Details
| Original language | English |
|---|---|
| Pages (from-to) | 279–293 |
| Number of pages | 15 |
| Journal | Biotechnology letters |
| Volume | 46 |
| Issue number | 2 |
| Early online date | 13 Feb 2024 |
| Publication status | Published - Apr 2024 |
Abstract
Purpose: 3D cell culture and hypoxia have been demonstrated to increase the therapeutic effects of mesenchymal stem/stromal cells (MSCs)-derived extracellular vesicles (EVs). In this study, a process for the production of MSC-EVs in a novel 3D bioreactor system under normoxic and hypoxic conditions was established and the resulting EVs were characterized. Methods: Human adipose-derived MSCs were seeded and cultured on a 3D membrane in the VITVO® bioreactor system for 7 days. Afterwards, MSC-EVs were isolated and characterized via fluorescence nanoparticle tracking analysis, flow cytometry with staining against annexin V (Anx5) as a marker for EVs exposing phosphatidylserine, as well as CD73 and CD90 as MSC surface markers. Results: Cultivation of MSC in the VITVO® bioreactor system demonstrated a higher concentration of MSC-EVs from the 3D bioreactor (9.1 × 10 9 ± 1.5 × 10 9 and 9.7 × 10 9 ± 3.1 × 10 9 particles/mL) compared to static 2D culture (4.2 × 10 9 ± 7.5 × 10 8 and 3.9 × 10 9 ± 3.0 × 10 8 particles/mL) under normoxic and hypoxic conditions, respectively. Also, the particle-to-protein ratio as a measure for the purity of EVs increased from 3.3 × 10 7 ± 1.1 × 10 7 particles/µg protein in 2D to 1.6 × 10 8 ± 8.3 × 10 6 particles/µg protein in 3D. Total MSC-EVs as well as CD73 −CD90 + MSC-EVs were elevated in 2D normoxic conditions. The EV concentration and size did not differ significantly between normoxic and hypoxic conditions. Conclusion: The production of MSC-EVs in a 3D bioreactor system under hypoxic conditions resulted in increased EV concentration and purity. This system could be especially useful in screening culture conditions for the production of 3D-derived MSC-EVs.
Keywords
- 3D cell culture, Bioreactors, Extracellular vesicles, Hypoxia, Mesenchymal stem cells
ASJC Scopus subject areas
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
- Chemical Engineering(all)
- Bioengineering
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
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In: Biotechnology letters, Vol. 46, No. 2, 04.2024, p. 279–293.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Dynamic cultivation of human mesenchymal stem/stromal cells for the production of extracellular vesicles in a 3D bioreactor system
AU - Almeria, Ciarra
AU - Weiss, René
AU - Keck, Maike
AU - Weber, Viktoria
AU - Kasper, Cornelia
AU - Egger, Dominik
N1 - Funding Open Access funding enabled and organized by Projekt DEAL. The authors declare that no funds, grants, or other support were received during the preparation of this manuscript.
PY - 2024/4
Y1 - 2024/4
N2 - Purpose: 3D cell culture and hypoxia have been demonstrated to increase the therapeutic effects of mesenchymal stem/stromal cells (MSCs)-derived extracellular vesicles (EVs). In this study, a process for the production of MSC-EVs in a novel 3D bioreactor system under normoxic and hypoxic conditions was established and the resulting EVs were characterized. Methods: Human adipose-derived MSCs were seeded and cultured on a 3D membrane in the VITVO® bioreactor system for 7 days. Afterwards, MSC-EVs were isolated and characterized via fluorescence nanoparticle tracking analysis, flow cytometry with staining against annexin V (Anx5) as a marker for EVs exposing phosphatidylserine, as well as CD73 and CD90 as MSC surface markers. Results: Cultivation of MSC in the VITVO® bioreactor system demonstrated a higher concentration of MSC-EVs from the 3D bioreactor (9.1 × 10 9 ± 1.5 × 10 9 and 9.7 × 10 9 ± 3.1 × 10 9 particles/mL) compared to static 2D culture (4.2 × 10 9 ± 7.5 × 10 8 and 3.9 × 10 9 ± 3.0 × 10 8 particles/mL) under normoxic and hypoxic conditions, respectively. Also, the particle-to-protein ratio as a measure for the purity of EVs increased from 3.3 × 10 7 ± 1.1 × 10 7 particles/µg protein in 2D to 1.6 × 10 8 ± 8.3 × 10 6 particles/µg protein in 3D. Total MSC-EVs as well as CD73 −CD90 + MSC-EVs were elevated in 2D normoxic conditions. The EV concentration and size did not differ significantly between normoxic and hypoxic conditions. Conclusion: The production of MSC-EVs in a 3D bioreactor system under hypoxic conditions resulted in increased EV concentration and purity. This system could be especially useful in screening culture conditions for the production of 3D-derived MSC-EVs.
AB - Purpose: 3D cell culture and hypoxia have been demonstrated to increase the therapeutic effects of mesenchymal stem/stromal cells (MSCs)-derived extracellular vesicles (EVs). In this study, a process for the production of MSC-EVs in a novel 3D bioreactor system under normoxic and hypoxic conditions was established and the resulting EVs were characterized. Methods: Human adipose-derived MSCs were seeded and cultured on a 3D membrane in the VITVO® bioreactor system for 7 days. Afterwards, MSC-EVs were isolated and characterized via fluorescence nanoparticle tracking analysis, flow cytometry with staining against annexin V (Anx5) as a marker for EVs exposing phosphatidylserine, as well as CD73 and CD90 as MSC surface markers. Results: Cultivation of MSC in the VITVO® bioreactor system demonstrated a higher concentration of MSC-EVs from the 3D bioreactor (9.1 × 10 9 ± 1.5 × 10 9 and 9.7 × 10 9 ± 3.1 × 10 9 particles/mL) compared to static 2D culture (4.2 × 10 9 ± 7.5 × 10 8 and 3.9 × 10 9 ± 3.0 × 10 8 particles/mL) under normoxic and hypoxic conditions, respectively. Also, the particle-to-protein ratio as a measure for the purity of EVs increased from 3.3 × 10 7 ± 1.1 × 10 7 particles/µg protein in 2D to 1.6 × 10 8 ± 8.3 × 10 6 particles/µg protein in 3D. Total MSC-EVs as well as CD73 −CD90 + MSC-EVs were elevated in 2D normoxic conditions. The EV concentration and size did not differ significantly between normoxic and hypoxic conditions. Conclusion: The production of MSC-EVs in a 3D bioreactor system under hypoxic conditions resulted in increased EV concentration and purity. This system could be especially useful in screening culture conditions for the production of 3D-derived MSC-EVs.
KW - 3D cell culture
KW - Bioreactors
KW - Extracellular vesicles
KW - Hypoxia
KW - Mesenchymal stem cells
UR - http://www.scopus.com/inward/record.url?scp=85185116181&partnerID=8YFLogxK
U2 - 10.1007/s10529-024-03465-4
DO - 10.1007/s10529-024-03465-4
M3 - Article
VL - 46
SP - 279
EP - 293
JO - Biotechnology letters
JF - Biotechnology letters
SN - 0141-5492
IS - 2
ER -