【作者】 左连富； 王静； 刘亮； 李金梅； 刘江惠； 郭建文；

【Author】 Zuo Lian-Fu Wang Jing Liu Liang Li Jin-Mei Liu Jiang-Hui Guo Jian-Wen the fourth hospital of Hebei medical university & Hebei provincial tumor hospital, Shijiazhuang, 050011, China

【机构】 河北医科大学大学第四医院暨河北省肿瘤医院肿瘤研究所；

【摘要】 目的:观察青蒿琥酯(Artesunate,Art)对人食管癌Eca 109细胞株及裸鼠移植瘤的抑瘤作用,并进一步探讨 Art对肿瘤细胞的周期阻滞作用。方法:体外培养人食管癌 Eca109细胞株,常规分离正常人外周血淋巴细胞,MTT法测定不同浓度Art对Eca109细胞及淋巴细胞增殖的影响,流式细胞术(FCM)测定肿瘤细胞的细胞周期变化,构建荷瘤裸鼠模型,观察Art对裸鼠人食管癌移植瘤的抑制情况。结果:MTT结果显示1μmol／L、10μmol／L、100μmol／ L Art均能显著抑制Eca109细胞的增殖(P<0．01),IC50 为(68．80±0．76)μmol／L,最大抑制率可达(78．31±1．48) ％,抑瘤水平与顺铂相当(74．27±0．41)％。Eca109细胞生长活力随Art浓度的增加以及作用时间的延长而降低,Art的抑制作用呈剂量,时间依赖性。Art对ConA诱导的淋巴细胞的生长增殖仅有轻微的抑制作用,抑制率约为2％～8％,表明各实验浓度的Art对正常免疫功能影响较小,顺铂对淋巴细胞的抑制率约为15％。流式细胞术检测结果显示,不同浓度Art对Eca109细胞的细胞周期的影响亦不同。对照组处于 S期的细胞多达41．8％,体现了肿瘤细胞生长增殖旺盛的特点。1 μ mol／L、10μmol／LArt可使S期细胞明显减少,细胞的生长被阻滞在G0+G1期,说明低浓度Art可有效阻止肿瘤细胞的基因组DNA的合成。100μmol／L Art亦可使 S期细胞数目减少,但其作用主要表现为将细胞阻滞在 G2+M期,说明高浓度Art不仅阻止肿瘤细胞的基因组DNA 的合成,还可显著抑制肿瘤细胞的分裂增殖。肿瘤细胞接种于裸鼠一周后,实验各组裸鼠均观察到有Eca 109细胞移植瘤生长,瘤体呈椭圆形或圆形,经组织病理学检查为鳞状细胞癌,可见明显角化细胞、癌细胞排列成巢状结构。实验结束时,各实验组肿瘤的体积及重量均明显小于对照组(P< 0．05),100 mg／kg、200 mg／kg、300 mg／kgArt组体积抑瘤率分别为32．6％、76．4％、18．0％,重量抑瘤率分别为 11．4％、33．2％、21．6％,顺铂组体积抑瘤率为48．3％,重量抑瘤率为39．7％。实验结果显示,Art的抑瘤作用并不随药物浓度的增大而增强,中浓度Art的抑瘤效果最佳,其抑瘤作用与顺铂相当。Art各组用药前后生长、饮食、活动等情况良好,未观察到血尿和便血,体重变化无显著性差异,顺铂组用药后体重明显减轻,并出现进食减少,行动迟缓,皮肤干涩等现象。实验结果表明,各实验浓度Art对裸鼠的毒副作用较小,而顺铂的毒副作用较大。结论:Art可显著抑制肿瘤细胞及裸鼠移植瘤的生长,且无明显的毒副作用,将肿瘤细胞阻滞于G0+G1及G2+M期,是Art发挥抑瘤作用的重要机制。更多还原

【Abstract】 Objective: To observe the effects of Art on the proliferation of human esophageal carcinoma Eca109 cell line and transplantation tumors of nude mice, further explore the cell cycle arrest effect of Art on tumor cells. Methods: Human esophageal carcinoma Eca109 cell and healthy human peripheral blood lymphocytes were cultured, the inhibitory effects of Art on cell proliferation were determined by MTT method, the changes of cell cycle of tumor cells were assayed by flow cytometry (FCM), the models of nude mice that bear tumors were established, the inhibitory effects of Art on the transplantated human esophageal carcinoma of nude mice were observed. Results: MTT analysis indicated that Art with the concentration of 1 μmol/L, 10 μmol/L, 100 μmol/L could significantly inhibit the proliferation of Eca109 cells(P < 0.01), and IC50 value was (68.80 ± 0.76) μmol/L, the inhibitory rate could reach at (78.31 ± 1.48)%, which was equivalent to that of PPD(74.27 ± 0.41)%. The inhibitory effect of Art was dose and time dependent Compared with PPD, Art had weaker inhibitory effect on the proliferation of lymphocytes, the inhibitory rate was about 2%-8%. FCM analysis of DNA content indicated that Art with different concentrations had different effect on the cell cycle. The majority of Eca109 cells from the control group located at S phase(41.8%), a characteristic of tumor cells. Of note, 1 μmol/L, 10 μmol/L Art could arrest the vast majority of Eca109 cells within G0+G1 phases with significantly decreased distribution at S phase. Interestingly, when the concentration of Art was up to 100 μmol/L, the cell cycle distribution changed greatly, most cells were arrested within G2+M phases with decreased distribution at S phase. Transplantation tumors were observed in all groups, and were confirmed to be esophageal squamous cell carcinoma by histological method. Tumor volume and weight of groups which were administrated with Art were smaller than those of control group, the maximum inhibitory rate was 76.4%. Conclusion: Art could inhibit the proliferation of tumor cell and transplantation tumor of nude mice with no apparent side effects, to arrest majority of tumor cells at G0+G1 and G2+M phases is the important mechanism of antitumor effects of Art.更多还原