【作者】 王晓宇；

【导师】 金英花；

【作者基本信息】 吉林大学 ， 生化与分子生物学， 2010， 硕士

【摘要】 恶性肿瘤现已经成为威胁人类健康和生命,导致死亡的主要原因之一。目前,化学疗法,放射性治疗,免疫疗法是肿瘤术后治疗的方法,其中化学治疗是辅助术后治疗的主要手段。治疗肿瘤的方法都是利用药物诱导肿瘤细胞发生凋亡起作用的。化疗药物的疗效在一定范围内与药物剂量成正比。随着用药剂量的增加,机体产生毒副作用的风险也相应增加。发现有协同作用的抗肿瘤药物可以使药物在治疗安全窗内产生更好的疗效。本研究中,我们使用天然成分人参皂苷Rh2(G-Rh2)和桦木酸(Bet A)同时作用于人肝癌细胞HepG2和人肺癌细胞A549,并且广泛的测定与分析人参皂苷Rh2(G-Rh2)和桦木酸(Bet A)对肿瘤细胞的杀伤效果。我们采用细胞形态学测定、流式细胞仪检测分析、caspase-3酶活性测定分析、免疫印记法分析等实验技术,证明了人参皂苷Rh2(G-Rh2)和桦木酸(Bet A)两者可诱导癌症细胞发生凋亡并且二者同时作用于癌症细胞时对癌症细胞的促凋亡作用更加明显。随着药物联合作用时间的延长,细胞内发生明显的caspase-9前体活化,PARP断裂、caspase-3激活及凋亡。研究结果表明人参皂苷Rh2(G-Rh2)能有效提高肿瘤细胞对桦木酸(Bet A)的敏感性。更多还原

【Abstract】 At present, cancer is a serious threat to human and animal health, which is a serious threat to the health of people. Fragnent, the main treatment for cancer is surgery, supplemented by radiotherapy, chemotherapy and other methods. The efficacy of chemotherapy is proportional to the dose within a certain range. As the increases the risk of side effects occurred a corresponding increase in the tumor cells to chemotherapeutic drugs, tolerance, often lead to treatment failure. Found a synergistic effect of anticancer drugs can make drugs safe in therapeutic doses to generate better results. Most of the anti-tumor treatment, mainly including chemotherapy through induced apoptosis, and to prevent the body from drug tolerance, two and multi-drug combination therapy in the treatment of malignant tumors is better than a single anti-cancer drug. Because of synergies between the various drugs help to enlarge the original weak death signal. Therefore, the drug combination treatment may be a strong therapeutic strategy.Ginsenoside Rh2 (ginsenoside Rh2, G-Rh2) is extracted from ginseng with natural active, the study confirmed that the composition has a strong anti-tumor activity. According to reports, G-Rh2 can induce human breast cancer cells, human melanoma cells, human prostate cancer cells, human leukemia cells, human liver cancer cells and other tumor cells. Ginsenoside Rh2 also induces tumor cell differentiation, enhance immunity, and improve the ability of anti-tumor effect.Betulinic acid [3β-Hydroxy-lup-20(29)-en-28-oic acid] is a lup-type pentacyclic triterpenoids. Betulin acid is the carboxylic acid derivatives of C-28 of betulinol. However, betulinic acid with more people expected pharmacological activity which diference with betulin, such as anti-inflammatory, anti-cancer, anti-HIV and so on. Betulinic acid has anti-tumor, anti-HIV, anti-inflammatory effects, anti-malarial in vitro and other diseases.In this study, we found ginsenoside Rh2 significantly increased sensitivity to betulinic acid in A549 cells and HepG2 cells. Through analysis of cell morphology of two cell lines joining the two drugs, the morphological change significantly, there bubble membrane, nuclear condensation phenomena such as apoptosis. DAPI nuclear staining can be more easily observed phenomenon. Based on the above phenomenon, we analyzed by flow cytometry showed that, cells in the Sub-G1 significantly increased the percentage of the number when G-Rh2 and Bet A join to A549 cells and HepG2 cells simultaneously,. Caspase-3 activity analysis showed that, cells joined G-Rh2 and Bet A at the same time increased the activity of caspase-3, compared with G-Rh2 and Bet A were added alone, and the activity of caspase-3 increased significantly as the time. Using Western-Blot analysis PARP that is substrate of caspase-3 , the results show that the fracture of PARP when G-Rh2 and Bet A added at the same time increased significantly. Using Western-Blot analysis of caspase-9 activation, the results showed that activation of caspase-9 that G-Rh2 and Bet A all in tumor cells is higher than G-Rh2 and Bet A alone in A549 cells and HepG2 cells, and with time, activation is more evident.In summary, we found that ginsenoside Rh2 can improve the A549 cells and HepG2 cells to betulinic acid sensitivity, and confirmed that the result is caused by tumor cells through programmed cell death occurred.更多还原