Dexmedetomidine inhibits ferroptosis of human renal tubular epithelial cells by activating the Nrf2/HO-1/GPX4 pathway

doi:10.12122/j.issn.1673-4254.2024.06.14

Abstract

Abstract:

Objective To investigate the protective effect of dexmedetomidine (DEX) against erastin-induced ferroptosis in human renal tubular epithelial cells (HK-2 cells) and explore the underlying mechanism. Methods HK-2 cells were treated with erastin alone or in combination with different concentrations (2.5, 5.0 and 10 μmol/L) of DEX, and the changes in cell viability were observed using CCK-8 assay. To explore the mechanism by which DEX inhibits erastin-induced ferroptosis, HK-2 cells were treated with erastin, erastin+10 μmol/L DEX, or erastin+10 μmol/L DEX+ML385 (a Nrf2 inhibitor), after which the cell viability was assessed. The level of intracellular Fe²⁺ was detected by cell ferrous iron colorimetric assay kit, and flow cytometry was performed to detect reactive oxygen species (ROS); MDA and reduced glutathione assay kits were used to detect the contents of MDA and GSH in the cells; The expressions of Nrf2, HO-1 and GPX4 proteins were detected by Western blotting. Results Erastin treatment significantly inhibited the viability of the cells, decreased GSH content, and increased intracellular levels of Fe²⁺, ROS and MDA. The combined treatment with 10 μmol/L DEX markedly increased the viability of the cells, increased GSH content, reduced the levels of Fe²⁺, ROS and MDA, and upregulated the protein expressions of Nrf2, HO-1 and GPX4 in the cells. The application of ML385 obviously blocked the protective effect of DEX and caused significant inhibition of the Nrf2/HO-1/GPX4 pathway, decreased the cell viability and GSH content, and increased the levels of Fe²⁺, ROS and MDA in HK-2 cells. Conclusion The protective effect of DEX against erastin-induced ferroptosis of HK-2 cells is probably mediated by activation of the Nrf2/HO-1/GPX4 pathway to inhibit oxidative stress.

Key words: dexmedetomidine, erastin, HK-2 cells, ferroptosis, Nrf2/HO-1/GPX4 pathway

Fangyuan ZHANG, Gang LIU. Dexmedetomidine inhibits ferroptosis of human renal tubular epithelial cells by activating the Nrf2/HO-1/GPX4 pathway[J]. Journal of Southern Medical University, 2024, 44(6): 1135-1140.

Figures/Tables 5

Fig.1 Effect of DEX on viability of HK-2 cells treated with erastin (Mean±SD, n=3). A: Effects of different concentrations of erastin on viability of HK-2 cells. ***P<0.001 vs 0 μmol/L. B: DEX at different concentrations increases viability of erastin-treated HK-2 cells. C: ML385 attenuates the protective effect of DEX in erastin-treated HK-2 cells. ***P<0.001 vs Control group; #P<0.05, ###P<0.001 vs Erastin group; &&&P<0.001 vs Erastin+DEX group.

Fig.2 Effect of DEX on intracellular Fe2+ level in HK-2 cells treated with erastin (Mean±SD, n=3). ***P<0.001 vs Control group; ###P<0.001 vs Erastin group; &P<0.05 vs Erastin+DEX group.

Fig.3 Effect of DEX on protein expressions of Nrf2, HO-1 and GPX4 in HK-2 cells treated with erastin (Mean±SD, n=3). A: Western blotting results. B-D: Expression levels of Nrf2, HO-1 and GPX4 proteins in HK-2 cells with different treatments. **P<0.01, ***P<0.001 vs Control group; ###P<0.001 vs Erastin group; &&&P<0.001 vs Erastin+DEX group.

Fig.4 Effect of DEX on ROS level in HK-2 cells treated with erastin (Mean±SD, n=3). ***P<0.001 vs Control group; ###P<0.001 vs Erastin group; &&&P<0.001 vs Erastin+DEX group.

Fig.5 Effect of DEX on MDA and GSH contents in HK-2 cells treated with Erastin (Mean±SD, n=3). A: MDA content in HK-2 cells with different treatment. B: GSH content in HK-2 cells with different treatment. ***P<0.001 vs Control group; ###P<0.001 vs Erastin group; &P<0.05, &&&P<0.001 vs Erastin+DEX group.

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