Effects of M2-type macrophages and GKT137831 on oxidative stress in hepatic stellate cells
10.3760/cma.j.cn501113-20231120-00206
- VernacularTitle:M2?型巨噬细胞与?GKT137831?对肝星状细胞氧化应激的影响
- Author:
Ruge SUN
1
;
Jing LI
;
Fengjun SHEN
Author Information
1. 山西医科大学第一医院消化内科，太原 030001
- Keywords:
Hepatic fibrosis;
Hepatic stellate cells;
M2-type macrophages;
Oxidative stress;
Reactive oxygen species
- From:
Chinese Journal of Hepatology
2024;32(3):201-207
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects of reduced nicotinamide adenine dinucleotide phosphooxidase 4 (NOX4) inhibitors GKT137831 and M2-type macrophages on oxidative stress markers NOX4, nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) in the rat hepatic stellate cell line (HSC-T6).Methods:Rat bone marrow macrophages were extracted and induced using interleukin (IL)-4 to differentiate them into M2 phenotype macrophages. HSC-T6 activation was performed with 5 μg/L transforming growth factor β1 (TGF-β1). The proliferation condition of HSC-T6 cells stimulated by the NOX4 inhibitor GKT137831 at a concentration gradient of 5 to 80 μmol/L after 48 hours was detected using the Cell Counting Kit-8 (CCK-8) assay. The optimal drug concentration was chosen and divided into an HSC co-culture group (the control group) and five experimental groups: the TGF-β1 stimulation group, the TGF-β1 +GKT137831 stimulation group, the M2-type macrophage + HSC co-culture group, the M2-type macrophage +TGF-β1 stimulation group, and the M2-type + TGF-β1 + GKT137831 stimulation group. Reactive oxygen species (ROS) production level was detected in each cell using the DCFH-DA probe method. NOX4, α-smooth muscle actin (α-SMA), Nrf2, and HO-1 levels in each group of HSC cells were detected using the qRT-PCR method and the Western blot method. The t-test was used to compare the two groups. The one-way ANOVA method was used to compare multiple groups. Results:Intracellular ROS increased significantly following TGF-β1 stimulation. ROS relative levels in each cell group were 1.03±0.11, 3.88±0.07, 2.90±0.08, 0.99±0.06, 3.30±0.05, 2.21±0.11, F ?=?686.1, P ?=?0.001, respectively. The mRNA and protein expressions of NOX4, α-SMA, Nrf2, and HO-1 were significantly increased ( P ?
