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. 2014 Jan 27;124(2):543–552. doi: 10.1172/JCI71858

Figure 4. PF4 has a direct effect on limiting Th17 differentiation.

Figure 4

(A) Transgenic expression of PF4 normalizes plasma IL-17 in Pf4–/– mice (n = 5–7, ± SD; *P < 0.01 vs. Pf4–/–). (B) PF4 directly inhibits Th17 differentiation in vitro. CD4+ T cells were cultured in Th0, Th1, or Th17 conditions in the presence of control or PF4 (1 μg/ml). After 3 days representative Th1 (Tbx21) and Th17 (Rorc) gene expression was determined (representative of 3 separate experiments with the same expression pattern). (C) T cells were cultured in Th17 conditions in the presence of buffer or PF4. IL-17–positive cells (flow cytometry) and IL-17 in the supernatant (ELISA) were measured (n = 4; mean ± SD. *P < 0.03 vs control). (D) Platelet PF4 limits Th17 differentiation. Naive CD4+ T cells were incubated in Th0 or Th17 conditions with buffer, WT,or Pf4–/– platelets. IL-17–positive T cells were quantified 4 days later. WT, but not Pf4–/– platelets, inhibited Th17 differentiation (n = 4; mean ± SD, *P < 0.03). (E and F) PF4 blocks TGF-β–induced Smad 2/3 phosphorylation. Jurkat T cells (E) or naive mouse CD4+ T cells (F) were incubated with control buffer, TGF-β (2 ng/ml), or TGF-β and PF4 (1 μg/ml). p-Smad2/3 was determined by Western blot and quantified by densitometry (n = 3; mean ± SD, *P < 0.01 vs. TGF-β only).