蛇床子素通过过氧化物酶体增殖物活化受体γ抑制乳腺癌MCF-7细胞增殖的研究

Osthole inhibits the proliferation of breast cancer MCF-7 cells via activating peroxisome proliferator-activated receptor γ

目的 研究蛇床子素对乳腺癌细胞MCF-7增殖和凋亡的影响并探讨其可能的作用机制.方法 分别以蛇床子素0、25、50、100、150、200 μmol/L干预MCF-7细胞;MTT法检测蛇床子素对细胞增殖的抑制程度;HE染色,在光学显微镜下观察蛇床子素干预后细胞的形态学改变;Annexin V-PI双染流式细胞术分析蛇床子素处理对细胞凋亡的影响,分别用反转录聚合酶链反应（RT-PCR）和Western blot法检测过氧化物酶体增殖物活化受体γ（PPARγ）和法尼醇X受体（FXR） mRNA和蛋白的表达.结果 蛇床子素干预72 h后对MCF-7细胞的增殖产生明显抑制,且有一定的剂量依赖趋势,蛇床子素200 μmol/L组对MCF-7细胞增殖的抑制率最高,为73.0％.光学显微镜下观察到蛇床子素干预72 h后MCF-7细胞数目较少,细胞核浓染及凋亡小体出现,并且呈现明显的剂量依赖关系.流式细胞术分析发现蛇床子素干预72 h后,与对照组相比,蛇床子素浓度达到50 μmol/L以上时,MCF-7早期凋亡细胞比例增加（P＜0.01）,尤其是蛇床子素200 μmol/L组达到（46.2±9.0）％;当蛇床子素浓度达到100 μmol/L以上时,中晚期凋亡率较对照组也升高（P＜0.05,P＜ 0.01）,尤其是蛇床子素200 μmol/L组达到（39.2±5.7）％,和MTT实验结果相吻合.另外,RT-PCR和Western Blot结果显示蛇床子素可上调PPARγ和FXR mRNA和蛋白的表达（P＜0.01）.结论 蛇床子素可抑制乳腺癌MCF-7细胞的增殖并可促进其凋亡,这些作用可能是通过调节与PPARγ和FXR介导的与细胞生长和代谢有关的基因来实现的.

Objective To investigate the effect of osthole on the proliferation and apoptosis of breast cancer cell line MCF-7 and its potential mechanisms.Methods Breast cancer cell line MCF-7 was treated by osthole 0,25,50,100,150 and 200 μmol/L respectively.MTT method was used to detect cell survival rate.HE staining was used to observe morphological changes,Annexin V-PI flow cytometry was used to analyze cell apoptosis,and RT-PCR and Western blot method were used to detect the mRNA and protein expression of peroxisome proliferator-activated receptor γ （PPARγ） and farnesoid X receptor （FXR）,respectively.Results MTT assay showed that strong cytotoxicity of cell line MCF-7 was induced after administration of osthole for 72 h in a dose-dependent manner.Especially,the maximum inhibitory rate,73.0 % appeared in the 200 μmol/L group.HE staining showed that the number of MCF-7 cells decreased,hyperchromatic nuclei and apoptotic bodies appeared after treatment with osthole for 72 h in a significant dose-effect manner.Flow cytometric analysis revealed that osthole could induce extensive apoptosis in MCF-7 cultures after treatment for 72 h compared with normal group （P 〈 0.05,P 〈 0.01）.In particular,when the concentration of osthole reached 50 μmol/L,the proportion of early apoptotic cells was significantly increased in a dose-dependent manner （P 〈 0.01）,especially.The maximum apoptosis rate （46.2±9.0） % appeared in the 200 μmol/L group,which was consistent with the results obtained from MTT assays.Moreover,osthole could significantly increased PPARγand FXR mRNA and protein expressions （P 〈 0.01）.Conclusion These data suggest that osthole could inhibit the proliferation of breast cancer MCF-7 cells and promote its apoptosis,which might be associated with the regulation of PPARγ and FXR-mediated target genes involved in cell growth and metabolism.