TLR4在龙血竭抑制肝癌细胞生长中的作用

Role of TLR4 in the inhibitory effect of Resina Draconis on human hepatoma cell

目的探讨龙血竭是否通过调控toll样受体4(TLR4)基因的表达对肝癌细胞起到抑制作用。方法首先通过CCK8法检测龙血竭对人肝癌细胞系HepG2和HepG2.2.15增殖的影响,并且确定后续实验的最适用药浓度;其次,将肝癌细胞系HepG2和HepG2.2.15分别分为对照组和上药组,上药组在培养基中加入龙血竭,对照组仅加入等量细胞培养基。进一步通过实时荧光定量PCR法、Western-Blot法检测各组细胞TLR4基因的表达水平。结果CCK8法显示,加入龙血竭后,HepG2和HepG2.2.15两组细胞系生长速度明显减慢,IC50值分别为(97.78±8.19)μg/mL和(66.89±5.86)μg/mL;经实时荧光定量PCR检测,与对照组相比,HepG2细胞系上药组TLR4 mRNA表达水平下降[(1.00±0.25)vs(0.32±0.07)],差异有统计学意义(P<0.05);与对照组相比,HepG2.2.15细胞系上药组TLR4 mRNA表达水平亦下降[(1.00±0.21)vs(0.29±0.06)],差异有统计学意义(P<0.05);经Western-blot检测,HepG2和HepG2.2.15两种细胞系上药组的TLR4的蛋白表达水平均明显下降[(0.89±0.05)vs(0.68±0.06)、(1.11±0.04)vs(0.56±0.08)],差异均有统计学意义(P<0.05)。结论龙血竭可能通过调控TLR4基因表达以抑制人肝癌细胞系HepG2和HepG2.2.15的生长。

Objective To discuss the role of toll-like receptor-4(TLR4)in the inhibitory effect of Resina Draconis on human hepatoma cells.Methods Firstly,effects of Resina Draconis on proliferation of human hepatoma cell line HepG2 and HepG2.2.15 were detected by Cell Counting Kit-8 assay,and the optimal inhibitory concentration was determined.Secondly,the human hepatoma HepG2 and HepG2.2.15 cells cultured in vitro were divided into experimental group(with Resina Draconis added in the culture)and control group(with the same amount of cell culture medium added)respectively.Then,RT-qPCR analysis and Western-Blot analysis were used to detect the level of TLR4 expression.Results CCK-8 assay showed that the proliferation of human hepatoma HepG2 and HepG2.2.15 cells in experimental groups with Resina Draconis were suppressed when compared to control groups,and the 50%inhibiting concentration were(97.78±8.19)μg/mL and(66.89±5.86)μg/mL,respectively.Compared with control groups,RT-qPCR analysis showed that mRNA level of TLR4 in the experimental group of HepG2 and HepG2.2.15 cells were significantly decreased with(1.00±0.25)vs(0.32±0.07),(1.00±0.21)vs(0.29±0.06),respectively(all P<0.05).Compared with control groups,Western-blot analysis showed that protein level of TLR4 in the experimental groups of HepG2 and HepG2.2.15 cells were both significantly decreased with(0.89±0.05)vs(0.68±0.06),(1.11±0.04)vs(0.56±0.08),respectively(all P<0.05).Conclusion Resina Draconis may inhibit the growth of human hepatoma HepG2 and HepG2.2.15 cells by mediating the expression of TLR4.