title: Sulfatides trigger increase of cytosolic free calcium and enhanced expression of tumor necrosis factor-alpha and interleukin-8 mRNA in human neutrophils. Evidence for a role of L-selectin as a signaling molecule.
abstract: Sulfatides have been established recently as ligands for L-selectin, and we investigated whether they trigger transmembrane signals through ligation of L-selectin. We found that sulfatides trigger the increase of cytosolic free calcium in neutrophils and that this effect was strictly dependent on sulfation of the galactose ring, as non-sulfated galactocerebrosides were not stimulatory. Chymotrypsin and phorbol 12-myristate 13-acetate treatment of neutrophils caused shedding of L-selectin, but not of class I major histocompatibility complex antigens or beta 2 integrins, and blunted the capability of neutrophils to respond to sulfatides with an increase of cytosolic free calcium. Four different anti-L-selectin antibodies (DREG-200, LAM1/3, LAM1/6, and LAM1/10), but not four control antibodies directed against different surface molecules of neutrophils, also triggered an increase of cytosolic free calcium. The anti-L-selectin antibodies were stimulatory both if used in a soluble form, after cross-linking with anti-mouse F(ab')2 fragments, and immobilized to protein A of Staphylococcus aureus through the Fc fragment. With immobilized antibodies, an increase of cytosolic free calcium was found also by plating neutrophils on antibodies bound to protein A-coated coverslips and monitoring the increase of cytosolic free calcium by fluorescence microscopy. Both sulfatides and anti-L-selectin antibody effects were not inhibited by pertussis toxin, thus indicating that a pertussis toxin-sensitive GTP-binding protein was not involved in signal transduction. Sulfatides also triggered an increase of tumor necrosis factor-alpha and interleukin-8 mRNAs in neutrophils. Also to act as stimuli of cytokine mRNA expression, sulfatides required sulfation of the galactose ring, as non-sulfated galactocerebrosides were not stimulatory, and depended on expression of L-selectin, as shedding of this molecules induced by chymotrypsin blunted their effects. These findings suggest that L-selectin can transduce signals activating selective cell function.
authors.affiliation: Institute of General Pathology, University of Verona, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Laudanna
authors.forename: G
authors.initials: G
authors.lastname: Constantin
authors.forename: P
authors.initials: P
authors.lastname: Baron
authors.forename: E
authors.initials: E
authors.lastname: Scarpini
authors.forename: G
authors.initials: G
authors.lastname: Scarlato
authors.forename: G
authors.initials: G
authors.lastname: Cabrini
authors.forename: C
authors.initials: C
authors.lastname: Dechecchi
authors.forename: F
authors.initials: F
authors.lastname: Rossi
authors.forename: M A
authors.initials: MA
authors.lastname: Cassatella
authors.forename: G
authors.initials: G
authors.lastname: Berton
mesh_terms: D002118:Calcium / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000502:physiology*; D002918:Chymotrypsin / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics*; D019041:L-Selectin; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D015398:Signal Transduction; D013433:Sulfoglycosphingolipids / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics*
chemical_list: D015815:Cell Adhesion Molecules; D016209:Interleukin-8; D012333:RNA, Messenger; D013433:Sulfoglycosphingolipids; D014409:Tumor Necrosis Factor-alpha; D019041:L-Selectin; D002918:Chymotrypsin; D002118:Calcium