Dihydromyricetin-mediated inhibition of the Notch1 pathway induces apoptosis in QGY7701 and Hep G2 hepatoma cells

AIM To investigate whether Dihydromyricetin(DHM) inhibits cell proliferation and promotes apoptosis by downregulating Notch1 expression.METHODS The correlation between Notch1 and Hes1(a Notch1 target molecule) expression in hepatoma samples was confirmed by q RT-PCR. In addition, MTT assays, flow cytometry and TUNEL analysis showed that DHM possessed strong anti-tumor properties, evidenced not only by reduced cell proliferation but also by enhanced apoptosis in QGY7701 and Hep G2 hepatocellular carcinoma(HCC) cells. The expressions of Notch1, Hes1, Bcl-2 and Bax were determined by Western blot.RESULTS Among the tested samples(n = 64), the expression levels of Notch1(75% of patients) and Hes1(79.7% of patients) m RNA in tumor tissues were higher than in the normal liver tissues. There was a negative correlation between the expression of Notch1 and the degree of differentiation and positively correlated with the Alpha Fetal Protein concentration. The viability of HCC cells treated with DHM was significantly inhibited in a dose and time-dependent manner. Apoptosis was induced in Hep G2 and QGY7701 cell lines following 24 h of DHM treatment. After treatment with DHM, the protein expression of Notch1 was downregulated, the apoptosis-related protein Bax was upregulated and Bcl2 was downregulated. Notch1 si RNA further enhanced the anti-tumor properties of DHM. CONCLUSION Notch1 is involved in the development of HCC and DHM inhibits cell proliferation and promotes apoptosis by down-regulating the expression of Notch1.

芍药苷通过抑制Notch-1信号通路影响肝癌细胞增殖、侵袭和凋亡的机制研究

目的探讨芍药苷对肝癌细胞增殖、侵袭和凋亡的影响及其机制。方法使用不同浓度(0、25、50、75、100、150μmol/L)芍药苷处理人肝癌细胞系HepG2,采用CCK-8法检测细胞增殖活性。根据芍药苷浓度将HepG2细胞株分为四组:空白对照组(0μmol/L)、芍药苷低剂量组(25μmol/L)、芍药苷中剂量组(50μmol/L)和芍药苷高剂量组(75μmol/L)。采用流式细胞仪检测细胞凋亡情况,Transwell法细胞侵袭迁移能力,Western blot法测定Notch-1和Hes-1蛋白水平。结果使用不同浓度(0、25、50、75、100、150μmol/L)芍药苷处理HepG2细胞24 h后,细胞增殖活性呈剂量依赖性降低[(83.34±8.06)%、(74.46±7.34)%、(63.59±2.51)%、(55.93±4.05)%、(39.99±6.06)%比100%,P<0.05]。当不同浓度(0、25、50、75μmol/L)芍药苷处理HepG2细胞24 h后,细胞穿膜数逐渐降低[(64.33±5.13)个、(49.67±3.51)个、(30.00±2.00)个比(85.33±5.51)个,P<0.05],细胞凋亡率逐渐升高[(13.37±2.57)%、(22.64±2.19)%、(28.53±1.37)%比(2.43±0.93)%,P<0.05],Notch-1蛋白表达水平逐渐降低[(0.75±0.05)、(0.55±0.06)、(0.39±0.04)比(1.13±0.15),P<0.05],Hes-1蛋白表达水平也逐渐降低[(0.55±0.07)、(0.38±0.04)、(0.17±0.03)比(0.80±0.07),P<0.05]。结论芍药苷呈剂量依赖性抑制肝癌细胞HepG2增殖、侵袭,并可促进细胞凋亡,其机制可能与抑制Notch-1信号通路相关。

姜黄素对肝癌细胞凋亡及TNF-ɑ、IL-1β、IL-6炎性因子影响的机制研究

目的探讨姜黄素对肝癌细胞凋亡及肿瘤坏死因子α(TNF-ɑ)、白介素1β(IL-1β)、白介素-6(IL-6)炎性因子的影响及机制。方法采用5、10、20、40、80、160μmol/L的姜黄素处理人肝癌Hep G2细胞24、48 h后,CCK8实验检测细胞的增殖情况,计算IC50值;50μmol/L的姜黄素处理Hep G2细胞48 h后,流式细胞仪检测细胞凋亡情况;Western blot检测TNF-ɑ、IL-1β、IL-6、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved Caspase-3)、Notch1、Hes1蛋白表达情况。结果细胞抑制率随着姜黄素作用时间及浓度的增加显著升高。根据IC50值选择50μmol/L的姜黄素作为研究对象;以50μmol/L的姜黄素处理肝癌细胞的凋亡率及Cleaved Caspase-3蛋白表达明显高于对照组,TNF-ɑ、IL-1β、IL-6、Notch1、Hes1蛋白表达均明显低于对照组(P﹤0.01)。结论姜黄素可呈时间及剂量依赖式抑制人肝癌Hep G2细胞增殖,可诱导细胞的凋亡,可通过降低TNF-ɑ、IL-1β、IL-6的表达达到抗炎作用,其机制与抑制Notch1信号通路有关。