雷公藤甲素对人骨肉瘤细胞阿霉素耐药株的逆转作用及机制研究  

Reversal effect and mechanism of triptolide on adriamycin-resistant human osteosarcoma cell line

作　　者：高峰 王秀会[1] 夏胜利 周小小[1] 易存国 付备刚 沈超 王明辉[1] GAO Feng;WANG Xiuhui;XIA Shengli;ZHOU Xiaoxiao;YI Cunguo;FU Beigang;SHEN Chao;WANG Minghui(Zhoupu Hospital Affiliated to Shanghai Medical College,Shanghai 201318,China)

机构地区：[1]上海健康医学院附属周浦医院上海市浦东新区周浦医院,上海201318

出　　处：《陕西医学杂志》2021年第6期648-651,656,共5页Shaanxi Medical Journal

基　　金：上海市科学技术委员会科研计划项目(16ZR1431600);上海市浦东新区卫生健康委员会临床特色学科建设计划项目(PWYTS2018-02);上海健康医学院师资人才百人库项目(IPBRK-18-07)。

摘　　要：目的:探讨雷公藤甲素(TPL)体外给药对人骨肉瘤细胞阿霉素(ADR)耐药株(U2OS/ADR)多药耐药(MDR)的逆转作用及可能机制。方法:使用人骨肉瘤细胞株U2OS,利用浓度递增法建立耐药骨肉瘤细胞U2OS/ADR模型。设置空白组、对照组、TPL组和维拉帕米阳性对照组(VER组)。CCK-8法测定TPL处理对U2OS/ADR细胞增殖和ADR敏感性的影响。以罗丹明123(Rh123)作为内标,采用流式细胞术评估细胞P-糖蛋白(P-gp)外排作用。实时定量PCR(qRT-PCR)检测TPL对MDR1基因转录的影响。Western blot检测TPL对磷酸化磷酸肌醇-3-激酶(p-PI3K)、磷酸化蛋白激酶B(p-AKT)、MDR1和肺耐药相关蛋白(LRP)表达的影响。结果:与对照组相比,TPL组使ADR对U2OS/ADR细胞生长的抑制作用提高(P<0.05)。TPL组Rh123阳性信号计数高于对照组(P<0.001)。与对照组相比,TPL组MDR1、LRP、p-PI3K和p-AKT蛋白表达水平及MDR1基因mRNA水平下降(均P<0.05)。结论:TPL具有抑制U2OS/ADR细胞生长、提高U2OS对ADR敏感性的作用,其机制可能与抑制PI3K/AKT信号通路活化、下调U2OS/ADR细胞MDR1和LRP基因表达、抑制P-gp对细胞毒物质的外排有关。Objective:To explore the reversal effect and possible mechanism of triptolide(TPL)in vitro on the multidrug resistance(MDR)of human osteosarcoma cell lines resistant to adriamycin(ADR)(U2OS/ADR).Methods:U2OS was used to establish a drug-resistant osteosarcoma cell model U2OS/ADR by increasing concentration scheme.The blank group,control group,TPL group and verapamil positive control group(VER group)were set up.CCK-8 test was used to determine the effects of TPL treatment on U2OS/ADR cell proliferation and ADR sensitivity.Rhodamine 123(Rh123)was used as an internal standard,and flow cytometry was used to evaluate cellular P-glycoprotein(P-gp)efflux.Real-time quantitative PCR(qRT-PCR)was used to detect the effect of TPL on the transcription of MDR1.Western blot was used to detect the effects of TPL on p-PI3K,p-AKT,MDR1 and LRP expression levels.Results:Compared with the control group,the TPL group increased the inhibitory effect of ADR on U2OS/ADR cell growth(P<0.05).The positive signal count of Rh123 in the TPL group was higher than that in the control group(P<0.001).Compared with the control group,the protein expression levels of MDR1,LRP,p-PI3K and p-AKT and the mRNA level of MDR1 gene decreased in the TPL group(all P<0.05).Conclusion:TPL can inhibit the growth of U2OS/ADR cells and increase the sensitivity of U2OS to ADR.The mechanism may be related to the inhibition of PI3K/AKT signal pathway activation,down-regulation of MDR1 and LRP gene expression in U2OS/ADR cells,and inhibition of the efflux of cytotoxic substances by P-gp.

关 键 词：雷公藤甲素 多药耐药基因 人骨肉瘤细胞阿霉素耐药株 PI3K通路 P-糖蛋白 肺耐药相关蛋白 

分 类 号：R738.1[医药卫生—肿瘤]