Loxosceles SPIDER VENOM INDUCES METALLOPROTEINASE MEDIATED CLEAVAGE OF MCP/CD46 AND MHCI AND INDUCES PROTECTION AGAINST C-MEDIATED LYSIS

van den Berg(1,2), C.W.; Andrade, R.M.G.(2), Magnoli, F.C.(2), Marchbank, K.J.(3), Tambourgi, D.V.(2)

(1)Department of Pharmacology, Therapeutics and Toxicology, University of Wales, College of Medicine, UK, (2)Laboratório de Imunoquímica, Instituto Butantan, Brazil and (3)Department of Medical Biochemistry, University of Wales, College of Medicine, UK.

We have recently shown that sphingomyelinase D toxins from the spider Loxosceles intermedia induce Complement (C) dependent haemolysis of autologous erythrocytes by induction of cleavage of cell surface glycophorins through activation of a membrane bound metalloproteinase. The aim of this study was to investigate the effects of these toxins on C-regulator expression and the C-resistance of nucleated cells. Cells were incubated with Loxosceles venom/toxins and the expression of C-regulators was assessed by flow cytometry. A reduced expression of membrane cofactor protein (MCP) was observed, while expression of decay accelerating factor (DAF) and CD59 was not affected. Analysis of other cell surface molecules showed a reduced expression of MHCI. Western blotting showed that a truncated form of MCP was released into the supernatant. Release could be prevented by inhibitors of metalloproteinases of the adamalysin family but not by inhibitors specific for matrix metalloproteinases. Cleavage of MCP was induced close to or in the membrane as demonstrated by the cleavage of transmembrane chimeras of CD59 and MCP. Although the venom/toxins induced a release of MCP, the C-susceptibility was decreased. The mechanism of this induction of resistance may involve a change in membrane fluidity induced by the sphingomyelinase activity of the toxin/venom and/or involvement of membrane bound proteases. The identity of the metalloproteinase(s) activated by the spider venom and the role in pathology of Loxoscelism remains to be established.

Supported by the Wellcome Trust as a Collaborative Research Initiative Grant to DVT and CWB and a Royal Society Travel Grant to CWB

CORRESPONDENCE TO:

van den Berg, C.W., Laboratório de Imunoquímica, Instituto Butantan, Avenida Vital Brasil, 1500, São Paulo, 05503-900, SP, Brazil, E.mail: vandenbergcw@yahoo.com