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Intravital Imaging of Blood Flow and HSPC Homing in Bone Marrow Microvessels

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Stem Cell Mobilization

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2017))

Abstract

Two-photon intravital microscopy (2P-IVM) is an advanced imaging technique that allows the visualization of dynamic cellular behavior deeply inside tissues and organs of living animals. Due to the deep tissue penetration, imaging of highly light-scattering tissue as the bone becomes feasible at subcellular resolution.

To better understand the influence of blood flow on hematopoietic stem and progenitor cell (HSPC) homing to the bone marrow (BM) microvasculature of the calvarial bone, we analyzed blood flow dynamics and the influence of flow on the early homing behavior of HSPCs during their passage through BM microvessels. Here, we describe a 2P-IVM approach for direct measurements of red blood cell (RBC) velocities in the BM microvasculature using repetitive centerline scans at the level of individual arterial vessels and sinusoidal capillaries to obtain a detailed flow profile map. Furthermore, we explain the isolation and enrichment of HSPCs from long bones and the transplantation of these cells to study the early homing behavior of HSPCs in BM sinusoids at cellular resolution. This is achieved by high-resolution spatiotemporal imaging through a chronic cranial window using transgenic reporter mice.

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Acknowledgments

We thank F. Winkler for sharing his expertise on the imaging setup for head immobilization. This work was supported by the Max Planck Society, the University of Münster, the DFG cluster of excellence “Cell in Motion,” and the European Research Council (AdG 339409 AngioBone).

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Authors and Affiliations

  1. Tissue Morphogenesis, Max Planck Institute for Molecular Biomedicine, Münster, Germany

    Jonas Stewen & Maria Gabriele Bixel

Authors
  1. Jonas Stewen
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  2. Maria Gabriele Bixel
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Corresponding author

Correspondence to Maria Gabriele Bixel .

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Editors and Affiliations

  1. Department of Internal Medicine II, Center for Medical Research, University of Tübingen, Tübingen, Germany

    Gerd Klein

  2. Institute of Transfusion Medicine and Immunology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany

    Patrick Wuchter

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Stewen, J., Bixel, M.G. (2019). Intravital Imaging of Blood Flow and HSPC Homing in Bone Marrow Microvessels. In: Klein, G., Wuchter, P. (eds) Stem Cell Mobilization. Methods in Molecular Biology, vol 2017. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9574-5_9

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  • DOI: https://doi.org/10.1007/978-1-4939-9574-5_9

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9573-8

  • Online ISBN: 978-1-4939-9574-5

  • eBook Packages: Springer Protocols

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