Letter | Published:

Endothelial tubes assemble from intracellular vacuoles in vivo

Naturevolume 442pages453456 (2006) | Download Citation



The formation of epithelial tubes is crucial for the proper development of many different tissues and organs, and occurs by means of a variety of different mechanisms1. Morphogenesis of seamless, properly patterned endothelial tubes is essential for the development of a functional vertebrate circulatory system, but the mechanism of vascular lumenization in vivo remains unclear. Evidence dating back more than 100 years has hinted at an important function for endothelial vacuoles in lumen formation2. More than 25 years ago, in some of the first endothelial cell culture experiments in vitro, Folkman and Haudenschild described “longitudinal vacuoles” that “appeared to be extruded and connected from one cell to the next”3,4, observations confirmed and extended by later studies in vitro showing that intracellular vacuoles arise from integrin-dependent and cdc42/Rac1-dependent pinocytic events downstream of integrin–extracellular-matrix signalling interactions5,6,7,8,9,10. Despite compelling data supporting a model for the assembly of endothelial tubes in vitro through the formation and fusion of vacuoles, conclusive evidence in vivo has been lacking, primarily because of difficulties associated with imaging the dynamics of subcellular endothelial vacuoles deep within living animals. Here we use high-resolution time-lapse two-photon imaging of transgenic zebrafish to examine how endothelial tubes assemble in vivo, comparing our results with time-lapse imaging of human endothelial-cell tube formation in three-dimensional collagen matrices in vitro. Our results provide strong support for a model in which the formation and intracellular and intercellular fusion of endothelial vacuoles drives vascular lumen formation.

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Change history

  • 07 July 2006

    The incorrect pdf version of this paper was posted online at publication. This was corrected 7 July 2006.


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We thank J. Faske for technical assistance, G. Martin for help in constructing the mRFP1-expressing human ECs, K. Tanegashima for assistance with western blot analysis, R. Tsien for providing the mRFP1 vector, and I. B. Dawid for critical reading of this manuscript. This work was supported in part by a grant from the NIH to G.E.D. B.M.W. is supported by the intramural program of the NICHD.

Author information


  1. Laboratory of Molecular Genetics

    • Makoto Kamei
    •  & Brant M. Weinstein
  2. Microscopy and Imaging Core, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, 20892, USA

    • Louis Dye
  3. Department of Pathology, Texas A&M University System Health Science Center, 208 Reynolds Medical Building, College Station, Texas, 77843-1114, USA

    • W. Brian Saunders
    • , Kayla J. Bayless
    •  & George E. Davis


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Competing interests

Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Corresponding author

Correspondence to Brant M. Weinstein.

Supplementary information

  1. Supplementary Methods

    Detailed descriptions of materials and methods used in this study. (DOC 68 kb)

  2. Supplementary Movie Legends

    Detailed legends for the Supplementary Movies. (DOC 52 kb)

  3. Supplementary Movie 1

    Time-lapse DIC imaging of cultured human umbilical vein endothelial cells forming, collapsing and fusing vacuoles. (MOV 2667 kb)

  4. Supplementary Movie 2

    Time-lapse DIC imaging of a single cultured human umbilical vein endothelial cell forming vacuoles that merge and expand into a highly enlarged vacuolar space. (MOV 2752 kb)

  5. Supplementary Movie 3

    Time-lapse 2-photon imaging of an embryonic zebrafish intersegmental vessel, with small vacuoles forming, collapsing and fusing. (MOV 4425 kb)

  6. Supplementary Movie 4

    Time-lapse 2-photon imaging of an embryonic zebrafish intersegmental vessel forming an expanded vacuolar/lumenal space. (MOV 4311 kb)

  7. Supplementary Movie 5

    Time-lapse 2-photon imaging of a lumenizing intersegmental vessel in a zebrafish embryo. (MOV 9109 kb)

  8. Supplementary Movie 6

    Time-lapse DIC imaging of a lumenizing intersegmental vessel in a zebrafish embryo. (MOV 7939 kb)

  9. Supplementary Movie 7

    Time-lapse DIC imaging of cultured human umbilical vein endothelial cells forming an intercellular space. (MOV 1783 kb)

  10. Supplementary Movie 8

    Time-lapse DIC imaging of cultured human umbilical vein endothelial cells forming an intercellular space. (MOV 9671 kb)

  11. Supplementary Movie 9

    Time-lapse epifluorescence imaging of cultured human umbilical vein endothelial cells forming an intercellular space without mixing of their respective cytoplasmic contents. (MOV 8120 kb)

  12. Supplementary Movie 10

    Time-lapse 2-photon imaging of red quantum-dot-injected embryonic zebrafish trunk vessels. (MOV 7650 kb)

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