Elevated CD154 (CD40 ligand) synthesis in T-cells from allergic patients after nonspecific stimulation in vitro

The interaction of the CD154 molecule (CD40 ligand, gp39) on activated T-cells with the CD40 antigen on B-cells seems to play a key role in immunoglobulin class switching. We aimed to compare the capacity of intracellular CD154 expression after nonspecific stimulation with phorbol-12-myristate-13-acetate and ionomycin on separated T-cells from allergic patients and healthy donors. We analyzed blood from 104 patients allergic to grass pollen, house dust mites or birch pollen, and from 44 healthy donors. Lymphocytes were isolated using a density gradient and B-cells were extracted by magnet-activated cell separation (MACS) using anti-CD19 microbeads. Cells were nonspecifically stimulated for 5 h, permeabilized and stained with anti-CD154 for fluorescence-activated cell sorter analysis. It was found that stimulation induced a 1.4% increase of intracellular CD154+ T-cells; a 4.6% increase of mean channel fluorescence of all T-cells from healthy donors; a 6.1% increase in intracellular CD154+ T-cells; and a 28.1% increase of mean channel fluorescence of all T-cells from allergic patients. The data demonstrated an elevated capability of B-cell independent CD154 synthesis in T-cells from allergic patients when compared to healthy individuals. It is possible that the enhanced IgE production of B-cells from allergic patients might be partly due to the phenomena described.