The characteristics of the interleukin (IL) 5‐reactive splenic B cell population of C57BL/6 nu/nu mice, with respect to IL5/IL2 reactivity, cell surface phenotype, VH gene family usage, autoreactivity and the structure of the IL5 receptor (IL 5R), 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 IL 5R+ B cell population. The cell surface phenotype of the IL 5R+ splenic B cells is IgM+, B220+, Ly‐1− and IL 2R p55−. Upon activation with IL 5 this cell surface phenotype changes, in that a vast majority of the B cells then express the p55 chain of the IL 2R, whereas the level of IL 5R 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 IL 5R 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 M IL5‐binding sites are present on IL5‐activated splenic B cells, and both high‐ and low‐affinity IL 5R 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. Thus, we conclude that the structure of the IL 5R on splenic B cells is identical to that on the IL5‐dependent B13 cell line, the IL5R+ splenic B cell population does not differ from the vast majority of the rest of the splenic B cell pool with respect to surface phenotypes (except for expressing IL 5R), VH gene family utilization and autoreactivity.
ASJC Scopus subject areas
- Immunology and Allergy