摘要
目的观察白花丹素对子宫肌瘤细胞自噬和细胞凋亡的影响,并探讨其调控机制。方法子宫肌瘤细胞分为4组:空白对照组(常规培养)、PB组(5μmol·L^(-1)白花丹素处理)、PB+miR-NC mimic组(5μmol·L^(-1)白花丹素处理+miR-NC mimic处理)、PB+miR-21 mimic(5μmol·L^(-1)白花丹素处理+miR-21 mimic处理);四甲基偶氮唑蓝(MTT)实验检测细胞活力变化;蛋白质印迹法(Western blot)检测LC3-磷脂酰乙醇胺缀合物(LC3Ⅱ)、自噬体吞噬蛋白Beclin-1、p62以及裂解的半胱氨酸天冬氨酸蛋白酶-3(Cleaved-Caspase-3)和B淋巴细胞瘤-2相关X蛋白(Bax)蛋白水平的变化情况;流式细胞术检测子宫肌瘤细胞凋亡率的变化情况;实时荧光定量聚合酶链反应法检测微小RNA-21(miR-21)的表达情况。结果空白对照组、PB组、PB+miR-NC mimic组、PB+miR-21 mimic组的miR-21表达水平分别为1.00±0.14、0.43±0.06、0.39±0.05、0.68±0.09;细胞活力分别为(100.00±8.76)%、(49.67±5.98)%,(50.39±7.06)%、(72.39±9.01)%;LC3Ⅱ/LC3Ⅰ蛋白表达水平分别为0.33±0.05、0.88±0.12、0.83±0.09、0.51±0.08;Beclin-1蛋白表达水平分别为0.25±0.04、0.75±0.09、0.71±0.10、0.42±0.06;p62蛋白水平分别为0.91±0.14、0.46±0.06、0.41±0.08、0.79±0.09;凋亡率分别为(6.21±0.69)%、(20.71±3.11)%、(23.06±3.04)%、(10.42±2.11)%;Cleaved-caspase-3蛋白表达水平分别为0.31±0.04、0.76±0.09、0.80±0.12、0.52±0.07;Bax蛋白表达水平分别为0.35±0.05、0.83±0.13、0.79±0.09、0.42±0.07;空白对照组的上述指标与PB组比较,PB+miR-NC mimic组的上述指标与PB+miR-21 mimic组比较,差异均有统计学意义(均P<0.05)。结论白花丹素可通过下调miR-21的表达促进子宫肌瘤细胞自噬并诱导凋亡。
Objective To observe the effects of plumbagin on autophagy and apoptosis of uterine fibroid cells,and to explore its regulatory mechanism.Methods Uterine fibroid cells were divided into four groups:Control group(conventional culture),PB group(5μmol·L^(-1)plumbagin treatment),PB+miR-NC mimic group(5μmol·L^(-1)plumbagin treatment+transfected with miR-NC mimic),PB+miR-21 mimic group(5μmol·L^(-1)plumbagin treatment+transfected with miR-21 mimic);methyl thiazolyl tetrazolium(MTT)was used to detect cell viability;the protein levels of LC3Ⅱ/LC3Ⅰ,Beclin-1,p62,cleaved-caspase-3 and B-cell lymphoma 2-associated X protein(Bax)were detected by Western blot;the apoptosis rate of uterine leiomyoma cells was detected by flow cytometry;the expression of microRNA-21(miR-21)was detected by real-time fluorescence quantitative polymerase chain reaction.Results The expression levels of miR-21 in control group,PB group,PB+miR-NC mimic group and PB+miR-21 mimic group were 1.00±0.14,0.43±0.06,0.39±0.05,0.68±0.09,respectively;the cell viability were(100.00±8.76)%,(49.67±5.98)%,(50.39±7.06)%,(72.39±9.01)%,respectively;the protein expression levels of LC3Ⅱ/LC3Ⅰwere 0.33±0.05,0.88±0.12,0.83±0.09,0.51±0.08,respectively;the expression levels of Beclin-1 protein were 0.25±0.04,0.75±0.09,0.71±0.10,0.42±0.06,respectively;the protein levels of P62 were 0.91±0.14,0.46±0.06,0.41±0.08,0.79±0.09,respectively;the apoptosis rates were(6.21±0.69)%,(20.71±3.11)%,(23.06±3.04)%,(10.42±2.11)%;cleaved-caspase-3 protein expression levels were 0.31±0.04,0.76±0.09,0.80±0.12,0.52±0.07,respectively;the expression levels of Bax protein were 0.35±0.05,0.83±0.13,0.79±0.09,0.42±0.07;the above indexes showed statistically significant differences between control group and PB group,PB+miR-NC mimic group and PB+miR-21 mimic group(P<0.05).Conclusion Plumbagin can promote autophagy and induce apoptosis of uterine leiomyoma cells by down regulating the expression of miR-21.
作者
梁秀兰
郑玉丹
LIANG Xiu-lan;ZHENG Yu-dan(Departmentof Gynecology,Xingtai People's Hospital,Xingtai 054000,Hebei Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2023年第6期814-818,共5页
The Chinese Journal of Clinical Pharmacology
