咖啡醇抑制结肠癌的生物学功效

发布时间：2018-05-19 03:35

  本文选题：咖啡醇 + 结肠癌　； 参考：《昆明医科大学》2017年硕士论文

【摘要】：[目的]本研究从体外细胞水平出发,探寻咖啡醇对结肠癌细胞HCT116增殖的影响及其量效关系,通过检测自噬标志物确定咖啡醇诱导结肠癌细胞产生自噬性死亡,进而通过检测不同自噬通路上的关键靶基因深入探索咖啡醇诱导自噬的关键通路和作用机理。通过探索咖啡醇抑制结肠癌的生物学功效,有助于进一步开发咖啡醇资源,使咖啡醇在结肠癌的防控中发挥作用,对于推动药物治疗的发展,促进基础研究对慢性病预防控制的关键作用有重大意义。[方法]培养细胞HCT116,加入不同浓度咖啡醇(OμM、10μM、20μM、40μM、80μM、160μM)与细胞孵育48h后,通过MTT实验检测咖啡醇对结肠癌细胞HCT116增殖的影响及其量效关系,通过透射电镜检测自噬标志物,进而运用Western Blot检测咖啡醇诱导的LC3B-II的表达变化情况,最后使用实时荧光定量PCR检测不同自噬通路上的关键靶基因。[结果]1.MTT的结果表明,随着咖啡醇浓度的不断增加,细胞活性受到了抑制,即咖啡醇能够抑制结肠癌细胞的增殖。2.将HCT116细胞用不同浓度咖啡醇(OμM、20μM、40μM)处理24h后,用40μM咖啡醇孵育的细胞在电镜下观察可见线粒体发生变形,有双层膜环绕的自噬体,这意味着咖啡醇诱导HCT116细胞发生了细胞自噬。3.Western印迹检测自噬标志物LC3B-Ⅱ。研究结果显示,随着咖啡醇浓度的不断增加,LC3B-Ⅰ条带痕迹逐渐减淡,而LC3B-Ⅱ则逐渐加强,推测咖啡醇到达一定浓度时会引起结肠癌细胞发生细胞自噬。4.随着咖啡醇浓度逐渐增加,Beclinl和mTOR的表达变化水平无统计学意义,推测结肠癌细胞HCT116的自噬发生可能不是通过这两条通路,具体的关键靶点还需进一步研究和探讨。[结论]1.咖啡醇能抑制结肠癌细胞增殖。2.一定浓度的咖啡醇能诱导HCT116细胞发生自噬。3.不同咖啡醇浓度下Beclinl和mTOR的相对表达水平变化无统计学意义,提示咖啡醇诱导的细胞自噬可能不是通过这两条通路,具体的关键靶点还需进一步研究。
[Abstract]:[objective] to explore the effect of caffeine on the proliferation of colon cancer cell line HCT116 and its dose-response relationship, and to determine the autophagic death induced by caffeine by detecting autophagy markers. Furthermore, the key pathway and mechanism of caffeine induced autophagy were explored by detecting the key target genes in different autophagy pathways. By exploring the biological effects of caffeine on colon cancer, it is helpful to further develop the resources of coffee alcohol, to make caffeine play an important role in the prevention and control of colon cancer, and to promote the development of drug therapy. It is of great significance to promote basic research for the prevention and control of chronic diseases. [methods] cultured cells were incubated with HCT116, 10 渭 MU 10 渭 MU 10 渭 MU (10 渭 M) and 40 渭 M (40 渭 M), 80 渭 MU (160 渭 M) for 48 h. The effects of caffeine on the proliferation of HCT116 and its dose-effect relationship were detected by MTT assay, and the autophagy markers were detected by transmission electron microscopy (TEM). Western Blot was used to detect the changes of LC3B-II expression induced by caffeine, and real-time quantitative PCR was used to detect the key target genes in different autophagy pathways. [results] the results of 1.MTT showed that with the increasing of caffeine concentration, cell activity was inhibited, that is, caffeine could inhibit the proliferation of colon cancer cells. After HCT116 cells were treated with different concentrations of caffeine O 渭 M (20 渭 M) 40 渭 M for 24 h, the cells incubated with 40 渭 M caffeine showed that mitochondria deformed and there was a double layer membrane surrounded autophagy under electron microscope. This means that caffeine induces autophagy in HCT116 cells. 3. Western blot detection of autophagy marker LC3B- 鈪，

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