Table 4 Published experimental studies detailing BM-MSCs post-thaw differentiation potential
From: The impact of cryopreservation on bone marrow-derived mesenchymal stem cells: a systematic review
Study | Species | Results post-thaw | Method of assessment |
|---|---|---|---|
Human | |||
 Bruder et al. [91] | Human | No effect on osteogenic differentiation ability | Cell re-plated for one passage post-thaw then re-plated and incubated with osteogenic supplements; quantification of alkaline phosphatase activity |
 Hirose et al. [41] | Human | No effect on osteogenic differentiation ability | Incubation with osteogenic media for 25 days; quantitative fluorescence analysis of calcein uptake |
 Kotobuki et al. [35] | Human | No effect on osteogenic differentiation ability | Incubation with osteogenic medium for 2 weeks; calcium and alkaline phosphatase activity staining |
 Kotobuki et al. [92] | Human | No effect on osteogenic differentiation ability | Incubation in osteogenic media for 2 weeks; quantification of alkaline phosphatase activity and calcein uptake |
 Xiang et al. [93] | Human | No effect on adipogenic or neuro genic differentiation ability | Cells at P15 post-thaw incubated in adipogenesis medium for 12 days; Oil Red O staining |
Cells at P15 post-thaw incubated in neurogenesis induction medium for 1 or 6Â days; fluorescent staining and RT-qPCR | |||
 Zhao et al. [94] | BM-MSC (human with chronic myeloid leukaemia) | No effect on differentiation ability | Incubation in osteogenic medium for 21 days; Von Kossa staining and RT-qPCR |
Incubation in adipogenic media for 14Â days; Oil Red staining and RT-qPCR | |||
Incubation in neurogenic medium; Immunocytochemistry and western blotting for NF, GFAP and GalC | |||
Endothelial differentiation for 2Â weeks; immunohistochemical and western blotting for CD31 and vWF | |||
 Liu et al. [28] | Human | Serum-free reduced-DMSO freezing solution gives comparable differentiation to 10% DMSO | Incubation in osteogenic or adipogenic media for 2 weeks, and chondrogenic media for 3 weeks |
 Doan et al. [95] | Human | No effect on adipogenic differentiation ability | Incubation in adipogenic medium for 2–3 weeks; Oil Red staining |
 Ginis et al. [50] | Human | No effect on osteogenic differentiation ability | Incubation with osteogenic media; quantification of alkaline phosphatase activity on days 7 and 14 after incubation as well as flow cytometry analysis at day 14 after incubation of alkaline phosphatase surface expression |
Quantification of calcium deposition at day 21 after incubation | |||
 Mamidi et al. [33] | Human | No effect on tri-lineage differentiation ability | Incubation in osteogenic differentiation media for 3 weeks; Alizarin red staining |
Incubation in adipogenic differentiation media for 3Â weeks; Oil Red O staining | |||
Incubation in chondrogenic differentiation media for 3Â weeks; Alcian blue staining | |||
 Matsumura et al. [26] | Human | No effect on tri-lineage differentiation ability | Incubation in osteogenic differentiation media for 14 days; Alizarin red staining and alkaline phosphatase activity |
Incubation in adipogenic differentiation media for 14Â days; Oil Red O staining and GPDN activity | |||
Incubation in chondrogenic differentiation media for 14Â days; Alcian blue staining | |||
 Kumazawa et al. [37] | Human | No effect on adipogenic or osteogenic differentiation ability | Incubation in osteogenic medium for 1, 2, and 3 weeks then alkaline phosphatase activity, calcium levels, alizarin red staining and RT-qPCR |
Incubation in adipogenic medium for 1, 2, and 3Â weeks; Oil Red O staining | |||
 Luetzkendorf et al. [40] | Human | No effect on adipogenic and osteogenic differentiation ability | Incubation in diff media until morphological signs of differentiation were visible (10–15 days) |
 Lechanteur et al. [34] | Human | No effect on differentiation ability | NA |
 Yuan et al. [52] | Human (BM-MSC engineered to express TRAIL) | No effect on tri-lineage differentiation ability | Differentiation procedures performed using StemPro differentiation kits according to manufacturer’s instructions |
Other species | |||
 Liu et al. [29] | Rat, mouse and calf | No effect on adipogenic or osteogenic differentiation ability | Incubation with adipogenic media for 2 weeks; Oil Red O staining and alkaline phosphatase activity expression staining with BCIP/NBT |
Incubation with adipogenic or osteogenic media for 2Â weeks then Oil Red O staining and alkaline phosphatase activity expression staining with BCIP/NBT | |||
 Naaldijk et al. [27] | Rat | In general, no difference in differentiation was observed (qualitative observation); Quantification of ALP: ALP activity is lower at ‘high’ (> 5%) levels of DMSO compared to solutions with a higher HES 450 content | Incubation with osteogenesis, adipogenesis and chondrogenic media for 2 weeks |
Quantification using alkaline phosphatase assay | |||
 Li et al. [96] | Dog | No effect on osteogenic differentiation ability | Incubation in osteogenic media for 5, 10 and 15 days; alkaline phosphatase activity measurement |
Incubation in osteogenic media for 21Â days; number of mineralized nodules determined | |||
 Zhu et al. [46] | Dog | No effect on osteogenic differentiation ability | Incubation with osteogenic media (21 days); measurement of alkaline phosphatase activity at 5, 10 and 15 days and Von Kossa staining and nodules counting at day 21 |
 Edamura et al. [36] | Dog | No effect on neurogenic differentiation ability | Incubation with neurogenic media for 6 h then RT-qPCR |
 Tokumoto et al. [48] | Monkey | Cryopreserved cells had a slightly higher ALP enzyme activity than non-cryopreserved cells | Incubation with osteogenic media for 4, 8 and 12 days; quantification of alkaline phosphatase enzyme activity |
 Nitsch et al. [97] | Monkey | No effect on adipogenic or osteogenic differentiation ability | Incubation with adipogenic media for 5 weeks; oil-red O staining |
Incubation with osteogenic medium for 21Â days; Von Kossa staining | |||
 Lauterboeck et al. [49] | Monkey | Significant decrease in oil droplet formation | Incubation with adipogenic medium for 20 days; Oil Red O staining |
No difference in osteogenic differentiation ability | Incubation with osteogenic induction medium for 3Â weeks; Alizarin red staining | ||
 Heino et al. [39] | Minipig | Cells lost their osteogenic differentiation potential | Cells incubated in osteogenic medium; stained for alkaline phosphatase activity (tested 9 days post-thaw) |