Elimination of Self-Reactive B Lymphocytes Proceeds in Two Stages: Arrested Development and Cell Death

Cell 1993, 72:325-335

By Suzanne B. Hartley, Michael P. Cooke, David A. Fulcher, Alan W. Harris, Suzanne Cory, Antony Basten, and Christopher C. Goodnow

In normal individuals, B cells that recognize self-antigens undergo a screening and censoring process to eliminate their ability to produce autoantibodies. Two key mechanisms have been identified to play a major role: clonal anergy, which functionally silences B cells without killing them and outright clonal deletion.

The elimination of self-reactive B cells is triggered by the binding of self-antigen to surface IgM molecules on immature B cells of the bone marrow. Much less is known about the induction of clonal anergy (tolerance). In a series of papers in this volume, the authors have investigated the elimination of self-reactive B cells in transgenic mouse models. In a single-transgenic (ST) model, B cells are engineered to express an antibody to hen egg lysozyme (HEL). In a double-transgenic (DT) model, in addition to producing antibodies to HEL, the mice are engineered to also express or secrete the HEL antigen.

In the ST mice, mature B cells expressing anti-HEL occur at a high frequency, whereas in the DT mice they are absent. Nonetheless, the DT mice did have normal numbers of immature HEL-reactive B cells which expressed low levels of IgM. Labeling studies of these immature HEL-positive B cells from both ST and DT mice suggested that they were rapidly turning over (half life of one day) and were completely replaced within three days. Since the DT mice produced few mature B cells, their immature cells must have been dying in one to three days. In cell culture studies, ST or DT immature B cells, in the absence of HEL, increased IgM expression 20-fold in 37 hours suggesting a maturation or activation of the cells. Other markers also increased. By contrast, in the presence of HEL, B cells from DT mice did not increase IgM or any of the other markers and B cells from ST animals that had increased IgM, showed a 20-fold decrease in IgM to look very similar to the DTs.

In a subsequent series of studies, the authors constructed transgenic mice whose B cells contained the bcl-2 gene expressed in their B cells to make them immortal (ST* or DT*). In ST* animals, immature B cells were found in the bone marrow but not in other lymphoid organs. By contrast, DT* animals had even more immature B cells in the marrow, which were also found in the spleen, blood or lymph nodes. When these cells were then placed in culture in the presence of a nonspecific B cell activator, they then expressed IgM and began to mature.

These findings suggest that constant exposure to antigen leads to arrested development of immature B cells and eventually to their death. The data argue that this is a two step process. In both ST and DT mice, immature B cells were found in the marrow, but were not dead. This could also be mimicked in culture. Furthermore, constant exposure to bcl-2 inhibited death, but did not block the cell's arrested development as immature B cells. Two other observations are worth noting. The lack of expression of differentiation makers on arrested immature cells may have limited their ability to target other immune organs. Also, the fact that immature B cells could be activated in culture suggests that they were not yet committed to irreversible programmed cell death.