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Maintenance of human germinal center B cells in vitro
JD Pound and J Gordon
Department of Immunology, Medical School, University of Birmingham, UK.
The ability to maintain germinal center (GC) B cells in culture should
facilitate studies on the molecular and cellular events which accompany
affinity maturation and the generation of memory in T-dependent responses.
We have investigated the ability of cytokines to maintain human tonsillar
GC B cells (IgD-/CD39-/CD38+/CD77+) in the "CD40 culture system". In the
absence of added cytokines, CD40 monoclonal antibody held on
CD32-transfected L cells effectively sustained DNA synthesis in GC B cells
for a maximum 3 to 4 days. Of the following cytokines (interleukin-1 beta
[IL-1 beta], IL-2, IL-3, IL-4, IL-6, IL- 7, IL-10, and stem cell factor),
only IL-2 and IL-4 provided a significant enhancement to DNA synthesis in
the CD40 culture system; this was modest and short-term. Following a study
on the cooperative activity between pairs of cytokines, triple combinations
were identified that could maintain high levels of GC B-cell stimulation
for at least 10 days. IL-10 was a common component of these synergistic
cytokine cocktails, which were IL-10 + IL-4 + IL-7; IL-10 + IL-3 + IL- 7;
IL-10 + IL-1 beta + IL-2; IL-10 + IL-1 beta + IL-3, and IL-10 + IL-3 +
IL-6. Culture of GC B cells with these cytokine combinations resulted in a
net increase in viable cell numbers of 50% to 100% whereas total cell
numbers increased up to fourfold. Cells recovered from these cultures
retained a GC B-cell phenotype with a significant proportion being
CD38+/CD44-, features characteristic of centroblasts. Studies with
metabolically inactive CD32-L cells supported a role for stromal
cell-derived soluble factors in maintaining GC B cells in vitro.
Volume 89,
Issue 3,
pp. 919-928,
02/01/1997
Copyright © 1997 by The American Society of Hematology
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