摘要
目的探讨维生素K2对Apo E-/-小鼠动脉粥样硬化内膜钙化和Toll样受体2(TLR2)及TLR4表达的影响。方法 18只6周龄雄性Apo E-/-小鼠,随机分为模型组、维生素K2干预组及对照组,每组6只。模型组及维生素K2干预组小鼠予高脂饮食,对照组小鼠予普通饮食喂养。维生素K2干预组在高脂饮食基础上同时予维生素K2灌胃,每天1次,共12周。小鼠19周龄时处死,测定血清脂质水平;HE染色观察主动脉组织形态学变化,Von Kossa染色观察主动脉钙化;测定血管钙含量和碱性磷酸酶活性判断血管钙化程度;免疫组织化学法、实时定量PCR(qRT-PCR)分别检测主动脉TLR2、TLR4蛋白和mRNA的表达。结果模型组小鼠主动脉HE染色可见内膜显著增厚,有典型的动脉粥样硬化斑块形成,Von Kossa染色斑块纤维帽下可见灶状黑色钙化团块,维生素K2干预组小鼠主动脉可见粥样硬化斑块,但Von Kossa染色斑块内未见明显黑色钙盐沉积;定量分析显示,维生素K2干预组小鼠主动脉血管壁钙含量和碱性磷酸酶活性均明显低于模型组(P<0.01);免疫组织化学染色显示TLR2、TLR4主要表达于粥样硬化斑块内,qRT-PCR证实模型组小鼠主动脉TLR2、TLR4表达均显著高于对照组(P<0.05),维生素K2干预组小鼠主动脉TLR2、TLR4 mRNA及蛋白水平较模型组明显降低,差异有统计学意义(P<0.05)。小鼠主动脉钙含量与TLR2 mRNA表达呈正相关(r=0.77,P<0.001),与TLR4 mRNA表达亦呈正相关(r=0.79,P<0.001)。结论维生素K2可降低高脂饮食Apo E-/-小鼠主动脉中钙含量和碱性磷酸酶活性,减少主动脉血管壁TLR2、TLR4的表达,维生素K2抑制内膜钙化的作用可能与下调TLR2、TLR4的表达有关。
Aim To assess the effects of vitamin K on calcification and the expression of Toll-like receptor 2( TLR2) as well as Toll-like receptor 4( TLR4) in aortic artery of Apo E- /-mice. Methods Eighteen 6-week-old male Apo E- /-mice were randomly divided into model group,interventional group and control group. Model group and interventional group were fed with high-fat diet,while control group were fed with normal diet. Mice in interventional group were administered orally with vitamin K2,once a day for 12 weeks. All mice were sacrificed when the mice were 19 weeks old,blood was collected and plasma triglyceride( TG),total cholesterol( TC),high density lipoprotein cholesterol( HDLC),low density lipoprotein cholesterol( LDLC) were measured. Aortic sections were stained with hematoxylin and eosin( HE) or Von Kossa,and observed under microscope. Calcium content and alkaline phosphatase activity of aorta were measured to determine the degree of vascular calcification. The expressions of TLR2 and TLR4 protein and mRNA levels in aotic sections of mice were detected by immunohistochemisty and qRT-PCR respectively. Results HE staining demonstrated significant thickening of the intima,with typical atherosclerotic plaque formation in model group mice,and von Kossa staining showed spotty black clumps of aortic calcification under the fibrous cap plaque,while intervention group had atherosclerotic plaques without the formation of significant calcium deposits; The quantitative analysis showed that aortic vascular wall calcium and alkaline phosphatase activity were significantly lower in the intervention group than that of the model group( P < 0. 01); Immunohistochemical staining showed that TLR2,TLR4 mainly expressed in atherosclerotic plaques,qRT-PCR confirmed that the expression of TLR2 and TLR4 was significantly higher in the model group mice aorta( P < 0. 05),while the expression level of TLR2 and TLR4 mRNA or protein were significantly lower in intervention group aortic than in the model group. The expression level of TLR2 and
出处
《中国动脉硬化杂志》
CAS
北大核心
2015年第3期217-223,共7页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金(81370408)
江苏省自然科学基金(BK20131246)
江苏省卫生厅项目(Q201308)
镇江市社会发展项目(SH2013024)
