Ox-HDL通过调控caspase-1对po4小鼠动脉粥样硬化斑块炎症介质及细胞焦亡的影响

Influence of ox-HDL on inflammatory mediators and pyroptosis of atherosclerotic plaque through regulating caspase-1 in po4 mice

目的探讨氧化修饰高密度脂蛋白(ox-HDL)调控caspase-1对po4小鼠动脉粥样硬化斑块炎症介质及细胞焦亡的影响。方法高脂饲料(胆固醇1.25%、脂肪基础饲料83.25%、0.125%氯化胆盐)饲养至12周,建立po4动脉粥样硬化小鼠模型,检测血清高密度脂蛋白(HDL)、低密度脂蛋白(LDL)、总胆固醇(TC)水平确定小鼠血脂水平,大体油红O染色确定纤维粥样斑块的形成,油红染色见斑块中央大片的红色脂滴池存在、且有纤维组织覆盖,说明动脉硬化小鼠模型构建成功。12周龄po4动脉粥样硬化小鼠随机编号分为对照组(n=8)和实验组(n=8),对照组小鼠经尾静脉注射生理盐水,剂量为14 ml/kg体重;实验组小鼠经尾静脉注射等体积0.9%氯化钠配置的ox-HDL,浓度100μg/ml。两组均2次/周注射,共注射6周。6周后,获取小鼠主动脉并依次采用大体油红O染色检测纤维粥样斑块的形成、TUNEL法检测细胞焦亡情况、免疫组化检测主动脉根部核苷酸结合寡聚化结构域样受体家族热蛋白结构域3炎性小体(NLRP3)、caspase-1表达水平、免疫沉淀技术检测ox-HDL与caspase-1的共表达作用。结果实验组主动脉斑块面积、TUNEL阳性细胞及NLRP3、caspase-1表达水平均大于对照组(P<0.05)。Ox-HDL、HDL与caspase-1共转染小鼠细胞,结果ox-HDL组荧光活性低于HDL组(P<0.05),HDL组和caspase-1组荧光活性差异无统计学意义(P>0.05)。结论Ox-HDL可通过介导caspase-1促进斑块进展及诱导NLRP3炎性小体活化,加剧斑块炎性反应和细胞焦亡。

Objective To discuss the influence of oxidized high-density lipoprotein-cholesterol(ox-HDL)on inflammatory mediators and pyroptosis of atherosclerotic plaque through regulating caspase-1 in po4 mice.Methods The model of atherosclerotic po4 mouse was established through feeding with high-fat diet containing 1.25%cholesterol,83.25%basic feed and 0.125%bile salt chloride for 12 weeks.The blood lipid level was determined through detecting serum high-density lipoprotein(HDL),low-density lipoprotein(LDL)and total cholesterol(TC).The formation of atherosclerotic plaques was determined by gross oil red O staining,which indicated that the model was successfully established.All po4 mice of atherosclerosis aged 12 weeks old were randomly divided into control group and test group(each n=8).The control group received normal saline injection(14 ml/kg)through tail vein,and test group received the injection in equal volume of ox-HDL(100μg/ml)containing sodium chloride(0.9%)through tail vein twice a week for 6 weeks.The formation of atherosclerotic plaques was detected by using gross oil red O staining,pyroptosis status was detected by using TUNEL assay,expression levels of inflammasome of nucleotide-binding oligomerization domain-like receptor 3(NLRP3)and caspase-1 were detected by using immunohistochemistry,and co-expression of ox-HDL and caspase-1 was detected by using immunoprecipitation technique.Results The aortic plaque area,TUNEL positive cells and expression levels of NLRP3 and caspase-1 were all higher in test group than those in control group(P<0.05).The results of co-transfection of ox-HDL,HDL and caspase-1 showed that fluorescence activity was lower in ox-HDL group than that in HDL group(P<0.05),and had no statistical significance between HDL group and caspase-1 group(P>0.05).Conclusion Ox-HDL can promote plaque progression and induce the activation of NLRP3 inflammasomes through mediating caspase-1,thus aggravae plaque inflammation and pyroptosis.