Inhibitory Effects and Mechanism of Anemoside B 4 on Hepatocellular Carcinoma Huh- 7 Cells and Tumor-bearing Nude Mice
- VernacularTitle:白头翁皂苷B4对肝癌细胞Huh-7及其荷瘤裸鼠肿瘤的抑制作用及机制研究
- Author:
Shuyi XUE
1
;
Mingchun LI
2
;
Qing MIAO
2
;
Yu ZHOU
2
Author Information
1. Dept. of Pharmacology，School of Pharmacy，Qingdao University，Shandong Qingdao 266021，China;Dept. of Pharmacy，No. 971 Hospital of the Navy of PLA，Shandong Qingdao 266071，China
2. Dept. of Pharmacy，No. 971 Hospital of the Navy of PLA，Shandong Qingdao 266071，China
- Publication Type:Journal Article
- Keywords:
Anemoside B4;
Hepatocellular carcinoma Huh-7;
Tumor-bearing nude mice;
Inhibitory effect;
Mechanism
- From:
China Pharmacy
2019;30(5):601-607
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE： To study the anti-tumor effect of anemoside B4 （AB4） on hepatocellular carcinoma Huh-7 and tumor-bearing nude mice and its mechanism. METHODS： Huh-7 cells were divided into AB4 treatment group， negative control group （constant volume of culture medium） and positive control group （5-fluorouracil 50 μmol/L）. The inhibitory rate of Huh-7 cell proliferation was tested by MTT after treated with 0， 3， 6， 12， 25， 50， 100， 200， 400， 800， 1 600 μmol/L AB4 for 12， 24， 36， 48 h； IC50 were calculated. The number of clone cells was evaluated by clone formation tests after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h. The apoptosis rate of Huh-7 cells was tested by Annexin Ⅴ-FITC/PI double staining after treated with 50 μmol/L AB4 for 24 h. The expression of apoptotic proteins such as Bcl-2， Bax， Caspase-3， Cleaved Caspase-3 and Cleaved PARP were tested by Western blot after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h. Tumor-bearing nude mice were randomly divided into negative control group （normal saline）， positive control group （5-fluorouracil 50 mg/kg）， AB4 lwo-dose， medium-dose and high-dose groups （25， 50， 100 mg/kg）， with 3 mice in each group. They were given relevant medicine intraperitoneally， once a day， for consecutive 19 d. The growth of tumor was observed， and anti-tumor rate was also calculated. HE staining was used to observe the morphology of tumor cells. RESULTS： The inhibition rate of AB4 on Huh-7 cell proliferation increased with the increase of concentration of AB4， but it did not increase significantly after reaching 50 μmol/L； it increased with the increase of time， but it did not increase significantly after 24 h. The IC50 of AB4 was （56.76±1.756） μmol/L. Compared with negative control group， the number of clone cells was decreased significantly after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h， while the protein expression of Bcl-2 was decreased significantly （P＜0.05）； the apoptotic rate， the protein expression of Bax， Caspase-3， Cleaved Caspase-3 and Cleaved PARP were increased significantly （P＜0.05 or P＜0.01）， there was no statistical significance， compared with positive control group. Compared with negative control group， the volume of tumor was decreased significantly in AB4 low-dose， medium-dose and high-dose groups， positive control group （P＜0.05）； the outline of tumor cells become more and more blurred； there were nuclear pyknosis， unclear nucleoplasm and nuclear fragmentation； its anti-tumor rates were 25.93％， 39.15％， 46.26％， 42.83％， respectively. CONCLUSIONS： AB4 inhibits Huh-7 cells and tumor-bearing mice， the mechanism of which may be associated with up-regulating the proportion of Bax/Bcl-2， activating Caspase-3， cracking PARP and inducing apoptosis.
