Phospho-ERK1/2 levels in CD4⁺-naive and central memory T cells at day 30 were positively correlated with acute GVHD. PBMCs from allo-HSCT recipients were stimulated with or without PMA and ionomycin, and phosphorylation of ERK1/2 was analyzed by flow cytometry (gating on CD4⁺, CD8⁺, and CD19⁺ cells). Closed circles, patients with GVHD; open circles, patients without GVHD. (A) Representative histograms showing CD4⁺ T cells from patients with and without GVHD are presented. Aggregated percentages of cells from respective cell subsets expressing phospho-ERK1/2 are shown. Samples stimulated with PMA and ionomycin are shown in gray, and unstimulated samples are shown in red. Percentages of CD4⁺ T cells (B), CD8⁺ T cells (C), CD19⁺ B cells (D), CD4⁺-naive T cells (E), CD4⁺ central memory T cells (F), and CD4⁺ effector memory T cells (G) (n = 10 per group; data are expressed as means ± SEMs). (H) Clinical course of GVHD, immunosuppressants used and phospho-ERK1/2 levels in CD4⁺ T cells at days 30 and 60. *P < .05; **P < .01 (unpaired 2-tailed Student t test). n.s., not significant.