THE MECHANISM OF TOLERANCE PRODUCED IN RATS TO SHEEP ERYTHROCYTES. II. THE PLAQUE-FORMING CELL AND ANTIBODY RESPONSE TO MULTIPLE INJECTIONS OF ANTIGEN BEGUN AT BIRTH.

An active immune response to sheep erythrocytes was demonstrated in rats made "tolerant" to sheep erythrocytes by twice-weekly antigen injections beginning on the day of birth. Groups of tolerant rats were sacrificed 4 days after they had received 5 to 42 antigen injections; spleens were sampled for plaque-forming (antibody-releasing) cells and sera were titrated for antibody to sheep erythrocytes using a sensitive "plate hemolysin" technique. During the 3rd week of life and after the 5th antigen injection, the tolerant rats had an immune response equivalent to that of rats of similar age which had received a single antigen injection, but spleens contained only about one-tenth as many plaque-forming cells as adults animals receiving similar antigen injections. Continued antigen injections produced a marked decline and stabilization of this relatively small population of antibody-forming cells; however, the number of plaque-forming cells in the tolerant rats remained considerably elevated above the numbers of plaque-forming cells present in the spleens of non-immunized animals. The sera from all but 1 tolerant rat had demonstrable antibody to sheep erythrocytes in low titer. A progressive recovery of the plaque-forming cell response and rise in antibody titers occurred in adult tolerant rats when the interval between the last 2 antigen injections was increased from 3 days to 14 or 28 days. The decline and stabilization of numbers of plaque-forming cells occurring with continued injections after the 3rd week of life paralleled a similar decline and stabilization in rats receiving similar antigen injections as adults. Also, the recovery of the plaque-forming cell and antibody response of tolerant animals paralleled the recovery observed when the interval between injections was increased in rats receiving similar antigen injections as adults. These findings suggested that the same mechanism controlled numbers of antibody-forming cells in tolerant and normally responsive adult animals. Repeated closely spaced antigen injections presumably interfered with either cell division or maturation of antibody-forming cells. As the interval between injections was increased, additional antibody-forming cells matured or were formed through cell division. Relatively constant antigenic stimulation provided a mechanism for controlling or limiting the response of antibody-forming cells. The mechanism controlling or limiting the response of antibody-forming cells would not account for the stabilization of numbers of antibody-forming cells at high levels for normal animals and at low levels for the tolerant animals. Passive immunization of growing rats with homologous anti-sheep erythrocyte serum markedly inhibited the plaque-forming cell response of growing rats. It was proposed that antibody produced by the small population of antibody-forming cells in the tolerant rats provided a feedback or homeostatic mechanism which inhibited transformation of potential antibody-forming cells to antibody-forming cells. Thus, tolerance to sheep erythrocytes was induced and maintained by two mechanisms. One mechanism, dependent on relatively constant antigenic stimulation, limited or controlled the numbers of antibody-forming cells. The other, dependent on the production of small quantities of antibody by a few antibody-forming cells, limited or controlled the transformation of potential antibody-forming cells to antibody-forming cells.