Expression of CD5 and CD38 by human CD5− B cells: Requirement for special stimuli
Authors: Zupo Simona , Dono Mariella , Massara Rosanna , Taborelli Giuseppe , Chiorazzi Nicholas , Ferrarini Manlio 
Published: March 2007

In this study the mode of expression of CD5 by human tonsillar CD5− B cells after stimulation with different agents was investigated. Resting B cells were separated into CD5+ and CD5− cells and the two cell fractions exposed to phorbol 12‐myristate 13‐acetate (PMA). CD5− B cells expressed CD5 and maximum CD5 expression was achieved after approximately 60 h of culture. Based upon the proportions of cells that express CD5 as well as those of the cells surviving in culture, it was calculated that 15–25 % of the total CD5− B cells were induced to express CD5. Unlike CD5− B cells, CD5+ B cells proliferated vigorously in response to PMA as assessed by [3H] thymidine incorporation and cell cycle analysis in vitro. However, the expression of CD5 by CD5− B cells was not related to the selective expansion of some CD5+ B cells left over as contaminant cells since this occurred in the absence of cell proliferation. Upon exposure to PMA, CD5− B cells remained in the G0‐G1 phases of the cell cycle and did not express the Ki67 antigen or incorporate [3H] thymidine. Furthermore, mitomycin C treatment of the CD5− B cells did not prevent CD5 expression. Phenotypic studies disclosed that CD5+ B cells but not CD5− B cells expressed CD39. This finding offered the opportunity to carry out an additional control experiment. Separation of the two populations according to the expression of CD39 confirmed the finding obtained by fractionating the cells into CD5+ and CD5− B cells. The cells induced to express CD5 also expressed CD38 that was not detected on resting CD5− B cells. In this respect, the CD5− B cells that converted into CD5+ cells (inducible CD5+ B cells) resembled the cells from the CD5+ B cell fractions that up‐regulated CD5 and also expressed CD38 upon exposure to PMA alone. Another example of coordinate expression of these two antigens was the finding that exposure to PMA in the presence of recombinant interleukin‐4 (rIL‐4) resulted in inhibition of the expression of CD5 and CD38. Although virtually all of the tonsillar CD5− B cells expressed the CD69 activation marker, no cells other than those co‐expressing CD5 and CD38 were induced to express CD5 by PMA alone. Resting CD5− B cells failed to express CD5 and/or CD38 when cultured with PMA in the presence of EL4 T cells and IL‐4‐free T cell supernatants. Although this combination of stimuli induced a vigorous cell proliferation, the failure to express CD5 and CD38 was not related to cell cycling, since mitomicyn C‐treated CD5− B cells also failed to express CD5 or CD38 when exposed to PMA in the presence of EL4 cells with or without T cell supernatants. Thus, exposure to T cells alone was sufficient to down‐regulate CD5 and CD38 expression. Collectively, the above findings indicate that mature CD5− B cells can follow distinct pathways of differentiation depending upon the nature of the stimuli encountered, and that CD5 expression may identify a special B cell subset or a particular stage of B cell differentiation.