Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells
Extracellular vesicles (EVs) have been recently identified as vital components of cell-based therapies based on the observation that conditioned media from cultured stromal cells reproduce some of the beneficial effects of intact cells. In order to obtain clinically active EVs derived from Mesenchym...
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MDPI AG
2020-02-01
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| Series: | Cells |
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| Online Access: | https://www.mdpi.com/2073-4409/9/2/422 |
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| author | Maria Antònia Forteza-Genestra Miquel Antich-Rosselló Javier Calvo Antoni Gayà Marta Monjo Joana Maria Ramis |
| author_facet | Maria Antònia Forteza-Genestra Miquel Antich-Rosselló Javier Calvo Antoni Gayà Marta Monjo Joana Maria Ramis |
| author_sort | Maria Antònia Forteza-Genestra |
| collection | DOAJ |
| description | Extracellular vesicles (EVs) have been recently identified as vital components of cell-based therapies based on the observation that conditioned media from cultured stromal cells reproduce some of the beneficial effects of intact cells. In order to obtain clinically active EVs derived from Mesenchymal Stromal Cells (MSCs) different procedures have been reported in the literature. Usually, non-confluent cells are incubated with culture medium for 48 h either with EV-depleted Fetal Bovine Serum (FBS) or without FBS. Our aim was to compare the effects of EVs isolated by ultracentrifugation from human umbilical cord MSC conditioned media obtained using these two conditions: with EV-depleted FBS (UC) or without FBS (UC<sub>w/o</sub>) on the mRNA expression levels of extracellular matrix related genes using the mouse chondrogenic cell line ATDC-5. We observed a deleterious effect on chondrogenic cells treated with UC<sub>w/o</sub>, showing higher mRNA expression levels of different metalloproteinases and decorin (<i>Dcn</i>) and lower collagen (<i>Col1a1</i> and <i>Col2a1</i>) and aggrecan (<i>Acan</i>) mRNA levels. To elucidate whether this deleterious effect was induced by the EVs or by any proteins co-purified in the EV pellet, we used size exclusion chromatography (SEC) to further purify the EV pellet, obtaining an EV enriched fraction (EV or EV<sub>w/o</sub>) and a protein enriched fraction (Prot or Prot<sub>w/o</sub>). Our results pointed that the negative effect on the chondrogenic cell line was due to the contaminant proteins coisolated with the EVs by ultracentrifugation and not from the EVs themselves. Thus, these results highlight the importance of working with well purified EV preparations to specifically achieve their therapeutic effect. |
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| last_indexed | 2024-03-12T08:28:25Z |
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| spelling | doaj.art-100265b603c9490d8a153090ca21ee022023-09-02T17:56:02ZengMDPI AGCells2073-44092020-02-019242210.3390/cells9020422cells9020422Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal CellsMaria Antònia Forteza-Genestra0Miquel Antich-Rosselló1Javier Calvo2Antoni Gayà3Marta Monjo4Joana Maria Ramis5Cell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainCell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainCell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainCell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainCell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainCell Therapy and Tissue Engineering Group, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Ctra Valldemossa km 7.5, 07122 Palma, SpainExtracellular vesicles (EVs) have been recently identified as vital components of cell-based therapies based on the observation that conditioned media from cultured stromal cells reproduce some of the beneficial effects of intact cells. In order to obtain clinically active EVs derived from Mesenchymal Stromal Cells (MSCs) different procedures have been reported in the literature. Usually, non-confluent cells are incubated with culture medium for 48 h either with EV-depleted Fetal Bovine Serum (FBS) or without FBS. Our aim was to compare the effects of EVs isolated by ultracentrifugation from human umbilical cord MSC conditioned media obtained using these two conditions: with EV-depleted FBS (UC) or without FBS (UC<sub>w/o</sub>) on the mRNA expression levels of extracellular matrix related genes using the mouse chondrogenic cell line ATDC-5. We observed a deleterious effect on chondrogenic cells treated with UC<sub>w/o</sub>, showing higher mRNA expression levels of different metalloproteinases and decorin (<i>Dcn</i>) and lower collagen (<i>Col1a1</i> and <i>Col2a1</i>) and aggrecan (<i>Acan</i>) mRNA levels. To elucidate whether this deleterious effect was induced by the EVs or by any proteins co-purified in the EV pellet, we used size exclusion chromatography (SEC) to further purify the EV pellet, obtaining an EV enriched fraction (EV or EV<sub>w/o</sub>) and a protein enriched fraction (Prot or Prot<sub>w/o</sub>). Our results pointed that the negative effect on the chondrogenic cell line was due to the contaminant proteins coisolated with the EVs by ultracentrifugation and not from the EVs themselves. Thus, these results highlight the importance of working with well purified EV preparations to specifically achieve their therapeutic effect.https://www.mdpi.com/2073-4409/9/2/422extracellular vesiclesultracentrifugationsize exclusion chromatographyatdc-5 cell linegene expressioncollagenhuman umbilical cord mesenchymal stromal cellsfbsconditioned mediapurity |
| spellingShingle | Maria Antònia Forteza-Genestra Miquel Antich-Rosselló Javier Calvo Antoni Gayà Marta Monjo Joana Maria Ramis Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells Cells extracellular vesicles ultracentrifugation size exclusion chromatography atdc-5 cell line gene expression collagen human umbilical cord mesenchymal stromal cells fbs conditioned media purity |
| title | Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells |
| title_full | Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells |
| title_fullStr | Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells |
| title_full_unstemmed | Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells |
| title_short | Purity Determines the Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells |
| title_sort | purity determines the effect of extracellular vesicles derived from mesenchymal stromal cells |
| topic | extracellular vesicles ultracentrifugation size exclusion chromatography atdc-5 cell line gene expression collagen human umbilical cord mesenchymal stromal cells fbs conditioned media purity |
| url | https://www.mdpi.com/2073-4409/9/2/422 |
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