CAJ | 학술논문

目的探讨姜黄素对恶性黑素瘤体外抗癌作用的分子机制.方法体外培养的人黑素瘤细胞株A375和C8161分为实验组和对照组.实验组经不同浓度姜黄素处理一定时间,对照组采用二甲基亚砜处理.采用噻唑蓝法检测姜黄素对A375和C8161细胞增殖的影响,体外侵袭实验检测姜黄素对A375和C8161细胞侵袭的影响,流式细胞仪检测姜黄素对A375和C8161细胞周期的影响,Western印迹检测姜黄素对A375和C8161细胞AKT/mTOR信号通路相关蛋白表达的影响.结果噻唑蓝法显示,姜黄素分别在5～ 15 mg/L和5 ～ 10 mg/L浓度时,对A375和C8161细胞抑制作用呈量效关系;在0～48h呈时效关系,与对照组相比差异有统计学意义(P＜ 0.001).其24 h的IC50分别为10 mg/L和5 mg/L.体外侵袭试验显示,10 mg/L和5 mg/L姜黄素分别作用于A375和C8161细胞72 h,可明显抑制细胞侵袭(P＜ 0.001).流式细胞仪检测显示,10 mg/L和5 mg/L姜黄素分别作用于A375和C8161细胞24 h,细胞周期阻滞于G2/M期;A375细胞G2/M期比例为35.00％±3.54％,与对照组120.80％±7.46％相比,差异有统计学意义(P＜ 0.001);C8161细胞G2/M期比例为19.33％±4.04％,与对照组85.00％±9.53％相比,差异有统计学意义(P＜ 0.001).Western印迹检测显示,分别经10 mg/L和5 mg/L姜黄素作用于后,A375和C8161细胞AKT/mTOR信号通路相关蛋白表达水平下降.结论姜黄素抑制人黑素瘤细胞株A375和C8161增殖及侵袭机制可能与其诱导细胞周期阻滞及抑制AKT/mTOR信号通路活化有关.
목적 탐토강황소대악성흑소류체외항암작용적분자궤제.방법 체외배양적인흑소류세포주A375화C8161분위실험조화대조조.실험조경불동농도강황소처리일정시간,대조조채용이갑기아풍처리.채용새서람법검측강황소대A375화C8161세포증식적영향,체외침습실험검측강황소대A375화C8161세포침습적영향,류식세포의검측강황소대A375화C8161세포주기적영향,Western인적검측강황소대A375화C8161세포AKT/mTOR신호통로상관단백표체적영향.결과 새서람법현시,강황소분별재5～ 15 mg/L화5 ～ 10 mg/L농도시,대A375화C8161세포억제작용정량효관계;재0～48h정시효관계,여대조조상비차이유통계학의의(P＜ 0.001).기24 h적IC50분별위10 mg/L화5 mg/L.체외침습시험현시,10 mg/L화5 mg/L강황소분별작용우A375화C8161세포72 h,가명현억제세포침습(P＜ 0.001).류식세포의검측현시,10 mg/L화5 mg/L강황소분별작용우A375화C8161세포24 h,세포주기조체우G2/M기;A375세포G2/M기비례위35.00％±3.54％,여대조조120.80％±7.46％상비,차이유통계학의의(P＜ 0.001);C8161세포G2/M기비례위19.33％±4.04％,여대조조85.00％±9.53％상비,차이유통계학의의(P＜ 0.001).Western인적검측현시,분별경10 mg/L화5 mg/L강황소작용우후,A375화C8161세포AKT/mTOR신호통로상관단백표체수평하강.결론 강황소억제인흑소류세포주A375화C8161증식급침습궤제가능여기유도세포주기조체급억제AKT/mTOR신호통로활화유관.
Objective To explore molecular mechanisms underlying the in vitro counteracting effect of curcumin on malignant melanoma.Methods Cultured A375 and C8161 human melanoma cells were cultivated in vitro,and randomly divided into several test groups and a control group to be treated with different concentrations of curcumin and dimethyl sulfoxide respectively for different durations.Then,methyl thiazolyl tetrazolium (MTT) assay,Transwell assay,flow cytometry and Western blot were performed to evaluate the effect of curcumin on the proliferation,invasion and cell cycle of,as well as expressions of AKT/mTOR signaling pathway-related proteins in A375 and C8161 cells respectively.Statistical analysis was carried out by using t test.Results MTT assay showed that the treatment with curcumin of 5-35 mg/L for 24-96 hours significantly inhibited the proliferation of both A375 and C8161 cells compared with that with dimethyl sulfoxide (all P ＜ 0.001),and the inhibitory effect was in a dose-dependent manner within the range of 5-15 mg/L for A375 cells and within the range of 5-10 mg/L for C8161 cells,and in a time-dependent manner from 0 to 48 hours for both cells.After treatment for 24 hours,the 50％ inhibitory concentration (IC50) of curcumin against A375 cells and C 8161 cells was 10 mg/L and 5 mg/L respectively.Transwell assay demonstrated that the invasion of A375 and C8161 cells was significantly suppressed by 72-hour treatment with curcumin at 10 mg/L and 5 mg/L respectively (both P ＜ 0.001).Flow cytometry showed that the cell cycle of A375 and C8161 cells was arrested at G2/M phase after 24-hour treatment with curcumin at 10 mg/L and 5 mg/L respectively,with significant differences in the proportion of A375 cells and C8161 cells in G2/M phase between the test group and control group (A375 cells:35.00％ ± 3.54％ vs.120.80％ ± 7.46％,P＜ 0.001;C8161 cells:19.33％ ± 4.04％ vs.85.00％ ± 9.53％,P ＜ 0.001).Western blot revealed that the expressions of AKT/mTOR signaling pathway-related proteins were decreased in A375 and C8161 cells after 24-hour treatment with 10 mg/L and 5 mg/L curcumin respectively.Conclusion Curcumin can inhibit the proliferation and invasion of A375 and C8161 cells,likely by blocking cell cycle and inhibiting activation of the AKT/mTOR signaling pathway.