摘要
目的研究紫铆因对骨肉瘤细胞凋亡及自噬的影响,并探讨自噬在紫铆因抗骨肉瘤中的作用。方法分别以不同浓度紫铆因作用骨肉瘤细胞24h和30μmol/L紫铆因作用骨肉瘤细胞不同时间,使用CCK-8试剂盒测定细胞活性变化;Western blot法检测凋亡相关蛋白Bax、Bcl-2、Cleaved caspase3及自噬相关蛋白p62、LC3、Beclin-1的表达情况;TUNEL染色检测骨肉瘤细胞的凋亡情况;透射电子显微镜检测自噬泡的产生;最后分别用自噬激动剂和自噬抑制剂预处理后再加入紫铆因作用于骨肉瘤细胞,并重复上述实验。结果紫铆因作用后,骨肉瘤细胞的活性明显下降,Bax、Cleaved caspase3、Beclin-1、LC3 Ⅱ的表达增加,而Bcl-2、p62的表达降低,骨肉瘤细胞出现明显的凋亡现象,透射电子显微镜观察到紫铆因处理组有较多自噬泡产生;使用自噬抑制剂预处理骨肉瘤细胞后,紫铆因对骨肉瘤的凋亡效果被逆转,而使用自噬激动剂预处理骨肉瘤细胞后,紫铆因使骨肉瘤细胞凋亡的效果进一步增强。结论紫铆因可以降低骨肉瘤细胞活性并促进其凋亡;同时紫铆因可以激活自噬,促进骨肉瘤细胞凋亡。
Objective To investigate the effect of butein on apoptosis and autophagy of osteosarcoma MG63 cells and its mechanism.Methods Human osteosarcoma MG63 cells were treated with butein at different concentrations for various times.Cell viability was measured by CCK-8 assay,the expression of apoptosis-related proteins Bax,BcI-2,Cleaved caspase3 and autophagy-related proteins p62,LC3 and Beclin-1 were detected by Western blot,the apoptosis of MG63 cells waere detected by TUNEL staining,the production of autophagic bubbles was observed by transmission electron microscopy.The osteosarcoma MG63 cells were pretreated with autophagy agonists and autophagy inhibitors before butein was added for treatment,the above exprements were repeated to explore the role of autophagy in anti-osteosarcoma of butein.Results After treatment with butein,the viability of MG63 cells were decreased significantly.Western blot results showed that the expression of Bax,Cleaved caspase3,Beclin-1 and LC3 Ⅱ were increased,while the expression of Bcl-2 and p62 were decreased.The results of TUNEL staining showed that there was significant apoptosis in osteosarcoma cells and many autophagic bubbles were observed with transmission electron microscopy in the butein treatment group.Pretreatment of osteosarcoma cells with autophagy inhibitor,the effect of butein on the apoptosis of osteosarcoma was reversed,while pretreatment of osteosarcoma cells with autophagy agonists,the effect of butein on the apoptosis of osteosarcoma was further enhanced.Conclusion Butein can inhibit the viability and induce apoptosis of osteosarcoma MG63 cells,which is associated with the activation of autophagy.
出处
《浙江医学》
CAS
2017年第13期1072-1076,1096,I0002,共7页
Zhejiang Medical Journal
基金
温州市科技计划项目(Y20150066)
关键词
紫铆因
骨肉瘤
凋亡
自噬
Butein
Osteosarcoma
Autophagy
Apoptosis
