Clarke S H, McCray S K
Department of Microbiology and Immunology, University of North Carolina at Chapel Hill 27599.
Eur J Immunol. 1993 Dec;23(12):3327-34. doi: 10.1002/eji.1830231240.
Five to fifteen percent of peritoneal B1 (CD5+) cells from unmanipulated mice produce antibodies that bind bromelain-treated mouse red blood cells and the hapten phosphatidylcholine (PtC). The majority of these B cells express either of two VH/V kappa gene combinations, VH12/V kappa 4 or VH11/V kappa 9. Both the VH11 and VH12 genes are rearranged to JH1 and encode third complementarity determining regions (CDR3) of restricted length and sequence. These and other observations argue strongly that PtC-specific B1 cells are antigen selected. To determine when selection of PtC-specific B1 cells begins in mice we have used the polymerase chain reaction to amplify VH12-D-JH1 rearrangements from livers of fetal and neonatal mice, and determined the CDR3 encoding sequences of individual clones. We find an unusually low ratio of productive (P) to non-productive (NP) rearrangements (0.4-1.0) at both developmental stages. P rearrangements in day 1 neonates are biased in D gene use and in the sequence and length of their deduced VHCDR3. These biases are similar to those of PtC-specific B1 cells in the adult peritoneum. D gene use and CDR3 length and sequence are significantly less biased among VH12 P rearrangements 2 to 3 days earlier in the day 18 fetal liver. We suggest that this rapid change in repertoire is due to positive ligand selection that is dependent on the sequence of VHCDR3. We suggest further that the majority of VH12-expressing cells are not ligand selected and consequently undergo programmed cell death. The evidence of restriction in day 1 neonatal livers and the low P/NP ratio in the fetus suggests that selection of VH12-expressing cells begins before birth.
未经过处理的小鼠腹膜B1（CD5 +）细胞中有5%至15%会产生能与菠萝蛋白酶处理过的小鼠红细胞和半抗原磷脂酰胆碱（PtC）结合的抗体。这些B细胞中的大多数表达两种VH/Vκ基因组合中的一种，即VH12/Vκ4或VH11/Vκ9。VH11和VH12基因都重排至JH1，并编码长度和序列受限的第三互补决定区（CDR3）。这些以及其他观察结果有力地表明，PtC特异性B1细胞是经过抗原选择的。为了确定PtC特异性B1细胞在小鼠体内何时开始选择，我们使用聚合酶链反应从胎儿和新生小鼠的肝脏中扩增VH12-D-JH1重排，并确定单个克隆的CDR3编码序列。我们发现在两个发育阶段，有生产性（P）重排与无生产性（NP）重排的比例异常低（0.4 - 1.0）。出生第1天的新生儿中P重排在D基因使用以及推导的VHCDR3的序列和长度方面存在偏向。这些偏向与成年腹膜中PtC特异性B1细胞的偏向相似。在出生第18天胎儿肝脏中，提前2至3天的VH12 P重排在D基因使用以及CDR3长度和序列方面的偏向明显较小。我们认为这种谱系的快速变化是由于依赖于VHCDR3序列的阳性配体选择。我们进一步认为，大多数表达VH12的细胞没有经过配体选择，因此会经历程序性细胞死亡。出生第1天新生儿肝脏中的限制证据以及胎儿中低P/NP比例表明，表达VH12的细胞的选择在出生前就已开始。
