哈萨克药阿尔泰瑞香不同提取物对人食管癌Eca-109细胞的体外抑制作用

发布时间：2018-01-10 22:03

本文关键词：哈萨克药阿尔泰瑞香不同提取物对人食管癌Eca-109细胞的体外抑制作用　出处：《新疆医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:采用现代细胞分子生物学实验技术,初探阿尔泰瑞香提取物抗食管癌Eca-109细胞作用机制,为哈萨克药阿尔泰瑞香的后期研究提供依据。方法:将阿尔泰瑞香三种提取物不同浓度,依次作用于人食管癌Eca-109细胞,采用四甲基偶氮唑盐法(MTT法)测定Eca-109细胞的增殖抑制率;通过计算细胞增殖抑制率,分析阿尔泰瑞香提取物对人食管癌细胞系Eca-109细胞增殖活性的影响。应用倒置荧光显微镜技术,观察细胞生长状况及形态学变化。运用流式细胞术,检测细胞周期及细胞凋亡率,探讨阿尔泰瑞香对人食管癌Eca-109细胞的增殖抑制的作用及机制。采用蛋白免疫印迹技术及实时荧光定量PCR技术对细胞内的PPARγ基因的表达水平进行分析,探讨阿尔泰瑞香不同提取物抗人食管癌Eca-109细胞的作用机制及与基因表达调控的关系。结果:阿尔泰瑞香三种提取物对人食管癌Eca-109细胞具有明显的抑制细胞生长的作用,与对照组比较,差异有统计学意义(P㩳0.05)。人食管癌Eca-109细胞经阿尔泰瑞香三种提取物处理后细胞形态发生变化。用阿尔泰瑞香三种提取物不同浓度处理人食管癌Eca-109细胞后处于S期的细胞数显著升高,跟空白对照组比较,差异具有统计学意义(P㩳0.05);G0/G1期的细胞数明显下降,与空白对照组比较,差异亦具有统计学意义(P㩳0.05)。人食管癌Eca-109细胞经阿尔泰瑞香三种提取物不同浓度处理后,检测细胞凋亡率,结果与空白对照组比较药物干预后癌细胞凋亡率明显增大,差异有统计学意义(P㩳0.05)。人食管癌Eca-109细胞经阿尔泰瑞香提取物不同浓度处理不同时间(24 h,48 h,72 h)后采用荧光定量PCR法检测细胞内PPARγmRNA表达量,结果经三种提取物处理癌细胞后细胞内PPARγmRNA表达量均显著增加(P㩳0.05)。阿尔泰瑞香乙酸乙酯提取物干预24 h,48 h,72 h后细胞内PPARγmRNA表达量增高(P㩳0.05)。阿尔泰瑞香正己烷、甲醇提取物干预48 h、72 h细胞内PPARγmRNA表达量增高(P㩳0.05),而干预24 h后PPARγmRNA表达量未见明显变化(P0.05)。Western Blot结果显示经阿尔泰瑞香提取物干预后细胞内PPARγ蛋白表达增高(P㩳0.05),干预48 h后的上调作用较强。结论:阿尔泰瑞香提取物抑制人食管癌Eca-109细胞增殖,阻滞细胞周期,诱导细胞凋亡,上调细胞内PPARγ基因表达。
[Abstract]:Objective: to explore the mechanism of anti-esophageal cancer Eca-109 cells by using modern cell molecular biological techniques. Methods: three kinds of extracts of Artemisia artemiana were used in human esophageal carcinoma Eca-109 cells in different concentrations. The proliferation inhibition rate of Eca-109 cells was measured by MTT assay. By calculating the inhibition rate of cell proliferation, the effect of the extract of Artemisia tenuifolia on the proliferation of human esophageal carcinoma cell line Eca-109 was analyzed. The inverted fluorescence microscopy was used. Cell cycle and apoptosis rate were detected by flow cytometry. To investigate the inhibitory effect and mechanism of Artemisia on the proliferation of human esophageal carcinoma Eca-109 cells. Expression of PPAR 纬 gene in human esophageal carcinoma Eca-109 cells by Western blot and real-time fluorescence quantitative PCR. Level analysis. To investigate the mechanism of different extracts of Daphne artemiana on human esophageal carcinoma Eca-109 cells and its relationship with gene expression regulation. The three extracts of Artemisia artemiana have obvious inhibitory effect on the growth of human esophageal carcinoma Eca-109 cells. Compared with the control group, the difference was statistically significant. 0.05). The morphological changes of human esophageal carcinoma Eca-109 cells treated with three extracts of Artemisia artemianum were observed in different concentrations of human esophageal carcinoma Eca-109 cells. The number of cells in S phase was significantly increased. Compared with the blank control group, the difference was statistically significant. The number of cells in the G _ 0 / G _ 1 phase of 0.05G _ 0 / G _ 1 decreased significantly, and the difference was statistically significant compared with the blank control group. The apoptosis rate of human esophageal carcinoma Eca-109 cells was measured after treated with different concentrations of three extracts of Artemisia tenuifolia. Results compared with the blank control group, the apoptosis rate of cancer cells increased significantly after drug intervention, and the difference was statistically significant. Human esophageal carcinoma Eca-109 cells were treated with different concentrations of Artemisia tenuifolia extract for 24 h or 48 h. After 72 h, the expression of PPAR 纬 mRNA was detected by fluorescence quantitative PCR assay. Results after treated with three kinds of extracts, the expression of PPAR 纬 mRNA in cancer cells increased significantly. The expression of PPAR 纬 mRNA in the cells was increased after treatment with ethyl acetate extract for 24 h and 48 h for 72 h. The expression of PPAR 纬 mRNA in the cells was increased after 48 h of treatment with methanol extract. 0.05). However, the expression of PPAR 纬 mRNA did not change significantly after 24 h intervention (P 0.05). The results of Western Blot showed that the expression of PPAR 纬 protein in the cells was increased after the intervention of the extract of Artemisia tenuifolia. Conclusion: the extract of Daphne altae inhibits the proliferation of human esophageal carcinoma Eca-109 cells, blocks cell cycle and induces apoptosis. The expression of PPAR 纬 gene was up-regulated.
【学位授予单位】：新疆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R29

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