兰州大学机构库 >学院待认领
漆黄素对膀胱癌细胞体内外作用的研究
Alternative TitleIn vitro and in vivo antitumor effects of fisetin on bladder cancer cells
李晶
Thesis Advisor王志平
2010-05-27
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name博士
Keyword膀胱 癌症 漆黄素 凋亡 细胞周期 动物模型
Abstract目的:本课题研究漆黄素对膀胱癌细胞的抑制作用,并进一步探讨其体内外作用机制,为漆黄素作为一种高效无毒的天然抗癌药物治疗膀胱癌提供有效的实验依据。方法: MTT法和克隆形成实验观察细胞存活率。Hoechst 33258染色检测细胞凋亡。FCM定量检测细胞周期变化和细胞凋亡率,RT-PCR检测了细胞中Bcl-2和Bax mRNA表达。Western blot检测凋亡调节蛋白的表达。本研究建立了大鼠膀胱癌原位动物模型。通过免疫组织化学法测定PCNA和TUNEL法测定细胞凋亡,探讨漆黄素在体内的抑瘤作用机制。结果: MTT实验结果和克隆形成实验显示,漆黄素抑制人膀胱癌细胞系的增殖;Hoechst 33258染色观察到凋亡的形态学变化;流式细胞术表明漆黄素使细胞周期停滞于G0/G1期。RT-PCR和Western blot的检测结果显示漆黄素下调了Bcl-2的表达,而上调了Bax的表达水平。漆黄素也调节了凋亡相关蛋白的表达。体内实验结果表明,漆黄素抑制和延迟了大鼠膀胱癌的形成和发展,同时诱导了大鼠膀胱癌细胞的凋亡。 结论:漆黄素对人膀胱癌细胞系体外增殖抑制作用,可能是通过对细胞周期分布的作用和诱导细胞凋亡而抑制膀胱癌细胞增殖作用。体内实验结果表明,漆黄素抑制和延迟大鼠膀胱癌细胞的形成与发展,同时诱导了癌细胞凋亡。预示了漆黄素是一种有效的预防与治疗膀胱癌的天然植物药。
Other AbstractObjective:Bladder cancer is one of the most common malignancies and tumor recurrence is a major clinical concern. Unfortunately, current chemotherapeutics are insufficient. Therefore, there is an obvious urgent need for novel, effective and safe therapies to prevent both recurrence and progression. The present study was carried out to investigate the effect of fisetin on cell cycle arrest and the induction of apoptosis in human bladder cancer cell lines and the mechanism of intravesical activity in a rat bladder tumor model. Methods:The MTT assay and clonal growth assay was used to determine cell viability. To determine whether the decrease in cell viability after fisetin treatment was the result of induction of apoptosis in bladder cancer cells, Hoechst 33258 fluorescence staining was performed to detect the morphology of cells. Meanwhile, apoptosis of the cancer cell lines was assessed by flow cytometric analysis after propidium iodide staining and the apoptotic-associated proteins were measured by western blot. RT-PCR was used to detect the expression of Bax and Bcl-xL mRNA. In addition, this study established a rat bladder carcinoma animal model induced by MNU. We observed the pathology changes of bladder by the HE staining. The tumor cell proliferation and apoptosis in a rat bladder carcinogenesis model were evaluated by proliferating cell nuclear antigen and terminal deoxyribonucleotide transferase-mediated nick-end labeling method. Results: Our results indicated that fisetin had cytotoxic effects and elicited a dose- and time-dependent inhibition of bladder cancer cell proliferation. Our data demonstrated that treatment of fisetin (60-100 μM; 48 h) was found to bring about the induction of apoptosis in bladder cancer cells, as evidenced by Hochest 33258 staining and sub-G1 peak of apoptotic markers detection in T24 and EJ cells exposed to fisetin. Our results also clearly showed that, in fisetin-treated T24 and EJ cells, apoptosis was accompanied by an increase in the percent of cells in G0/G1 and reduction in G2/M indicating the cell cycle arrest in G0/G1. G1-phase arrest was associated with a marked decrease in the protein expression of cyclin D1, cyclin E, CDK4 and CDK2 and the increase in the protein expression of p53 and p21. There was also induction of mitochondrial release of cytochrome c into cytosol. In addition, the study also found that exposure of human bladder cancer cells to fisetin could upregulated the expression of Bax and B...
URL查看原文
Language中文
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/222875
Collection学院待认领
Affiliation临床医学院
Recommended Citation
GB/T 7714
李晶. 漆黄素对膀胱癌细胞体内外作用的研究[D]. 兰州. 兰州大学,2010.
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