肝素和阿霉素对人鼻咽癌CNE2细胞增殖与凋亡的协同作用

摘要

AIM:To measure the effect of addition of heparin to adriamycin (ADM) on cell proliferation and apoptosis in CNE2 cells and investigate the possible molecular mechanisms of heparin and ADM interactions.METHODS:Cell viability and cell cycle were determined by MTT assay and flow cytometry.Apoptosis was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)and agarose gel electrophoresis.The expression of Bax,Bcl-2,p53,and p21 was examined by Westem blot.RESULTS:ADM (5 mg/L) alone inhibited the growth of CNE2 cells,which was magnified when heparin was added.ADM elicited typical apoptotic morphologic changes.Compared with ADM or heparin alone,ADM plus heparin obviously enhanced the number of TUNEL-positive cells from 12.6 % ± 1.1% to 65.7 % ±1.3 %,and the DNA ladder was more clearly observed.After exposure to different concentrations of heparin ( with or without ADM) for 24 h,CNE2 cells were accumulated in G0/G1 phase.There was a decrease in the number of cells in S phase by the combined heparin and ADM treatment compared to heparin or ADM alone.The ratio of Bax/Bcl-2 was elevated,and p53 and p21 mRNA were over-expression.CONCLUSION:Heparin and ADM appear to interact in a synergistic manner,which may be related to the over-expression of p53 and p21 mRNA and the elevated ratio of Bax/Bcl-2 mRNA.%目的:观察肝素联合阿霉素对人鼻咽癌细胞的=增殖与凋亡的影响及其可能的分子机制.方法:用MTT法、流式细胞术观察肝素联合阿霉素对人鼻咽癌细胞增殖及细胞周期的影响;而用TUNEL、琼脂糖凝胶电泳、Western blot等方法观察肝素与阿霉素联合应用对人鼻咽癌细胞凋亡的作用.结果:肝素与阿霉素联合应用后对鼻咽癌细胞的生长抑制及其凋亡具有显著增强作用,同阿霉素单独应用于诱导细胞凋亡相比,低剂量的肝素与阿霉素联合应用后发现:TUNEL阳性细胞从12.6%±1.1%增加到65.7%±1.3%;G0/G1期细胞阻止,S期细胞明显减少,DNA"梯形"变化更加明显;Bax/Bcl-2的比率及p53和p21基因的表达明显升高.结论:肝素与阿霉素对人鼻咽癌细胞在抗增殖及促凋亡方面有协同作用,这可能与Bax/Bcl-2比率的升高及p53和p21的表达上调有关.