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Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells

BACKGROUND: Synovial mesenchymal stem cells (MSCs) are an attractive cell source for cartilage and meniscus regeneration. The optimum cryopreservation medium has not been determined, but dimethylsulfoxide (DMSO) should be excluded, if possible, because of its toxicity. The purposes of our study were...

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Autores principales: Fujisawa, Ryota, Mizuno, Mitsuru, Katano, Hisako, Otabe, Koji, Ozeki, Nobutake, Tsuji, Kunikazu, Koga, Hideyuki, Sekiya, Ichiro
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612159/
https://www.ncbi.nlm.nih.gov/pubmed/31279341
http://dx.doi.org/10.1186/s12891-019-2700-3
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author Fujisawa, Ryota
Mizuno, Mitsuru
Katano, Hisako
Otabe, Koji
Ozeki, Nobutake
Tsuji, Kunikazu
Koga, Hideyuki
Sekiya, Ichiro
author_facet Fujisawa, Ryota
Mizuno, Mitsuru
Katano, Hisako
Otabe, Koji
Ozeki, Nobutake
Tsuji, Kunikazu
Koga, Hideyuki
Sekiya, Ichiro
author_sort Fujisawa, Ryota
collection PubMed
description BACKGROUND: Synovial mesenchymal stem cells (MSCs) are an attractive cell source for cartilage and meniscus regeneration. The optimum cryopreservation medium has not been determined, but dimethylsulfoxide (DMSO) should be excluded, if possible, because of its toxicity. The purposes of our study were to examine the possible benefits of higher concentrations of serum and the effectiveness of 100% serum (without DMSO) for the cryopreservation of synovial MSCs. METHODS: Human synovium was harvested from the knees of four donors with osteoarthritis during total knee arthroplasty. Synovial MSCs (8 × 10(5) cells) were suspended in 400 μL medium and used as a Time 0 control. The same number of synovial MSCs was also suspended in 400 μL α-MEM medium containing 10% fetal bovine serum (FBS) (5% DMSO, and 1% antibiotic), 95% FBS (and 5% DMSO), or 100% FBS (no DMSO) and cryopreserved at − 80 °C for 7 days. After thawing, the cell suspensions (1.5 μL; 3 × 10(3) cells) were cultured in 60 cm(2) dishes for 14 days for colony formation assays. Additional 62.5 μL samples of cell suspensions (1.25 × 10(5) cells) were added to tubes and cultured for 21 days for chondrogenesis assays. RESULTS: Colony numbers were significantly higher in the Time 0 and 95% FBS groups than in the 10% FBS group (n = 24). Colony numbers were much lower in the 100% FBS group than in the other three groups. The cell numbers per dish reflected the colony numbers. Cartilage pellet weights were significantly heavier in the 95% FBS group than in the 10% FBS group, whereas no difference was observed between the Time 0 and the 95% FBS groups (n = 24). No cartilage pellets formed at all in the 100% FBS group. CONCLUSION: Synovial MSCs cryopreserved in 95% FBS with 5% DMSO maintained their colony formation and chondrogenic abilities to the same levels as observed in the cells before cryopreservation. Synovial MSCs cryopreserved in 100% FBS lost their colony formation and chondrogenic abilities. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12891-019-2700-3) contains supplementary material, which is available to authorized users.
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spelling pubmed-66121592019-07-16 Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells Fujisawa, Ryota Mizuno, Mitsuru Katano, Hisako Otabe, Koji Ozeki, Nobutake Tsuji, Kunikazu Koga, Hideyuki Sekiya, Ichiro BMC Musculoskelet Disord Research Article BACKGROUND: Synovial mesenchymal stem cells (MSCs) are an attractive cell source for cartilage and meniscus regeneration. The optimum cryopreservation medium has not been determined, but dimethylsulfoxide (DMSO) should be excluded, if possible, because of its toxicity. The purposes of our study were to examine the possible benefits of higher concentrations of serum and the effectiveness of 100% serum (without DMSO) for the cryopreservation of synovial MSCs. METHODS: Human synovium was harvested from the knees of four donors with osteoarthritis during total knee arthroplasty. Synovial MSCs (8 × 10(5) cells) were suspended in 400 μL medium and used as a Time 0 control. The same number of synovial MSCs was also suspended in 400 μL α-MEM medium containing 10% fetal bovine serum (FBS) (5% DMSO, and 1% antibiotic), 95% FBS (and 5% DMSO), or 100% FBS (no DMSO) and cryopreserved at − 80 °C for 7 days. After thawing, the cell suspensions (1.5 μL; 3 × 10(3) cells) were cultured in 60 cm(2) dishes for 14 days for colony formation assays. Additional 62.5 μL samples of cell suspensions (1.25 × 10(5) cells) were added to tubes and cultured for 21 days for chondrogenesis assays. RESULTS: Colony numbers were significantly higher in the Time 0 and 95% FBS groups than in the 10% FBS group (n = 24). Colony numbers were much lower in the 100% FBS group than in the other three groups. The cell numbers per dish reflected the colony numbers. Cartilage pellet weights were significantly heavier in the 95% FBS group than in the 10% FBS group, whereas no difference was observed between the Time 0 and the 95% FBS groups (n = 24). No cartilage pellets formed at all in the 100% FBS group. CONCLUSION: Synovial MSCs cryopreserved in 95% FBS with 5% DMSO maintained their colony formation and chondrogenic abilities to the same levels as observed in the cells before cryopreservation. Synovial MSCs cryopreserved in 100% FBS lost their colony formation and chondrogenic abilities. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12891-019-2700-3) contains supplementary material, which is available to authorized users. BioMed Central 2019-07-06 /pmc/articles/PMC6612159/ /pubmed/31279341 http://dx.doi.org/10.1186/s12891-019-2700-3 Text en © The Author(s). 2019 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research Article
Fujisawa, Ryota
Mizuno, Mitsuru
Katano, Hisako
Otabe, Koji
Ozeki, Nobutake
Tsuji, Kunikazu
Koga, Hideyuki
Sekiya, Ichiro
Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title_full Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title_fullStr Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title_full_unstemmed Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title_short Cryopreservation in 95% serum with 5% DMSO maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
title_sort cryopreservation in 95% serum with 5% dmso maintains colony formation and chondrogenic abilities in human synovial mesenchymal stem cells
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612159/
https://www.ncbi.nlm.nih.gov/pubmed/31279341
http://dx.doi.org/10.1186/s12891-019-2700-3
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