A lectin model for unresponsiveness. I. Reversible effects of high doses of concanavalin A on spontaneous and induced DNA synthesis

A mitogen model for unresponsiveness has been studied. In 72 hr cultures the lowest supraoptimal dose of Concanavalin (Con A) showing no stimulation of rat thymocytes above background was 160 μg/ml. This dose was also found to inhibit substantially (50%) the high spontaneous DNA synthesis by thymocytes during the first 24 hr of culture. This effect was most significant 12 to 24 hr after addition of high dose Con A. It was not necessary for the cells to remain continuously with high dose Con A, since washing out unbound Con A after 1 hr still led to reduced [³H] thymidine incorporation at 24 or 72 hr. Furthermore, high dose Con A treatment suppressed DNA synthesis when added at 48 hr to cultures optimally stimulated with Con A. Interaction with high dose Con A appears to inhibit the ability of cells to initiate new rounds of DNA synthesis without blocking ongoing DNA replication. Removal of Con A with α methyl D mannopyranoside (mannose) led to only partial reversal of these effects after 1 hr although 'unresponsiveness' was reversible at 15 min. Suppression was not due to cytotoxic or agglutinating effects of the high dose Con A. Two x 10⁶ thymocytes exposed to 160 μg of Con A for 1 hr bound 1.7 μg, 86% of which was elutable with mannose. There was no reduction in the ability of high dose treated cells to bind a subsequent optimal dose of Con A. Since Con A binds to essentially all thymocytes, this system is useful in following early events in the induction of lectin 'unresponsiveness' as a model for immunologic tolerance.