摘要
目的探讨冠心宁(GXN)调控动脉粥样硬化(AS)斑块稳定性的分子机制。方法制备含有GXN药物的血清,利用氧化低密度脂蛋白(ox-LDL)诱导血管平滑肌细胞(VSMC)构建AS体外模型,Western blot和qPCR检测VSMC表型转换情况。通过沉默肿瘤坏死因子α诱导蛋白3(TNFAIP3),检测GXN对VSMC表型转换的影响。构建凋亡信号调节激酶1(ASK1)过表达载体,并利用Lip3000转染进细胞,检测GXN是否通过凋亡信号调节激酶1/c-Jun氨基末端激酶(ASK1/JNK)通路发挥作用。结果与对照组比较,ox-LDL组的细胞表型转换,表现为I型胶原蛋白(COLIA1和COLIA2)、TNFAIP3和α-平滑肌肌动蛋白(α-SMA)的表达水平降低(均P<0.01),基质金属蛋白酶(MMP)2、MMP9、MMP13、骨桥蛋白(OPN)、ASK1磷酸化与ASK1比值(p-ASK1/ASK1)、JNK磷酸化与JNK比值(p-JNK/JNK)升高(均P<0.01);GXN处理后VSMC表型转换为收缩型,表现为与ox-LDL组相比COLIA1、COLIA2和α-SMA水平增高(均P<0.01),MMP2、MMP9、MMP13、OPN、TNFAIP3的表达水平、p-ASK1/ASK1以及p-JNK/JNK都降低(均P<0.01)。沉默TNFAIP3后,与ox-LDL+10%GXN+sh-NC组相比,COLIA1、COLIA2、TNFAIP3和α-SMA水平降低(均P<0.01),MMP2、MMP9、MMP13、OPN、p-ASK1/ASK1以及p-JNK/JNK都升高(均P<0.01)。过表达ASK1后,与ox-LDL+10%GXN+oe-NC组相比,COLIA1、COLIA2和α-SMA的表达水平降低(均P<0.01),MMP2、MMP9、MMP13、OPN、p-ASK1/ASK1以及p-JNK/JNK都升高(均P<0.01)。结论GXN可能通过TNFAIP3调控ASK1/JNK通路介导VSMC表型转换,从而起到稳定动脉硬化斑块的作用。
Objective To explore the molecular mechanism of Guanxinning(GXN)regulating the stability of atherosclerotic(AS)plaques.Methods Serum containing GXN was prepared.Vascular smooth muscle cells(VSMCs)induced with oxidized low-density lipoprotein(ox-LDL)were used to construct the in vitro AS model.The phenotypic transformation of VSMCs was detected by Western blot and qPCR.The effect of GXN on VSMCs phenotypic transformation was examined by silencing tumor necrosis factorα-induced protein 3(TNFAIP3).In addition,an apoptosis signalregulating kinase 1(ASK1)overexpression vector was constructed and transfected into cells with Lip3000 to detect whether GXN exerts its effects through the ASK1/c-Jun N-terminal kinase(ASK1/JNK)pathway.Results Compared with the control group,the ox-LDL group showed a phenotypic transformation of cells characterized by decreased expression levels of type I collagen(COLIA1 and COLIA2)andα-smooth muscle actin(α-SMA)(all P<0.01),and increased expression levels of matrix metalloproteinases(MMP)2,MMP9,MMP13,osteopontin(OPN),TNFAIP3,phosphorylated ASK1/ASK1 ratio(p-ASK1/ASK1),and phosphorylated JNK/JNK ratio(p-JNK/JNK)(all P<0.01).After GXN treatment,VSMCs transformed to a contractile phenotype,indicated by increased levels of COLIA1,COLIA2,TNFAIP3 andα-SMA compared to the ox-LDL group(all P<0.01),and decreased levels of MMP2,MMP9,MMP13,OPN,p-ASK1/ASK1,and p-JNK/JNK(all P<0.01).After TNFAIP3 silencing,compared to the ox-LDL+10%GXN+sh-NC group,the levels of COLIA1,COLIA2,TNFAIP3 andα-SMA decreased(all P<0.01),while the expression levels of MMP2,MMP9,MMP13,OPN,p-ASK1/ASK1,and p-JNK/JNK increased(all P<0.01).Overexpression of ASK1 resulted in decreased levels of COLIA1,COLIA2 andα-SMA(all P<0.01),and increased expression levels of MMP2,MMP9,MMP13,OPN,p-ASK1/ASK1,and p-JNK/JNK compared to the ox-LDL+10%GXN+oe-NC group(all P<0.01).Conclusion GXN may stabilize atherosclerotic plaques by regulating the phenotypic transformation of VSMCs through the TNFAIP3-ASK1/JNK pathway.
作者
毛萍
吕方超
徐晨凯
唐礼江
Mao Ping;Lyu Fangchao;Xu Chenkai;Tang Lijiang(Department of Caridiology,Zhejiang Hospital,Hangzhou 310013,China)
出处
《心脑血管病防治》
2024年第10期5-9,18,共6页
CARDIO-CEREBROVASCULAR DISEASE PREVENTION AND TREATMENT
基金
浙江省医学会临床医学科研专项基金(2022ZYC-Z14)。
