Short-Term Evaluation of Cellular Fate in an Ovine Bone Formation Model
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animal Experiments
2.2. Selection of STRO-4-Positive MSCs
2.3. CM-Dil Labelling
2.4. MNP Labelling of STRO-4-Positive oMSCs
2.5. Encapsulation of oMSCs within an ECM Gel Construct for In Vivo Delivery
2.6. Assessment of Cellular Viability by LDH Staining
2.7. Histology
2.8. Immunocytochemistry
2.9. Quantification of C-Reactive Protein Levels by ELISA
2.10. Statistical Analysis
3. Results
3.1. Implants Remained Intact with No Degradation Observed at Either 2 or 7 Days Post-Implantation
3.2. Significant Loss in Cell Viability Determined at 7 Days Post-Implantation
3.3. No Adverse Inflammatory Response and No Elevation in Non-Haem Iron Level Detected at Days 2 and 7
3.4. Cellular Infiltration within the ECM Construct Is Observed for All Groups
3.5. Osteocalcin Expression by Implanted Cells Is Observed in Day 7 ECM Constructs from All Three Groups
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Group | Cells | MNPs | Magnet | CM-Dil Stain | Number of Defects | Time Point |
---|---|---|---|---|---|---|
1 (MNPs + magnet) | + | + | + | Yes | 6 | 2 days |
2 (Cell only) | + | − | − | Yes | 6 | 2 days |
3 (MNPs − magnet) | + | + | − | Yes | 6 | 2 days |
4 (MNPs + magnet) | + | + | + | Yes | 6 | 7 days |
5 (Cell only) | + | − | − | Yes | 6 | 7 days |
6 (MNPs − magnet) | + | + | − | Yes | 6 | 7 days |
Primary Antibody | Product Code | Secondary Antibody |
---|---|---|
Anti-Stro-4 (20 µg/mL) | Gift from Professor Andrew Zannettino | Invitrogen, A21236 (10 µg/mL) |
Anti-CD45 (10 µg/mL) | WS0544B-100 (Kingfisher Biotech) | Invitrogen, A21236 (10 µg/mL) |
Anti-dextran (1 µg/mL) | 60026 (Stemcell Technologies) | Invitrogen, A21236 (10 µg/mL) |
Anti-osteocalcin | ab13420 (Abcam) | Invitrogen, A21236 (10 µg/mL) |
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Markides, H.; Foster, N.C.; McLaren, J.S.; Hopkins, T.; Black, C.; Oreffo, R.O.C.; Scammell, B.E.; Echevarria, I.; White, L.J.; El Haj, A.J. Short-Term Evaluation of Cellular Fate in an Ovine Bone Formation Model. Cells 2021, 10, 1776. https://doi.org/10.3390/cells10071776
Markides H, Foster NC, McLaren JS, Hopkins T, Black C, Oreffo ROC, Scammell BE, Echevarria I, White LJ, El Haj AJ. Short-Term Evaluation of Cellular Fate in an Ovine Bone Formation Model. Cells. 2021; 10(7):1776. https://doi.org/10.3390/cells10071776
Chicago/Turabian StyleMarkides, Hareklea, Nicola C. Foster, Jane S. McLaren, Timothy Hopkins, Cameron Black, Richard O. C. Oreffo, Brigitte E. Scammell, Iria Echevarria, Lisa J. White, and Alicia J. El Haj. 2021. "Short-Term Evaluation of Cellular Fate in an Ovine Bone Formation Model" Cells 10, no. 7: 1776. https://doi.org/10.3390/cells10071776
APA StyleMarkides, H., Foster, N. C., McLaren, J. S., Hopkins, T., Black, C., Oreffo, R. O. C., Scammell, B. E., Echevarria, I., White, L. J., & El Haj, A. J. (2021). Short-Term Evaluation of Cellular Fate in an Ovine Bone Formation Model. Cells, 10(7), 1776. https://doi.org/10.3390/cells10071776