Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage
Abstract Extracellular vesicles (EVs) are widely investigated for their implications in cell-cell signaling, immune modulation, disease pathogenesis, cancer, regenerative medicine, and as a potential drug delivery vector. However, maintaining integrity and bioactivity of EVs between Good Manufacturi...
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BMC
2025-01-01
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| Series: | Journal of Biological Engineering |
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| Online Access: | https://doi.org/10.1186/s13036-024-00470-z |
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| author | Alan B. Dogan Spencer R. Marsh Rachel J. Tschetter Claire E. Beard Md R. Amin L. Jane Jourdan Robert G. Gourdie |
| author_facet | Alan B. Dogan Spencer R. Marsh Rachel J. Tschetter Claire E. Beard Md R. Amin L. Jane Jourdan Robert G. Gourdie |
| author_sort | Alan B. Dogan |
| collection | DOAJ |
| description | Abstract Extracellular vesicles (EVs) are widely investigated for their implications in cell-cell signaling, immune modulation, disease pathogenesis, cancer, regenerative medicine, and as a potential drug delivery vector. However, maintaining integrity and bioactivity of EVs between Good Manufacturing Practice separation/filtration and end-user application remains a consistent bottleneck towards commercialization. Milk-derived extracellular vesicles (mEVs), separated from bovine milk, could provide a relatively low-cost, scalable platform for large-scale mEV production; however, the reliance on cold supply chain for storage remains a logistical and financial burden for biologics that are unstable at room temperature. Herein, we aim to characterize and engineer a freeze-dried, mEV formulation that can be stored at room temperature without sacrificing structure/bioactivity and can be reconstituted before delivery. In addition to undertaking established mEV assays of structure and function on our preparations, we introduce a novel, efficient, high throughput assay of mEV bioactivity based on Electric Cell Substrate Impedance Sensing (ECIS) in Human dermal fibroblast monolayers. By adding appropriate excipients, such as trehalose and tryptophan, we describe a protective formulation that preserves mEV bioactivity during long-term, room temperature storage. Our identification of the efficacy of tryptophan as a novel additive to mEV lyophilization solutions could represent a significant advancement in stabilizing small extracellular vesicles outside of cold storage conditions. |
| format | Article |
| id | doaj-art-8e0da3fdd1c145a3828186186e72420f |
| institution | Kabale University |
| issn | 1754-1611 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | BMC |
| record_format | Article |
| series | Journal of Biological Engineering |
| spelling | doaj-art-8e0da3fdd1c145a3828186186e72420f2025-01-19T12:26:33ZengBMCJournal of Biological Engineering1754-16112025-01-0119111610.1186/s13036-024-00470-zStabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storageAlan B. Dogan0Spencer R. Marsh1Rachel J. Tschetter2Claire E. Beard3Md R. Amin4L. Jane Jourdan5Robert G. Gourdie6Virginia Tech Carilion School of MedicineFralin Biomedical Research Institute at Virginia Tech CarilionMaterials Science and Engineering, Virginia TechFralin Biomedical Research Institute at Virginia Tech CarilionFralin Biomedical Research Institute at Virginia Tech CarilionFralin Biomedical Research Institute at Virginia Tech CarilionFralin Biomedical Research Institute at Virginia Tech CarilionAbstract Extracellular vesicles (EVs) are widely investigated for their implications in cell-cell signaling, immune modulation, disease pathogenesis, cancer, regenerative medicine, and as a potential drug delivery vector. However, maintaining integrity and bioactivity of EVs between Good Manufacturing Practice separation/filtration and end-user application remains a consistent bottleneck towards commercialization. Milk-derived extracellular vesicles (mEVs), separated from bovine milk, could provide a relatively low-cost, scalable platform for large-scale mEV production; however, the reliance on cold supply chain for storage remains a logistical and financial burden for biologics that are unstable at room temperature. Herein, we aim to characterize and engineer a freeze-dried, mEV formulation that can be stored at room temperature without sacrificing structure/bioactivity and can be reconstituted before delivery. In addition to undertaking established mEV assays of structure and function on our preparations, we introduce a novel, efficient, high throughput assay of mEV bioactivity based on Electric Cell Substrate Impedance Sensing (ECIS) in Human dermal fibroblast monolayers. By adding appropriate excipients, such as trehalose and tryptophan, we describe a protective formulation that preserves mEV bioactivity during long-term, room temperature storage. Our identification of the efficacy of tryptophan as a novel additive to mEV lyophilization solutions could represent a significant advancement in stabilizing small extracellular vesicles outside of cold storage conditions.https://doi.org/10.1186/s13036-024-00470-zMilk-derived EVsExtracellular vesiclesLyophilizationTrehaloseTryptophanRoom temperature storage |
| spellingShingle | Alan B. Dogan Spencer R. Marsh Rachel J. Tschetter Claire E. Beard Md R. Amin L. Jane Jourdan Robert G. Gourdie Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage Journal of Biological Engineering Milk-derived EVs Extracellular vesicles Lyophilization Trehalose Tryptophan Room temperature storage |
| title | Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage |
| title_full | Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage |
| title_fullStr | Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage |
| title_full_unstemmed | Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage |
| title_short | Stabilizing milk-derived extracellular vesicles (mEVs) through lyophilization: a novel trehalose and tryptophan formulation for maintaining structure and Bioactivity during long-term storage |
| title_sort | stabilizing milk derived extracellular vesicles mevs through lyophilization a novel trehalose and tryptophan formulation for maintaining structure and bioactivity during long term storage |
| topic | Milk-derived EVs Extracellular vesicles Lyophilization Trehalose Tryptophan Room temperature storage |
| url | https://doi.org/10.1186/s13036-024-00470-z |
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