Emerging human B cell repertoire. Influence of developmental stage and interindividual variation.

Abstract

In B cell precursors developing in fetal lymphopoietic tissue, the selection of VH, DH, and JH gene segments for initial H chain gene assembly is biased. The present study was designed to determine whether these biases persist in fully developed human fetal B cells and to examine specificities encoded by the favored elements. B cells were prepared from two sites representing different stages of development, i.e., fetal liver as a source of cells newly generated in that lymphopoietic environment and fetal spleen as a source of more mature cells, potentially subject to selective environmental factors. The expressed repertoires were sampled by two methods. EBV transformation so binding and structure could be examined simultaneously and generation of cDNA from individual, sorted, unstimulated B cells. We found that mature B cells in liver and secondary lymphoid tissue exhibit the same degree of bias in VH use we previously reported in lymphopoietic tissue of the same gestational age. However, the pattern of DH and JH use more nearly resembled that of the adult, suggesting that some constraints imposed by the rearrangement process are normalized rapidly. Sequences recovered from EBV-transformed clones were not distinguishable from transcripts recovered from single cells by direct amplification. Among antibodies expressed by the EBV clones, binding to self-Ag was common, binding profiles varied, and, in contrast to mice, there was little relationship between specificity and VH element. Interestingly, the two individuals studied differed in the VH element most commonly used. One resembled previously studied fetal repertoires in that VH56p1 encoded about 20% of expressed antibodies, whereas the other did not express VH56p1 and used VH26 in 25% of expressed antibodies. This was found to reflect a lack of the genomic VH56p1 allele, suggesting that genetic variation at the VH locus may significantly influence the emerging human antibody repertoire.