Lectin histochemistry for in situ profiling of rat colon sialoglycoconjugates

The growing interest in glycoconjugates expressed and released by the epithelium of the intestinal mucosa is tightly related to the multiple functional roles attributed to sialic acid and its derivatives. In the present work, biotin and HRP conjugated lectins were used to detect the sialylation pattern and to identify specific structural features of sialoderivatives in the rat colon. In particular, the occurrence and distribution of sialic acids linked alpha2,6 to D-Gal/D-GalNAc and alpha2,3 to D-Gal were directly demonstrated with SNA and MAL II binding, respectively. In addition, in order to by-pass the specificity problems of SNA and MAL II as histochemical reagents, as well as to look for additional and complementary information about acetylation degree and sites, we combined sialidase digestion, potassium hydroxide deacetylation, and differential periodate oxidation with PNA and DBA binding. The data showed the distribution and structure of sialic acid-beta-D-Gal(1-3)-D-GalNAc and sialic acid-D-GalNac sequences, which proved to be widely distributed as cellular components or secretory products in surface goblet cells and crypt cells of the colonic epithelium. A high degree of O-acetylation, with acetyl groups mainly at 9 and 4 positions, was found, showing an increasing gradient from the proximal to distal portion of the colon. These results, which largely reproduce the sialylation pattern in other species, contribute new insights in defining the tissue specific expression of sialoderivatives in the colonic mucosa, and testify to their high heterogeneity which the wide range of sialic acid functional correlates in the intestinal tract depend on.