Glutathione Induces Keap1 S-Glutathionylation and Mitigates Oscillating Glucose-Induced β-Cell Dysfunction by Activating Nrf2
Abstract
:1. Introduction
2. Materials and Methods
2.1. In Vivo Studies
2.2. Cell Culture and Treatments
2.3. Pancreatic Islet Isolation and Primary Culture
2.4. Glucose-Stimulated Insulin Secretion (GSIS) Assay
2.5. RNA Isolation and Real-Time Quantitative PCR
2.6. Western Blotting
2.7. Co-Immunoprecipitation and Determination of S-Glutathionylation of Keap1
2.8. Construction of Keap1 Cysteine Mutants
2.9. Immunofluorescence Staining
2.10. GSH/GSSG Assay
2.11. Detection of Mitochondrial ROS Level
2.12. Apoptosis Assay
2.13. Statistical Analysis
3. Results
3.1. GSH Prevents the Inhibition of Nrf2, Grx1, and HO-1 Expressions in Islets of Rats Induced by LOsG
3.2. GSH Prevents OsG-Induced β-Cell Failure
3.3. GSH Activates Nrf2 Pathway in OsG-Treated β-Cells
3.4. GSH Promotes Nrf2-Related Antioxidant Enzyme Expression to Attenuate Oxidative Stress in OsG-Treated β-Cells and Primary Islets
3.5. Inhibition of Nrf2 Abrogates the Cytoprotective Effects of GSH in OsG-Treated β-Cells
3.6. GSH Increases the S-Glutathionylation of Keap1 to Activate the Nrf2 Pathway in OsG-Treated β-Cells
3.7. The S-Glutathionylation of Keap1 at Cys273 and Cys288 Caused the Activation of the Keap1/Nrf2 Pathway in GSH-Treated β-Cells under OsG Condition
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Target Genes | Forward Primer (5′ to 3′) | Reverse Primer (5′ to 3′) | Accession No. |
---|---|---|---|
Ho-1 | CCCAGAGGCTGTGAACTCTG | GGGGAAAGCAGTCATGGTCA | NM_012580.2 |
Nqo1 | GAAAGGATGGGAGGTGGTCG | GCTCCCCTGTGATGTCGTTT | NM_017000.3 |
Gclc | TGATTGAAGGGACACCTGGC | TGTGCTCTGGCAGTGTGAAT | NM_012815.2 |
Grx1 | CGTGGTCTCCTGGAATTTGTG | AAGACCCGAGGAACTGTTCTTG | NM_022278.1 |
Nrf2 | TAGATCTTGGGGTAAGTCGAGA | CTCTTGTCTCTCCTTTTCGAGT | NM_001399173.1 |
Ins2 | TGGAAGCTCTCTACCTGGTGT | GTGCCAAGGTCTGAAGGTCAC | NM_019130.2 |
Ins1 | CTACACACCCAAGTCCCGTC | CCAAGGTCTGAAGATCCCCG | NM_019129.3 |
Ngn3 | GTCAGAGACTGTCACACCCC | TGGAACTGAGCACTTCGTGG | NM_021700.1 |
MafA | TTCTGGAGAGCGAGAAGTGC | CGCGCTCACAGAAAGAAGTC | XM_017603453.1 |
Pdx1 | CGGACATCTCCCCATACG | AAAGGGAGATGAACGCGG | NM_022852.3 |
β-actin | TTTAATGTCACGCACGATTTCCC | CCCATCTATGAGGGTTACGC | NM_031144.3 |
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Chen, X.; Zhou, Q.; Chen, H.; Bai, J.; An, R.; Zhang, K.; Zhang, X.; An, H.; Zhang, J.; Wang, Y.; et al. Glutathione Induces Keap1 S-Glutathionylation and Mitigates Oscillating Glucose-Induced β-Cell Dysfunction by Activating Nrf2. Antioxidants 2024, 13, 400. https://doi.org/10.3390/antiox13040400
Chen X, Zhou Q, Chen H, Bai J, An R, Zhang K, Zhang X, An H, Zhang J, Wang Y, et al. Glutathione Induces Keap1 S-Glutathionylation and Mitigates Oscillating Glucose-Induced β-Cell Dysfunction by Activating Nrf2. Antioxidants. 2024; 13(4):400. https://doi.org/10.3390/antiox13040400
Chicago/Turabian StyleChen, Xiufang, Qian Zhou, Huamin Chen, Juan Bai, Ruike An, Keyi Zhang, Xinyue Zhang, Hui An, Jitai Zhang, Yongyu Wang, and et al. 2024. "Glutathione Induces Keap1 S-Glutathionylation and Mitigates Oscillating Glucose-Induced β-Cell Dysfunction by Activating Nrf2" Antioxidants 13, no. 4: 400. https://doi.org/10.3390/antiox13040400
APA StyleChen, X., Zhou, Q., Chen, H., Bai, J., An, R., Zhang, K., Zhang, X., An, H., Zhang, J., Wang, Y., & Li, M. (2024). Glutathione Induces Keap1 S-Glutathionylation and Mitigates Oscillating Glucose-Induced β-Cell Dysfunction by Activating Nrf2. Antioxidants, 13(4), 400. https://doi.org/10.3390/antiox13040400