Mechanism of Inducing Ferroptosis in Hepatocellular Carcinoma Cells by Shugan Quyu Jiedu Prescription Based on p53/SLC7A11/GPX4 Pathway
10.13422/j.cnki.syfjx.20232024
- VernacularTitle:基于p53/SLC7A11/GPX4通路研究疏肝祛瘀解毒方诱导肝癌细胞铁死亡的作用机制
- Author:
Xiaojun CAI
1
;
Renyi YANG
2
;
Zhibin WANG
3
;
Yilin GONG
1
;
Ke WANG
4
;
Lizhu LIN
4
;
Chong ZHONG
4
;
Jing LI
1
Author Information
1. The First Affiliated Hospital of Hunan University of Chinese Medicine, Changsha 410007, China
2. School of Integrated Chinese and Western Medicine, Hunan University of Chinese Medicine, Changsha 410208, China
3. Hunan University of Chinese Medicine, Changsha 410208, China
4. The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou 510405, China
- Publication Type:Journal Article
- Keywords:
Shugan Quyu Jiedu prescription;
hepatocellular carcinoma;
ferroptosis;
proliferation
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2024;30(8):74-82
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo investigate the effect of Shugan Quyu Jiedu prescription (SGQYJDF) on inducing ferroptosis in hepatocellular carcinoma cells based on the tumor protein 53 (p53)/solute carrier family 7 member 11 (SLC7A11)/glutathione peroxidase 4 (GPX4) pathway. MethodMHCC97H cells were divided into the blank serum group (10% blank serum medium), SGQYJDF-containing serum low concentration group (5% SGQYJDF-containing serum and 5% blank serum medium), SGQYJDF-containing serum medium concentration group (7.5% SGQYJDF-containing serum and 2.5% blank serum medium), SGQYJDF-containing serum high concentration group (10% SGQYJDF-containing serum medium) and sorafenib group (sorafenib concentration of 10 μmol·L-1 in 10% blank serum medium). After 24 hours of intervention, the cell survival rate was detected by cell counting kit-8 (CCK-8) assay. The cell proliferation ability was detected by 5-ethynyl-2′-deoxyuridine (EdU) staining. The intracellular ferrous ion (Fe2+) level was detected by ferrous ion fluorescent probe (FerroOrange) staining. The intracellular malondialdehyde (MDA) and glutathione (GSH) levels were detected by colorimetric assays. The ultrastructure of mitochondria was observed by transmission electron microscopy. The expression levels of ferroptosis-related proteins p53, SLC7A11 and GPX4 were detected by Western blot. ResultIn terms of cell viability, compared with the blank serum group, the SGQYJDF group showed a dose-dependent decrease in the survival rate of MHCC97H cells. Effect of the medium and high concentrations of SGQYJDF on the survival rate of MHCC97H cells were significantly decreased (P<0.01). Additionally, the results of the EdU assay showed that both the medium and high concentrations of SGQYJDF were able to inhibit the proliferation ability of MHCC97H cells (P<0.05, P<0.01). Regarding the biochemical indicators of ferroptosis, compared to the blank serum group, the medium and high concentrations of SGQYJDF were able to dose-dependently increase the intracellular Fe2+ level (P<0.01). The low, medium, and high concentrations of SGQYJDF were able to dose-dependently decrease the level of GSH in MHCC97H cells (P<0.01) and increase the level of MDA in the cells (P<0.05, P<0.01). In terms of pathway-related protein expression, compared to the blank serum group, the medium and high concentrations of SGQYJDF could significantly increase the expression of p53 (P<0.01). The low, medium, and high concentrations of SGQYJDF could significantly decrease the expression of GPX4 (P<0.01). The high concentration of SGQYJDF could decrease the expression of SLC7A11 (P<0.01). In terms of the cell morphology of ferroptosis, compared with the blank serum group, transmission electron microscopy revealed that the low concentration of SGQYJDF caused mitochondrial deformation, while the medium and high concentrations of SGQYJDF resulted in reduced mitochondrial volume, increased double-layer membrane density, and decreased mitochondrial cristae. These features were similar to those of sorafenib-induced ferroptosis. Furthermore, compared with the sorafenib group, the high concentration of SGQYJDF showed no statistically significant differences in cell survival rate, proliferation ability, Fe2+ level, MDA level, and GSH level. ConclusionThe results suggest that SGQYJDF may induce ferroptosis and inhibit proliferation in hepatocellular carcinoma MHCC97H cells by upregulating the expression of p53, suppressing the expressions of GPX4 and SLC7A11, downregulating the level of GSH, and leading to the accumulation of intracellular Fe2+ and MDA.