新疆软紫草提取物对HepG2细胞凋亡的影响及其抗小鼠原位肝癌的作用

Effects of extracts from Arnebia euchroma (Royle) Johnst on apoptosis of HepG2 and its anti-hepatocellular carcinoma in mice

目的研究新疆软紫草根[Arnebia euchroma(Royle)Johnst,AE]不同提取物诱导人肝癌细胞株(HepG2)的凋亡作用,并通过小鼠原位肝癌模型验证其抗肝癌作用,初步探究AE提取物的抗肝癌作用。方法首先采用MTT法和Annexin V-FITC/PI双染色法检测AE各提取物对HepG2细胞的抗增殖以及促凋亡作用,采用Western blot法检测了细胞中Bcl-2凋亡家族蛋白的表达水平,综合上述实验结果,筛选出促凋亡作用明显的提取物进行抗小鼠原位肝癌的实验,评价了给药前后原位肝癌小鼠的脏器指数、肝功能指标及组织切片。结果随着AE提取物极性的降低,对HepG2细胞抗增殖、促凋亡作用增强,AE石油醚部位(AEP)抗增殖和诱导凋亡的作用最明显;在AEP剂量为1.56μg·L^(-1)时,Bcl-2蛋白水平明显下降(P<0.05),且呈剂量依赖性,Bad蛋白表达水平在AEP剂量为3.125μg·L^(-1)时明显上调(P<0.05)。动物实验验证了AEP可以改善肝功能,减缓肝癌发展进程的作用。结论研究表明AEP对肝细胞癌的发展具有一定的抑制作用,发挥该作用与调节Bcl-2家族蛋白有关。

Aim To study the apoptosis of human hepatoma cell line(HepG2)induced by different polar parts of Arnebia euchroma(Royle)Johnst(AE)and to verify its anti-hepatoma effect by a mouse orthotopic liver cancer model so as to explore the anti-cancer effect of AE extract.Methods Firstly,MTT method and Annexin V-FITC/PI double staining method were used to detect the anti-proliferative and pro-apoptotic effects of each polar part of AE on HepG2 cells,and Western blot was used to detect the expression of Bcl-2 apoptosis family proteins incells.Based on the above experimental results,the effective parts with significant pro-apoptotic effect were screened out for anti-in situ liver cancer experiments in mice,and the organ indexes,liver function indexes and tissue sections of mice with orthotopic liver cancer before and after administration were evaluated.Results With the decrease of the polarity of AE extract,the anti-proliferation and pro-apoptotic effects on HepG2 cells were enhanced,and the anti-proliferation and apoptosis-inducing effects of AE petroleum ether fraction(AEP)were the most significant.When AEP dose was 1.56μg·L^(-1),the protein level of Bcl-2 decreased significantly in a dose-dependent manner(P<0.05),and the expression level of Bad protein was significantly up-regulated when AEP dose was 3.125μg·L^(-1)(P<0.05).Animal experiments verified the effect of AEP on improving liver function and slowing down the development of liver cancer.Conclusions AEP has a certain inhibitory effect on the development of hepatocellular carcinoma,which is related to the regulation of Bcl-2 family proteins.