Induction of Immunity and Tolerance in Vitro by Hapten Protein Conjugates

Abstract

A critical stage in the induction of antibody responses is the interaction of antigen molecules with the surface receptors of immunocompetent cells. This interaction may result in either immunity, or its opposite, immune tolerance. The differences between the pattern of antigen binding to nonthymus-dependent lymphocytes (B cells) 1 in immunity and tolerance are not fully understood. Recently, attempts were made to study this problem in tissue culture, using dinitrophenylated polymeric flagellin (DNP POL) as antigen (1). The interaction of antigen molecules with cell membranes may be readily studied in dissociated lymphoid cell populations in vitro, where antigen concentrations are relatively uniform and catabolism negligible compared with tile situation in vivo (2). Furthermore, since tile response to DNP POL in vitro does not require the participation of either thymus-dependent lymphocytes (T cells) (1, 3) carrier-reactive cells, or macrophages (4, 5), only the pattern of reaction of DNP POL molecules with B cells determines the ensuing immunological events. It was found that the number of 2,4-dinitrophenyl (DNP) haptenic groups present per monomeric unit of the DNP POL molecule (i.e. the epitope density) dictated its capacity to induce either immunity or tolerance (1). Antigen molecules with few DNP groups immunized, but never induced tolerance, whereas highly conjugated DNP POL induced tolerance, but never immunity (1). Based on these observations, a mechanism of B celt signal discrimination between immunity and tolerance was proposed, based on the number

In vitro tolerance induction of neonatal murine B cells.

“…The evidence presented in this study also confirms previous reports that demonstrated that soluble antigens may bind to mature B cells and even inhibit stimulation, but, if the antigen is removed before stimulation, the B cells remain unaffected (45,46). Therefore, it appears that the "one signal" mode (antigen binding to B cells) is irrelevant to mature B cells but tolerogenic to developing B cells (Table IV).…”

INDUCTION OF B CELL TOLERANCE IN VITRO TO 2,4-DINITROPHENYL COUPLED TO A COPOLYMER OF D-GLUTAMIC ACID AND D-LYSINE (DNP-D-GL)

“…In the present context, its main usefulness was to yield higher plaque numbers in control cultures of spleen cells from nude mice, and it is of interest that the nonresponsiveness caused by DNP-D-GL could be maintained despite the addition of this adjuvant-like material. Table I also shows that concentrations of DNP-D-GL of 10 ng/ml and above gave significant impairment of DNP- (8). However, this would require that DNP-I)-GL remain at the lymphocyte surface and not be removed by "capping" and receptor pinocytosis (13, 14), since DNP-FLA was present continuously during the 3 day challenge period.…”

“…Of course, each molecule carries many DNP residues, so one could envisage efficient receptor cross-linking with DNP-D-GL. Yet highly DNP-substituted albumins and globulins are not good tolerogens in vitro (2,8). A careful stud)-of tolerogenicity with hapten substitution rate as the key variable would be revealing.…”

“…Modifications as previously described from our laboratories (6)(7)(8)12) were introduced for measuring hapten-or proteinspecific PFC. Each result cited is the mean of 4-16 cultures.…”

Ultrastructural Radioautographic Studies on Capping and Endocytosis of Anti‐immunoglobulin by Mouse Spleen Lymphocytes

“…This is further suggested by the results of Greaves and Baumingcr (1972), who found that phytohacipaggiutiiiin and pokeweed mitogen coupled to Sepharose beads stimulated T and B cells, although the mitogens could not be taken into the cell. The degree of binding and extent of cross-linking between receptors may provide the method of signal discrimination between immunity and tolerance, as suggested by Feldmann (1972). Binding of the Ag to its receptor may then cause a conformational change which results in the activation of a membrane-associated enzyme, such as aden)l cyclase, to produce a "second messenger" carrying the signal to the nucleus.…”