ApoE基因敲除小鼠动脉粥样硬化模型构建

Construction of atherosclerosis model in ApoE knockout mice

目的:采用载脂蛋白E基因敲除((ApoE-/-)雄鼠,给予高脂饲料喂养,构建动脉粥样硬化模型,观察血脂变化及主动脉全长动脉粥样硬化斑块形成。方法:选取饲养于SPF屏障环境内8周龄的载脂蛋白E基因敲除((ApoE-/-)雄鼠18只,随机分成对照组(9只)和实验组(9只)。实验前对两组雄鼠进行鼠尾基因型鉴定,并随机抽取6只,测得8周龄对照组小鼠血脂。实验组给予高脂饲料(脂肪15%、胆固醇1.25%、0.5%胆酸盐);对照组予普通饲料,饮水不限。分别在喂养0周、8周时眼眶采血,测血脂。两组小鼠均在饲养8周处死,处死前12 h禁食不禁水,取主动脉全长,油红O染色,观察主动脉病理变化。成功建立AS模型后,将小鼠分为AS+生理盐水(NS)组及AS+辛伐他汀(ST)组(各9只),分别在给药4周及8周时眼眶取血,测血清中血脂浓度变化。结果:对小鼠鼠尾基因型鉴定确定(ApoE基因敲除雄鼠均为阳性纯合子小鼠,且Neo cassette已被去除。通过对(ApoE基因敲除雄鼠实验组及对照组血脂比较,表现为TG、TC和LDL-C显著升高(P<0.05),HDI-C(P<0.05)显著降低;实验组高脂饲料喂养8周后,(ApoE基因敲除雄鼠主动脉形成明显的AS斑块,对照组则未见明显AS斑块。AS小鼠给予NS和ST后,检测表明ST给药时间增长,TC、TG、LDL-C均有降低趋势。其中,TC及LDL-C给药ST 8周后有显著性差异(P<0.05),小鼠主动脉油红O染色显示斑块脂质沉积较NS组有所减少,面积减小,凸向管腔程度也明显降低(即狭窄程度降低)。结论:(ApoE基因敲除雄鼠高脂饲料喂养8周后,血脂明显异常,且形成明显的AS斑块,AS模型成功构建;AS小鼠给予ST(8周)则血脂浓度明显降低,且主动脉油红染色显示斑块明显少于对照组,表明该模型可用于抗动脉粥样硬化药物药效检测。

Objective： The （ApoE gene knockout（（ApoE-/-） male mice were given high fat diet,construct the model of atherosclerosis,observe the changes of blood lipid and aortic atherosclerotic plaque formation. Methods： 18 mice were randomly divided into control group（n = 9） and experimental group（n = 9）,which were fed into the SPF barrier for at least 8 weeks. Before the experiment,two groups of male mice were used to identify the genotypes of the tail,and 6 mice were randomly selected,and the blood lipid of normal control group（n = 8 weeks） were measured. The experimental group was given high fat diet（15% fat,1. 25% cholesterol,0. 5% bile salts）; the control group was given ordinary feed,drinking water is not limited. At 0 weeks and 8 weeks,blood samples were collected and blood lipids were measured. Two groups of mice were killed at the end of feeding for 8 weeks,before the death of fasting for 12 hours. The aortic was observed by stained with oil red O. Results： （ApoE gene knockout male mice were all positive homozygous mice,and Neo cassette had been removed. Based on the experimental group and control group of mice blood lipid,the expressions of TG,TC and LDL-C were significantly increased（P〈0. 05）,HDI-C（P〈0. 05） decreased significantly; the experimental group with high fat diet for 8 weeks,（ApoE knockout male mice aorta significantly the formation of AS plaque,while the control group had no obvious AS plaque. The AS model was successfully established,the mice were divided into AS ＋ NS group and AS ＋ ST group,respectively take the orbital blood at 4 and 8 weeks to detect the changes of serum lipid levels,with the time growth,TC,TG,LDL-C level of the AS ＋ NS group were significantly increased and were higher than that of AS ＋ ST group. Among them,TC and LDL-C in group AS ＋ ST（8 weeks） group was significantly lower than AS ＋ NS group,the results were significant differences（P〈0. 05）,and simvastatin Group 8 week mice aorta oil red O staining showed that the plaque lipid deposition little and smaller than in group NS,and the area decreased,the plaque convex to the lumen also decreased（to reduce the degree of stenosis）. Conclusion： （ApoE knockout male mice fed with high fat diet showed abnormal blood lipid,and formed AS plaques,while the control group did not show significant AS plaques. The 8 week old （ApoE knockout male mice on the high-fat diet for 8 weeks,can be successfully constructed AS model; （ApoE gene knockout male mice were given ST（8 weeks）,the serum lipid concentrations were significantly decreased,and aortic plaque oil red staining showed significantly less than AS ＋ NS group,and there is no significant increase compared with the onset stage,which indicated that ST had good effect on lipid regulation and anti AS in treatment.