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Theranostics 2023; 13(5):1470-1489. doi:10.7150/thno.77597 This issue Cite

Research Paper

Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro

Maria Elisabetta F. Palamà1, Simona Coco2, Georgina M. Shaw3, Daniele Reverberi4, Maddalena Ghelardoni1, Paola Ostano5, Giovanna Chiorino5, Laura Sercia6, Luana Persano6, Maria Cristina Gagliani1, Katia Cortese1, Dario Pisignano6,7, Josephine Mary Murphy3, Chiara Gentili1✉

1. Department of Experimental Medicine, University of Genoa, Genoa, Italy
2. Lung Cancer Unit, IRCCS Ospedale Policlinico San Martino, Genoa, Italy
3. Regenerative Medicine Institute, National University of Ireland Galway, Galway, Ireland
4. U.O. Molecular Pathology, IRCCS Ospedale Policlinico San Martino, Genoa, Italy
5. Cancer Genomics Lab, Fondazione Edo ed Elvo Tempia, Biella, Italy
6. Institute of Nanoscience (CNR-NANO), Pisa, Italy
7. Department of Physics, University of Pisa, Pisa, Italy

✉ Corresponding author: Prof. Chiara Gentili. chiara.gentiliit

Citation:
Palamà MEF, Coco S, Shaw GM, Reverberi D, Ghelardoni M, Ostano P, Chiorino G, Sercia L, Persano L, Gagliani MC, Cortese K, Pisignano D, Murphy JM, Gentili C. Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro. Theranostics 2023; 13(5):1470-1489. doi:10.7150/thno.77597. https://www.thno.org/v13p1470.htm
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Abstract

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Rationale: Mesenchymal stromal cells (MSCs)-derived extracellular vesicles (EVs) emerged as an innovative strategy for the treatment of chronic disorders such as osteoarthritis (OA). Biological activity of EVs is generally driven by their cargo, which might be influenced by microenvironment. Therefore, pre-conditioning strategies, including modifications in culture conditions or oxygen tension could directly impact on MSCs paracrine activity. In this study we selected an appropriate preconditioning system to induce cells to perform the most suitable therapeutic response by EV-encapsulated bioactive factors.

Methods: A xeno-free supplement (XFS) was used for isolation and expansion of MSCs and compared to conventional fetal bovine serum (FBS) culture. Bone Marrow-derived MSCs (BMSCs) were pre-conditioned under normoxia (20% O2) or under hypoxia (1% O2) and EVs production was evaluated. Anti-OA activity was evaluated by using an in vitro inflammatory model. miRNA content was also explored, to select putative miRNA that could be involved in a biological function.

Results: Modulation of IL-6, IL-8, COX-2 and PGE2 was evaluated on hACs simultaneously treated with IL-1α and BMSC-derived EVs. FBS-sEVs exerted a blunt inhibitory effect, while a strong anti-inflammatory outcome was achieved by XFS-sEVs. Interestingly, in both cases hypoxia pre-conditioning allowed to increase EVs effectiveness. Analysis of miRNA content showed the upregulation in XFS-hBMSC-derived EVs of miRNA known to have a chondroprotective role, such as let-7b-5p, miR-17, miR-145, miR-21-5p, miR-214-3p, miR-30b-5p, miR-30c-5p. Activated pathways and target genes were investigated in silico and upregulated miRNAs functionally validated in target cells. MiR-145 and miR-214 were found to protect chondrocytes from IL-1α-induced inflammation and to reduce production of pro-inflammatory cytokines.

Conclusions: XFS medium was found to be suitable for isolation and expansion of MSCs, secreting EVs with a therapeutic cargo. The application of cells cultured exclusively in XFS overcomes issues of safety associated with serum-containing media and makes ready-to-use clinical therapies more accessible.

Keywords: extracellular vesicles, osteoarthritis, mesenchymal stromal cells, microRNA, xeno free medium

Citation styles

APA
Palamà, M.E.F., Coco, S., Shaw, G.M., Reverberi, D., Ghelardoni, M., Ostano, P., Chiorino, G., Sercia, L., Persano, L., Gagliani, M.C., Cortese, K., Pisignano, D., Murphy, J.M., Gentili, C. (2023). Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro. Theranostics, 13(5), 1470-1489. https://doi.org/10.7150/thno.77597.

ACS
Palamà, M.E.F.; Coco, S.; Shaw, G.M.; Reverberi, D.; Ghelardoni, M.; Ostano, P.; Chiorino, G.; Sercia, L.; Persano, L.; Gagliani, M.C.; Cortese, K.; Pisignano, D.; Murphy, J.M.; Gentili, C. Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro. Theranostics 2023, 13 (5), 1470-1489. DOI: 10.7150/thno.77597.

NLM
Palamà MEF, Coco S, Shaw GM, Reverberi D, Ghelardoni M, Ostano P, Chiorino G, Sercia L, Persano L, Gagliani MC, Cortese K, Pisignano D, Murphy JM, Gentili C. Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro. Theranostics 2023; 13(5):1470-1489. doi:10.7150/thno.77597. https://www.thno.org/v13p1470.htm

CSE
Palamà MEF, Coco S, Shaw GM, Reverberi D, Ghelardoni M, Ostano P, Chiorino G, Sercia L, Persano L, Gagliani MC, Cortese K, Pisignano D, Murphy JM, Gentili C. 2023. Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a “therapeutic” miRNA cargo ameliorating cartilage inflammation in vitro. Theranostics. 13(5):1470-1489.

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