【作者】 王启文；

【导师】 李才；

【作者基本信息】 吉林大学 ， 病理学与病理生理学， 2009， 硕士

【摘要】 肺癌是当前世界上严重危害人类健康的恶性肿瘤之一,其发病率及死亡率很高,目前尚缺乏有效的治疗方法。近年国内外大量研究结果显示,他汀类药物具有多向性效应,除了降低血脂作用外,对多种肿瘤具有抑制细胞增殖、诱导细胞分化或凋亡、抑制血管生成、降低肿瘤细胞侵袭转移能力等作用,很可能成为一种具有应用前景的抗肿瘤药物。然而,有关氟伐他汀对小鼠Lewis肺癌抑制作用及其机制的研究尚未见报道。本研究应用移植性小鼠Lewis肺癌模型给予氟伐他汀处理后,观察对移植性肿瘤生长和肺转移的抑制作用;在体外观察氟伐他汀对小鼠Lewis肺癌细胞增殖和凋亡的影响。实验结果表明,氟伐他汀对小鼠Lewis肺癌移植瘤的生长和肺转移有明显抑制作用,且呈剂量和时间依赖方式;氟伐他汀与环磷酰胺联合应用对Lewis肺癌移植瘤生长和肺转移的抑制效果,明显优于单独应用氟伐他汀或环磷酰胺。体外研究结果表明,氟伐他汀呈剂量和时间依赖方式抑制Lewis肺癌细胞增殖;在体外氟伐他汀能诱导Lewis肺癌细胞凋亡,这种诱导细胞凋亡的作用是通过甲羟戊酸途径介导的。更多还原

【Abstract】 Lung carcinoma is one of the most common malignant tumors which severely harm the mankind health, both morbidity and mortality are high, and the effective therapeutic modalities remain insufficient.Statins, 3-hydroxy-3-methylgluyaryl coenzyme A reductase inhibitors, are the most effective medication for lowering cholesterol, cardiovascular morbidity and mortality. Fluvastatin is one of statins. In the recent years, accumulating evidence suggests that statins have pleiotropic effects beyond reducing cholesterol synthesis. Statins have been shown to inhibit proliferation, induce differentiation and apoptosis, reduce angiogenesis and invasion capability in several tumor cells, whereas have no side effects on the normal cells, suggesting their potent applications as an antitumor reagent. The antitumor potential of statins and its mechanism have become one of research hot spots nowadays. However, the inhibitory effects of fluvastatin on the growth and apoptosis of murine Lewis lung carcinoma and their mechanisms have not been reported yet.In the present work, the effect of Flu on the inhibition of tumor growth and pulmonary metastasis in a Lewis lung carcinoma murine model was studied. After Lewis lung carcinoma-bearing mice were administrated Flu at different doses for 15 days, the volume and mass weight of implanted tumors were measured, tumor weight inhibitory rates were calculated, lung weight and pulmonary metastasis nodules were counted, the inhibitory rates of pulmonary metastasis were calculated. The techniques of cytology, cytochemistry and flow cytometry were used to investigate the effects of fluvastatin on proliferation and apoptosis of Lewis lung carcinoma cells.The main results of this study are as follows:1. Flu showed significantly inhibitory effects on the growth of implanted tumors of Lewis lung carcinoma in mice. The inhibitory effects were increased with elevating dose of fluvastatin and extending action time, indicating a dose- and time-dependent manner, and the action of the high dose of fluvastatin was most obvious. As compared with treatment with fluvastatin or cyclophosphamide alone, combined treatment with both fluvastatin and cyclophosphamide resulted in more significant growth inhibition on implanted tumors.2. The results revealed that fluvastatin exerted suppressive action on pulmonary metastasis of Lewis lung carcinoma model. As compared with control, the lung weight and pulmonary metastatic foci were obvioursly decreased, and inhibitory rates of pulmonary metastasis were markedly elevated in the groups at high and middling doses of fluvastatin, CYC and Flu+CYC. The inhibitory effects of combined treatment with both fluvastatin and cyclophosphamide were more significant.3. After Lewis lung carcinoma cells were treated with 1,5, 10 and 20μmol/L of fluvastatin for 1, 2, 3, and 4 days in vitro, the cell growth activity was significantly suppressed. The inhibitory effects were increased with elevating concentration of fluvastatin and extending treated periods, suggesting a concentration- and time-dependent manner.4. The results of CCK-8 assay indicated that Flu showed significantly suppressive effect on the proliferation of Lewis lung carcinoma cells. In the same time-point of treatment, the cell proliferation inhibitory rate was elevated with increasing concentrations of fluvastatin, displaying a concentration-dependent manner, among them, the effects were more obvious at 10 and 20μmol/L of fluvastatin. In the same concentration of fluvastatin, the cell proliferation inhibitory rate was elevated with periods treated with fluvastatin, indicating a time-dependent manner, the inhibitory effects were more potent in fluvastatin-treated for 48 and 72 hours.5. Flow cytometry assays with Annexin V-FITC/PI double staining exhibited that compared with control, the apoptotic cells were increased after Lewis lung cancer cells were exposed to 20μmol/L of fluvastatin for 48 h (4.37% vs 12.83%, P<0.01), indicating fluvastatin induced apoptosis of Lewis lung carcinoma cells. Fluvastatin-induced cell apoptosis was reversed by coexposure to both fluvastatin and mevalonate (4.37% vs 5.89%, P>0.05), suggesting that the apoptosis inducing effect of fluvastatin was mediated through mevalonate pathway. 6. Electron microscopic study exhibited that after Lewis lung carcinoma cells were treated with fluvastatin (10μmol/L), the diminished cell body, condensed cytoplasm, dilated endoplasmic reticulum and vacuolization, as well as nuclear chromatin condensation with lumping and aggregating under the nuclear membrane and the apoptotic body were observed, showing the morphological changes of fluvastatin-induced apoptosis.The conclusions of this study are as follows:1. Fluvastatin exhibits significantly inhibitory effects on the growth and pulmonary metastasis of implanted tumors of Lewis lung carcinoma in mice in a dose- and time-dependent manner.2. The combined treatment with both fluvastatin and cyclophosphamide can produce more significant inhibitory effects on growth and pulmonary metastasis of implanted tumors as compared with treatment with fluvastatin or cyclophosphamide alone.3. Fluvastatin significantly inhibits proliferation of Lewis lung carcinoma cells in vitro, and the inhibitory effects are increased with elevating concentrations of fluvastatin and treated periods, suggesting a concentration- and time-dependent manner.4. Fluvastatin treatment is able to induce apoptosis of Lewis lung carcinoma cells in vitro, the apoptosis inducing effect of fluvastatin was mediated through mevalonate pathway.The creative points of the present study are: it is demonstrated that fluvastatin can suppress the growth and pulmonary metastasis of implanted tumors of Lewis lung carcinoma in mice, the combined treatment with both fluvastatin and cyclophosphamide can produce more significant inhibitory effects; fluvastatin inhibits proliferation of Lewis lung carcinoma cells in vitro in a concentration- and time-dependent manner, and induces apoptosis of Lewis lung carcinoma cells.更多还原