Simultaneous observation of capillary nets and tenascin in intestinal villi

In order to reveal the biological role of capillaries in a tissue, it is desirable to study the three-dimensional distribution of capillary nets in relation to tissue architecture. However, the simultaneous observation of the three-dimensional distribution of these nets and other substances has been rarely performed to date. In the present study, we have developed a novel method for investigating the three-dimensional distribution of capillary nets simultaneously with that of extracellular matrix components, such as tenascin, using the confocal laser scanning microscope.

Adult male mice were perfused with TRITC-labelled gelatin. After perfusion, the intestine was irrigated with chilled fixative containing paraformaldehyde and picric acid, dissected, and returned to the same fixative. The intestine was further sectioned and indirectly immunostained for tenascin using an FITC-labelled antibody.

The three-dimensional distribution of capillary nets and tenascin in villi was simultaneously observed on stereo pairs of pseudo-colored and superimposed images. Tenascin was distributed at the basement membrane zone and in the underlying connective tissue but absent in some regions where capillary nets were running just beneath the epithelium. Substances other than tenascin also can be examined in correlation with capillary nets.

This method will be useful for investigating the biological role of capillary nets.