Human B cell activation and cell cycle progression: stimulation with anti‐μ and Staphylococcus aureus Cowan strain I

The responses of resting human B lymphocytes to a variety of activation signals were studied. Human tonsillar B lymphocytes were separated according to size by countercurrent elutriation. The small B lymphocytes were then stimulated in vitro with various concentrations of anti‐μ antibody in the presence or absence of B cell growth factor (BCGF) or with Staphylococcus aureus Cowan strain I (SAC). Cellular volume changes and RNA synthesis were measured over the first 24 h of stimulation and were similar with either 15 μg/ml of anti‐μ, 100 μg/ml of anti‐μ, or SAC. In the subsequent 24 h, however, substantial increases occurred in the amount of RNA synthesis and cell enlargement only in those cultures stimulated with 100 μg/ml of anti‐μ or SAC, but not in the cultures stimulated with 15 μg/ml of anti‐μ. The addition of BCGF to those cultures stimulated with 15 μg/ml of anti‐μ did not alter the increases in cellular volume and RNA synthesis found 24 h after stimulation with anti‐μ alone. However, over the subsequent 24 h, the presence of BCGF in culture enhanced both B cell volume changes and RNA synthesis, when compared to cultures stimulated with 15 μg/ml of anti‐μ alone. In addition, BCGF enhanced DNA synthesis in cultures stimulated with low and high concentrations of anti‐μ. DNA content changes following stimulation with anti‐μ, anti‐μ plus BCGF, and SAC were also measured using propidium iodide staining and flow cytometry. Optimal concentrations of anti‐μ induced 20% of the resting B cells to enter S phase, while optimal concentrations of anti‐μ plus BCGF or SAC induced approximately 40%. Finally, prestimulation of resting B cells for 24 h with a low concentration of anti‐μ, sufficient for cell enlargement but not S phase progression, allowed for rapid entrance of the prestimulated B cells into S phase when a high concentration of anti‐μ or SAC was added. These findings suggest the existence of a control point in the progression of human B cells through the cell cycle. This control point is located in the G₁ phase of the cycle and is reached 24 to 36 h after a surface immunoglobulin‐mediated stimulus.