Mitogen-induced B-cell differentiation in Xenopus laevis

Spleen cells from the South African clawed toad Xenopus laevis proliferate vigorously upon pokeweed mitogen (PWM) stimulation, as measured by 3H-thymidine uptake. The peak response is observed after 6-8 days of culture, and both light- and electron-microscopy examinations of stimulated cells reveal the presence of a large number of lymphoblasts. Beside proliferation, PWM induces the in vitro differentiation of a large proportion of lymphoblasts into immunoglobulin (Ig)-producing plasmablasts, as shown by direct immunofluorescence. On average, 25% of the lymphoblasts contain cytoplasmic high-molecular-weight Ig (IgM) at days 8-10, whereas less than 1% of the lymphoblasts have cytoplasmic low-molecular-weight Ig (IgY). Ig's of mitogen-stimulated cells were labeled biosynthetically and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE). The results confirm that Xenopus lymphoblasts induced in vitro synthesize Ig polypeptide chains. These are assembled into monomers of H2L2 units or polymerized hexameric forms. Both the monomer and hexamer forms are secreted. Two Ig heavy chains with apparent molecular weights (MW) of 74 and 81 daltons (kd) are detected in the cell lysates. The secreted chains have respectively an MW which is 3 kd and 5 kd greater than the intracellular forms. The two-dimensional gel electrophoretic pattern of PWM-induced Ig's is as heterogeneous as that of serum IgM. We conclude that PWM induces the in vitro proliferation and polyclonal differentiation of a large proportion of splenic B lymphocytes into plasmablasts.