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目的建立人舌鳞癌卡铂耐药细胞株Tca8113/CBP,探讨甲基莲心碱Nef对Tca8113/CBP卡铂耐药性的逆转作用及其机制。方法采用逐步递增卡铂浓度、体外间歇诱导法建立Tca8113/CBP耐卡铂细胞株;MTT法检测Nef对Tca8113/CBP卡铂耐药性的逆转作用;RT-PCR法检测Nef作用前后Tca8113/CBP内耐药相关基因肺耐药蛋白LRP与成纤维细胞肌型原肌球蛋白FMT的表达水平;免疫荧光观察Nef作用前后细胞形态结构变化。结果历时5个月建立的Tca8113/CBP细胞株对卡铂耐药性稳定,耐药指数为4.06。Nef对Tca8113/CBP的卡铂耐药性有逆转作用,逆转倍数达到2.12倍;与Tca8113/CBP细胞比较,经Nef干预后,Tca8113/CBP细胞内LRP与FMT mRNA表达水平明显降低(P<0.01),细胞内F-actin的含量也明显下降,排列整齐,分布趋于均匀,接近非耐药Tca8113细胞水平。结论 Nef对Tca8113/CBP的卡铂耐药性有逆转作用,其机制可能与干预Tca8113/CBP细胞内LRP和FMT表达,增加Tca8113/CBP细胞内卡铂的含量,改变微丝结构,降低细胞黏附能力,干扰细胞增殖周期有关。
Objective To establish a human tongue squamous cell carcinoma cell line Tca8113 / CBP resistant to Tca8113 / CBP and its mechanism. Methods Tca8113 / CBP resistant carboplatin cell lines were established by increasing the concentration of carboplatin stepwise and intermittently in vitro. The reversal effect of Nef on the resistance of Tca8113 / CBP carboplatin was detected by MTT assay. The expression of Tca8113 / CBP The expression level of lung resistance protein LRP and fibromuscal tropomyosin FMT in endometriosis-resistant genes were detected by immunofluorescence. The morphological changes of cells before and after Nef were observed by immunofluorescence. Results The Tca8113 / CBP cell line established after 5 months showed stable resistance to carboplatin with a resistance index of 4.06. Compared with Tca8113 / CBP cells, the expression of LRP and FMT mRNA in Tca8113 / CBP cells was significantly decreased after Nef treatment (P <0.01) ), The content of F-actin in the cells also decreased significantly, arranged neatly, the distribution tended to be uniform, close to the level of non-resistant Tca8113 cells. Conclusions Nef can reverse the resistance of carboplatin in Tca8113 / CBP. The mechanism may be related to intervene the expression of LRP and FMT in Tca8113 / CBP, increase the content of carboplatin in Tca8113 / CBP, change the structure of microfilaments and decrease the cell adhesion Ability to interfere with cell cycle.