Immuno- and Enzyme-histochemistry of HRP for Demonstration of Blood Vessel Permeability in Mouse Thymic Tissues by “In Vivo Cryotechnique”

It is difficult to understand the in vivo permeability of thymic blood vessels, but “in vivo cryotechnique” (IVCT) is useful to capture dynamic blood flow conditions. We injected various concentrations of horseradish peroxidase (HRP) with or without quantum dots into anesthetized mice via left ventricles to examine architectures of thymic blood vessels and their permeability at different time intervals. At 30 sec after HRP (100 mg/ml) injection, enzyme reaction products were weakly detected in interstitium around some thick blood vessels of corticomedullary boundary areas, but within capillaries of cortical areas. At 1 and 3 min, they were more widely detected in interstitium around all thick blood vessels of the boundary areas. At 10 min, they were diffusely detected throughout interstitium of cortical areas, and more densely seen in medullary areas. At 15 min, however, they were uniformly detected throughout interstitium outside blood vessels. At 30 min, phagocytosis of HRP by macrophages was scattered throughout the interstitium, which was accompanied by decrease of HRP reaction intensity in interstitial matrices. Thus, time-dependent HRP distributions in living mice indicate that molecular permeability and diffusion depend on different areas of thymic tissues, resulting from topographic variations of local interstitial flow starting from corticomedullary areas.