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为探讨双依他尼酰乙二胺(EDEA)对人耐顺铂卵巢癌细胞(COC1/DDP)的抗肿瘤活性,本实验用CCK-8细胞增殖实验检测顺铂(DDP)及双依他尼酰乙二胺对细胞生长的抑制作用,用流式细胞术检测细胞凋亡率,Western Blot检测细胞内Bcl-2家族凋亡相关蛋白表达,瑞氏染色观察细胞形态。结果显示,EDEA对人正常细胞HEK293、LO2的低毒剂量接近1.2μmol/L,DDP对上述细胞低毒剂量约1.0μmol/L。分别作用COC1/DDP细胞24 h、48 h、72 h后,EDEA的半抑制浓度IC_(50)均为0.67μmol/L,但DDP对应IC_(50)分别为52.9μmol/L、18.1μmol/L和15.2μmol/L。在浓度均为1.0μmol/L时,EDEA作用COC1/DDP细胞48 h后其凋亡率接近44%,而DDP作用此细胞48h后凋亡率仅约4%。与单用1.0μmol/L DDP处理相比,1.0μmol/L EDEA作用48 h后Bcl-2等抗凋亡蛋白下调,Bax等促凋亡蛋白上调,且p-Akt表达被抑制,细胞形态发生凋亡样改变。结论,EDEA对人正常细胞低毒剂量与DDP相当但对COC1/DDP有显著生长抑制作用,且药效远强于相同浓度DDP;EDEA对耐顺铂卵巢癌细胞COC1的生长抑制作用与p-Akt表达抑制和细胞凋亡增强相关。
In order to investigate the antitumor activity of dual-EDTA on human cisplatin-resistant ovarian cancer cells (COC1 / DDP), CCK-8 cell proliferation assay was used to detect the effects of cisplatin (DDP) The inhibitory effect of NADPH on cell growth was detected by flow cytometry. The apoptosis-related proteins of Bcl-2 family were detected by Western Blot. Wright’s stain was used to observe the cell morphology. The results showed that the low dose of EDEA to human normal cells HEK293, LO2 was close to 1.2μmol / L, DDP low dose of about 1.0μmol / L. The half inhibitory concentration (IC 50) of EDEA was 0.67μmol / L, while the corresponding IC 50 of DDP were 52.9μmol / L and 18.1μmol / L for 24 h, 48 h and 72 h, respectively And 15.2 μmol / L. At a concentration of 1.0 μmol / L, the apoptosis rate of COC1 / DDP cells treated with EDEA for 48 h was 44%, while the apoptotic rate of DDP treated cells was only about 4% after 48 h. Compared with 1.0μmol / L DDP treatment alone, the anti-apoptotic protein such as Bcl-2 was down-regulated after Bcl-2 treatment with 1.0μmol / L EDEA for 48h, the pro-apoptotic proteins such as Bax were upregulated and the expression of p-Akt was inhibited. Apoptosis-like changes. CONCLUSION: EDEA has a low toxic dose equivalent to DDP but a significant inhibitory effect on COC1 / DDP in human normal cells, and its potency is much higher than that of DDP. EDEA inhibits the growth of COC1 in cisplatin-resistant ovarian cancer cells and p- Inhibition of Akt expression is associated with increased apoptosis.