One of the greatest enigmas of modern biology is how the geometry of muscular and skeletal structures are created and how their development is controlled during growth and regeneration. Scaling and shaping of vertebrate muscles and skeletal elements has always been enigmatic and required an advanced technical level in order to analyse the cell distribution in 3D. In this work, synchrotron X-ray computed microtomography (µCT) and chemical contrasting has been exploited for a quantitative analysis of the 3D-cell distribution in tissues of a developing salamander (Pleurodeles waltl) limb - a key model organism for vertebrate regeneration studies. We mapped the limb muscles, their size and shape as well as the number and density of cells within the extracellular matrix of the developing cartilage. By using tomographic approach, we explored the polarity of the cells in 3D, in relation to the structure of developing joints. We found that the polarity of chondrocytes correlates with the planes in joint surfaces and also changes along the length of the cartilaginous elements. Our approach generates data for the precise computer simulations of muscle-skeletal regeneration using cell dynamics models, which is necessary for the understanding how anisotropic growth results in the precise shapes of skeletal structures.

Central European Institute of Technology Brno University of Technology Brno Czech Republic

Department of Cellular and Molecular Biology Karolinska Institutet Solna 171777 Stockholm Sweden

Department of Physiology and Pharmacology Karolinska Institutet Solna 171777 Stockholm Sweden

Department of Physiology and Pharmacology Karolinska Institutet Solna 171777 Stockholm Sweden Department of Molecular Neurosciences Medical University Vienna Vienna Austria

Department of Physiology and Pharmacology Karolinska Institutet Solna 171777 Stockholm Sweden Department of Orthopaedics Xiangya Hospital Central South University Changsha Hunan Province China

Elettra Sincrotrone Trieste S C p A Basovizza Trieste Italy

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$a A quantitative analysis of 3D-cell distribution in regenerating muscle-skeletal system with synchrotron X-ray computed microtomography / $c M. Tesařová, L. Mancini, A. Simon, I. Adameyko, M. Kaucká, A. Elewa, G. Lanzafame, Y. Zhang, D. Kalasová, B. Szarowská, T. Zikmund, M. Novotná, J. Kaiser,

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$a One of the greatest enigmas of modern biology is how the geometry of muscular and skeletal structures are created and how their development is controlled during growth and regeneration. Scaling and shaping of vertebrate muscles and skeletal elements has always been enigmatic and required an advanced technical level in order to analyse the cell distribution in 3D. In this work, synchrotron X-ray computed microtomography (µCT) and chemical contrasting has been exploited for a quantitative analysis of the 3D-cell distribution in tissues of a developing salamander (Pleurodeles waltl) limb - a key model organism for vertebrate regeneration studies. We mapped the limb muscles, their size and shape as well as the number and density of cells within the extracellular matrix of the developing cartilage. By using tomographic approach, we explored the polarity of the cells in 3D, in relation to the structure of developing joints. We found that the polarity of chondrocytes correlates with the planes in joint surfaces and also changes along the length of the cartilaginous elements. Our approach generates data for the precise computer simulations of muscle-skeletal regeneration using cell dynamics models, which is necessary for the understanding how anisotropic growth results in the precise shapes of skeletal structures.

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$a Mancini, Lucia $u Elettra-Sincrotrone Trieste S.C.p.A., Basovizza, Trieste, Italy.

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$a Simon, Andras $u Department of Cellular and Molecular Biology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden.

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$a Adameyko, Igor $u Department of Physiology and Pharmacology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden. Department of Molecular Neurosciences, Medical University Vienna, Vienna, Austria.

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$a Kaucká, Markéta $u Department of Physiology and Pharmacology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden. Department of Molecular Neurosciences, Medical University Vienna, Vienna, Austria.

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$a Elewa, Ahmed $u Department of Cellular and Molecular Biology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden.

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$a Lanzafame, Gabriele $u Elettra-Sincrotrone Trieste S.C.p.A., Basovizza, Trieste, Italy.

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$a Zhang, Yi $u Department of Physiology and Pharmacology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden. Department of Orthopaedics, Xiangya Hospital, Central South University, Changsha, Hunan Province, China.

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$a Kalasová, Dominika $u Central European Institute of Technology, Brno University of Technology, Brno, Czech Republic.

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$a Szarowská, Bára $u Department of Physiology and Pharmacology, Karolinska Institutet, Solna, 171777, Stockholm, Sweden.

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$a Zikmund, Tomáš $u Central European Institute of Technology, Brno University of Technology, Brno, Czech Republic.

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$a Novotná, Marie $u Central European Institute of Technology, Brno University of Technology, Brno, Czech Republic.

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$a Kaiser, Jozef $u Central European Institute of Technology, Brno University of Technology, Brno, Czech Republic. kaiser@fme.vutbr.cz.

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