chondroitin-sulfates and 1-2-dipalmitoyl-3-phosphatidylethanolamine

The effects of glycosaminoglycans (GAG) on cell-to-substratum adhesion and neurite elongation were examined in primary cultures of fetal rat hippocampal neurons using tissue culture dishes coated with GAGs coupled to dipalmitoylphosphatidylethanolamine (PE), a novel probe for biological functions of GAGs. Both chondroitin sulfate conjugate to PE (CS-PE) and hyaluronic acid conjugate to PE (HA-PE) promoted neurite elongation from neurons in a dose-dependent manner when immobilized onto polylysine-coated dishes at various concentrations up to 1.0 microg/ml. The coating of CS-PE or HA-PE at a concentration higher than 1.0 microg/ml resulted in failure of neurite extension and adhesion of neurons to the substrata. In contrast, heparin conjugate to PE (HP-PE) did not exert any effects on neurite elongation or on cell attachment at these concentrations. These findings suggest that GAGs serve as a modulator for neurite elongation during neuronal network formation in the developing central nervous system.

Topics: Animals; Cell Adhesion; Cells, Cultured; Chondroitin Sulfates; Fetus; Glycosaminoglycans; Hippocampus; Hyaluronic Acid; Molecular Structure; Neurites; Neurons; Phosphatidylethanolamines; Rats

Heparin and chondroitin sulfate disaccharides have been investigated by high-performance (HP) TLC and liquid secondary-ion mass spectrometry (LSIMS) after conversion to neoglycolipid derivatives by reductive-amination with an aminolipid (dihexadecyl phosphatidylethanolamine, DHPE). Mobility on HPTLC was largely determined by the number of sulfate groups present, but was also influenced by the position of sulfate, monosaccharide composition and linkage. The mass spectra acquired directly from the TLC plate provided quasimolecular and fragment ions from which composition, including sulfate content, and sequence information was obtained at high sensitivity. Lipid DHPE conjugation and TLC-LSIMS were performed to analyse products of the oxymercuration reaction used to cleave unsaturated uronic acid (delta UA) residues from glycosaminoglycan (GAG) fragments produced by enzymatic degradation with glycan lyases. Previously the identification of the product from delta UA and the integrity of the remaining structures from oligosaccharides larger than disaccharide have not been made. Multiple and characteristic products of the cleaved delta UA were detected and these can be used for identification of terminal delta UA and its sulfate content. It was established with several disaccharides and a tetrasaccharide that glycosidic linkages and O- and N-sulfate groups are preserved in the remaining structures after removal of delta UA. These results indicate that the oxymercuration reaction will be applicable to generating series of GAG fragments containing unmodified sequences for biological activity studies.

Topics: Carbohydrate Sequence; Chondroitin Sulfates; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Disaccharides; Glycolipids; Glycosaminoglycans; Heparin; Mass Spectrometry; Mercury; Methylation; Molecular Sequence Data; Oxidation-Reduction; Phosphatidylethanolamines; Sequence Analysis; Uronic Acids

Adherence of Plasmodium falciparum-infected erythrocytes to cerebral postcapillary venular endothelium is believed to be a critical step in the development of cerebral malaria. Some of the possible receptors mediating adherence have been identified, but the process of adherence in vivo is poorly understood. We investigated the role of carbohydrate ligands in adherence, and we identified chondroitin sulfate (CS) as a specific receptor for P. falciparum-infected erythrocytes. Parasitized cells bound to Chinese hamster ovary (CHO) cells and C32 melanoma cells in a chondroitin sulfate-dependent manner, whereas glycosylation mutants lacking chondroitin sulfate A (CSA) supported little or no binding. Chondroitinase treatment of wild-type CHO cells reduced binding by up to 90%. Soluble CSA inhibited binding to CHO cells by 99.2 +/- 0.2% at 10 mg/ml and by 72.5 +/- 3.8% at 1 mg/ml, whereas a range of other glycosaminoglycans such as heparan sulfate had no effect. Parasite lines selected for increased binding to CHO cells and most patient isolates bound specifically to immobilized CSA. We conclude that P. falciparum can express or expose proteins at the surface of the infected erythrocyte that mediate specific binding to CSA. This mechanism of adherence may contribute to the pathogenesis of P. falciparum malaria, but has wider implications as an example of an infectious agent with the capacity to bind specifically to cell-associated or immobilized CS.

Topics: Animals; Cell Adhesion; Cell Adhesion Molecules; Cells, Cultured; Child; CHO Cells; Chondroitin Sulfates; Cricetinae; Cricetulus; Endothelium, Vascular; Erythrocytes; Glycosylation; Heparitin Sulfate; Host-Parasite Interactions; Humans; Malaria, Falciparum; Melanoma; Phosphatidylethanolamines; Plasmodium falciparum; Receptors, Cell Surface; Tumor Cells, Cultured; Umbilical Veins