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目的:研究香加皮单体成分宝藿苷Ⅰ在体内外对人食管癌细胞Eca109的增殖抑制作用及其对细胞凋亡的影响。方法:采用柱层析和高效液相色谱等逐步分离法对香加皮抗肿瘤活性成分进行分离和纯化,应用薄层层析和电喷质谱分析进行成分鉴定;采用MTT法分析不同浓度宝藿苷Ⅰ对食管癌细胞株Eca109增殖的抑制作用;应用FCM分析细胞周期变化和凋亡率变化;皮下注射Eca109细胞建立裸鼠移植瘤动物模型,通过肌肉注射给药检测宝藿苷Ⅰ的体内抗肿瘤效果;Western印迹法检测移植瘤组织凋亡相关基因survivin的表达变化。结果:宝藿苷Ⅰ可明显抑制Eca109细胞的增殖(P<0.05),并呈浓度及时间依赖性。经宝藿苷Ⅰ作用48h后,随着药物浓度的增加(12.5、25.0和50.0μg/mL),Eca109细胞G0/G1期明显增加(P<0.05),S期和G2/M期细胞明显减少(P<0.05);经宝藿苷Ⅰ作用后,Eca109细胞的凋亡率随药物作用时间的延长(24、48和72h)和宝藿苷浓度的增加(0、12.5、25.0和50.0μg/mL)而明显升高(P<0.01);Eca109裸鼠移植瘤经宝藿苷Ⅰ(15mg/kg)治疗后,肿瘤生长受到明显抑制(P<0.01),生长抑制率达(60.9±0.16)%;survivin的蛋白表达明显降低(P<0.05)。结论:香加皮单体成分宝藿苷Ⅰ不仅在体外对Eca109细胞的增殖具有显著的抑制作用,在体内也有较好的抗食管癌效果。其抗瘤机制可能与阻滞Eca109细胞周期发展和诱导凋亡相关基因survivin的表达降低有关。
OBJECTIVE: To study the inhibitory effect of rubigoside Ⅰ, a monomer of Xiangnong Pi, on the proliferation of human esophageal cancer cell line Eca109 in vitro and in vivo and its effect on apoptosis. METHODS: The antitumor active components of Xiangkapi were separated and purified by step-by-step separation methods such as column chromatography and high performance liquid chromatography. The constituents were identified by TLC and electrospray ionization mass spectrometry. Glycosides Ⅰ on the proliferation of esophageal cancer cell line Eca109; FCM analysis of changes in cell cycle and apoptosis rate; subcutaneous injection of Eca109 cells in nude mice xenograft tumor model, Tumor effect. The expression of survivin in the tumor tissue was detected by Western blotting. Results: Po-glycoside Ⅰ significantly inhibited the proliferation of Eca109 cells (P <0.05) in a concentration-and time-dependent manner. After 48 hours of treatment, the G0 / G1 phase of Eca109 cells was significantly increased (P <0.05) with the increase of drug concentration (12.5,25.0 and 50.0μg / mL), while the cells in S phase and G2 / M phase were significantly decreased (P0.05) .Apoptosis rate of Eca109 cells increased with the increase of the action time (24, 48 and 72h) and the concentration of treasure glycosides (0,12.5,25.0 and 50.0μg / (P <0.01). The tumor growth of Eca109 xenografts was significantly inhibited (P <0.01) and the growth inhibition rate was (60.9 ± 0.16) %. The protein expression of survivin was significantly decreased (P <0.05). CONCLUSION: Botrytis licorice I, a component of Xiangkapi, not only has a significant inhibitory effect on the proliferation of Eca109 cells in vitro, but also has good anti-esophageal cancer effect in vivo. The anti-tumor mechanism may be related to blocking the development of Eca109 cell cycle and decreasing the expression of apoptosis-related gene survivin.