Human Umbilical Cord Mesenchymal Stem Cells Protect against Renal Ischemia-Reperfusion Injury by Secreting Extracellular Vesicles Loaded with miR-148b-3p That Target Pyruvate Dehydrogenase Kinase 4 to Inhibit Endoplasmic Reticulum Stress at the Reperfusion Stages
Abstract
:1. Introduction
2. Results
2.1. Isolation and Identification of HucMSC-EVs
2.2. miR-148b-3p Participates in the Protective Effect of HucMSC-EVs on Renal Ischemia
2.3. miR-148b-3p Regulates PDK4 and Participates in the Protective of I/R Injury
2.4. PDK4 Aggravated the I/R Injury in HK-2 Cells by ATF-6 Pathway of ER Stress
2.5. HucMSC-EVs Alleviate Kidney I/R Injury In Vivo
3. Discussion
4. Materials and Methods
4.1. Animals, Cells and Reagents
4.2. Cell Culture Experiments
4.3. Cell Transfection
4.4. Animal Experiments
4.5. Quantitative Real-Time Polymerase Chain Reaction
4.6. Dual-Luciferase Reporter Assay
4.7. Biochemical Analysis
4.8. HE Staining
4.9. Immunohistochemical Staining
4.10. Small Extracellular Vesicles Isolation
4.11. Small Extracellular Vesicles Identification
4.12. Nanoparticle Tracking Analysis (NTA)
4.13. Transmission Electron Microscope (TEM)
4.14. Tracking of Small Extracellular Vesicles and Internalization Assay
4.15. Flow Cytometric Analysis
4.16. Western Blotting
4.17. Statistical Analysis
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Genes | Primer Sequence |
---|---|
miR-148b-3p | 5′-CGCGTCAGTGCATCACAGAA-3′ |
U6 | 5′-ATGGACTATCATATGCTTACCGAT-3′ |
PDK4 | as:5′-CUAUAGACUCAGAAGACAAAG-3′ ss: 5′-CUUUGUCUUCUGAGUCUAUAG-3′ |
β-actin | Forward:5′-AAACGTGCTGCTGACCGAG-3′ Reverse: 5′-TAGCACAGCCTFFATAGCAAC-3′ |
human DEDD | Forward:5′-GCCCTACGTCACCCTCAAG-3′ Reverse: 5′-GCCTTGGTTCTGGATCACTG-3′ |
human KIT | Forward:5′-AAAGGCAGAAGCCACCAACA-3′ Reverse: 5′-AGGAGCGGTCAACAAGGAAA-3′ |
human PIK3R3 | Forward:5′-GGCTTGCCTACCCTGTTCAT-3′ Reverse: 5′-CGGCCTCTCCACTTCACATT-3′ |
human PDK4 | Forward:5′-TGGAGCATTTCTCGCGCTAC-3′ Reverse: 5′-TCCACCAAATCCATCAGGCT-3′ |
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Shi, W.; Zhou, X.; Li, X.; Peng, X.; Chen, G.; Li, Y.; Zhang, C.; Yu, H.; Feng, Z.; Gou, X.; et al. Human Umbilical Cord Mesenchymal Stem Cells Protect against Renal Ischemia-Reperfusion Injury by Secreting Extracellular Vesicles Loaded with miR-148b-3p That Target Pyruvate Dehydrogenase Kinase 4 to Inhibit Endoplasmic Reticulum Stress at the Reperfusion Stages. Int. J. Mol. Sci. 2023, 24, 8899. https://doi.org/10.3390/ijms24108899
Shi W, Zhou X, Li X, Peng X, Chen G, Li Y, Zhang C, Yu H, Feng Z, Gou X, et al. Human Umbilical Cord Mesenchymal Stem Cells Protect against Renal Ischemia-Reperfusion Injury by Secreting Extracellular Vesicles Loaded with miR-148b-3p That Target Pyruvate Dehydrogenase Kinase 4 to Inhibit Endoplasmic Reticulum Stress at the Reperfusion Stages. International Journal of Molecular Sciences. 2023; 24(10):8899. https://doi.org/10.3390/ijms24108899
Chicago/Turabian StyleShi, Wei, Xiang Zhou, Xinyuan Li, Xiang Peng, Guo Chen, Yang Li, Chunlin Zhang, Haitao Yu, Zhenwei Feng, Xin Gou, and et al. 2023. "Human Umbilical Cord Mesenchymal Stem Cells Protect against Renal Ischemia-Reperfusion Injury by Secreting Extracellular Vesicles Loaded with miR-148b-3p That Target Pyruvate Dehydrogenase Kinase 4 to Inhibit Endoplasmic Reticulum Stress at the Reperfusion Stages" International Journal of Molecular Sciences 24, no. 10: 8899. https://doi.org/10.3390/ijms24108899