Occurrence of heparin in circulating basophil-like cells in the hemolymph of the invertebrate chordate Styela plicata (Chordata-Tunicata)*
Cintia M. de Barrosa,b, Leonardo R. de Andradeb,c, Silvana Allodib,c, Christian Viskovd, Pierre A. Mourier d, Moisés C.M. Cavalcantea, Anita H. Straus e, Helio K. Takahashie, and Mauro S. G. Pavãoa
aLaboratório de Tecido Conjuntivo, Hospital Universitário Clementino Fraga Filho and Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ, CEP 21941-590, Brasil; bPrograma de Pós-Graduação em Ciências Morfológicas, Instituto de Ciências Biomédicas (ICB), CCS, UFRJ, Rio de Janeiro, RJ, CEP 21941-590, Brasil; cDepartamento de Histologia e Embriologia, ICB, CCS, UFRJ, Rio de Janeiro, RJ, CEP 21941-590, Brasil; d Sanofi-aventis, Centre de Recherche de Paris, Unité de Glycochimie, Batiment Lavoisier 13, Quai Jules Guesde 94400 Vitry-sur Seine, France; eDepartamento de Bioquímica, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brasil.
The hemolymph of ascidians (Chordata-Tunicata) contains different types of hemocytes embedded in liquid plasma. In the present study, heparin and a sulfated heteropolysaccharide were purified from the hemolymph of the ascidian Styela plicata. Chemical analysis indicated that the heteropolysaccharide is composed by glucose (~60%) and galactose (~40%), and is highly sulfated. High-performance liquid chromatography of the disaccharides formed after exhaustive degradation with specific lyases revealed that the hemolymph heparin is composed mainly by the disaccharides ∆UA(2SO4)-1®4-b-D-GlcN(SO4) (39.7%) and ∆UA(2SO4)-1®4-b-D-GlcN(SO4)(6SO4) (38.2%). Small amounts of the 3-O-sulfated disaccharides ∆UA(2SO4)-1®4-b-D-GlcN(SO4)(3SO4) (9.8%) and ∆UA(2SO4)-1®4-b-D-GlcN(SO4)(3SO4)(6SO4) (3.8%) were also detected. These 3-O-sulfated disaccharides were demonstrated to be essential for the binding of the hemocyte heparin to antithrombin III. Electron microscopy techniques were used to characterize the ultrastructure of the hemocytes and to localize heparin and histamine in these cells. At least five cell types were recognized and classified as: univacuolated and multivacuolated cells, amebocytes, hemoblasts and granulocytes. Immunocytochemistry showed that heparin and histamine co-localize in intracellular granules of only one type of hemocyte, the granulocyte. These results show for the first time the presence of heparin in a circulating cell in an invertebrate chordate and may contribute for the understanding of the evolution of the immune system in this phylum.
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