MAP3K3介导NF-κB信号通路在卵巢癌化疗抵抗中作用的初步研究

发布时间：2018-05-08 22:42

本文选题：卵巢癌 + MAP3K3　；参考：《石河子大学》2017年硕士论文

【摘要】：目的:干扰MAP3K3表达和/或阻断NF-κB信号通路,检测MAP3K3在卵巢癌细胞中的表达,结合NF-κB信号通路相关因子及凋亡相关蛋白的表达变化,分析和探讨MAP3K3介导NF-κB信号通路在卵巢癌化疗抵抗中的作用。方法:分别用特异性干扰MAP3K3表达质粒和MAP3K3真核表达质粒转染内源性高表达和/或低表达MAP3K3的卵巢癌细胞系SKOV3和OV2008,采用Western Blot和qRT-PCR检测转染效率;运用MTT方法检测干扰前后卵巢癌细胞对顺铂、紫杉醇和TNF-α敏感性的变化,TUNEL法检测细胞凋亡;通过使用NF-κB信号通路特异性阻断剂QNZ,观察顺铂处理后NF-κB信号通路关键因子和凋亡相关因子表达的变化情况。结果:(1)MAP3K3蛋白和mRNA在SKOV3细胞中内源性高表达,在OV2008细胞中低表达,OV2008细胞对顺铂的敏感性显著高于SKOV3细胞;(2)在SKOV3中敲低MAP3K3表达后,顺铂诱导细胞凋亡的数目显著增加,细胞对顺铂敏感性增强,生长速度减慢,同时NF-κB信号通路相关因子p-P65、p-IκBα和抗凋亡蛋白Bcl-2表达下调,促凋亡蛋白cleaved-caspase3、BAX表达上调;(3)在OV2008中过表达MAP3K3后,顺铂诱导细胞凋亡的数目显著减少,细胞对顺铂的敏感性减弱,生长加快,同时NF-κB信号通路关键因子p-P65、p-IκBα和抗凋亡蛋白Bcl-2表达上调,促凋亡蛋白cleaved-caspase3、BAX表达下调;(4)在SKOV3中抑制MAP3K3表达后细胞对紫杉醇和TNF-α的敏感性增强;在OV2008中过表达MAP3K3后细胞对紫杉醇和TNF-α的敏感性减弱;阻断NF-κB信号通路,两株细胞对紫杉醇和TNF-α的敏感性增强。结论:MAP3K3在卵巢癌细胞系中表达水平的差异可能与卵巢癌细胞的化疗抵抗相关。MAP3K3高表达促进了NF-κB信号通路激活以及下游抗凋亡蛋白的表达,抑制促凋亡蛋白表达,降低了化疗药物对卵巢癌细胞的诱导凋亡能力,提示MAP3K3可能是通过介导NF-κB信号通路促进卵巢癌细胞发生化疗抵抗。
[Abstract]:Aim: to interfere with the expression of MAP3K3 and / or block NF- 魏 B signaling pathway, detect the expression of MAP3K3 in ovarian cancer cells, and combine the changes of NF- 魏 B signal pathway related factors and apoptosis-related proteins. To investigate the role of NF- 魏 B signaling pathway mediated by MAP3K3 in chemotherapeutic resistance of ovarian cancer. Methods: ovarian cancer cell lines SKOV3 and OV2008 were transfected with specific interfering MAP3K3 expression plasmids and MAP3K3 eukaryotic expression plasmids. The transfection efficiency was detected by Western Blot and qRT-PCR. The sensitivity of ovarian cancer cells to cisplatin, paclitaxel and TNF- 伪 was detected by MTT method. Apoptosis was detected by Tunel method. The expression of key factors and apoptosis-related factors in NF- 魏 B signaling pathway after cisplatin treatment was observed by using QNZ, a specific inhibitor of NF- 魏 B signaling pathway. Results the endogenous expression of MAP3K3 protein and mRNA in SKOV3 cells was significantly higher than that in OV2008 cells. The sensitivity to cisplatin in OV2008 cells was significantly higher than that in SKOV3 cells (P < 0.05). The number of apoptosis induced by cisplatin increased significantly when MAP3K3 expression was low in SKOV3. At the same time, the expression of NF- 魏 B signaling pathway related factor p-P65, p-I 魏 B 伪 and anti-apoptotic protein Bcl-2 was down-regulated, and the expression of cleaved-caspase3mBAX was up-regulated in OV2008. The number of apoptosis induced by cisplatin was significantly reduced, the sensitivity of cells to cisplatin was weakened, and the growth rate was accelerated. Meanwhile, the expression of p-P65 p-I 魏 B 伪 and anti-apoptotic protein Bcl-2 was up-regulated in NF- 魏 B signaling pathway. After inhibiting the expression of MAP3K3 in SKOV3, the sensitivity of cells to paclitaxel and TNF- 伪 was enhanced, the sensitivity of cells to paclitaxel and TNF- 伪 was weakened after overexpression of MAP3K3 in OV2008, and the signal pathway of NF- 魏 B was blocked. The sensitivity of the two cells to paclitaxel and TNF- 伪 was enhanced. Conclusion the difference of the expression level of protein MAP3K3 in ovarian cancer cell line may be related to the chemotherapeutic resistance of ovarian cancer cell line. The overexpression of MAP3K3 promotes the activation of NF- 魏 B signaling pathway and the expression of downstream anti-apoptotic protein, and inhibits the expression of pro-apoptotic protein. The ability of apoptosis induced by chemotherapeutic drugs was reduced, suggesting that MAP3K3 may promote chemotherapeutic resistance by mediating NF- 魏 B signaling pathway.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.31

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