杨梅素通过促进DRP1依赖性线粒体分裂诱导人卵巢癌SKOV3细胞凋亡

doi:10.13481/j.1671-587x.20180503

摘要/Abstract

摘要： 目的：观察杨梅素与mdivi-1分别或联合作用于人卵巢癌SKOV3细胞后细胞凋亡和线粒体分裂情况，探讨杨梅素诱导SKOV3细胞凋亡的机制。方法：体外培养SKOV3细胞，随机分为对照组、mdivi-1组、杨梅素组和联合给药组，mdivi-1组以50 μmol·L^-1 mdivi-1作用1 h后更换普通细胞培养液继续培养23 h，杨梅素组以50 g·L^-1杨梅素作用24 h，联合给药组以50 μmol·L^-1 mdivi-1作用1 h后更换含50 g·L^-1杨梅素的细胞培养液继续培养23 h。MTT法检测各组细胞存活率；Muse^®凋亡检测试剂盒检测各组细胞凋亡率；蛋白免疫印迹法（Western blotting）观察细胞色素C（Cyt C）、半胱氨酸天冬氨酸蛋白酶3（caspase3）、动力相关蛋白1（DRP1）和分裂蛋白1（FIS1）表达水平；MitoTracker^® Red荧光探针特异性标记线粒体观察各组细胞线粒体分裂情况。结果：与对照组比较，杨梅素组细胞存活率明显降低（P<0.05）；与杨梅素组比较，联合给药组细胞存活率明显升高（P<0.05）。与对照组比较，杨梅素组细胞凋亡率升高（P<0.05）；与杨梅素组比较，联合用药组细胞凋亡率明显降低（P<0.05）。与对照组比较，杨梅素组细胞中Cyt C和caspase3蛋白表达水平明显升高（P<0.05）；与杨梅素组比较，联合用药组Cyt C和caspase3蛋白表达水平明显降低（P<0.05）。与对照组比较，杨梅素组细胞线粒体分裂程度增强；与杨梅素组比较，联合用药组线粒体分裂程度下降。与对照组比较，杨梅素组细胞中DRP1和FIS1蛋白表达水平升高（P<0.05）；与杨梅素组比较，联合用药组DRP1和FIS1蛋白表达水平降低（P<0.05）。结论：杨梅素可通过促进DRP1依赖性线粒体分裂诱导人卵巢癌SKOV3细胞凋亡。

关键词: 杨梅素, mdivi-1, 动力相关蛋白1, 线粒体分裂, 细胞凋亡, SKOV3细胞

Abstract: Objective:To observe the apoptosis and mitochondrial fission of human ovarian cancer SKOV3 cells after treated by myricetin and dynamin related protein 1 (DRP1) inhibitor mdivi-1 alone or combined, and to explore the mechanism of myricetin in inducing the apoptosis of SKOV3 cells. Methods:The SKOV3 cells were cultured in vitro and randomly divided into control group, mdivi-1 group, myricetin group and combined group.The cells in mdivi-1 group were treated with 50 μmol·L^-1 madivi-1 for 1 h followed by common culture medium for 23 h;the cells in myricetin group were treated with 50 g·L^-1 myricetin for 24 h;the cells combined group were treated with 50 μmol·L^-1 midiv-1 for 1 h followed by 50 g·L^-1 myricetin for 23 h. The survival rates of cells in various groups were detected by MTT assay. The apoptoic rates of cells in various groups were detected by Muse^® apoptosis detection kit. The expression levels of Cyt C, caspase3, DRP1 and FIS1 were observed by Western blotting method. The mitochondrial fission of cells in various groups was observed with MitoTracker^® Red. Results:Compared with control group, the survival rate of cells in myricetin group was decreased significantly (P<0.05); compared with myricetin group, the survival rate of cells in combined group was increased significantly(P<0.05). Compared with control group, the apoptotic rate of cells in myricetin group was increased (P<0.05); compared with myricetin group, the apoptotic rate of cells in combined group was decreased (P<0.05). Compared with control group, the expression levels of Cyt C and caspase3 proteins in the cells in myricetin group were increased(P<0.05); compared with myricetin group, the expression levels of Cyto C and caspase3 proteins in the cells in combined group were decreased(P<0.05). Compared with control group,the degree of mitochondrial fission of the cells in myricetin group was increased;compared myricetin group,the degree of mitochondrial fission of the cells in combined group was decreased.Compared with control group, the expression levels of DRP1 and FIS1 proteins in the cells in myricetin group were increased (P<0.05); compared with myricetin group, the expression levels of DRP1 and FIS1 proteins in the cells in combined group were decreased (P<0.05). Conclusion:Myricetin can induce the apoptosis of human ovarian cancer SKOV3 cells by promoting the DRP1-dependent mitochondrial fission.

Key words: myricetin, mdivi-1, dynamin related protein 1, mitochondrial fission, apoptosis, SKOV3 cells

中图分类号:

R73-3

于洋, 徐路, 刘师兵, 李松岩, 徐冶. 杨梅素通过促进DRP1依赖性线粒体分裂诱导人卵巢癌SKOV3细胞凋亡[J]. 吉林大学学报(医学版), 2018, 44(05): 903-907.

YU Yang, XU Lu, LIU Shibing, LI Songyan, XU Ye. Induction effect of myricetin on apoptosis of human ovarian cancer SKOV3 cells by promoting DRP1-dependent mitochondrial fission[J]. Journal of Jilin University(Medicine Edition), 2018, 44(05): 903-907.

参考文献

[1] Xiao S, Mu ZQ, Cheng CR, et al. Three new biflavonoids from the branches and leaves of Cephalotaxus oliveri and their antioxidant activity[J]. Nat Prod Res, 2018,15:1-7.
[2] Tian Y, Puganen A, Alakomi HL, et al. Antioxidative and antibacterial activities of aqueous ethanol extracts of berries, leaves, and branches of berry plants[J]. Food Res Int, 2018,106:291-303.
[3] Sharma K, Mahato N, Lee YR. Systematic study on active compounds as antibacterial and antibiofilm agent in aging onions[J]. J Food Drug Anal, 2018, 26(2):518-528.
[4] Khalfallah A, Berrehal D, Bensouici C, et al. Flavonoids, cytotoxic, antioxidant and antibacterial activities of Evax pygmaea[J]. Pharm Biol, 2017, 55(1):2292-2296.
[5] Hariri BM, McMahon DB, Chen B, et al. Plant flavones enhance antimicrobial activity of respiratory epithelial cell secretions against Pseudomonas aeruginosa[J]. PLoS One, 2017, 12(9):e0185203.
[6] Taverniti V, Dalla Via A, Minuzzo M, et al. In vitro assessment of the ability of probiotics, blueberry and food carbohydrates to prevent S. pyogenes adhesion on pharyngeal epithelium and modulate immune responses[J]. Food Funct, 2017, 8(10):3601-3609.
[7] Choi S, Youn J, Kim K, et al. Apigenin inhibits UVA-induced cytotoxicity in vitro and prevents signs of skin agingin vivo[J]. Int J Mol Med, 2016, 38(2):627-634.
[8] Wei J, Zhang G, Zhang X, et al. Anthocyanins from black chokeberry (aroniamelanocarpa elliot) delayed aging-related degenerative changes of brain[J]. J Agric Food Chem, 2017, 65(29):5973-5984.
[9] Martínez-Pérez C, Ward C, Turnbull AK, et al. Antitumour activity of the novel flavonoid Oncamex in preclinical breast cancer models[J]. Br J Cancer, 2016, 114(8):905-916.
[10] Autret A, Martin SJ. Bcl-2 family proteins and mitochondrial fission/fusion dynamics[J]. Cell Mol Life Sci, 2010, 67(10):1599-1606.
[11] Meyer JN, Hartman JH, Mello DF. Mitochondrial Toxicity[J]. Toxicol Sci, 2018, 162(1):15-23.
[12] Cabrera-Serrano M, Junckerstorff RC, Atkinson V, et al. Novel CHKB mutation expands the megaconial muscular dystrophy phenotype[J]. Muscle Nerve, 2015, 51(1):140-143.
[13] Zhao C, Chen Z, Qi J, et al. Drp1-dependent mitophagy protects against cisplatin-induced apoptosis of renal tubular epithelial cells by improving mitochondrial function[J]. Oncotarget, 2017, 8(13):20988-21000.
[14] Manchanda R, Legood R, Antoniou AC, et al. Changing the risk threshold for surgical prevention of ovarian cancer[J]. BJOG, 2018, 125(5):541-544.
[15] 于洋, 刘亚萌, 吴少花, 等. 杨梅素对人卵巢癌SKOV3细胞的凋亡诱导作用[J]. 吉林大学学报:医学版, 2015, 41(5):902-906.
[16] Sehrawat A, Croix CS, Baty CJ, et al. Inhibition of mitochondrial fusion is an early and critical event in breast cancer cell apoptosis by dietary chemopreventative benzyl isothiocyanate[J]. Mitochondrion, 2016, 30:67-77.
[17] Chen SD, Zhen YY, Lin JW, et al. Dynamin-related protein 1 promotes mitochondrial fission and contributes to the hippocampal neuronal cell death following experimental status epilepticus[J].CNS Neurosci Ther, 2016, 22(12):988-999.
[18] Huang G, Massoudi D, Muir AM, et al. WBSCR16 is a guanine nucleotide exchange factor important for mitochondrial fusion[J]. Cell Rep, 2017, 20(4):923-934.
[19] Fan LF, He PY, Peng YC, et al. Mdivi-1 ameliorates early brain injury after subarachnoid hemorrhage via the suppression of inflammation-related blood-brain barrier disruption and endoplasmic reticulum stress-based apoptosis[J]. Free Radic Biol Med, 2017,112:336-349.
[20] Hu C, Huang Y, Li L. Drp1-dependent mitochondrial fission plays critical roles in physiological and pathological progresses in mammals[J]. Int J Mol Sci, 2017, 18(1):E144.
[21] Li GB, Fu RQ, Shen HM, et al. Polyphyllin I induces mitophagic and apoptotic cell death in human breast cancer cells by increasing mitochondrial PINK1 levels[J]. Oncotarget, 2017, 8(6):10359-10374.
[22] Han XJ, Yang ZJ, Jiang LP, et al. Mitochondrial dynamics regulates hypoxia-induced migration and antineoplastic activity of cisplatin in breast cancer cells[J]. Int J Oncol, 2015,46(2):691-700.