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Self-protection against triptolide-induced toxicity in human hepatic cells via Nrf2-ARE-NQO1 pathway
Ling-Ling Zhou1, Cong Zhou1,2, Xiao-Wen Liang3
1School of Pharmacy, The First Clinical Medical College, Nanjing University of Chinese Medicine, Nanjing, 210023, China.
Objective:
To find the signaling pathway of triptolide (TP)-induced liver injury and to reveal whether NF-E2-related factor 2 (Nrf2) plays an important role in cellular self-protection.
Methods:
The L-02 and HepG2 cells were cultured and treated with various concentrations of TP. The cell viability was observed, and the cell medium was collected for detecting the aspartate aminotransferase (ALT), alanine aminotransferase (AST), lactate dehydrogenase (LDH), superoxide dismutase (SOD) and L-glutathione production (GSH) levels. Nrf2 and its downstream target NAD(P)H: quinine oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) expression, the nuclear translocation of Nrf2, and the binding ability of Nrf2 and antioxidant response element (ARE) were also identified. Meanwhile, shRNA was used to silence Nrf2 in L-02 cells to find out whether Nrf2 plays a protective role.
Results:
The viability of the L-02 and HepG2 cells treated with TP decreased in a doseand time-dependent manner, and TP (20-80 μg/mL) markedly induced the release of ALT, AST and LDH (P<0.05 or P<0.01), reduced the levels of SOD and GSH (P<0.01), and increased the intracellular reactive oxygen species. Meanwhile, TP augmented the Nrf2 expression in L-02 and HepG2 cells (P<0.05 or P<0.01), induced Nrf2 nuclear translocation, increased the Nrf2 ARE binding activity, and increased HO-1 and NQO1 expressions. Nrf2 knockdown revealed a more severe toxic effect of TP (P<0.05 or P<0.01).
Conclusions:
Human hepatic cells treated with TP induced oxidative stress, and led to cytotoxicity. Self-protection against TP-induced toxicity in human hepatic cells might be via Nrf2-ARE-NQO1 transcriptional pathway.
Insights
Triptolide (TP) induces liver injury through oxidative stress. Cellular self-protection against TP toxicity involves the Nrf2-ARE-NQO1 pathway, highlighting Nrf2
Area of Science:
- Hepatology and Toxicology
- Molecular Biology
- Cellular Signaling
Background:
- Triptolide (TP) is a compound known to induce liver injury.
- Understanding the molecular mechanisms of TP-induced hepatotoxicity is crucial for developing protective strategies.
- The role of cellular antioxidant defense systems, particularly the Nrf2 pathway, in mitigating drug-induced liver injury requires further investigation.
Purpose of the Study:
- To elucidate the signaling pathway underlying triptolide (TP)-induced liver injury.
- To determine the involvement of NF-E2-related factor 2 (Nrf2) in cellular self-protection against TP toxicity.
Main Methods:
- Cultured L-02 and HepG2 human hepatic cells were treated with varying concentrations of TP.
- Assessed cell viability and measured liver injury markers (ALT, AST, LDH) and oxidative stress indicators (SOD, GSH, reactive oxygen species).
- Investigated Nrf2 expression, nuclear translocation, and binding activity to the antioxidant response element (ARE), along with downstream targets (NQO1, HO-1).
- Utilized shRNA to silence Nrf2 and evaluate its protective role.
Main Results:
- TP treatment decreased cell viability and increased liver injury markers (ALT, AST, LDH) in a dose- and time-dependent manner.
- TP induced oxidative stress by increasing reactive oxygen species and decreasing SOD and GSH levels.
- TP upregulated Nrf2 expression, promoted its nuclear translocation, enhanced Nrf2-ARE binding, and increased the expression of NQO1 and HO-1.
- Nrf2 knockdown exacerbated TP-induced cytotoxicity.
Conclusions:
- TP induces cytotoxicity in human hepatic cells via oxidative stress.
- The Nrf2-ARE-NQO1 transcriptional pathway plays a significant role in the self-protection of human hepatic cells against TP-induced toxicity.
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