ASPECTS OF THE IMMUNE RESPONSE AGAINST TOXIC PROTEIN IRRADIATED WITH 60CO GAMMA RAYS

CAPRONI P.(1), BAPTISTA J. A.(1), SPENCER P. J.(1), HIGA O. Z.(1), ANDRADE JR. H. F.(2), NASCIMENTO N.(1)

(1)Laboratório de Química de Proteínas, Centro de Biologia Molecular, Instituto de Pesquisas Energéticas e Nucleares, (2)IMTSP - Instituto de Medicina Tropical de São Paulo, Laboratório de Protozoologia

Ionizing radiation has been successfully employed to modify the immunological properties  of  biomolecules. Very promising results were obtained when crude animal venoms, as well as isolated toxins, were treated with gamma rays, yielding toxoids with good immunogenicity. The obtention of modified antigens with lower toxicity and preserved or improved immunogenicity would be useful. Ionizing radiation has proven to be a powerfull tool to attenuate snake venoms toxicity without affecting and even increasing their immunogenic properties. However, little is known about the modifications that irradiated molecules undergo and even less about the immunological response that such antigens elicit. In the present work, we investigated the immunological behavior of bothropstoxin-I, a K49 phospholipase, before and after irradiation. Structural modifications of the toxin were investigated by SDS-PAGE. Aiming to compare the toxicity of the between native and irradiated forms of the toxin, an in vitro citotoxicity assay, using CHO cells, was performed. Isogenic mice were immunized with either the native or the irradiated toxin.The circulating antibodies were isotyped and titrated by ELISA. According to our data, irradiation promoted structural modifications in the toxin, characterized by higher molecular weight forms of the protein (aggregates and oligomers). The citotoxicity assay showed that the modified toxin was 5 folds less toxic than its native counterpart. Our data indicate that irradiated toxins were immunogenic and the antibodies elicited by them were able to recognise the native toxin in ELISA. These results indicate that irradiation of toxic proteins can promote significant modifications in their yours structures, but still retain many of the original antigenic and immunological properties of native proteins. Also, our data indicate that the irradiated protein induced higher titers of IgG2a and IgG2b, suggesting that Th1 cells were predominantly involved in the immune response.

KEY WORDS: Immunological ionizing radiation, bothropstoxin-1.

FINANCIAL SUPPORT: CNPq

CORRESPONDENCE TO:

Priscila Caproni, Laboratório de Química de Proteínas,  Centro de Biotecnologia, IPEN/CNEN, São Paulo, Av. Professor Lineu Prestes, 2242, CEP 05508-900, São Paulo, SP. Email: pricaproni@hotmail.com