摘要
目的:探讨不同浓度的二十二碳六烯酸(docosahexaenoic acid,DHA)对人肝癌细胞HepG2生长增殖抑制及促进凋亡的作用及其机制,为其应用于肝癌的药物预防和治疗提供实验及理论依据.方法:以DHA浓度0μg/mL为阴性对照,设置浓度为15、30、45、60、75μg/mL的实验组.采用CCK-8和流式细胞术检测不同浓度DHA对HepG2细胞生长的相对抑制率及凋亡情况,并运用荧光定量PCR及Western blot分别从基因和蛋白质水平分析不同浓度DHA作用前后β-连环蛋白(β-catenin)及C-myc表达量的变化.结果:CCK-8实验表明DHA在体外能抑制H e p G2细胞生长,在D H A作用浓度为0-45μg/mL,作用时间24 h时,实验组与对照组细胞吸光度(A)值差异有显著的统计学意义(P<0.01),实验组内两两比较差异有统计学意义(P<0.01),细胞生长抑制率最高达90.7%.继续增加药物浓度或作用时间,差异无统计学意义.流式细胞术发现DHA可促进HepG2细胞凋亡,实验组与对照组细胞凋亡率差异有显著的统计学意义(P<0.01),实验组内两两比较差异有统计学意义(P<0.01).荧光定量PCR显示DHA可下调HepG2细胞中C-myc基因表达水平,实验组与对照组C-myc基因相对表达量的差异有显著的统计学意义(P<0.01),实验组内两两比较差异有统计学意义(P<0.01);实验组与对照组及各实验者组之间β-catenin基因的相对表达量差异无统计学意义.Western blot显示DHA可降低HepG2细胞β-catenin、C-myc蛋白质的表达量.结论:DHA对人肝癌HepG2细胞有明显的生长增殖抑制及促进凋亡的作用,与其降低β-catenin蛋白质水平进而下调C-myc基因表达水平有关.
AIM： To investigate the effects of docosahexaenoic acid （DHA） of different concentrations on the proliferation and apoptosis of human hepatocelluar carcinoma HepG2 cells, and to explore the possible molecular mechanisms involved. METHODS： HepG2 cells were cultured in vitro and treated with different concentrations of DHA （15, 30, 45, 60, 75μg/mL） for different durations, with untreated cells as controls. The relative rate of reduced growth of HepG2 cellswas detected by the CCK-8 method. Flow cytometry assay was applied to determine the rate of HepG2 cell apoptosis. After DHA treatment, the levels of β-catenin and C-myc mRNAs and proteins were measured by real-time PCR and Western blot, respectively. RESULTS： In the concentration range from 0 to 45μg/mL and with a treatment time of 24 h, DHA significantly inhibit the growth of HepG2 cells in vitro, and significant differences in the absorbance （A value） were observed between the experimental group and control group （P 〈 0.01）, and between each two concentrations in the experimental group. If drug concentration or action time was increased, the results showed no statistically significant differences. Flow cytometric analysis indicated that DHA promotes the apoptosis of HepG2 cells. There were significant differences in the apoptosis rate between the experimental group and control group （P 〈 0.01）, and between each two concentrations in the experimental group. Real-time PCR detected low levels of C-myc expression in HepG2 cells treated with DHA, and significant differences in C-myc expression were observed between the experimental group and control group （P 〈 0.01） and between each two concentrations in the experimental group. There was no significant difference in β-catenin relative expression between the experimental group and control group, or between different concentrations in the experimental group. Western blot analysis demonstrated that DHA could decrease the protein expression of β-catenin and C-myc in HepG2 cells. CONCLUSION： DHA promotes apoptosis and inhibits proliferation of HepG2 cells possibly via mechanism associated with down-regulating the protein expression of β-catenin and the mRNA expression of C-myc.
出处
《世界华人消化杂志》
CAS
北大核心
2014年第13期1834-1840,共7页
World Chinese Journal of Digestology
作者简介
李衍彦，青岛大学医学院肝胆外科硕士研究生，主要从事肝脏肿瘤外科治疗方面的研究
通讯作者：曹景玉，主任医师，266003，青岛市黄岛区五台山路1677号，青岛大学附属医院（黄岛院区）cjy7027@163com
