Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells
Mesenchymal stem cells (MSCs) are progenitors for bone-forming osteoblasts and lipid-storing adipocytes, two major lineages co-existing in bone marrow. When isolated in vitro, these stem cells recapitulate osteoblast or adipocyte formation if treated with specialised media, modelling how these linea...
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| Format: | Article |
| Language: | English |
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MDPI AG
2019-07-01
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| Series: | Biomimetics |
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| Online Access: | https://www.mdpi.com/2313-7673/4/3/48 |
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| author | Nigel De Melo Sarah McGinlay Robert Markus Laura Macri-Pellizzeri Michael E. Symonds Ifty Ahmed Virginie Sottile |
| author_facet | Nigel De Melo Sarah McGinlay Robert Markus Laura Macri-Pellizzeri Michael E. Symonds Ifty Ahmed Virginie Sottile |
| author_sort | Nigel De Melo |
| collection | DOAJ |
| description | Mesenchymal stem cells (MSCs) are progenitors for bone-forming osteoblasts and lipid-storing adipocytes, two major lineages co-existing in bone marrow. When isolated in vitro, these stem cells recapitulate osteoblast or adipocyte formation if treated with specialised media, modelling how these lineages interact in vivo. Osteogenic differentiation is characterised by mineral deposits accumulating in the extracellular matrix, typically assessed using histological techniques. Adipogenesis occurs with accumulation of intracellular lipids that can be routinely visualised by Oil Red O staining. In both cases, staining requires cell fixation and is thus limited to end-point assessments. Here, a vital staining approach was developed to simultaneously detect mineral deposits and lipid droplets in differentiating cultures. Stem cells induced to differentiate produced mixed cultures containing adipocytes and bone-like nodules, and after two weeks live cultures were incubated with tetracycline hydrochloride and Bodipy to label mineral- and lipid-containing structures, respectively. Fluorescence microscopy showed the simultaneous visualisation of mineralised areas and lipid-filled adipocytes in live cultures. Combined with the nuclear stain Hoechst 33258, this approach further enabled live confocal imaging of adipogenic cells interspersed within the mineralised matrix. This multiplex labelling was repeated at subsequent time-points, demonstrating the potential of this new approach for the real-time high-precision imaging of live stem cells. |
| first_indexed | 2024-12-13T04:42:20Z |
| format | Article |
| id | doaj.art-e5e260529cfc47d1ab8104446dce2a71 |
| institution | Directory Open Access Journal |
| issn | 2313-7673 |
| language | English |
| last_indexed | 2024-12-13T04:42:20Z |
| publishDate | 2019-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biomimetics |
| spelling | doaj.art-e5e260529cfc47d1ab8104446dce2a712022-12-21T23:59:18ZengMDPI AGBiomimetics2313-76732019-07-01434810.3390/biomimetics4030048biomimetics4030048Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem CellsNigel De Melo0Sarah McGinlay1Robert Markus2Laura Macri-Pellizzeri3Michael E. Symonds4Ifty Ahmed5Virginie Sottile6Wolfson STEM Centre, School of Medicine, The University of Nottingham, Nottingham NG7 2RD, UKWolfson STEM Centre, School of Medicine, The University of Nottingham, Nottingham NG7 2RD, UKSchool of Life Sciences, University of Nottingham, Nottingham NG7 2RD, UKWolfson STEM Centre, School of Medicine, The University of Nottingham, Nottingham NG7 2RD, UKThe Early Life Research Unit, Division of Child Health, School of Medicine, The University of Nottingham, Nottingham NG7 2RD, UKAdvanced Materials Group, Faculty of Engineering, The University of Nottingham, Nottingham NG7 2RD, UKWolfson STEM Centre, School of Medicine, The University of Nottingham, Nottingham NG7 2RD, UKMesenchymal stem cells (MSCs) are progenitors for bone-forming osteoblasts and lipid-storing adipocytes, two major lineages co-existing in bone marrow. When isolated in vitro, these stem cells recapitulate osteoblast or adipocyte formation if treated with specialised media, modelling how these lineages interact in vivo. Osteogenic differentiation is characterised by mineral deposits accumulating in the extracellular matrix, typically assessed using histological techniques. Adipogenesis occurs with accumulation of intracellular lipids that can be routinely visualised by Oil Red O staining. In both cases, staining requires cell fixation and is thus limited to end-point assessments. Here, a vital staining approach was developed to simultaneously detect mineral deposits and lipid droplets in differentiating cultures. Stem cells induced to differentiate produced mixed cultures containing adipocytes and bone-like nodules, and after two weeks live cultures were incubated with tetracycline hydrochloride and Bodipy to label mineral- and lipid-containing structures, respectively. Fluorescence microscopy showed the simultaneous visualisation of mineralised areas and lipid-filled adipocytes in live cultures. Combined with the nuclear stain Hoechst 33258, this approach further enabled live confocal imaging of adipogenic cells interspersed within the mineralised matrix. This multiplex labelling was repeated at subsequent time-points, demonstrating the potential of this new approach for the real-time high-precision imaging of live stem cells.https://www.mdpi.com/2313-7673/4/3/48live monitoringstem cellmineralisationlipid detection |
| spellingShingle | Nigel De Melo Sarah McGinlay Robert Markus Laura Macri-Pellizzeri Michael E. Symonds Ifty Ahmed Virginie Sottile Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells Biomimetics live monitoring stem cell mineralisation lipid detection |
| title | Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells |
| title_full | Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells |
| title_fullStr | Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells |
| title_full_unstemmed | Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells |
| title_short | Live Simultaneous Monitoring of Mineral Deposition and Lipid Accumulation in Differentiating Stem Cells |
| title_sort | live simultaneous monitoring of mineral deposition and lipid accumulation in differentiating stem cells |
| topic | live monitoring stem cell mineralisation lipid detection |
| url | https://www.mdpi.com/2313-7673/4/3/48 |
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