量PCR检测小鼠肝脏相关基因的m RNA表达水平,蛋白质印迹检测血浆中apo A-Ⅰ、PON1及血清淀粉样蛋白A(SAA)水平.结果显示:慢病毒感染小鼠可成功在血循环中检测到EpK,与Lv-GFP对照组比较,Lv-EpK组apoE-/-小鼠的血脂及脂蛋白分布、apo A-Ⅰ水平、PON1活性无明显改变,但EpK组小鼠的主动脉斑块面积较对照组显著减少(主动脉根部斑块面积(0.87±0.07)mm2 vs(1.03±0.08)mm2,P<0.05;主动脉胸腹段斑块占管腔比42.0%vs 55.8%,P<0.01).EpK可显著降低血中SAA水平,并抑制肝脏炎症因子TNFα和IL-6的表达.结果说明,EpK拟肽具有减退apoE-/-小鼠动脉粥样硬化斑块的作用,其机制可能与其发挥的抗炎作用有关.
We previously reported that a human apolipoprotein E mimetic peptide, designated EpK, enhanced the ability of HDL in mediating cholesterol efflux and suppressing LPS-induced proinflammatory cytokine expression in culture macrophage. The aim of this study was to investigate the impact of this peptide on atherosclerosis in apolipoprotein E deficient （apoE-/-） mice, employed lentivirus expression system to achieve the secretion of EpK in vivo. Eighteen female apoE-/-- mice at 11 months of age were randomly divided into two groups. All mice were injected with pWPI lentivirus （Lv-GFP control group） or pWPI/EpK lentivirus （Lv-EpK） from retro-orbital venous plexus and fed with a chow diet. Blood samples were collected from mice to determine plasma lipids levels and paraoxonase-1 （PON1） activity during eighteen weeks after lentiviral injection. The extent of atherosclerosis was examined using Oil Red O-stained cross-sections of the aorta root and by en face analysis of the aorta. The mRNA levels of inflammatory cytokines in liver were measured by quantitative real-time PCR and the target protein levels in plasma were detected by Western blot. The results showed that EpK peptide was successfully detected in blood by secreting expression in the liver. There were no obvious differences in plasma lipids levels, lipoprotein profile, apolipoprotein A I level and paraoxonase-1 （PON1） activity between two groups. However, the mean area of atherosclerotic lesions in Lv-EpK mice was significant reduced in aortic arch and total aorta, compared with Lv-GFP control groups （the lesion area of aortic root was （0.87±0.07） mm2 vs （1.03±0.08） mm2, P 〈 0.05; the percent area of aortic lesion area was 42% vs 55.8%, P 〈 0.01）. Moreover, expression of EpK resulted in significantly reduced plasma serum amyloid A （SAA） levels and the mRNA levels of tumor necrosis factor α （TNFα and interleukin-6 （IL-6） in mouse liver. These results suggest that the apoE mimetic peptide EpK, is able to reduce atherosclerotic plaque in apoEj mice, which the mechanism may be involved in its anti-inflammatory property.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2015年第9期833-842,共10页
Progress In Biochemistry and Biophysics
基金
国家自然科学基金(81270364)
湖北省卫生厅人才计划基金(JX6B60)资助项目~~
关键词
动脉粥样硬化
APOE
拟肽
炎症
脂代谢
atherosclerosis, apolipoprotein E, mimetic peptide, inflammation, lipid metabolism
