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目的研究抗肿瘤药物白桦脂酸(BA)对人食管鳞癌KYSE170细胞的抑制作用及抗肿瘤作用机制。方法采用不同质量浓度(0、5、10、20、40、60、80、100μg.mL-1)白桦脂酸处理KYSE170细胞,作用不同时间(24、48、72 h)后,噻唑蓝(MTT)法检测KYSE170细胞生长的短期抑制作用;不同质量浓度(0、5、20、40μg.mL-1)白桦脂酸作用于细胞24 h后以克隆形成实验检测药物对细胞生长的长期抑制作用;不同质量浓度白桦脂酸(10、60、100μg.mL-1)作用KYSE170细胞24、48 h后以流式细胞仪FITC-Annexin V/PI双染法检测细胞凋亡变化,不同质量浓度白桦脂酸(0、10、40μg.mL-1)作用细胞24 h后以流式细胞仪检测细胞周期的改变。结果噻唑蓝实验显示,不同质量浓度白桦脂酸对KYSE170细胞均有抑制作用,抑制效果呈时间和剂量依赖性,24、48、72 h的IC50分别为(56.81±2.56)、(39.73±2.77)和(29.28±3.05)μg.mL-1;克隆形成结果显示,0、5、20、40μg.mL-1药物作用细胞后,克隆形成率分别为(89.56+5.00)%、(61.00+2.03)%、(31.33+3.51)%和(15.33+2.33)%,随白桦脂酸质量浓度增大而明显降低(P<0.01)。凋亡结果显示,各实验组KYSE170细胞凋亡率随白桦脂酸质量浓度加大和药物作用时间的延长明显增高(P<0.05或0.01)。细胞周期结果显示,0、1、10μg.mL-1白桦脂酸作用细胞24 h后,随药物质量浓度增大,G1期细胞比例减少,S期细胞比例不断增加,实验组与空白组对比均有统计学差异(P<0.01)。结论白桦脂酸能抑制人食管鳞癌KYSE170细胞的生长,这一作用可能是通过诱导细胞凋亡、阻滞细胞停留在S期来实现的。
Objective To study the inhibitory effect of anti-tumor drug betulinic acid (BA) on human esophageal squamous cell carcinoma KYSE170 cells and its anti-tumor mechanism. Methods KYSE170 cells were treated with Betulinic acid at various concentrations (0, 5, 10, 20, 40, 60, 80, 100μg.mL-1) for 24 hours, 48 hours and 72 hours. MTT ) Assay was used to detect the short-term inhibition of KYSE170 cell growth. The long-term inhibition of cell growth was detected by colony formation assay after different concentrations of betulinic acid (0, 5, 20, 40μg.mL-1) The apoptosis of KYSE170 cells treated with different concentration of betulinic acid (10, 60, 100μg.mL-1) for 24h and 48h was detected by flow cytometry FITC-Annexin V / PI double staining method. Different concentrations of betulin After treated with acid (0, 10, 40μg.mL-1) for 24 h, the changes of cell cycle were detected by flow cytometry. Results The results of thiazolyl blue assay showed that betulinic acid inhibited the proliferation of KYSE170 cells in a time and dose dependent manner. The IC50 values at 24, 48 and 72 h were (56.81 ± 2.56) and (39.73 ± 2.77) And (29.28 ± 3.05) μg.mL-1, respectively. Clone formation results showed that the rates of colony formation were (89.56 + 5.00)%, (61.00 + 2.03) %, (31.33 + 3.51)% and (15.33 + 2.33)%, respectively, with the concentration of betulinic acid increasing (P <0.01). The results of apoptosis showed that the apoptosis rate of KYSE170 cells in each experimental group was significantly increased with the increase of the concentration of betulinic acid and the prolongation of the drug action time (P <0.05 or 0.01). The results of cell cycle showed that the percentage of cells in G1 phase decreased and the percentage of S phase cells increased with the increase of drug concentration 24 h after treatment with 0, 1, 10μg.mL-1 betulinic acid There was a significant difference (P <0.01). Conclusion Betulinic acid can inhibit the growth of human esophageal squamous cell carcinoma KYSE170 cells, which may be achieved by inducing apoptosis and arresting cells in S phase.