CpG DNA activation and plasma-cell differentiation of CD27- naive human B cells.
Unmethylated CpG DNA activation of naive CD27- B cells has been reported to require B-cell-receptor (BCR) cross-linking. We describe a culture system using CpG DNA with sequential steps for T-cell-independent activation of naive CD19+CD27- human peripheral blood B cells that induces efficient CD138+ plasma-cell differentiation. CD27+ and CD27- B cells were cultured in a 3-step system: (1) days 0 to 4: CpG, IL-2/10/15; (2) days 4 to 7: IL-2/6/10/15 and anti-CD40L; (3) days 7 to 10: IL-6/15, IFN-alpha, hepatocyte growth factor, and hyaluronic acid. Both CD27+ and CD27- B cells up-regulated intracytoplasmic TLR-9 following CpG DNA activation. CD27- B-cell activation required cell-cell contact. Both naive and memory B cells progressed to a plasma-cell phenotype: CD19lowCD20lowCD27+CD38+HLA-DRlow. Seventy percent of the CD27--derived CD138+ cells demonstrated productive V chain rearrangements without somatic mutations, confirming their origin from naive precursors. Plasma cells derived from CD27+ B cells were primarily IgG+, while those from CD27- B cells were IgM+. Our results indicate that under certain conditions, naive B cells increase TLR-9 expression and proliferate to CpG DNA stimulation without BCR signaling. In addition to its immunologic significance, this system should be a valuable method to interrogate the antigenic specificity of naive B cells.
Huggins
Jennifer
J
Department of Pathology, James P. Wilmot Cancer Center, University of Rochester Medical Center, NY 14642, USA.
Pellegrin
Tina
T
Felgar
Raymond E
RE
Wei
Chungwen
C
Brown
Miguel
M
Zheng
Bo
B
Milner
Eric C B
EC
Bernstein
Steven H
SH
Sanz
Ignacio
I
Zand
Martin S
MS
D001402:B-Lymphocytes / Q000166:cytology*; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D018899:CpG Islands; D016207:Cytokines / Q000494:pharmacology; D004247:DNA / Q000502:physiology*; D006801:Humans; D008213:Lymphocyte Activation; D010950:Plasma Cells / Q000166:cytology*; D051217:Toll-Like Receptor 9; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7
D016207:Cytokines; C495371:TLR9 protein, human; D051217:Toll-Like Receptor 9; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7; D004247:DNA