山杏叶乙酸乙酯提取物调控乳腺癌MCF7细胞增殖和凋亡的机制研究
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摘要
为探究山杏叶乙酸乙酯提取物对乳腺癌MCF7细胞株增殖和凋亡的影响,本研究采用CCK8法检测山杏叶提取物对MCF7细胞增殖的抑制作用,流式细胞仪检测细胞凋亡率,倒置荧光显微镜和流式细胞仪分别检测胞内活性氧(ROS)的水平变化,RT-PCR检测细胞周期及凋亡相关基因的表达情况,试剂盒检测caspase-3的活性。实验表明,山杏叶提取物可降低MCF7细胞存活率,促进细胞凋亡,增加胞内ROS水平。同时上调和Bax,下调Bcl-2,增强caspase-3活性,并降低CDK4、Cyclin E和Cyclin D1的表达。综上说明山杏叶提取物可通过调控周期蛋白的表达来抑制MCF7细胞的增殖,并通过caspase途径和提升ROS水平来诱导MCF7细胞的凋亡。
To explore the effect of ethyl acetate extract of wild apricot leaves on proliferation and apoptosis of breast cancer MCF7 cell,CCK8 assay was used to detect the inhibitory effect of extract of wild apricot leaves on the proliferation of MCF7 cells.Flow cytometry was used to detect the apoptosis rate.Inverted fluorescence microscopy and flow cytometry were used to detect the changes of intracellular reactive oxygen species(ROS).RT-PCR was used to detect the expression of cell cycle and apoptosis-related genes.Detection kit was used to measure caspase-3 activity.The results showed that extract of wild apricot leaves could reduce the survival rate of MCF7 cells,promote cell apoptosis and increase the level of intracellular reactive oxygen species(ROS).Further,it could up-regulate Bax,down-regulate Bcl-2,enhance caspase-3 activity,and reduce the expression of CDK4,Cyclin E and Cyclin D1.In conclusion,ethyl acetate extract of wild apricot leaves could inhibit the proliferation of MCF7 cells by regulating cyclin expressions,and induce apoptosis of MCF7 by caspase pathway and the up-regulating of ROS level.
To explore the effect of ethyl acetate extract of wild apricot leaves on proliferation and apoptosis of breast cancer MCF7 cell,CCK8 assay was used to detect the inhibitory effect of extract of wild apricot leaves on the proliferation of MCF7 cells.Flow cytometry was used to detect the apoptosis rate.Inverted fluorescence microscopy and flow cytometry were used to detect the changes of intracellular reactive oxygen species(ROS).RT-PCR was used to detect the expression of cell cycle and apoptosis-related genes.Detection kit was used to measure caspase-3 activity.The results showed that extract of wild apricot leaves could reduce the survival rate of MCF7 cells,promote cell apoptosis and increase the level of intracellular reactive oxygen species(ROS).Further,it could up-regulate Bax,down-regulate Bcl-2,enhance caspase-3 activity,and reduce the expression of CDK4,Cyclin E and Cyclin D1.In conclusion,ethyl acetate extract of wild apricot leaves could inhibit the proliferation of MCF7 cells by regulating cyclin expressions,and induce apoptosis of MCF7 by caspase pathway and the up-regulating of ROS level.
引文
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2 Zheng Y,Wu CX,Zhang ML.The epidemic and characteristics of female breast cancer in China[J].Chin Oncol(中国癌症杂志),2013,23:561-569.
3 Luo NA,Qu YQ,Dong R.The Treatment progress of the breast cancer[J].Prog Mod Biomed(现代生物医学进展),2015,15:160-166.
4 Wen X.Extraction of apricot kernel protein inhibiting human colon adenocarcinoma cell growth[D].Beijing:Beijing Forestry University(北京林业大学),2012.
5 Ding MM.Apricot kernel protein enzymatic hydrolysate significantly inhibits proliferation of human breast cancer cell[D].Beijing:Beijing Forestry University(北京林业大学),2016.
6 Liu S,Wang XQ,Liao GH,et al.Research progress on antitumor activity and mechanism of amygdalin[J].Shanghai JTradit Chin Med(上海中医药杂志),2017,51:99-101.
7 Peng YF,Li J,Li W,et al.Study on the betulinic acid,oleanolic acid and ursolic acid in leaf of Prunus sibirica L.[J].Hebei J Forestry Orchard Res(河北林果研究),2012,27:157-160.
8 Huang RY,Zhu SS,Tang JH,et al.Effect of selaginella doederleinii ethyl acetate extract on proliferation and migration of Hep-2 cells[J].Nat Prod Res Dev(天然产物研究与开发),2017,29:2024-2029.
9 Harmand PO,Duval R,Liagre B,et al.Ursolic acid induces apoptosis through caspase-3 activation and cell cycle arrest in Ha Cat cells[J].Int J Oncol,2003,23:105-12.
10 Hu CJ,Guo RZ.Association of Ras and its signal transduction pathway with skin tumors[J].J Med Postgrad(医学研究生学报),2012,25:771-773.
11 Lin YJ,Guo RZ,Wang HQ.Expression and signification of cell cycle regulation protein Cyclin D1-CDK4-p21 in scar cancer[J].J Med Postgrad(医学研究生学报),2014,27:923-927.
12 Song FH,Luo M,Fan N,et al.Combination of dihydroartemisinin and gefitinib inhibits cell cycle and migration of lung adenocarcinoma cells in vitro[J].Med J Chin Peop Arm Pol For(武警医学),2019,30:228-232.
13 Kuang SS,Zheng JL,Yang L,et al.Expression and significance of Cyclin E and related genes in Hepatocellular carcinoma[J].Chin Cancer(中国肿瘤),2016,25:143-149.
14 Li J,Gao H,Tian Z,et al.Effects of chronic heat stress on granulosa cell apoptosis and follicular atresia in mouse ovary[J].J Anim Sci Biotechnol,2017,8(1):57-67.
15 Park M,Choi Y,Choi H,et al.Wilms'tumor suppressor gene(WT1)suppresses apoptosis by transcriptionally downregulating Bax expression in immature rat granulosa cells[J].JOvarian Res,2014,7:118-128.
16 Lu M,Xia L,Hua H,et al.Acetyl-keto-beta-boswellic acid induces apoptosis through a death receptor 5-mediated pathway in prostate cancer cells[J].Cancer Res,2008,68:1180-1186.
17 Li YL,Wang GF,Peng H,et al.Research of inhibitory mechanism of salidroside B on proliferation and metastasis of gastric cancer SNU-1 cells[J].Chin J Immun(中国免疫学杂志),2017,33:1815-1827.
18 Mu W,Cheng XF,Zhang X,et al.Effect of hinokiflavone on the proliferation and apoptosis of hepatocellular carcinoma Hep G2 cell[J].Central South Pharmacy(中南药学),2019,17:399-404.
19 Mohammad G1,Alam K,Nawaz MI,et,al.Mutual enhancement between high-mobility group box-1 and NADPH oxidase-derived reactive oxygen species mediates diabetes-induced upregulation of retinal apoptotic markers[J].J Physiol Biochem,2015,71:359-372.
20 Wang X,Bai H,Zhang X,et al.Inhibitory effect of oleanolic acid on hepato-cellular carcinoma via ERK-p53-mediated cell cycle arrest and mitochondrial-dependent apoptosis[J].Carcinogenesis,2013,34:1323-1330.