TO901317促进MCF-7与MDA-MB-231人乳腺癌细胞凋亡的研究

发布时间：2018-03-08 10:37

本文选题：TO901317　切入点：乳腺癌细胞　出处：《南华大学》2015年硕士论文　论文类型：学位论文

【摘要】：研究背景乳腺癌是一种常见的恶性肿瘤,近几年有研究指出孤儿受体家族中的肝X受体α(LXRα)与乳腺癌的发生有着密切的关系,其中肝X受体激动剂TO901317尤其对LXRα具有高亲和力,可抑制多种肿瘤细胞包括乳腺癌的增殖。最近有文献进一步指出TO901317还能诱导前列腺癌和黑色素瘤细胞凋亡。目的研究肝X受体激动剂TO901317对人乳腺癌细胞凋亡的影响以及对细胞内LXRα、TTP和NF-κB p65等表达的作用,为乳腺癌的分子靶向治疗提供新的思路。方法1.使用不同浓度(0μM,10μM,20μM,40μM)TO901317处理MCF-7和MDA-MB-231人乳腺癌细胞不同时间(0h,12h,24h,48h),用MTT法观察细胞的活性。2.用流式细胞术以及Hoechst染色检测不同浓度(0μM,10μM,20μM,40μM)TO901317处理MCF-7和MDA-MB-231细胞不同时间(0h,12h,24h,48h)后细胞凋亡的表现。3.RT-q PCR检测TO901317处理MCF-7和MDA-MB-231细胞的量效组和时效组中LXRα和Bcl-2 m RNA的改变;Western blot检测LXRα和细胞凋亡相关蛋白Bax、Bcl-2、Caspase 3和Cleaved-caspase3等表达的差异。4.通过生物信息学软件Interactome 3D和STRING,在线预测了LXRα、TTP和NF-κB p65三者之间的关系。5.RT-PCR检测TO901317处理MCF-7细胞的量效组和时效组中TTP和NF-κB p65 m RNA表达的改变。结果随着TO901317处理浓度的增加和处理时间的延长,对细胞的活性抑制作用逐渐增强,凋亡现象逐渐明显。当细胞处理24h,浓度在20μM时,对细胞的活性抑制约为50%,且伴随明显的细胞凋亡形态,因此,TO901317量效组采纳24h作为处理时间,时效组采用20μM作为细胞处理浓度。与此同时,RT-q PCR检测发现,TO901317可上调LXRαm RNA水平,下调Bcl-2 m RNA表达。Western blot检测除有上述一致的趋势外,还进一步发现TO901317可促使凋亡蛋白Bax、Cleaved-caspase 3表达增多,进一步证实TO901317促进两种细胞凋亡。通过生物信息学在线预测结果显示,LXRα、TTP和NF-κB p65三者之间可能存有直接或间接的相互作用。RT-PCR检测发现TO901317可上调TTP m RNA表达,并下调NF-κB p65 m RNA水平。结论1.TO901317能够促使MCF-7和MDA-MB-231乳腺癌细胞凋亡;2.TO901317促使乳腺癌细胞凋亡的机制与细胞中LXRα/TTP/NF-κB p65通路相关。
[Abstract]:Background Breast cancer is a common malignant tumor. In recent years, it has been pointed out that liver X receptor 伪 (LXR 伪) in the orphan receptor family is closely related to the occurrence of breast cancer, in which liver X receptor agonist TO901317 has high affinity for LXR 伪. TO901317 can also induce apoptosis of prostate cancer and melanoma cells. Objective to study the effect of liver X receptor agonist TO901317 on apoptosis of human breast cancer cells. And the expression of LXR 伪 -TTP and NF- 魏 B p65 in cells. Methods 1. MCF-7 and MDA-MB-231 human breast cancer cells were treated with different concentrations of 0 渭 MU 10 渭 MU 10 渭 MU and 40 渭 M TO901317 at different times. The activity of cells was observed by MTT method. Flow cytometry and Hoechst staining were used to detect the cell activity. 2. Changes of LXR 伪 and Bcl-2 m RNA in MCF-7 and MDA-MB-231 cells treated with 10 渭 M ~ 10 渭 M ~ (10 渭 M) and 40 渭 M ~ (10 渭 m) of 0 渭 m ~ (10 渭 m) at different time after treatment with 10 渭 M ~ (10 渭 M) and 40 渭 M ~ (10 渭 M) TO901317). (3. RT-q PCR detection of LXR 伪 and Bcl-2 m RNA changes in MCF-7 and MDA-MB-231 cells treated with TO901317 at different time.) LXR 伪 and apoptosis-related proteins Baxcl-2 (Caspase 3 and Cleaved-caspase3) were detected by Western blot in the aging group and in the dose-effecting group of MCF-7 and MDA-MB-231 cells treated with TO901317. The relationship between LXR 伪 TTP and NF- 魏 B p65 was predicted online by using bioinformatics software Interactome 3D and STRING.5.The changes of TTP and NF- 魏 B p65m RNA expression in MCF-7 cells treated with TO901317 were detected by RT-PCR. The increase of TO901317 concentration and the prolongation of treatment time, When the cells were treated for 24 h and the concentration of 20 渭 M was 20 渭 M, the inhibition of cell activity was about 50 and accompanied by obvious apoptotic morphology, so TO901317 dose-effect group was treated with 24 hours as the treatment time. At the same time, TO901317 could up-regulate the level of LXR 伪 m RNA and down-regulate the expression of Bcl-2 m RNA. Western blot showed that TO901317 could increase the expression of apoptosis protein Baxon Cleaved-caspase 3. The results of online bioinformatics prediction showed that there might be direct or indirect interaction between LXR 伪 TTP and NF- 魏 B p65. RT-PCR analysis showed that TO901317 could up-regulate the expression of TTP m RNA. Conclusion TO901317 can induce apoptosis of MCF-7 and MDA-MB-231 breast cancer cells. 2. The mechanism of TO901317 inducing apoptosis of breast cancer cells is related to the LXR 伪 / TTP / NF- 魏 B p65 pathway.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R737.9

【相似文献】