组蛋白去乙酰化酶SIRT1调控氧化低密度脂蛋白诱导巨噬细胞凋亡的表达  

Effects and mechanism of histone deacetylase SIRT1 controlled macrophage apoptosis induced by oxidized low density lipoprotein

作　　者：李卉[1] 姜朝阳 刘岩[1] 张曼[1] LI Hui;JIANG Chaoyang;LIU Yan;ZHANG Man(Department of Cardiovascular Medicine,Central Hospital Affiliated to Shenyang Medical College,Shenyang 110024,Liaoning,China)

机构地区：[1]沈阳医学院附属中心医院心血管内科,辽宁沈阳110024

出　　处：《山东大学学报（医学版）》2022年第1期6-12,共7页Journal of Shandong University:Health Sciences

基　　金：沈阳市科学技术计划项目(20-205-4-031);辽宁省教育厅2020年度科学研究经费项目(SYYX202010);辽宁省普通本科高等学校校际合作项目(协同创新)(重大科研项目第185);沈阳医学院科学研究基金(20191005)

摘　　要：目的观察组蛋白H3赖氨酸残基9乙酰化(H3K9Ac)在氧化低密度脂蛋白(oxLDL)诱导的巨噬细胞凋亡模型中的表达,探讨组蛋白去乙酰化酶———炎症因子沉默信息调节蛋白1(SIRT1)对组蛋白乙酰化的影响,及其通过基因表观遗传学作用,经氧化物酶体增殖物激活受体γ(PPARγ)通路调控巨噬细胞凋亡的机制。方法培养BALB/c小鼠单核巨噬细胞(RAW264.7),并加入oxLDL构建巨噬细胞模型。将细胞分为对照组(加入双蒸水)和实验组(加入50μg/mL oxLDL),分别检测两组细胞凋亡及白细胞介素(IL-6)、SIRT1、H3K9Ac和PPARγ的蛋白表达水平。另外,将实验组细胞分别给予SIRT1兴奋剂(白藜芦醇,终浓度50 nmoL/L)和SIRT1抑制剂(尼克酰胺,终浓度50 nmoL/L),观察SIRT1过表达或抑制对oxLDL诱导巨噬细胞模型中细胞凋亡及SIRT1、H3K9Ac、PPARγ和磷酸化过氧化物酶体增殖物激活受体γ(Ser112位点)[pPPARγ(S112)]的蛋白表达水平的影响。采用Hoechst荧光凋亡染色法检测各组细胞凋亡;采用Western blotting法检测各组细胞IL-6、SIRT1、H3K9Ac、PPARγ和pPPARγ(S112)的蛋白表达。结果①实验组细胞凋亡数(84.88±5.89)高于对照组(7.13±3.31)(P<0.01)。实验组IL-6蛋白相对表达水平(0.50±0.01)高于对照组(0.20±0.02)(P<0.01)。实验组SIRT1蛋白相对表达水平(0.20±0.01)低于对照组(0.30±0.02)(P<0.01)。实验组H3K9Ac蛋白相对表达水平(0.32±0.02)高于对照组(0.22±0.02)(P<0.01)。实验组PPARγ蛋白相对表达水平(0.11±0.02)低于对照组(0.20±0.03)(P<0.01)。②SIRT1兴奋剂组细胞凋亡数(28.63±6.44)低于实验组(84.88±5.89)(P<0.01);SIRT1抑制剂组细胞凋亡数(266.88±35.10)高于实验组(84.88±5.89)(P<0.01)。SIRT1兴奋剂组SIRT1蛋白相对表达水平(0.27±0.03)高于实验组(0.20±0.01)(P<0.01);SIRT1抑制剂组SIRT1蛋白相对表达水平(0.10±0.01)低于实验组(0.20±0.01)(P<0.01)。SIRT1兴奋剂组H3K9Ac蛋白相对表达水平(0.21±0.02)低于实验组(0.Objective To observe the expression of histone H3 lysine residue 9 acetylation(H3K9Ac)in macrophage apoptosis model induced by oxidized low density lipoprotein(oxLDL),and to explore the controlling mechanism of histone deacetylase-inflammatory factor silencing information regulating protein1(SIRT1)on macrophage apoptosis by gene epigenetics,which was realized by peroxisome proliferator activated receptor γ(PPARγ)signaling pathway.Methods Mouse BALB/c macrophage cell line RAW264.7 was cultured with 50μg/mL oxLDL.The cells were divided into control group(treated with double distilled water)and experimental group(treated with 50μg/mL oxLDL).The number of apoptotic cells and protein expressions of interleukin(IL-6),SIRT1,H3K9Ac and PPARγ were detected.In addition,the experimental group was treated with SIRT1 stimulant(resveratrol,final concentration 50 nmoL/L)and SIRT1 inhibitor(nicotinamide,final concentration 50 nmoL/L).The apoptosis and protein expressions of SIRT1,H3K9Ac,PPARγ and phosphorylated peroxisome proliferator-activated receptorγ(Ser112 site)[pPPARγ (S112)]after SIRT1 overexpression or inhibition were detected.The cell apoptosis was detected with Hoechst fluorescence apoptosis staining.The protein expressions of IL-6,SIRT1,H3K9Ac,PPARγ and pPPARγ (S112)were detected with Western blotting.Results The number of apoptotic cells in the experimental group was higher than that in the control group[(84.88±5.89)vs(7.13±3.31),P<0.01].The relative protein expressions of IL-6 and H3K9Ac in the experimental group were higher than those in the control group[(0.50±0.01)vs(0.20±0.02),(0.32±0.02)vs(0.20±0.03),P<0.01],while the relative protein expressions of SIRT1 and PPARγ in the experimental group were lower than those in the control group[(0.20±0.01)vs(0.30±0.02),(0.11±0.02)vs(0.20±0.03),P<0.01].The number of apoptotic cells in the SIRT1 stimulant group was lower than that in the experimental group[(28.63±6.44)vs(84.88±5.89),P<0.01],while the number of apoptotic cells in the SIRT1 inhibitor gro

关 键 词：基因表观遗传学 组蛋白 组蛋白去乙酰化酶 乙酰化修饰 氧化物酶体增殖物激活受体γ 凋亡 

分 类 号：R392.12[医药卫生—免疫学]