Characterization of the interleukin 5‐reactive splenic B cell population

Abstract

The characteristics of the interleukin (IL) 5-reactive splenic B cell population of C57BL/6 nu/nu mice, with respect to IL 5/IL2 reactivity, cell surface phenotype, VH gene family usage, autoreactivity and the structure of the IL5 receptor (IL5R), were analyzed. It was found that 2%-4% of splenic B cells express relatively high levels of IL 5R as determined by the binding of the anti-IL 5R monoclonal antibody R52.120. Over 90% of the splenic B cells that mature to IgM secretion upon activation with IL5 are comprised in this small subpopulation of B cells. Moreover, the vast majority of splenic B cells that mature to IgM-secreting cells when activated by IL2 also reside in this IL5R+B cell population. The cell surface phenotype of the IL5R+ splenic B cells is IgM+, B220+, Ly-1- and IL2R p55-. Upon activation with IL5 this cell surface phenotype changes, in that a vast majority of the B cells then express the p55 chain of the IL2R, whereas the level of IL5R decreases. VH gene family usage in the IL5-activated splenic B cells was analyzed by in situ hybridization. VH gene family usage was found to be random and not different from the VH genes expressed in LPS-activated B cells. Hybridoma collections from IL5-activated splenic B cells and LPS-activated B cells were screened and compared for the production of autoantibodies and antibodies directed against the haptens (4-hydroxy-3-iodo-5-nitrophenyl)acetyl (NIP) and 2,4,6-trinitrophenyl (TNP). In both collections high, but not significantly different frequencies of autoantibody-(32% IL5, 31.4% LPS) and of anti-hapten antibody (27.8% IL5, 18.6% LPS)-producing hybridomas were found. The structure of the IL5R on IL5-activated B cells was analyzed by 125I-labeled IL5 binding and cross-linking. About 100 high-affinity (10(-11) M) and 1000 low-affinity (10(-9) M) IL5-binding sites are present on IL5-activated splenic B cells, and both high- and low-affinity IL5R are similar to those expressed on the IL5-dependent B13 cell line. Cross-linking of 125I-labeled IL5 to the receptors on IL5-activated B cells revealed one major IL5-binding protein of 45-50 kDa molecular mass and another minor binding protein of 130-140 kDa. The same IL5-binding proteins are present on the IL5-dependent B13 cell line.(ABSTRACT TRUNCATED AT 400 WORDS)