鱼藤素在诱导肺鳞癌细胞凋亡中的作用及相关机制研究

发布时间：2018-03-13 12:07

本文选题：鱼藤素　切入点：肺鳞癌　出处：《浙江大学》2016年博士论文　论文类型：学位论文

【摘要】：肺癌是目前全球范围内致死率居于首位的恶性肿瘤。尽管近年来对于肺癌治疗的研究取得了长足进展,特别是靶向药物的出现,更是给肺癌患者的治疗带来了新的曙光,但是相比于腺癌,肺鳞癌靶向治疗的探索却明显迟滞。目前临床治疗中尚不存在十分有效的肺鳞癌靶向药物,肺鳞癌潜在的分子靶点也仍在探索中,并且有多项临床研究结果显示,靶向药物在肺鳞癌患者的治疗中效果欠佳。因此,进一步探索潜在而有效的肺鳞癌治疗方法,改善肺鳞癌患者的预后势在必行。近年来,植物提取物鱼藤素开始逐渐受到人们的关注。有多项体内外研究结果显示,鱼藤素具有很强的抗肿瘤特性,可以抑制肿瘤细胞增殖,促进肿瘤细胞凋亡的发生。尽管这一作用已经在多种肿瘤中被研究证实,但是鱼藤素对于肺鳞癌的作用及相关机制却鲜有报道。本课题选用NCI-H520和SK-MES-1两种人肺鳞癌细胞株,分别进行细胞培养,通过CCK8、流式细胞凋亡检测、RT-PCR和western blot等技术分析鱼藤素对于肺鳞癌细胞增殖能力和凋亡的影响。并采用免疫共沉淀、细胞转染、流式细胞凋亡检测和western blot等技术探讨鱼藤素对于肺鳞癌细胞作用的相关机制和涉及的信号通路。同时,本课题还构建了裸鼠NCI-H520异种移植模型,通过体内实验进一步验证鱼藤素对于肺鳞癌的作用。本课题通过上述实验研究发现：鱼藤素可以显著增加肺鳞癌细胞的凋亡,且该促凋亡作用呈时间浓度依赖性。同时,鱼藤素还可以引起半乳糖凝集素-1(Galectin-1)表达水平的下降。过表达Galectin-1可以减弱鱼藤素对于肺鳞癌细胞的促凋亡作用,沉默Galectin-1蛋白表达可以增强肺鳞癌细胞对鱼藤素的敏感性。进一步研究发现,Galectin-1与H-Ras在肺鳞癌细胞内存在直接的相互作用。下调Galectin-1表达水平可以抑制抗凋亡Ras/Raf/ERK信号通路的激活。体内实验中,鱼藤素可以显著抑制移植瘤的生长,并降低Galectin-1表达水平。这些结果表明,鱼藤素可以通过下调Galectin-1表达水平来抑制Ras/Raf/ERK信号通路,从而诱导肺鳞癌细胞凋亡的发生。
[Abstract]:Lung cancer is the leading cause of death in the world. Although the research on lung cancer treatment has made great progress in recent years, especially the appearance of targeted drugs, it has brought a new dawn to the treatment of lung cancer patients. However, compared with adenocarcinoma, the exploration of targeted therapy for lung squamous cell carcinoma is obviously delayed. At present, there is no effective target drug for lung squamous cell carcinoma, and the potential molecular target of lung squamous cell carcinoma is still being explored. Many clinical studies have shown that targeted drugs are not effective in the treatment of lung squamous cell carcinoma. Therefore, it is imperative to further explore the potential and effective treatment of lung squamous cell carcinoma and improve the prognosis of patients with lung squamous cell carcinoma. In vivo and in vitro studies have shown that rotenin has strong anti-tumor properties and can inhibit the proliferation of tumor cells. Although this effect has been confirmed in many kinds of tumors, the effect of rotenin on lung squamous cell carcinoma and its related mechanism have been rarely reported. In this study, two kinds of human lung squamous cell carcinoma cell lines, NCI-H520 and SK-MES-1, were selected. The cell culture was carried out, and the effects of rotenin on the proliferation and apoptosis of lung squamous cell carcinoma cells were analyzed by CCK8, flow cytometry, RT-PCR and western blot. The cells were transfected by co-immunoprecipitation. Flow cytometry and western blot techniques were used to investigate the mechanism and signal pathway involved in the effect of rotenin on lung squamous cell carcinoma cells. At the same time, a xenotransplantation model of NCI-H520 in nude mice was constructed. The effect of rotenin on lung squamous cell carcinoma was further verified by in vivo experiments. In this study, we found that rotenin could significantly increase apoptosis of lung squamous cell carcinoma cells in a time and concentration-dependent manner. The overexpression of Galectin-1 could attenuate the apoptosis-inducing effect of catenin on lung squamous cell carcinoma cells. Silencing the expression of Galectin-1 protein can enhance the sensitivity of lung squamous cell carcinoma cells to rotenin. Further study shows that there is a direct interaction between Galectin-1 and H-Ras in lung squamous cell carcinoma cells. Down-regulation of Galectin-1 expression level can inhibit the anti-apoptotic Ras/Raf/ERK signal pathway. Activation of the pathway. In vivo experiments, These results suggest that rotenin can inhibit Ras/Raf/ERK signaling pathway by down-regulation of Galectin-1 expression and induce apoptosis in lung squamous cell carcinoma cells.
【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R734.2

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