Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Real-time in vivo imaging of platelets, tissue factor and fibrin during arterial thrombus formation in the mouse

Abstract

We have used confocal and widefield microscopy to image thrombus formation in real time in the microcirculation of a living mouse. This system provides high-speed, near-simultaneous acquisition of images of multiple fluorescent probes and of a brightfield channel. Vascular injury is induced with a laser focused through the microscope optics. We observed platelet deposition, tissue factor accumulation and fibrin generation after laser-induced endothelial injury in a single developing thrombus. The initiation of blood coagulation in vivo entailed the initial accumulation of tissue factor on the upstream and thrombus–vessel wall interface of the developing thrombus. Subsequently tissue factor was associated with the interior of the thrombus. Tissue factor was biologically active, and was associated with fibrin generation within the thrombus.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Imaging of platelets, fibrin and tissue factor in arterial thrombi using fluorescent antibodies.
Figure 2: Time course and localization of platelets and fibrin during thrombus formation.
Figure 3: Confocal fluorescence imaging of the distribution of tissue factor, platelets and fibrin in a thrombus.
Figure 4: Structural organization during thrombus development.
Figure 5: Schematic diagram of the confocal–widefield high-speed microscope used for real-time intravital microscopy of the microcirculation of a living mouse.

References

  1. 1

    Rosen, E.D. et al. Laser-induced noninvasive vascular injury models in mice generate platelet- and coagulation-dependent thrombi. Am. J. Pathol. 158, 1613–1622 (2001).

    CAS  Article  Google Scholar 

  2. 2

    Savage, B., Almus-Jacobs, F. & Ruggeri, Z.M. Specific synergy of multiple substrate–receptor interactions in platelet thrombus formation under flow. Cell 94, 657–666 (1998).

    CAS  Article  Google Scholar 

  3. 3

    Inoue, S. & Inoue, T. (eds.). Direct-view High Speed Confocal Scanner–the CSU-10 (Academic Press, New York, 2000).

    Google Scholar 

  4. 4

    Morrissey, J.H. Tissue factor: an enzyme cofactor and a true receptor. Thromb. Haemost. 86, 66–74 (2001).

    CAS  Article  Google Scholar 

  5. 5

    Morrissey, J.H., Fakhrai, H. & Edgington, T.S. Molecular cloning of the cDNA for tissue factor, the cellular receptor for the initiation of the coagulation protease cascade. Cell 50, 129–135 (1987).

    CAS  Article  Google Scholar 

  6. 6

    Spicer, E.K. et al. Isolation of cDNA clones coding for human tissue factor: primary structure of the protein and cDNA. Proc. Natl. Acad. Sci. USA 84, 5148–5152 (1987).

    CAS  Article  Google Scholar 

  7. 7

    Koyama, T. et al. Determination of plasma tissue factor antigen and its clinical significance. Br. J. Haematol. 87, 343–347 (1994).

    CAS  Article  Google Scholar 

  8. 8

    Fareed, J., Callas, D.D., Hoppensteads, D. & Bermes, E.W. Tissue factor antigen levels in various biological fluids. Blood Coagul. Fibrinolysis 6 (Suppl. 1), S32–S36 (1995).

    CAS  Article  Google Scholar 

  9. 9

    Zumbach, M. et al. Tissue factor antigen is elevated in patients with microvascular complications of diabetes mellitus. Exp. Clin. Endocrinol. Diabetes 105, 206–212 (1997).

    CAS  Article  Google Scholar 

  10. 10

    Giesen, P.L. et al. Blood-borne tissue factor: another view of thrombosis. Proc. Natl. Acad. Sci. USA 96, 2311–2315 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Ley, K. Gene-targeted mice in leukocyte adhesion research. Microcirculation 2, 141–150 (1995).

    CAS  Article  Google Scholar 

  12. 12

    Yang, J. et al. Targeted gene disruption demonstrates that P-selectin glycoprotein ligand 1 (PSGL-1) is required for P-selectin-mediated but not E- selectin-mediated neutrophil rolling and migration. J. Exp. Med. 190, 1769–1782 (1999).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank S. Inoué for introducing us to the Yokogawa CSU-10 high-speed confocal microscope, Eric Furie for digital image capture advice, and F. Castellino and E. Rosen for demonstrating the laser injury model. This work was supported by grants from the US National Institutes of Health (HL51926 and HL69435). The confocal microscope was obtained with partial support from the National Institutes of Health (S10RR15680).

Author information

Affiliations

Authors

Corresponding author

Correspondence to Bruce Furie.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Falati, S., Gross, P., Merrill-Skoloff, G. et al. Real-time in vivo imaging of platelets, tissue factor and fibrin during arterial thrombus formation in the mouse. Nat Med 8, 1175–1180 (2002). https://doi.org/10.1038/nm782

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing