摘要
目的探讨长链非编码RNA(lncRNA)结直肠肿瘤差别表达基因(CRNDE)对氧化型低密度脂蛋白(oxLDL)诱导的巨噬细胞凋亡和炎性因子分泌的影响及其机制。方法以0、25、50、100μmol/ml oxLDL处理小鼠巨噬细胞RAW264.724 h后,采用实时荧光定量PCR检测CRNDE表达。将RAW264.7巨噬细胞分为对照组(正常培养,n=3)、oxLDL组(以100μmol/ml oxLDL处理,n=3)、oxLDL+siNC组(转染阴性对照RNA后给予100μmol/ml oxLDL处理,n=3)和oxLDL+siCRNDE组(转染靶向CRNDE的小干扰RNA后给予100μmol/ml oxLDL处理,n=3),采用流式细胞仪检测细胞凋亡率,免疫印迹法检测B淋巴细胞瘤2基因(Bcl-2)、Bax、磷酸化NF-κB(p-NF-κB)P65、NF-κB抑制蛋白(IκBa)和磷酸化IκBa(p-IκBa)蛋白表达。结果0、25、50、100μg/ml oxLDL刺激后巨噬细胞中lncRNA CRNDE表达水平比较,有统计学差异(1.00±0.07 vs 1.52±0.23 vs 2.28±0.59 vs 3.56±0.78,P<0.01)。与对照组比较,oxLDL组、oxLDL+siNC组和oxLDL+siCRNDE组巨噬细胞中CRNDE表达、细胞凋亡率、白细胞介素6(IL-6)、TNF-α、Bax、p-NF-κB P65、p-IκBa蛋白表达明显升高,Bcl-2蛋白表达明显降低(P<0.05);oxLDL+siCRNDE组巨噬细胞中lncRNA CRNDE表达、细胞凋亡率、IL-6、TNF-α、Bax、p-NF-κB P65及p-IκBa蛋白表达明显低于oxLDL组,Bcl-2蛋白表达明显高于oxLDL组(P<0.05)。结论下调CRNDE表达可抑制oxLDL诱导的巨噬细胞凋亡和炎性因子分泌,其作用机制可能与抑制NF-κB信号通路的活化有关。
Objective To study the effect of lncRNA CRNDE gene on oxLDL-induced apoptosis of macrophages and secretion of inflammatory factors and its mechanism.Methods The expression of CRNDE gene was detected by RT-fluorescent qPCR after the macrophages were treated in 0μmol/ml oxLDL,25μmol/ml oxLDL,50μmol/ml oxLDL and 100μmol/ml oxLDL respectively for 24 h.The in vitro cultured macrophages were divided into control group(treated in normal culture,n=3),oxLDL group(treated in 100μmol/ml oxLDL,n=3),oxLDL+siNC group(transfected with NC-siRNA and treated in 100μmol/ml oxLDL,n=3)and oxLDL+siCRNDE gene group(treated in 100μmol/ml oxLDL after transfection with CRNDE-siRNA,n=3).The expression of CRNDE gene was detected by RT-fluorescent qPCR and flow cytometry respectively.The apoptosis of macrophages was assayed by flow cytometry.The expressions of Bcl-2,Bax,NF-κB P65,IκBa,p-NF-κB P65 and p-IκBa were detected by Western blot.Results The expression level of lncRNA CRNDE gene was significantly lower in macrophages treated in 0μg/ml oxLDL than in those treated in 25μmol/ml oxLDL,50μmol/ml oxLDL and 100μmol/ml oxLDL(1.00±0.07 vs 1.52±0.23 vs 2.28±0.59 vs 3.56±0.78,P<0.01).The apoptosis rate of macrophages and the expression levels of CRNDE gene,IL-6,TNF-α,Bax,p-NF-κB P65,p-IκBa were significantly higher while the expression level of Bcl-2 was significantly lower in oxLDL group,oxLDL+siNC group and oxLDL+siCRNDE group than in control group(P<0.05).The apoptosis rate of macrophages and the expression levels of CRNDE gene,IL-6,TNF-α,Bax,p-NF-κB P65,p-IκBa were significantly lower while the expression level of Bcl-2 was significantly higher in oxLDL+siCRNDE group than in oxLDL group(P<0.05).Conclusion Down regulating the expression of CRNDE gene suppresses oxLDL-induced apoptosis of macrophages and secretion of inflammatory factors by inhibiting the NF-κB signaling pathway activation.
作者
胡姗姗
杨贵丽
胡菁
田凡青
彭雯
Hu Shanshan;Yang Guili;Hu Jing;Tian Fanqing;Peng Wen(Department of Cardiology and neurology,Wuhan University of science and Technology Hospital,Wuhan 430065,China)
出处
《中华老年心脑血管病杂志》
CAS
北大核心
2020年第11期1197-1201,共5页
Chinese Journal of Geriatric Heart,Brain and Vessel Diseases
