Molecular single-cell analysis reveals that CD5-positive peripheral blood B cells in healthy humans are characterized by rearranged V-kappa genes lacking somatic mutation

B cells expressing the CD5 cell surface antigen are involved in certain B cell malignancies and autoimmune diseases, From studies in the mouse, it emerged that CD5(+) B cells represent a separate lineage of B lymphocytes that, in contrast to conventional (CD5(-)) B cells, are not driven into T cell-dependent immune responses in which rearranged variable (V) region genes are diversified by somatic hypermutation. Against this background it came as a surprise that human disease-involved CD5-positive autoreactive B cells as well as B cell chronic lymphocytic leukemias can harbor somatically mutated V region genes, Recent V gene analyses on CD5(+) B cells in healthy adults did not give rise to a clear picture about the fraction of somatically mutated among all CD5(+) B cells, In this work we used a molecular single-cell analysis to determine reliably the frequency of mutated CD5(+) B cells in healthy humans: single, kappa light chain-expressing CD5(+) peripheral blood B cells were isolated by flow cytometry, and rearranged V kappa genes were amplified by PCR, From one donor, CD5(+)CD19(+) B cells were analyzed, Since CD5(+) B cells were found among IgM(+)IgD(+) and IgM(+)IgD(-) cells (but almost not among class-switched cells) from two other donors, individual cells corresponding to these IgM-expressing subsets were investigated separately, The sequence analysis of rearranged V-kappa genes revealed that most if not all CD5(+) B cells in healthy humans carry unmutated V region genes, From one of the donors, a novel polymorphic J(kappa)2 gene segment was identified. To explain the discrepancy between the frequent occurrence of disease-associated somatically mutated CD5(+) B cells and the low incidence or absence of somatic mutation in normal CD5(+) B cells, we speculate that CD5(+) B cells usually do not participate in germinal center reactions, but if they occasionally do so, they may be at an increased risk to become involved in autoimmune diseases or B cell malignancies.