Improved Wound Healing and Skin Regeneration Ability of 3,2′-Dihydroxyflavone-Treated Mesenchymal Stem Cell-Derived Extracellular Vesicles
Abstract
:1. Introduction
2. Results
2.1. Screening Proliferative Effects of Various Flavonoids on MSCs
2.2. The Proliferative Effect of 3,2′-DHF on MSCs
2.3. Characterization and Enhanced Production of EVs from 3,2′-DHF-Treated MSCs
2.4. Anti-Inflammatory Effects of Fla-EVs
2.5. In Vitro Wound Closure Effect of Fla-EVs on Human Dermal Fibroblasts and Keratinocytes
2.6. Regeneration Effects of Fla-EVs in Excisional Skin Wound Model
3. Discussion
4. Materials and Methods
4.1. Cell Culture
4.2. Cell Viability Test
4.3. Isolation and Characterization of EVs
4.4. Western Blot
4.5. Transmission Electron Microscopy (TEM)
4.6. Flow Cytometry
4.7. RAW 264.7 Cell Anti-Inflammation Assay
4.8. Griess Assay
4.9. Enzyme-Linked Immunosorbent Assay (ELISA)
4.10. In Vitro Migration Assay (Scratch Assay)
4.11. Skin Excisional Wound-Healing Mouse Model
4.12. Histological Analysis
4.13. 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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Instrumentation | Product Number | Company |
---|---|---|
xMark™ microplate absorbance spectrophotometer | 681150 | Bio-Rad Laboratories |
High-speed centrifuge | Avanti J-E | Beckman Coulter |
Ultracentrifuge | Optima L-90K | Beckman Coulter |
ZetaView | TWIN PMX-220 | Particle Metrix |
iBright™ Imagers | CL-1000 | Invitrogen |
Transmission electron microscope | JEM-1010 | Nippon Denshi |
Flow cytometer | CytoFLEX | Beckman Coulter |
Antibody | Source | Company (Cat No.) |
---|---|---|
Recombinant anti-CD9 antibody | rabbit | Abcam (ab263023) |
Recombinant anti-CD81 antibody | rabbit | Abcam (ab109201) |
Calnexin rabbit mAb | rabbit | Cell signaling technology (2679) |
GM130 rabbit mAb | rabbit | Cell signaling technology (12480) |
p-Akt1 antibody | mouse | Santa Cruz Biotechnology (sc-293125) |
Akt1 polyclonal antibody | rabbit | Cusabio (CSB-PA000855) |
Phospho-MAPK3/MAPK1 (T202/Y204) antibody | rabbit | Cusabio (CSB-PA000749) |
p44/42 MAP kinase antibody | rabbit | Tebu-bio (B7074) |
β-actin antibody | mouse | Santa Cruz Biotechnology (sc-47778) |
Anti-rabbit IgG, HRP-linked antibody | goat | Santa Cruz Biotechnology (7074) |
Anti-mouse IgG, HRP-linked antibody | horse | Santa Cruz Biotechnology (7076) |
Antibody | Source | Company (Cat No.) |
---|---|---|
CD73 monoclonal antibody | mouse | Invitrogen (41-0200) |
Human/porcine/canine CD90/Thy1 antibody | sheep | R&D Systems (AF2067) |
CD105 monoclonal antibody | mouse | Invitrogen (MA5-11854) |
CD34 antibody, anti-human | mouse | Miltenyi Biotec (130-108-040) |
CD45 antibody, anti-human, REAfinity™ | human | Miltenyi Biotec (130-110-771) |
Goat anti-mouse IgG (H + L), Superclonal™ recombinant secondary antibody, Alexa Fluor™ 488 | goat | Invitrogen (A28175) |
Donkey F(ab’)2 anti-sheep IgG H&L (PE) preadsorbed | donkey | Abcam (ab7009) |
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Kim, S.; Shin, Y.; Choi, Y.; Lim, K.-M.; Jeong, Y.; Dayem, A.A.; Lee, Y.; An, J.; Song, K.; Jang, S.B.; et al. Improved Wound Healing and Skin Regeneration Ability of 3,2′-Dihydroxyflavone-Treated Mesenchymal Stem Cell-Derived Extracellular Vesicles. Int. J. Mol. Sci. 2023, 24, 6964. https://doi.org/10.3390/ijms24086964
Kim S, Shin Y, Choi Y, Lim K-M, Jeong Y, Dayem AA, Lee Y, An J, Song K, Jang SB, et al. Improved Wound Healing and Skin Regeneration Ability of 3,2′-Dihydroxyflavone-Treated Mesenchymal Stem Cell-Derived Extracellular Vesicles. International Journal of Molecular Sciences. 2023; 24(8):6964. https://doi.org/10.3390/ijms24086964
Chicago/Turabian StyleKim, Sehee, Yeokyung Shin, Yujin Choi, Kyung-Min Lim, Yeojin Jeong, Ahmed Abdal Dayem, Yoonjoo Lee, Jongyub An, Kwonwoo Song, Soo Bin Jang, and et al. 2023. "Improved Wound Healing and Skin Regeneration Ability of 3,2′-Dihydroxyflavone-Treated Mesenchymal Stem Cell-Derived Extracellular Vesicles" International Journal of Molecular Sciences 24, no. 8: 6964. https://doi.org/10.3390/ijms24086964