3-O-C_(12)-HSL通过抑制lncRNA CD48-AS和CD48的表达而阻碍Mo-DCs成熟  

3-O-C_(12)-HSL hampers the maturation of Mo-DCs by inhibiting the expression of lncRNA CD48-AS and CD48

作　　者：罗燕芬[1] 庄奇真 张轩[1] 陈茶[1] 刘杨 颜星星 肖倩[1] 李有强 LUO Yan-fen;ZHUANG Qi-zhen;ZHANG Xuan;CHEN Cha;LIU Yang;YAN Xing-xing;XIAO Qian;LI You-qiang(Department of Laboratory Medicine,Guangdong Provincial Hospital of Traditional Chinese Medicine,Guangzhou 501180,China;the Second Clinical Medical College of Guangzhou University of Traditional Chinese Medicine;Department of Laboratory Medicine,He Xian Memorial Hospital,Southern Medical University)

机构地区：[1]广东省中医院检验医学部,501180 [2]广州中医药大学第二临床医学院 [3]南方医科大学附属何贤纪念医院检验科

出　　处：《天津医药》2021年第12期1233-1239,共7页Tianjin Medical Journal

基　　金：国家自然科学基金资助项目(81601736);广东省中医药局中医药科研项目(20192027)。

摘　　要：目的筛选并阐明长链非编码RNA(lncRNA)CD48-AS与其反义互补分子CD48 m RNA在N-3-氧代十二烷酰-L-同型丝氨酸内酯(3-O-C_(12)-HSL)阻碍人单核细胞诱导的树突状细胞(Mo-DCs)成熟过程中发挥作用的机制。方法从健康人外周血中分离Mo-DCs,将未成熟的Mo-DCs细胞分为阴性对照组(0.1%DMSO)、脂多糖(LPS)阳性对照组(100μg/L的LPS)和实验组(100μg/L LPS+40μmol/L 3-O-C_(12)-HSL)进行lncRNA芯片检测。筛选lncRNA表达谱中差异表达5倍以上的自然反义lncRNA进行聚类分析,从中筛选差异具有统计学意义的自然反义lncRNA CD48-AS。未成熟的Mo-DCs分为阴性对照组、LPS阳性对照组以及LPS+C5、C10、C25实验组(分别加入5、10、25μmol/L的3-O-C_(12)-HSL)。利用荧光定量PCR检测lncRNA CD48-AS和反义分子CD48 mRNA在实验体系中的表达水平;通过生物信息学分析lncRNA CD48-AS与CD48反义互补区及其蛋白编码功能;通过核糖核酸酶保护实验(RPA)验证lncRNA CD48-AS是否与CD48形成二聚体,从而通过影响靶CD48的表达来发挥调控作用。最后检测lncRNA CD48-AS在细胞内的定位。结果 3-O-C_(12)-HSL处理Mo-DCs后lncRNA表达谱出现了特异性改变。3-O-C_(12)-HSL可下调由LPS诱导的Mo-DCs中lncRNA CD48-AS及其反义靶分子CD48的表达。生物信息学分析lncRNA CD48-AS不具备蛋白编码功能,为非编码RNA;lncRNA CD48-AS与CD48形成RNA二聚体从而减少RNA酶对CD48 mRNA的降解,使得CD48 mRNA表达增加;lncRNA CD48-AS在Mo-DCs中主要定位在细胞核中,细胞质中表达量较少。结论 3-O-C_(12)-HSL可通过下调lncRNA CD48-AS,进而影响CD48的表达来阻碍Mo-DCs的成熟。Objective To screen and clarify the mechanism of long non-coding RNA CD48-AS(lncRNA CD48-AS)and the antisense complementary molecule CD48 in the process of N-3-oxododecanoyl-L-homoserine lactone(3-O-C_(12)-HSL) hampering the maturation of human monocyte-induced dendritic cells(Mo-DCs).Methods Mo-DCs were extracted from the peripheral blood of healthy individuals,and the immature Mo-DCs were divided into three groups for lncRNA chip analysis:the negative control group(0.1% DMSO),the lipopolysaccharide(LPS) positive control group(100 μg/L LPS) and the treatment group(100 μg/L LPS+40 μmol/L 3-O-C_(12)-HSL).The natural antisense lncRNA expressed more than 5 times in the lncRNA expression profile were screened for Cluster analysis.The natural antisense lncRNA CD48-AS was screened.Immature Mo-DCs were divided into the negative control group,the LPS positive control group,and the 3-O-C_(12)-HSL treatment group(5,10,25 μmol/L 3-O-C_(12)-HSL).The expression of lncRNA CD48-AS and antisense molecule CD48 were measured using quantitative PCR in each group.The complementary regions of lncRNA CD48-AS and CD48 and the protein coding function of lncRNA CD48-AS were analyzed through bioinformatics.Whether lncRNA CD48-AS affected the expression of CD48 by forming a dimer with CD48 was verified through the ribonuclease protection experiment(RPA).Preliminary experiments were carried out on the localization of lncRNA CD48-AS in cells.Results The lncRNA expression profile of Mo-DCs showed specific changes after 3-O-C_(12)-HSL treatment.3-O-C_(12)-HSL down-regulated the expression of lncRNA CD48-AS and its antisense target molecule CD48 induced by LPS.Bioinformatics analysis showed that lncRNA CD48-AS was a non-coding RNA without protein coding function.lncRNA CD48-AS may form RNA dimers with CD48 to reduce the degradation of CD48 by RNase and increase the expression of CD48.lncRNA CD48-AS was mainly located in the nucleus in Mo-DCs,with less expression in cytoplasm.Conclusion 3-O-C_(12)-HSL can inhibit the maturation of Mo-DCs by

关 键 词：RNA 长链非编码 树突细胞 脂多糖类 CD48抗原 计算生物学 N-3-氧代十二烷酰-L-同型丝氨酸内酯 长链非编码RNA CD48-AS 

分 类 号：R392.12[医药卫生—免疫学]