Carbohydrate induced modulation of cell membrane I. Interaction of sialic acid with peripheral blood lymphocytes: A spin label study

Abstract

SiaIic acid is now known to serve as ligand for lymphocyte lectins known as selectins. Although its role as a ligand for adhesion molecules has been studied extensively, no studies have been performed to determine the physiological changes in lymphocytes after binding of sialic acid to lymphocyte lectin. We report for the first time that interaction of lymphocytes with siahc acid severely restricts the rotational mobility of the cell surface proteins as well as membrane lipids, as studied by EPR spectroscopy using spin probes. Binding of mucin totally immobilizes the lymphocyte membrane. Surprisingly the binding of sialic acid or mucin also immobilized the aqueous probe TEMPO, indicating an appreciable increase in cytoplasmic viscosity.

Normal but not altered mucins activate neutrophils

“…Carbohydrate-carbohydrate interactions, such as those involved in the glycosynapse (Hakomori 2002), are essential for the extravasation and tissue ingress of leucocytes. Mucins are decorated with epitopes whose ligands can be found on cells of the immune system (Hajela et al 1994;Handa et al 1997;Kim et al 1999;Prakobphol et al 1999;Leppanen et al 2002). We have shown that these epitopes are also common on normal human ocular mucins (Berry et al 1996(Berry et al , 2000(Berry et al , 2003Ellingham et al 1999).…”

Carbohydrate induced modulation of cell membrane: II. Spin label study of fluidity changes in peripheral blood lymphocyte membrane

“…We also studied the biophysical changes following lymphocyte-sialic acid interaction occurring at the lymphocyte membrane using EPR spectroscopy. Our preliminary studies indicated that binding of sialic acid or mucin to GPBLs markedly reduces the rotational mobility of membrane proteins as well as that of lipids [8]. The results obtained demanded a detailed investigation in this area as it was not clear whether this was a specific effect produced by interaction of lymphocytes with sialic acid or its derivatives only or was a generalized effect produced by the interaction of membrane lectins with their respective ligands.…”

“…GPBLs were isolated using Histopaque-1077, treated with various mono-, di-and oligosaccharides and spin labeled with either 5-DS, 16-DS, MECP and TEMPO as described earlier [7,8]. The carbohydrates and their complex forms used were glucose, N-acetylglucosamine, maltose, mannose, mannose-6-phosphate, galactose, maltotriose, fucoidan, hyaluronic acid, sialic acid and heparin.…”

“…Instrument settings employed were as follows: scan range 100 G, field set 3237 G, temperature 27°C, time constant 1 s, scan time 1 s, modulation amplitude 2 G for 16-DS and TEMPO and 1 G for 5-DS and MECP, modulation frequency 100 kHz, microwave power 5 mW, microwave frequency 9.01 GHz, receiver gain 2.5 × 10 4. The rotational correlation time T~ and order parameter $3 were calculated as described earlier [8]. The experiments were repeated 4 to 6 times and data were subjected to statistical analysis using the STAT-P-GW-BASIC Rev 1.02 programme.…”

Carbohydrate induced modulation of cell membrane. VI. Binding of exogenous lectin induces susceptibility of erythrocytes to free radical damage: a spin label study

“…Instrument setting was as follows: scan range 100 G; field set 3237 G; temperature 27°C; time constant 1 s; scan time 4 s; modulation amplitude 2 G; modulation frequency 100 kHz; microwave power 5 mW; microwave frequency 9.01 kHz; receiver gain 2.5 X10 4 . The rotational correlation time Tc, expressed in seconds, was calculated as described earlier [18].…”