Binding ofMonoclonal Antibody AA4 to Gangliosides on Rat Basophilic Leukemia Cells Produces Changes Similar to Those Seen with FCE Receptor Activation

The mAb AA4 binds to novel derivatives of the ganglioside GD1b on rat basophilic leukemia (RBL2H3) cells . Some of the gangliosides are located close to the high affinity IgE receptor (FCcRI), and binding of mAb AA4 inhibits FCERI-mediated histamine release . In the present study, mAb AA4 was found to bind exclusively to mast cells in all rat tissues examined . In vitro, within 1 min of mAb AA4 binding, the cells underwent striking morphologic changes . They lost their normal spindle shaped appearance, increased their ruffling, and spread over the surface of the culture dish . These changes were accompanied by a redistribution of the cytoskeletal elements, actin, tubulin, and vimentin, but only the actin was associated with the membrane ruffles . Binding of mAb AA4 also induces a rise in intracellular calcium, stimulates phosphatidyl inositol breakdown, and activates PKC . MAST cells and basophils contain a variety of inflammatory mediators which are released following cross-linking of high affinity IgE surface receptors (FcERI)' . The activation of the receptor results in a complex series of biochemical and morphological events (48) . Briefly, when FcERI are cross-linked, either by antigen on IgE-sensitized cells or by antibody directed against the receptor, there is stimulation of phospholipase C resulting in hydrolysis ofphosphatidyl inositol with the generation of 1,2 diacylglycerol and inositol 1,3,5 phosphate . Both the 1,2 diacylglycerol and the inositol 1,4,5 phosphate are thought to act as intracellular signals ; the 1,2 diacylglycerol by activating protein kinase C (PKC), and the inositol 1,4,5 phos1. Abbreviations used in this paper: DNP44-HSA, 44 molecules of 2,4-dinitrophenyl conjugated to 1 molecule of human serum albumin; EMEM, Eagle's minimum essential medium with Earl's salts ; FcERI, high affinity IgE receptor on basophils/mast cells ; PKC, protein kinase C; NBD,N-[7 Nitrobenz-2-oxa-1,3 diazol-4-yl)] ; RBL-2113, rat basophilic leukemia 2113