Microscopic bioimaging of blood flow and distribution of cancer cells in lungs is essential to analyze mechanism of lung metastasis. Such cancer metastasis has been well known to induce hypercoagulable states and thrombosis. In histopathological tissue sections, however, it has been difficult to capture rapid phenomenon of thrombus formation due to technical problems associated with much less retention of soluble serum components as well as dynamic histological features reflecting their living states. In this study, to achieve bioimaging of both hypercoagulable states and thrombosis induced by early metastasis of mouse B16-BL6 melanoma, "in vivo cryotechnique" (IVCT) was used, which retained soluble components at their original sites. Glutathione-coated quantum dots (QDs) were subsequently injected after melanoma cells via right ventricles to examine plasma flow with fluorescence emission. At 5s after the melanoma injection, melanoma cells were mostly stacked and intruded in alveolar capillaries with changing their shapes. Assembly of platelets initially appeared at 1min, and they aggregated around the stacked melanoma cells at 5min. Such aggregated platelets were immunopositive for both phospho-tyrosine 418 and 527 of Src, indicating their partial signal activation. Fibrin monomers were also immunolocalized around both melanoma cells and platelet aggregates, and massive immunoreaction deposits of fibrinogen were also detected near the same areas, but more strongly detected around the melanoma cells, indicating initial thrombus formation. In those areas, QDs were rarely detected, probably because of the lack of blood supply. Thus, IVCT revealed histopathological features of initial thrombosis under their circulatory conditions.
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