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Abstract Gene therapy has the potential to facilitate targeted expression of therapeutic proteins to promote cartilage regeneration in osteoarthritis (OA). The dense, avascular, aggrecan‐glycosaminoglycan (GAG) rich negatively charged cartilage, however, hinders their transport to reach chondrocytes in effective doses. While viral vector mediated gene delivery has shown promise, concerns over immunogenicity and tumorigenic side‐effects persist. To address these issues, this study develops surface‐modified cartilage‐targeting exosomes as non‐viral carriers for gene therapy. Charge‐reversed cationic exosomes are engineered for mRNA delivery by anchoring cartilage targeting optimally charged arginine‐rich cationic motifs into the anionic exosome bilayer by using buffer pH as a charge‐reversal switch. Cationic exosomes penetrated through the full‐thickness of early‐stage arthritic human cartilage owing to weak‐reversible ionic binding with GAGs and efficiently delivered the encapsulated eGFP mRNA to chondrocytes residing in tissue deep layers, while unmodified anionic exosomes do not. When intra‐articularly injected into destabilized medial meniscus mice knees with early‐stage OA, mRNA loaded charge‐reversed exosomes overcame joint clearance and rapidly penetrated into cartilage, creating an intra‐tissue depot and efficiently expressing eGFP; native exosomes remained unsuccessful. Cationic exosomes thus hold strong translational potential as a platform technology for cartilage‐targeted non‐viral delivery of any relevant mRNA targets for OA treatment.
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Anionic polymers amplify electrokinetic perfusion through extracellular matrices
Electrical stimulation (ES) promotes healing of chronic epidermal wounds and delays degeneration of articular cartilage. Despite electrotherapeutic treatment of these non-excitable tissues, the mechanisms by which ES promotes repair are unknown. We hypothesize that a beneficial role of ES is dependent on electrokinetic perfusion in the extracellular space and that it mimics the effects of interstitial flow. In vivo , the extracellular space contains mixtures of extracellular proteins and negatively charged glycosaminoglycans and proteoglycans surrounding cells. While these anionic macromolecules promote water retention and increase mechanical support under compression, in the presence of ES they should also enhance electro-osmotic flow (EOF) to a greater extent than proteins alone. To test this hypothesis, we compare EOF rates between artificial matrices of gelatin (denatured collagen) with matrices of gelatin mixed with anionic polymers to mimic endogenous charged macromolecules. We report that addition of anionic polymers amplifies EOF and that a matrix comprised of 0.5% polyacrylate and 1.5% gelatin generates EOF with similar rates to those reported in cartilage. The enhanced EOF reduces mortality of cells at lower applied voltage compared to gelatin matrices alone. We also use modeling to describe the range of thermal changes that occur during these electrokinetic experiments and during electrokinetic perfusion of soft tissues. We conclude that the negative charge density of native extracellular matrices promotes electrokinetic perfusion during electrical therapies in soft tissues and may promote survival of artificial tissues and organs prior to vascularization and during transplantation.
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- Award ID(s):
- 2033522
- PAR ID:
- 10409957
- Date Published:
- Journal Name:
- Frontiers in Bioengineering and Biotechnology
- Volume:
- 10
- ISSN:
- 2296-4185
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3389/fbioe.2022.983317
