Effect of Shipi Xiaoji Beverage（实脾消积饮）-containing Serum on Mitochondrial Dynamics Imbalance and Ferroptosis of Liver Cancer HepG2 Cells
10.13288/j.11-2166/r.2024.06.011
- VernacularTitle:实脾消积饮含药血清对肝癌HepG2细胞线粒体动力学平衡和铁死亡的影响
- Author:
Huiying JIAN
1
;
Kexiong LI
2
;
Puhua ZENG
2
Author Information
1. Hunan University of Chinese Medicine， Changsha， 410208
2. Hunan Province Hospital of Integrated Traditional Chinese and Western Medicine/ The Affiliated Hospital of Hunan Academy of Traditional Chinese Medicine
- Publication Type:Journal Article
- Keywords:
primary liver cancer;
Shipi Xiaoji Beverage （实脾消积饮）;
HepG2 cell;
mitochondria;
dynamics;
ferroptosis
- From:
Journal of Traditional Chinese Medicine
2024;65(6):609-617
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo explore the possible mechanism of Shipi Xiaoji Beverage （实脾消积饮） in treating primary liver cancer from the perspective of mitochondrial dynamics imbalance and ferroptosis. MethodsThe Shipi Xiaoji Beverage-containing serum was prepared， and liver cancer HepG2 cells were cultured in vitro. The blank group， control group， cisplatin group （10 µg/ml）， and 5%， 10%， and 15% medicated serum groups were set， with 4 duplicate holes in each group. After adding the corresponding medicinals to each group， the cell survival rate was calculated using the CCK-8 test after 24-hour culture at 37 ℃ with 5% CO2. Using transwell invasion assay， the number of cells that reached the upper chamber membrane through penetrating the Matrigel was calculated after culturing for 48 hours. The scratch migration rate was calculated after incubation for 24 h and 48 h in the scratch experiment， and the concentration of the medicated serum was screened for subsequent experiments. The control group， 10% medicated serum group （the screened concentration） and 10% medicated serum+Mdivi-1 group were set up. After culturing for 24 hours， the average fluorescence intensity of mitochondria and reactive oxygen species （ROS）， adenosine triphosphate （ATP） content and level were measured， and the protein expression of cytoplasmic and mitochondrial dynein-related protein 1 （Drp1） and mitochondrial phosphorylated dynamin-related protein 1 （p-Drp1） was detected. The control group， 10% medicated serum group， ferroptosis activator group （Erastin， 10 μmol/L） and 10% medicated serum+ferroptosis inhibitor group （fer-1，10 μmol/L） were set up， and after adding the corresponding medicinals to each group and culture for 24 hours， the intracellular glutathione （GSH）， malondialdehyde （MDA） content and ferrous ion level were detected， as well as the protein expression of solute carrier family 7 member 11 （SLC7A11） and glutathione peroxidase 4 （GPX4） in cells. ResultsCompared to those in the control group， the cell survival rate， invasion number， 24 h and 48 h scratch migration rate in all medicated serum groups and cisplatin group significantly decreased （P＜0.01）； as the concentration of medicated serum increased， the cell survival rate， invasion number and 48h scratch migration rate of each medicated serum group gradually decreased （P＜0.01）， and finally， 10% of Shipi Xiaoji Beverage-containing serum was selected for subsequent experimental concentration. Compared to the control group， the 10% medicated serum group had decreased average fluorescence intensity of mitochondria and ATP content and increased average fluorescence intensity of ROS （P＜0.05）； compared to the 10% medicated serum group， the 10% medicated serum+Mdivi-1 group had higher average fluorescence intensity of mitochondria and ATP content and lower average fluorescence intensity of ROS （P＜0.05 or P＜0.01）. Compared to those in the control group， GSH content and protein expression of SLC7A11 and GPX4 in other groups significantly decreased， while MDA content and ferrous ion level increased （P＜0.05 or P＜0.01）； compared to the ferroptosis activator group， the 10% medicated serum group and the 10% medicated serum+ferroptosis inhibitor group had decreased MDA content and ferrous ion level， and increased protein expression of SLC7A11 and GPX4 （P＜0.05 or P＜0.01）； the 10% medicated serum+ferroptosis inhibitor group had higher GSH level than the 10% medicated serum group （P＜0.01）. ConclusionShipi Xiaoji Beverage may inhibit the proliferation， infestation， migration of HepG2 cells by causing mitochondrial dynamics imbalance and inducing ferroptosis， thereby playing against liver cancer.
