摘要
目的:观察丹栀逍遥散含药血清对乳腺癌MDA-MB-231细胞的影响,探讨其作用机制是否与干预能量代谢相关。方法:6周龄SD雄性大鼠30只,随机分为空白组、丹栀逍遥散水煎剂组(8.99 g·kg^(-1))、西黄丸组(0.55 g·kg^(-1)),药物干预7 d,分离血清。噻唑蓝(MTT)比色法检测细胞活力,流式细胞术检测细胞周期,Annenxin V/碘化丙啶(PI)双染色法检测细胞凋亡,葡萄糖含量检测试剂盒检测细胞内葡萄糖含量,免疫荧光染色法检测细胞葡萄糖转运蛋白1(GLUT1)表达情况,Seahorse XFe细胞能量代谢分析仪检测细胞胞外酸化率(ECAR)和耗氧率(OCR),蛋白免疫印迹法(Western blot)检测细胞已激糖酶2(HK2),丙酮酸激酶M2型(PKM2),乳酸脱氢酶A(LDHA)表达。结果:与空白组比较,丹栀逍遥散水煎剂含药血清可抑制MDA-MB-231细胞增殖,15%含药血清干预48 h效果最佳(P<0.01)。与空白组比较,丹栀逍遥散水煎剂组细胞DNA暂停分裂期/DNA合成前期(G0/G1期)上调(P<0.01),DNA合成期(S期),DNA合成后期/细胞分裂期(G2/M期)下调(P<0.01);降低MDA-MB-231细胞GLUT1表达;降低MDA-MB-231细胞的基础糖酵解,糖酵解能力值,糖酵解储备值,基础呼吸值,腺苷三磷酸(ATP)产量,储备呼吸值(P<0.01);下调MDA-MB-231细胞HK2,PKM2,LDHA蛋白表达(P<0.05,P<0.01)。结论:丹栀逍遥散含药血清能够抑制乳腺癌MDA-MB-231细胞增殖、促进细胞凋亡、使细胞周期阻滞,其作用机制可能与其逆转乳腺癌MDA-MB-231细胞能量代谢重编程有关。
Objective:To observe the effect of Danzhi Xiaoyaosan-containing serum on MDA-MB-231 breast cancer cells,and to find out whether the action mechanism is related to its intervention in energy metabolism.Method:Thirty six-week-old male SD rats were randomly divided into the blank group,Danzhi Xiaoyaosan(8.99 g·kg^(-1))group,and Xihuangwan(0.55 g·kg^(-1))group.The serum was isolated after drug intervention for seven days.The cell viability was detected by methyl thiazolyl-tetrazolium(MTT)assay,the cell cycle by flow cytometry,and the apoptosis by Annenxin V/propidium iodide(PI)double staining.Following the determination of intracellular glucose content using the glucose testing kit,the expression of glucose transporter 1(GLUT1)was measured by immunofluorescence staining.Seahorse XFe cell energy metabolism analyzer was used to detect the extracellular acidification rate(ECAR)and oxygen consumption rate(OCR).The expression levels of hexokinase 2(HK2),pyruvate kinase M2(PKM2),and lactate dehydrogenase A(LDHA)were assayed by Western blot.Result:Compared with the blank group,Danzhi Xiaoyaosan-containing serum inhibited the proliferation of MDA-MB-231 cells,with the best effect observed after intervention with 15%Danzhi Xiaoyaosan-containing serum for 48 h(P<0.01),blocked the MDA-MB-231 cells in G0/G1 phase(P<0.01),and down-regulated the GLUT1 expression,basal glycolysis,glycolysis capacity,glycolytic reserve,basal respiration,adenosine triphosphate(ATP)production,spare respiratory capacity(P<0.01),as well as the protein expression of HK2,PKM2,and LDHA in MDA-MB-231 cells(P<0.05,P<0.01).Conclusion:Danzhi Xiaoyaosan-containing serum inhibits MDA-MB-231 cell proliferation,promotes apoptosis,and induces cell cycle arrest,which may be related to its reversal of energy metabolic reprogramming in MDA-MB-231 cells.
作者
徐爽
李然
刘立萍
XU Shuang;LI Ran;LIU Li-ping(College of Traditional Chinese Medicine(TCM),Liaoning University of TCM,Shenyang 110032,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2022年第3期8-14,共7页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金面上项目(81673876)。
关键词
乳腺癌
丹栀逍遥散
能量代谢重编程
糖酵解
breast cancer
Danzhi Xiaoyaosan
energy metabolic reprogramming
glycolysis
