Upregulation of DDR1 following the expression of LMP1 in primary human GC B cells. (A) Q-RT-PCR analysis of DDR1 mRNA levels following the expression of LMP1 in primary human GC B cells compared with cells transfected with an empty vector as a control. LMP1 expression was followed by the robust upregulation of DDR1 mRNA. Shown is 1 of 4 experiments performed in triplicate on different donor GC B cells. Normalized values were expressed relative to empty vector set to a value of 1. (B) FACS analysis of DDR1 expression in GC B cells transfected with LMP1 or an empty vector. Left panels show cells stained with only primary DDR1 antibody or rabbit secondary antibody and used as negative controls. Right panels show staining of cells with both antibodies following transfection with empty vector or LMP1. Cells transfected with empty vector show no change in DDR1 expression compared with cells stained with rabbit secondary antibody alone, indicating that DDR1 is not expressed in empty vector transfected GC B cells. LMP1 expression led to detectable DDR1 protein expression in almost a quarter of GC B cells. (C) DDR1 mRNA expression following the treatment of primary human GC B cells with CD40L (200 ng/mL) for 1 and 3 hours in comparison with unstimulated GC B cells for the same corresponding time points. Treatment with CD40L did not induce the upregulation of DDR1. In contrast, there was a robust increase in the expression of the known CD40 targets TNFAIP3 and ICAM1. Data shown is representative of 3 replicates with normalized values expressed relative to control set to a value of 1.