PRELIMINARY STUDIES ON THE INTERACTION OF TNF-ALPHA AND IFN-GAMMA WITH ALPHA-2-MACROGLOBULIN

The binding of I-125-labelled recombinant human TNF-alpha and IFN-gamma to isolated human blood alpha(2)-macroglobulin has been investigated using molecular sieving procedures and non-denaturing PA gel electrophoresis in combination with autoradiography. These studies revealed that both cytokines readily bind to the electrophoretically fast form of alpha(2)M generated by methylamine or protease treatment of this protein. PAGE/SDS gel investigations indicated that TNF-alpha bound non-covalently while the IFN-gamma interaction was covalent in nature. Preliminary competition studies also indicate that cold TNF-alpha and IL-2 are more effective than cold IFN-gamma at inhibiting the binding of labelled IFN-gamma to alpha(2)M. Bioassays revealed that "native" alpha(2)M or its derivatives at 2 mg/ml concentration did not impair the antiproliferative effects of TNF-alpha and IFN(gamma) on susceptible bladder tumour cell lines. Furthermore they did not interfere in the induction of Class II antigen expression by IFN(gamma) on inducible cell lines or in a 2-site ELISA assay for TNF.