Glycoconjugates with Neu5Ac(?2,6)Gal(?1,4)GlcNAc sequences: a selective lectin-histochemical property of Kurloff cells in guinea pig thymus

Abstract

The Kurloff cell (KC), a natural killer lymphocyte, contains a large (10-microns diameter) periodic acid-Schiff (PAS)-positive lysosome-like inclusion body called the Kurloff body (KB), which exhibits strong acid phosphatase activity. The presence of Sambucus nigra agglutinin (SNA)-reactive Neu5Ac(alpha 2,6)-D-Gal/Gal-NAc(beta 1,4)GlcNAc oligosaccharide sequences and the absence of the corresponding Neu5Ac(alpha 2,3) Maackia amurensis agglutinin (MAA)-reactive sequence in the major 35-kDa N-glycosylproteins of the complex or hybrid type extracted from purified KC were established by Western-lectin-blotting of cytosolic extracts from purified KC. Moreover, these SNA-reactive sequences, or at least part of them, were shown to be borne by sialidase-sensitive KC acid isophosphatases. Thymic sections rich in KC, from estrogenized guinea pigs were examined by affino-histochemistry with these sialic acid-reactive lectins. The SNA-reactivity of thymic sections was quasi-exclusively confined to KC clusters, whereas the whole thymic section was negative for MAA. KC were not SNA-reactive following preincubation and incubation with 200 mM lactose. When submitted to enzymatic or mild chemical desialylation processes, the SNA-reactivity of the KC clusters was enhanced. The SNA-reactivity of KC clusters was completely abolished following prolonged chemical desialylation, whereas the PAS-positivity of KB remained unchanged. Even after a prolonged sialidase treatment, this SNA-reactivity was only reduced. Moreover, after both these desialylation processes, KC developed a heavier Ricinus communis agglutinin-reactivity, thus confirming the presence of penultimate Gal residues in their abundant SNA-reactive oligosaccharide sequences Neu5Ac(alpha 2,6)Gal(beta 1,4)GlcNAc. Such a selective lectin histochemical property provides a marker for detecting KC.

Characterization of the non‐sulphated highly anionic kurloff cell glycoconjugates by lectin‐ and immunoblotting: Evidence for the presence of α(2,8)‐linked polysialic acid‐bearing molecules

“…To ascertain the presence of polysialosyl units, we tested the effect of endo N; after pretreatment of peak IV with endosialidase, anti-Men B antibody did not give any positive reaction (fig 6b). KC major glycoproteins, previously characterized as containing SNAreactive oligosaccharides (fig 6c) with ~t(2,6)-linked sialic acid [33,42] were used as negative control glycoproteins for anti- Men B (fig 6d). Crude tissue extracts from fetal mouse brain and immunopurified embryonic forms of N-CAM were used as positive control for the presence of (NeuAc~2,8) n sequences (fig 6e, f).…”

“…Using highly purified KC [29], soluble extracts obtained after dissociative extraction in either 4 M GnHCI or 7 M urea were subjected to ion exchange chromatography and the macromolecules responsible for these histochemical affinities were identified: i) unbound major 30-35 kDa glycoproteins designated as major KC GP [30,35] among which a fraction containing Neu5Ac(oc2,6)-D-Gal/Gal-NAc(~I,4)GIcNAc oligosaccharide sequences were shown to correspond to acid phosphatase subunits [33,42]; and it) protease-resistant chondroitin-4-sulphate proteoglycans eluted at 0.55 M NaCI [31,36] which correspond to PASpositivity and TB-metachromasia of the KB, respectively.…”

Expression of Gal beta 1,4GlcNAc alpha 2,6-sialyltransferase and alpha 2,6-linked sialoglycoconjugates in normal human and rat tissues.

“…In H35 rat hepatoma cells, dexamethasone increased a2.6-ST expression (45,46). Furthermore, Oulhaj et al (47) found that the natural killer T-cells in thymus of estrogenized or pregnant guinea pigs stained with SNA. In the present study, the decidual cells of human and rat mature placenta stained vividly with antia2,6-ST.…”

Introduction to the thymus

“…It would be a good idea in another topical issue to cover the application of other modern techniques to the thymus. These might include confocal microscopy, which is seldom used in this area, and lectin histochemistry, as we have done in our study on Kurloff cells at the corticomedullary junction of the thymus in the guinea-pigs (Oulhaj et al, 1993).…”