姜黄素通过调控超级增强子相关基因抑制骨肉瘤的增殖和迁移  

Curcumin inhibits the proliferation and migration of osteosarcoma by regulating the expression of super-enhancer-associated genes

作　　者：欧阳湛波 朱海宏 刘忠越 涂超[4] 屈健[1,2] 鲁琼[1,2] 徐敏[5] OUYANG Zhanbo;ZHU Haihong;LIU Zhongyue;TU Chao;QU Jian;LU Qiong;XU Min(Department of Pharmacy,Second Xiangya Hospital,Central South University,Changsha 410011;Institute of Clinical Pharmacy,Second Xiangya Hospital,Central South University,Changsha 410011;Department of Pharmacy,Yueyang Central Hospital,Yueyang Hunan 414000;Department of Orthopaedics,Second Xiangya Hospital,Central South University,Changsha 410011;Department of Critical Care Medicine,Second Xiangya Hospital,Central South University,Changsha 410011,China)

机构地区：[1]中南大学湘雅二医院药学部,长沙410011 [2]中南大学湘雅二医院临床药学研究所,长沙410011 [3]岳阳市中心医院药学部,湖南岳阳414000 [4]中南大学湘雅二医院骨科,长沙410011 [5]中南大学湘雅二医院重症医学科,长沙410011

出　　处：《中南大学学报（医学版）》2024年第4期541-552,共12页Journal of Central South University ：Medical Science

基　　金：湖南省财政厅省直单位补助基金(湘财建一指[2018] 92号)。

摘　　要：目的:超级增强子相关基因可能跟骨肉瘤进程息息相关,姜黄素对骨肉瘤等肿瘤具有一定的抑制作用。本研究旨在探讨姜黄素在体内外对骨肉瘤的影响,并研究姜黄素是否可以通过抑制超级增强子相关基因LIM衰老细胞抗原样结构域1(LIM and senescent cell antigen-like-containing domain 1,LIMS1)、富含半胱氨酸的酸性分泌蛋白(secreted protein acidic and rich in cysteine,SPARC)、含无菌α基序结构域蛋白4A(sterile alpha motif domain containing4A,SAMD4A)的表达来抑制骨肉瘤的进展。方法:用5~50μmol/L姜黄素分别处理人骨肉瘤细胞系(MG63细胞或U2OS细胞)24、48和72 h后,采用四甲基偶氮唑盐(methyl thiazolyl tetrazolium,MTT)法检测细胞活力。分别用二甲基亚砜(dimethyl sulfoxide,DMSO)或姜黄素(2.5、5.0μmol/L)孵育细胞7 d,采用克隆形成实验测定体外细胞增殖能力。用含DMSO或姜黄素(10、15μmol/L)处理细胞后,采用划痕愈合实验、transwell迁移实验评价细胞的迁移能力,实时反转录聚合酶链式反应(real-time reverse transcription PCR,real-time RT-PCR)和蛋白质印迹法检测细胞中LIMS1、SPARC、SAMD4A的mRNA和蛋白质的表达水平。建立骨肉瘤荷瘤裸鼠模型,用姜黄素灌胃14 d后评估姜黄素在体内对骨肉瘤体积和重量的影响。采用real-time RT-PCR检测12对骨肉瘤患者的癌组织和癌旁组织中LIMS1、SPARC、SAMD4A的mRNA表达水平。结果:不同浓度的姜黄素分别处理细胞24、48和72 h后,细胞活力均显著下降。与DMSO组相比,2.5μmol/L姜黄素组和5.0μmol/L姜黄素组细胞克隆形成率均明显下降(均P<0.01)。划痕愈合实验结果表明:与DMSO组比较,10μmol/L姜黄素组和15μmol/L姜黄素组细胞迁移率均明显下降,除10μmol/L姜黄素组24 h的U2OS细胞迁移率差异无统计学意义外(P>0.05),其余差异均有统计学意义(P<0.01或P<0.001)。Transwell迁移实验结果表明:与DMSO组比较,10μmol/L姜黄素组和15μmol/L姜黄�Objective:Super-enhancer-associated genes may be closely related to the progression of osteosarcoma,curcumin exhibits a certain inhibitory effect on tumors such as osteosarcoma.This study aims to investigate the effects of curcumin on osteosarcoma in vitro and in vivo,and to determine whether curcumin can inhibit the progression of osteosarcoma by suppressing the expression of super-enhancer-associated genes LIM and senescent cell antigen-like-containing domain 1(LIMS1),secreted protein acidic and rich in cysteine(SPARC),and sterile alpha motif domain containing 4A(SAMD4A).Methods:Human osteosarcoma cell lines(MG63 cells or U2OS cells)were treated with 5 to 50μmol/L curcumin for 24,48,and 72 hours,followed by the methyl thiazolyl tetrazolium(MTT)assay to detect cell viability.Cells were incubated with dimethyl sulfoxide(DMSO)or curcumin(2.5,5.0μmol/L)for 7 days,and a colony formation assay was used to measure in vitro cell proliferation.After treatment with DMSO or curcumin(10,15μmol/L),a scratch healing assay and a transwell migration assay were performed to evaluate cell migration ability.Real-time reverse transcription polymerase chain reaction(real-time RT-PCR)and Western blotting were used to detect mRNA and protein expression levels of LIMS1,SPARC,and SAMD4A in the cells.An osteosarcoma-bearing nude mouse model was established,and curcumin was administered via gavage for 14 days to assess the impact of curcumin on tumor volume and weight in vivo.Real-time RT-PCR was used to measure mRNA expression levels of LIMS1,SPARC,and SAMD4A in the cancer and adjacent tissues from 12 osteosarcoma patients.Results:After treating cells with different concentrations of curcumin for 24,48,and 72 hours,cell viability were all significantly decreased.Compared with the DMSO group,the colony formation rates in the 2.5μmol/L and 5.0μmol/L curcumin groups significantly declined(both P<0.01).The scratch healing assay showed that,compared with the DMSO group,the migration rates of cells in the 10μmol/L and 15μmol/L curcumin

关 键 词：骨肉瘤 姜黄素 超级增强子相关基因 增殖 迁移 

分 类 号：R285[医药卫生—中药学]