ANAPHYLACTIC AND CHEMOTACTIC RESPONSE OF MAMMALIAN-CELLS TO ZYMOSAN-ACTIVATED SHARK SERUM

Human anaphylatoxins, C3a and C5a, are small proteins which are genera ted by proteolytic cleavage of the complement components C3 and C5 dur ing activation of complement. Complement activity via the classical an d alternative pathways has been described for the nurse shark (Ginglym ostoma cirratum) and studies have demonstrated chemotactic activity of shark leucocytes in response to activated shark serum. However, anaph ylatoxins have not been reported in an elasmobranch. This investigatio n was undertaken to examine activated shark serum for anaphylatoxins b y studying the anaphylactic and chemotactic response of mammalian cell s. Shark (G. cirratum) serum, activated with zymosan (10 mg ml(-1)) fo r 3 h at 30 degrees C, was fractionated on Sephadex G-100 and G-25 col umns, and fractions were pooled and assayed fort spasmogenic and chemo tactic activity. In vitro chemotaxis assays were set up in blind well chemotaxis chambers using human leucocytes as responding cells. A sign ificant (P < 0.05) migratory response was obtained with several serum pools. Spasmogenic activity was measured by smooth muscle contraction of rat ileum, and was present in serum fractions containing low molecu lar size (< 17.5 kDa) proteins/peptides, which also induced chemotaxis of human leucocytes. No spasmogenic or chemotactic activity was demon strated in fractions of normal (nan-activated) shark serum. This study shows that mammalian cells respond to factors generated in activated shark serum and results suggest that the chemotactic and anaphylactic activity in shark serum might be due to a biologically active peptide( s) derived from the cleavage of complement component(s), perhaps analo gous to mammalian C3a and/or C5a. (C) 1997 Academic Press Limited.