Extracellular Vesicles and Cx43-Gap Junction Channels Are the Main Routes for Mitochondrial Transfer from Ultra-Purified Mesenchymal Stem Cells, RECs
Abstract
:1. Introduction
2. Results
2.1. Mitochondrial Transfer to ρ0 Cells from RECs in a Direct-Contact System
2.2. Mitochondrial Transfer via TNTs
2.3. Transfer of REC- and BMSC-Derived Mitochondria to ρ0 Cells via Cx43-Regulated GJCs
2.4. REC-Derived Mitochondrial Transfer to ρ0 Cells and mtDNA Recovery in the Noncontact System
2.5. Characterization of Exosomes
2.6. REC-EXO Contributes to the Donation of Mitochondria by RECs and BMSCs to ρ0 Cells
2.7. REC-EXO Contributes to the Restoration of Mitochondrial Function in ρ0 Cells
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Transfer of Mitochondria from RECs and BMSCs Using the Direct-Contact System
4.3. Transfer of Mitochondria from RECs and BMSCs Using the Noncontact System
4.4. Fluorescence Microscopy
4.5. Western Blotting (WB)
4.6. Isolation and Characterization of Exosomes
4.7. Analysis of Mitochondrial DNA
4.8. Transmission Electron Microscopy
4.9. Measurement of Mitochondrial Membrane Potential
4.10. Measurement of ROS Levels
4.11. Seahorse Analysis of Mitochondrial Function
4.12. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene Name | Primer Pairs | Product Size (bp) |
---|---|---|
COX1 for real-time PCR | Forward 5′-GCT ACC ATA ATC ATC GCT ATC-3′ | 155 |
Reverse 5′-GCT AAT ACA ATG CCA GTC AG-3′ | ||
HVR2 for real-time PCR | Forward 5′-CTA TGT CGC AGT ATC TGT CT-3′ | 86 |
Reverse 5′-AGT AAG TAT GTT CGC CTG TA-3′ | ||
COX1 for PCR | Forward 5′-ACA CGA GCA TAT TTC ACC TCC G-3′ | 336 |
Reverse 5′-GGA TTT TGG CGT AGG TTT GGT C-3′ | ||
HVR2 for PCR | Forward 5′-CTC ACG GGA GCT CTC CAT GC-3′ | 401 |
Reverse 5′-CTG TTA AAA GTG CAT ACC GCC A-3′ | ||
CH1 | Forward 5′-GGC TCT GTG AGG GAT ATA AAG ACA-3′ | 98 |
Reverse 5′-CAA ACC ACC CGA GCA ACT AAT CT-3′ | ||
ACTB | Forward 5′-TGG CAC CCA GCA CAA TGA A-3′ | 186 |
Reverse 5′-CTA AGT CAT AGT CCG CCT AGA AGC A-3′ |
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Yang, J.; Liu, L.; Oda, Y.; Wada, K.; Ago, M.; Matsuda, S.; Hattori, M.; Goto, T.; Ishibashi, S.; Kawashima-Sonoyama, Y.; et al. Extracellular Vesicles and Cx43-Gap Junction Channels Are the Main Routes for Mitochondrial Transfer from Ultra-Purified Mesenchymal Stem Cells, RECs. Int. J. Mol. Sci. 2023, 24, 10294. https://doi.org/10.3390/ijms241210294
Yang J, Liu L, Oda Y, Wada K, Ago M, Matsuda S, Hattori M, Goto T, Ishibashi S, Kawashima-Sonoyama Y, et al. Extracellular Vesicles and Cx43-Gap Junction Channels Are the Main Routes for Mitochondrial Transfer from Ultra-Purified Mesenchymal Stem Cells, RECs. International Journal of Molecular Sciences. 2023; 24(12):10294. https://doi.org/10.3390/ijms241210294
Chicago/Turabian StyleYang, Jiahao, Lu Liu, Yasuaki Oda, Keisuke Wada, Mako Ago, Shinichiro Matsuda, Miho Hattori, Tsukimi Goto, Shuichi Ishibashi, Yuki Kawashima-Sonoyama, and et al. 2023. "Extracellular Vesicles and Cx43-Gap Junction Channels Are the Main Routes for Mitochondrial Transfer from Ultra-Purified Mesenchymal Stem Cells, RECs" International Journal of Molecular Sciences 24, no. 12: 10294. https://doi.org/10.3390/ijms241210294