[Effects of M2-type macrophages and GKT137831 on oxidative stress in hepatic stellate cells].
Zhonghua Gan Zang Bing Za Zhi
; 32(3): 201-207, 2024 Mar 20.
Article
em Zh
| MEDLINE
| ID: mdl-38584100
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 ß1 (TGF-ß1). 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-ß1 stimulation group, the TGF-ß1 +GKT137831 stimulation group, the M2-type macrophage + HSC co-culture group, the M2-type macrophage +TGF-ß1 stimulation group, and the M2-type + TGF-ß1 + 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-ß1 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, Fâ =â 686.1, Pâ =â 0.001, respectively. The mRNA and protein expressions of NOX4, α-SMA, Nrf2, and HO-1 were significantly increased (Pâ <â 0.05). After the addition of GKT137831, ROS, and NOX4, α-SMA mRNA and protein expression were comparatively decreased in the TGF-ß1 stimulation group (Pâ <â 0.05), while mRNA and protein expressions of Nrf2 and HO-1 were increased (Pâ <â 0.05). The expression of ROS and NOX4, as well as α-SMA mRNA and protein, produced by HSC were significantly decreased in the co-culture group compared to the single culture group after TGF-ß1 stimulation (Pâ <â 0.05). After the addition of GKT137831, ROS, NOX4, α-SMA mRNA, and protein expression were further reduced in the co-culture group compared with the single culture group (Pâ <â 0.05), while the mRNA and protein expression of Nrf2 and HO-1 were further increased (Pâ <â 0.05).Conclusion:
NOX4 inhibitor GKT137831 can reduce RO, NOX4, and α-SMA levels while increasing Nrf2 and HO-1 levels in hepatic stellate cells. After M2-type macrophage co-culture, GKT137831 assists in lowering ROS, NOX4, and α-SMA levels while accelerating Nrf2 and HO-1 levels in hepatic stellate cells, which regulates the balance between oxidative stress and anti-oxidative stress systems, thereby antagonizing the fibrosis process.Palavras-chave
Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
Piridonas
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Pirazolonas
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Fator de Crescimento Transformador beta1
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Células Estreladas do Fígado
Idioma:
Zh
Ano de publicação:
2024
Tipo de documento:
Article