瓜蒌-薤白对ApoE－/－小鼠动脉粥样硬化过程中NLRP3炎症小体活化的影响

目的通过建立ApoE^-/-小鼠动脉粥样硬化(As)模型,探讨As病程不同时间点NOD样受体热蛋白结构域3(NLRP3)炎症小体表达水平的变化及瓜蒌-薤白的干预作用。方法将高脂饲养6、20、34周的ApoE^-/-小鼠均随机分为模型组(M1、M2、M3)和给药组6 g/(kg·d)](GX1、GX2、GX3),每组10只;另设C57BL/6J小鼠为空白组(C1、C2、C3)。空白组及模型组小鼠给予生理盐水灌胃,给药组小鼠每日给予相应药物灌胃,共4周。实验结束后处死小鼠,油红O染色评估主动脉斑块面积及形态;HE染色观察主动脉病理形态学变化;免疫组织化学法检测主动脉NLRP3表达;ELISA法检测血清中白细胞介素1β(IL-1β)和白细胞介素18(IL-18)水平;Western blot法检测主动脉组织中NLRP3、凋亡相关斑点样蛋白(ASC)、含半胱氨酸的天冬氨酸蛋白水解酶1(Caspase-1)的蛋白表达;qRT-PCR检测主动脉组织中NLRP3、ASC、Caspase-1的mRNA表达。结果在As病程进展过程中,模型组小鼠主动脉脂质累积和斑块面积显著增加,血清中IL-1β和IL-18的表达不断升高,NLRP3和ASC的蛋白及mRNA的表达均不断上调。Caspase-1的蛋白表达也呈上升趋势,但M2与M3组间的比较无统计学差异。与模型组相比,给药组各时间点小鼠主动脉的脂质累积和斑块面积显著减少,血清中IL-1β和IL-18的水平降低;主动脉组织中N LRP3、ASC、Caspase-1蛋白和mRNA表达显著下调。结论NLRP3炎症小体参与了主动脉As的病变过程,瓜蒌-薤白可能通过调节As模型小鼠主动脉不同阶段NLRP3炎症小体的表达,从而发挥抗As的作用。

Effects of Trichosanthis Fructus-Allii Macrostemonis Bulbus on the activation of NLRP3 inflammasomes in ApoE-/-mice at different stages of atherosclerosis

Aim To observe the changes in the expression level of NOD-like receptor pyrin domain containing 3(NLRP3)inflammasomes at different stages of the course of atherosclerosis(As)and the intervention effect of Trichosanthis Fructus-Allii Macrostemonis Bulbus through establishment of the model of atherosclerosis in ApoE^-/-mice.Methods High-fat-fed ApoE^-/-mice for 6,20,34 weeks were randomly divided into model groups(M1,M2,M3)and medication groups(6g/(kg·d))(GX1,GX2,GX3),10 mice in each group.C57BL/6 mice were set as blank control group(C1,C2,C3).The mice in the blank control group and the model group were given normal saline by gavage,and the mice in the medication group were given the corresponding drugs by gavage daily for 4 weeks.After the experiment,the mice were sacrificed and the aortic plaque area and morphology were evaluated by oil red O staining.HE staining was used to observe the pathomorphological changes of the aorta.Immunohistochemical method was used to detect NLRP3 expression in the aorta.The levels of interleukin-1β(IL-1β)and interleukin-18(IL-18)in serum were detected by ELISA.Western blot was used to detect the protein expression of NLRP3,apoptosis-associated speck-like protein containing(ASC),cysteinyl aspartate specific proteinase-1(Caspase-1)in aortic tissue.qRT-PCR was used to detect the mRNA expression of NLRP3,ASC,Caspase-1 in aortic tissue.Results At different stages of the course of As,lipid accumulation and plaque area in aorta of mice were significantly increased in model group,the levels of IL-1βand IL-18 in serum was continuously increased,and the protein and mRNA expressions of NLRP3 and ASC were continuously up-regulated,the protein expression of Caspase-1 also showed an upward trend,but there was no statistical significance between M2 and M3.Compared with the model group,the lipid accumulation and plaque area in aorta of mice at different stages were significantly decreased in medication group,and the levels of IL-1βand IL-18 in serum were decreased;The protein and the mRNA expression of NLRP3,ASC and Caspase-1 in aortic tissues were significantly down-regulated.Conclusions NLRP3 inflammasome was involved in the pathological process of aorta As.Trichosanthis Fructus-Allii Macrostemonis Bulbus drug parican regulate the expression of NLRP3 inflammasomes at different stages in the aorta of As model mice,and thus play a role in protecting As.