Mechanism of b-lymphocyte activation: failure to obtain evidence of a direct role of the Ig receptors in the triggering process

Abstract

Experiments were designed to test two hypotheses of B-cell activation by antigen: the cross-linking concept, postulating that a suitable degree of antigen-induced cross-linking of the Ig receptors is sufficient for immunocyte triggering, and the two-signal hypothesis, suggesting that a first signal delivered by antigen interacting with the Ig receptors followed by a second signal given by, for example, a polyclonal B-cell activator is necessary for activation. The results did not support either of these hypotheses. Thus, the hapten FITC coupled to human serum albumin and human gammaglobulin in different conjugation ratios failed to activate B cells, whether the hapten-protein conjugates were soluble or precipitated, whether the experiments were carried out in the presence or absence of different concentrations of sera from different species, and irrespective of the day of assay. Furthermore, the same FITC-protein conjugates or FITC itself coupled to Sepharose particles failed to induce a specific anti-FITC response, even though a range of 10-9-fold concentrations of FITC were used. In contrast, FITC coupled to lipopolysaccharide (LPS) regularly induced a primary anti-FITC response in all the above systems, whether FITC-LPS was soluble or coupled to Sepharose particles. The conjugation ratio of FITC to LPS was within the range of epitope densities used with FITC-protein conjugates. Analogous studies were performed with the above compounds and, in addition, NNP-cap and fowl gammaglobulin, added alone or together with LPS to lymphocyte cultures. In no case did the antigen plus LPS give a better specific anti-FITC response than LPS alone, irrespective of the culture conditions, the epitope densities, the physical form of the conjugates, and whether they were bound to Sepharose particles or not, although this would be expected in terms of the two-signal concept. The results are compatible with the one nonspecific signal hypothesis, ascribing a passive role to the Ig receptors and an active triggering function to thymus-independent antigens. Therefore, the ability to trigger B cells directly will depend on the nature of the carrier, triggering being achieved if the carrier is a polyclonal B-cell activator; the epitope density and the degree of cross-linking of Ig receptors are unimportant for delivering the triggering signal, although they can facilitate the binding of the conjugate to the specific B cells.