Expression of BLNK and Syk is increased in cGVHD B cells ex vivo. Relative mRNA expression of 84 genes known to be involved in lymphocyte activation were analyzed in CD27⁺ and CD27⁻ B cells from patients with cGVHD (n = 3) and healthy donors (n = 3). Data indicate the fold regulation of each gene that met statistical criteria, depicted in (A) CD27⁺ or (B) CD27⁻ B cells from patients with cGVHD compared with healthy donors. Dashed lines indicate a greater than threefold increase (above line) or decrease (below line) in mRNA expression. Gray bars represent genes that are altered in both CD27⁺ and CD27⁻ B-cell subsets. The P value for all genes is <.05; Student t test of replicate 2^(−ΔCt) values for each gene in the cGVHD group compared with healthy as calculated by SAbioscience Web-Based Array Analysis. Protein expression by mean fluorescence intensity (MFI) of (C) BLNK and (D) Syk in total peripheral B cells isolated from patients without cGVHD (−cGVHD, open circles) and with cGVHD (+cGVHD, filled squares). Data are median ± range from 2 independent experiments. *P <.05. (E) Protein expression by MFI of BLNK and Syk in B cells isolated from healthy donors (filled triangles), plated (1.5 × 10⁶ cell/mL) and stimulated with α-IgM (50 μg/mL) or phosphate-buffered saline (PBS) as a control for 36 hours. Fold increase of α-IgM divided by PBS is depicted. *P = .003 (1-sample Student t test, theoretical mean = 1). Data are median ± range pooled from 2 independent experiments. *P < .05.