阿司匹林对来那度胺抗骨髓瘤效应的增敏作用及分子机制研究

发布时间：2018-05-29 13:19

本文选题：来那度胺 + 阿司匹林　；参考：《南昌大学》2015年硕士论文

【摘要】：目的:1、观察来那度胺(lenalidomide,LEN)和阿司匹林(aspirin,ASA)联合对骨髓瘤细胞株增殖、凋亡与细胞周期的影响,分析两者在骨髓瘤细胞株中的相互作用。2、检测LEN和ASA联合对骨髓瘤细胞VEGF蛋白及wnt/β-catenin信号通路蛋白表达的影响,探讨ASA对LEN抗骨髓瘤的增敏机制。方法:1、常规培养骨髓瘤细胞株(MM1.S及RPMI-8226),制作细胞生长曲线,选取对数生长期细胞进行后续实验。2、采用CCK-8法检测LEN和ASA联合用药对MM1.S细胞和RPMI-8226细胞增殖的影响。3、采用Annexin V-FITC/PI染色方法检测LEN和ASA联合用药对MM1.S细胞和RPMI-8226细胞凋亡的影响。4、采用流式细胞仪检测LEN和ASA联合用药对MM1.S细胞和RPMI-8226细胞周期的影响。5、Western Blot法检测LEN和ASA联合用药对MM1.S细胞和RPMI-8226细胞VEGF蛋白、β-catenin及下游分子cyclinD1蛋白表达的影响。结果:1、药物作用24~72 h后,LEN与ASA联合以时间依赖性抑制细胞增殖活性,且显著高于LEN(1μM)及ASA(2.5 mM)单药组。相互作用分析显示:LEN与ASA在抗MM细胞增殖中呈协同作用。2、药物作用48 h后,LEN与ASA联合组对MM1.S及RPMI-8226细胞凋亡诱导率显著高于LEN或ASA单药组。3、药物作用48 h后,与LEN或ASA单药组相比,LEN与ASA联合组诱导MM1.S及RPMI-8226细胞阻滞于G1期比率明显增高。4、药物作用48 h后,相比LEN或ASA单药组,两者联合后显著下调了MM1.S及RPMI-8226细胞VEGF蛋白表达。5、在MM1.S与RPMI-8226细胞中,LEN作用72 h后诱导胞核内β-catenin蛋白及其下游cyclin D1蛋白表达上调,而ASA促使胞浆β-catenin磷酸化,导致胞核内β-catenin蛋白其下游分子cyclin D1蛋白表达下降。结论:1、阿司匹林与来那度胺在抑制骨髓瘤细胞增殖中呈协同作用;2、阿司匹林通过下调骨髓瘤细胞株VEGF表达对来那度胺发挥协同抗骨髓瘤作用;3、阿司匹林通过促进细胞浆β-catenin磷酸化,从而阻止β-catenin入细胞核发挥对来那度胺的化疗增敏作用。
[Abstract]:Objective: to observe the effects of the combination of renalidomide en and aspirin on the proliferation, apoptosis and cell cycle of myeloma cell line. To investigate the effect of LEN and ASA on the expression of VEGF protein and wnt/ 尾 -catenin signal pathway protein in myeloma cell line, and to explore the mechanism of ASA on anti-myeloma. Methods: cell growth curves were prepared by routine culture of myeloma cell lines MM1.S and RPMI-8226. Logarithmic growth phase cells were selected for follow-up experiment. CCK-8 assay was used to detect the effect of combined use of LEN and ASA on the proliferation of MM1.S cells and RPMI-8226 cells. Annexin V-FITC/PI staining was used to detect the apoptosis of MM1.S cells and RPMI-8226 cells treated with LEN and ASA. The effect of combined use of LEN and ASA on the cell cycle of MM1.S and RPMI-8226 was detected by flow cytometry. The effects of LEN and ASA on the expression of VEGF protein, 尾 -catenin and cyclinD1 in MM1.S and RPMI-8226 cells were detected by Western Blot assay. Results the cell proliferation activity was inhibited in a time-dependent manner by the combination of Len and ASA for 24 ~ 72 h, and was significantly higher than that in LEN(1 渭 M and ASA(2.5 mm) groups. The results of interaction analysis showed that ASA and WLEN had synergistic effects on the proliferation of MM cells. After 48 h of drug treatment, the apoptotic induction rate of MM1.S and RPMI-8226 cells in the combination group was significantly higher than that in the LEN or ASA group, and 48 h after the drug treatment, the apoptosis induction rate of MM1.S and RPMI-8226 cells in the combination group was significantly higher than that in the single drug LEN or ASA group. Compared with LEN or ASA monotherapy group, the ratio of MM1.S and RPMI-8226 cell arrest in G1 phase was significantly increased in ASA combined with ASA group. After 48 h of treatment, compared with LEN or ASA group, the ratio of MM1.S and RPMI-8226 cell arrest was significantly higher than that of LEN or ASA group. In combination, the expression of VEGF protein in MM1.S and RPMI-8226 cells was significantly down-regulated. The expression of 尾 -catenin protein and its downstream cyclin D1 protein were up-regulated in the nucleus of MM1.S and RPMI-8226 cells after 72 h of treatment with en, and the phosphorylation of 尾 -catenin in the cytoplasm was induced by ASA. The expression of 尾 -catenin protein in the nucleus decreased in the downstream of cyclin D1 protein. Conclusion: 1, aspirin and renalidomide have synergistic effect on inhibiting the proliferation of myeloma cells. Aspirin plays a synergistic role in inhibiting myeloma by down-regulating the expression of VEGF in myeloma cell line. Aspirin can promote the proliferation of myeloma cells by promoting the proliferation of myeloma cells. Cytoplasmic 尾 -catenin phosphorylation, In order to prevent 尾-catenin into the nucleus to play the role of chemosensitization of leinadamide.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R733.3

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