The tolerance inducing properties in rats of bacterial flagellin cleaved at the methionine residues

Abstract

Flagellin (molecular weight 40,000) from Salmonella adelaide was reacted with CNBr to give mainly four peptide fragments, A, B, C and D, the largest of which (fragment A, molecular weight 18,000) retained all the serological activity of flagellin itself. Mixtures of the fragments or individual fragments were injected into neonatal or adult rats, either as a single dose in saline or in adjuvant, or in multiple doses in saline. Antibody titres, measured by bacterial immobilization, were compared with those given after an injection of flagellin. Fragment A and the complete CNBr digest of flagellin (the `digest') were both immunologically active whereas fragments B, C and D were inactive.

When given as a single injection in saline into adult rats, the `digest' or fragment A was much less efficient than flagellin at inducing either a primary antibody response or the production of primed cells for a secondary antibody response. In contrast, the `digest' or fragment A was effective at triggering primed cells to give a secondary antibody response. These findings were consistent with an interpretation given in detail elsewhere relating to `in vivo' localization of labelled flagellin and fragment A in rat lymph nodes to the immune response. Fragment A and flagellin were equally immunogenic when injected in Freund's complete adjuvant.

Rats given a course of injection of the `digest' starting on the day of birth became almost completely tolerant to either flagellin or polymerized flagellin. Daily injections of the `digest' or fragment A for 4 weeks or longer into adult rats resulted in a significant degree of tolerance to flagellin and to polymerized flagellin. Adult rats made tolerant in this way responded normally to BSA injected in Freund's complete adjuvant and to a slightly decreased extent to sheep RBC.

It was concluded that by a process of partial degradation, a highly immunogenic substance, polymerized flagellin, had been converted into a preparation with strong tolerance-inducing properties. The relevance of this approach to transplantation antigens was discussed.