洛铂诱导宫颈癌SiHa细胞凋亡与增强射线敏感性的研究

发布时间：2018-03-20 18:39

  本文选题：洛铂　切入点：射线敏感性　出处：《西南医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:探讨洛铂抑制宫颈癌SiHa细胞系(cervical cancer cell line SiHa,SiHa)增殖、诱导细胞凋亡及增强细胞射线敏感性的作用及机制。方法:1.以0、1、2.5、5、10、20μmol/L浓度的洛铂对SiHa细胞的作用24、48和72 h,利用四氮唑溴盐比色试验[3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetra-zoliumromide,MTT]检测洛铂对宫颈癌SiHa细胞增殖率的影响。2.选取对数期生长的SiHa细胞接受洛铂处理,依次接受0、2、4、6、8 Gy剂量的X射线照射48小时,利用单克隆形成实验检测洛铂对SiHa细胞的射线敏感性影响。3.依照处理方式的不同,依次分为空白组、洛铂组(5μmol/L)、照射组(6 Gy)和洛铂(5μmol/L)+照射组(6 Gy)。利用流式细胞术检测洛铂、射线以及洛铂联合射线对SiHa细胞凋亡率的影响。4.利用Western blotting检测洛铂对促凋亡蛋白Bax和抑凋亡蛋白Bcl-2在SiHa细胞中表达的影响。结果:1.MTT结果得出不同浓度的洛铂处理SiHa细胞24、48和72 h的IC 50值。2.单击多靶模型拟合单克隆形成实验数据得出空白组和洛铂处理组的D0值分别为:3.588 Gy,2.226 Gy;空白组和洛铂组Dq值分别为:0.792 Gy,0.053 Gy;洛铂在SiHa细胞中的SER值为1.612。3.流式细胞结果检测空白组、洛铂组、射线组和洛铂联合射线组的凋亡率分别为:4.44±2.07%,30.50±3.17%,43.08±5.93%和84.68±4.90%。4.Western blotting检测发现洛铂组、射线组相对空白组促凋亡蛋白Bax表达显著上调,抑凋亡蛋白Bcl-2显著下调。洛铂联合射线组相对洛铂组、射线组进一步显著上调Bax和下调Bcl-2的表达。结论:1.洛铂能够显著的抑制宫颈癌SiHa细胞的增殖,随着洛铂药物浓度增大和作用时间的延长SiHa细胞的增殖率逐渐下降。2.洛铂能显著增强宫颈癌SiHa细胞的射线敏感性,洛铂处理后的SiHa细胞在射线下的存活分数显著低于未经洛铂处理的SiHa细胞。3.洛铂和放射照射分别能诱导宫颈癌SiHa细胞凋亡,并且洛铂联合射线能够更加显著促进SiHa细胞凋亡。4.洛铂和放射照射作用于宫颈癌SiHa细胞时,洛铂和放射照射分别能够提高促凋亡蛋白Bax的表达水平。洛铂同步放射照射能提高促凋亡蛋白Bax的表达水平,且显著高于单独使用洛铂或放射照射时。5.洛铂和放射照射作用于宫颈癌SiHa细胞时,洛铂和放射照射分别能够降低抑凋亡蛋白Bcl-2的表达水平。洛铂同步放射照射能降低抑凋亡蛋白Bcl-2的表达水平,且显著低单独使用洛铂或放射照射时。6.洛铂通过诱导增加促凋亡蛋白Bax的表达水平、抑制抑凋亡蛋白Bcl-2的表达水平,来实现对宫颈癌SiHa细胞的放疗增敏作用。
[Abstract]:Objective: to investigate the inhibitory effect of Loxplatin on the proliferation of cervical carcinoma SiHa cell line, Cervical cancer cell line SiHaSiHa. Effects and mechanisms of inducing apoptosis and enhancing radiosensitivity. Methods the proliferation rate of Loxoplatin on SiHa cells of cervical cancer was detected by MTT assay for 24 h, 48 h and 72 h, respectively. SiHa cells growing in logarithmic phase were treated with loplatin. After 48 hours of X-ray irradiation at a dose of 0, 2, 2, 4, 6 and 8 Gy, the effects of roplatin on the radiosensitivity of SiHa cells were detected by monoclonal formation assay. The cells were divided into blank group according to the different treatment methods. Luoplatin group (5 渭 mol / L) and roplatin (5 渭 mol / L) group (6 Gy) and roplatin (5 渭 mol / L) irradiation group (6 Gy). Flow cytometry was used to detect roplatin. Effects of radiation and Loxplatin combined radiation on apoptosis rate of SiHa cells .4.The effects of Loxplatin on the expression of apoptosis-promoting protein Bax and anti-apoptotic protein Bcl-2 in SiHa cells were detected by Western blotting. Results 1. The results showed that different concentrations of Lopplatin were treated with different concentrations of Lopplatin in SiHa cells. The IC50 value of SiHa cells at 24 h, 48 h and 72 h. Click on the multi-target model to fit the experimental data of monoclonal formation. The D0 values of blank group and loplatin treatment group were: 3.588 Gy / 2.226 Gy; DQ value of blank group and roplatin group were 0. 0.792 Gy / 0. 053 Gy; Lopplatin was in SiHa cells. The SER value in flow cytometry was 1.612.3. The apoptotic rates of Loxoplatin group, radiation group and Loxplatin combined radiation group were 30. 50 卤3. 17 卤43. 08 卤5.93% and 84. 68 卤4. 90% respectively. Western blotting analysis showed that the expression of apoptotic protein Bax was significantly up-regulated in Lopplatin group, and in the radiation group compared with the control group. The expression of Bax and Bcl-2 were further up-regulated and down-regulated in the radiation group compared with the roplatin group. Conclusion: 1. Lopplatin can significantly inhibit the proliferation of cervical cancer SiHa cells. The proliferation rate of SiHa cells decreased gradually with the increase of Lopplatin concentration and the prolonged time of treatment. Lopplatin could significantly enhance the radiosensitivity of cervical cancer SiHa cells. The survival fraction of SiHa cells treated with Lopplatin was significantly lower than that of SiHa cells without Lopplatin treatment. 3. Loxoplatin and radiation irradiation could induce apoptosis of cervical cancer SiHa cells, respectively. In addition, Loxoplatin combined with radiation can significantly promote apoptosis of SiHa cells. 4. loplatin and radiation irradiation on SiHa cells of cervical cancer. Lopplatin and radiation irradiation increased the expression of apoptosis-promoting protein Bax and the expression of pro-apoptotic protein Bax. And it was significantly higher than that in SiHa cells of cervical cancer treated by Loxplatin alone or radiation irradiation. Lopplatin and radiation could decrease the expression of Bcl-2, and Loxplatin combined with radiation could decrease the expression of Bcl-2. In addition, proapoptotic protein (Bax) expression and inhibition of Bcl-2 expression in cervical cancer SiHa cells were inhibited by low dose of Loxoplatin alone or radiation irradiation, so that the radiosensitization of cervical cancer SiHa cells could be achieved by increasing the expression of apoptosis-promoting protein Bax and inhibiting the expression of apoptosis-inhibiting protein Bcl-2.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R96

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