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ToF-SIMS study of differentiation of human bone-derived stromal cells: new insights into osteoporosis

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Abstract

Lipids have numerous important functions in the human body, as they form the cells’ plasma membranes and play a key role in many disease states, presumably also in osteoporosis. Here, the fatty acid composition of the outer plasma membranes of cells differentiated into the osteogenic and adipogenic direction is studied with surface-sensitive time-of-flight secondary ion mass spectrometry (ToF-SIMS). For data evaluation, principal component analysis (PCA) is applied. Human (bone-derived) mesenchymal stromal cells (hMSCs) from an osteoporotic donor and a control donor are compared to reveal differences in the fatty acid composition of the membranes. The chemical information is correlated to staining and real-time quantitative polymerase chain reaction (rt-qPCR) results to provide insight into the gene expression of several differentiation markers on the RNA level. Adipogenic differentiation of hMSCs from a non-osteoporotic donor correlates with increased relative intensities of all fatty acids under investigation. After osteogenic differentiation of non-osteoporotic cells, the relative mass signal intensities of unsaturated fatty acids such as oleic and linoleic acids are increased. However, the osteoporotic cells show increased levels of palmitic acid in the plasma membrane after exposure to osteogenic differentiation conditions, which correlates to an immature differentiation state relative to non-osteoporotic osteogenic cells. This immature differentiation state is confirmed by increased early osteogenic differentiation factor Runx2 on RNA level and by less calcium mineralization spots seen in von Kossa staining and ToF-SIMS images.

Time-of-flight secondary ion mass spectrometry is applied to analyze the fatty acid composition of the outer plasma membranes of cells differentiated into the adipogenic and osteogenic direction. Cells from an osteoporotic and a control donor are compared to reveal differences due to differentiation and disease stage of the cells.

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Acknowledgments

This study was funded by the Deutsche Forschungsgemeinschaft (DFG) as part of the Collaborative Research Centre Transregio 79 (SFB/TRR 79—subproject M5 in collaboration with subproject B11). Kaija Schaepe gratefully acknowledges the financial support of the Friedrich Naumann Foundation for Freedom. We thank Svenja Otto for the help with the data analysis and Alena-Svenja Wagner and Anne Hild for many helpful discussions. The authors thank Dan Graham, Ph.D., for developing the NESAC/BIO Toolbox used in this study and NIH grant EB-002027 for supporting the toolbox development.

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Authors and Affiliations

  1. Institute of Physical Chemistry, Justus Liebig University of Giessen, Heinrich-Buff-Ring 17, 35392, Giessen, Germany

    Kaija Schaepe, Tessa Bartges, Anja Henss, Marcus Rohnke & Jürgen Janek

  2. Institute of Veterinary Anatomy, Histology and Embryology, Justus Liebig University of Giessen, Frankfurter Strasse 98, 35392, Giessen, Germany

    Janina Werner, Kristina Glenske & Sabine Wenisch

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  1. Kaija Schaepe
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Correspondence to Marcus Rohnke.

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The authors declare that they have no conflict of interest.

Research involving human participants and/or animals

The experiments were approved by the local ethical committee of the Medical Faculty of the Justus Liebig University of Giessen (decision 05/06 and 106/06).

Informed consent

The femoral heads from patients, undergoing total hip arthroplasty due to primary osteoarthritis, were provided from Asklepios Clinic Lich GmbH (Lich, Germany). All patients gave their informed consent to provide the retrieved biological material for the purpose of scientific research.

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Schaepe, K., Werner, J., Glenske, K. et al. ToF-SIMS study of differentiation of human bone-derived stromal cells: new insights into osteoporosis. Anal Bioanal Chem 409, 4425–4435 (2017). https://doi.org/10.1007/s00216-017-0386-7

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