| Autor: |
Christiansen, D., Milland, J., Thorley, B. R., McKenzie, I. F. C., Mottram, P. L., Purcell, L. J., Loveland, B. E. |
| Předmět: |
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| Zdroj: |
Immunology; Mar96, Vol. 87 Issue 3, p348-354, 7p |
| Abstrakt: |
Human CD46 (membrane cofactor protein) is a type 1 glycoprotein that functions to protect autologous cells from complement-mediated damage by binding C3b and C4b for their factor I-mediated cleavage. We now describe the production and function of recombinant soluble CD46 (rsCD46), which was produced as a truncated form by mutagenesis using the splice overlap extension polymerase chain reaction, by inserting a translational stop codon into the CD46 cDNA at the junction of the transmembrane and extracellular domains. After transfection of an expression construct into 293-EBNA (Epstein-Barr nuclear antigen)-transformed cells, secretion of rsCIM6 protein was detected by immunoradiometric assay using monoclonal antibodies. Following a single-step immunoaffinity purification, the protein resolved as a single band of ∼ 56000 MW on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The purified rsCD46 (51 μg/ml) protected Chinese hamster ovary (CHO)cells from lysis initiated by a high titre rabbit anti-CHO antibody and complement from rabbit or human. The protection was specifically mediated by rsCD46 because the monoclonal antibody M 177, which blocks interaction between CD46 and C3b/C4b, abrogated the protection. The results demonstrate that rsCD46 is effective as a fluid-phase regulator of complement activation on cell surfaces, even when initiated by the classical complement pathway. The in vivo efficacy of rsCD46 was investigated using a mouse heart to rat xenograft model. Adminstration of a bolus injection of rsCD46 was effective at delaying hyperacute graft rejection. These data suggest that rsCD46 may have a role as a therapeutic agent. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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