van Maanen, J.C., Bach, F.C., Snuggs, J.W. et al. (4 more authors) (2025) Explorative study of modulatory effects of notochordal cell-derived extracellular vesicles on the IL-1β-induced catabolic cascade in nucleus pulposus cell pellets and explants. JOR SPINE, 8 (1). e70043. ISSN 2572-1143
Abstract
Background
Cell-free regenerative strategies, such as notochordal cell (NC)-derived extracellular vesicles (EVs), are an attractive alternative in developing new therapies for intervertebral disc (IVD) degeneration. NC-EVs have been reported to elicit matrix anabolic effects on nucleus pulposus cells from degenerated IVDs cultured under basal conditions. However, the degenerative process is exacerbated by pro-inflammatory cytokines contributing to the vicious degenerative cycle. Therefore, this study explores whether NC-EVs modulate interleukin (IL)-1β-mediated pro-inflammatory responses in the degenerating disc.
Methods
This study utilized two IL-1β induced pro-catabolic culture models; a dog 3D nucleus pulposus (NP) cell pellet culture and a human patient-derived, ex vivo NP tissue culture system. Porcine NC-EVs were generated from NC-conditioned medium by differential centrifugation followed by size exclusion chromatography. Donor matched EV-depleted media were generated by overnight ultracentrifugation, whereafter the EV-depleted NCCM supernatant was subjected to size exclusion chromatography. To investigate whether observed effects were EV-associated, NC-EVs conditions were compared to EV-depleted controls in the absence and presence of IL-1β.
Results
The size and concentration of NC-EVs were quantified by nanoparticle tracking analysis, which showed minimal donor variation and confirmed depletion of EVs in the EV-depleted media. In the IL-1β-induced catabolic cascade, the NC-EVs did not elicit anabolic effects at the matrix level nor did they rescue the pro-catabolic phenotype within dog pellets. Modification of the CCL2 secretion seemed to be context dependent in the human explants: where EVs treatment stimulated CCL2 secretion but in the presence of IL-1β this effect was counteracted. Secretion of IL-6 and C-X-C motif chemokine ligand 1 was significantly decreased in NC-EV + IL-1β vs. control+IL-1β but not compared to EV-depleted human explant controls. Altogether, this data provides evidence for a protective modulatory role of NC-EVs. Considering the homeostatic function EVs exert, inherently encompassing subtle biologic modifications, the current study may have lacked sufficient power to demonstrate statistical significance in a sample set with evident donor variation.
Conclusions
NC-EVs may modulate the production of specific cytokines and chemokines in human degenerate explants when the key pro-inflammatory cytokine IL-1β is present. Implementation of the technical EV-depleted controls in further studies is essential to robustly demonstrate that these effects are EV-mediated and not associated with other secreted factors co-isolated during EV-isolation.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2025 The Author(s). This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes. http://creativecommons.org/licenses/by-nc/4.0/ |
Keywords: | CCL2; degeneration; dog; human explant; intervertebral disc |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Medicine, Dentistry and Health (Sheffield) > School of Medicine and Population Health |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 04 Feb 2025 11:20 |
Last Modified: | 04 Feb 2025 11:20 |
Status: | Published |
Publisher: | Wiley |
Refereed: | Yes |
Identification Number: | 10.1002/jsp2.70043 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:222816 |