Figure 4.

P2rx4 is necessary for lung fibrosis. (A) qPCR for collagen genes and P2rx4 in primary mouse lung fibroblasts isolated from mice with fibroblast-specific P2rx4 deletion. N=3 mice. P values are for unpaired one-tailed t-tests. (B) qPCR for collagen genes in human (upper) and mouse (lower) primary lung fibroblasts treated with ATPγS. N=3 separate individuals and n=3 separate mice. P values are for unpaired two-tailed t-tests. (C) Left: Sirius red staining of lungs at 21 days post bleomycin injury in Col1a2-CreERT2: P2rx4 fl/fl or Col1a2-CreERT2 controls. Representative images are shown. Quantification is for n=3 mice. Scale bar=50μm. P value is for unpaired two-tailed t-test. Right: Hydroxyproline assays of whole lung collagen from mice with fibroblast-specific P2rx4 deletion with two different Cre drivers as indicated, and controls. N=3-10 as shown. P values are for one-way ANOVA followed by Sidak’s multiple comparison test. qPCR is for P2rx4 in fibroblasts isolated from Pdgfrb-Cre: P2rx4 fl/fl or Pdgfrb-Cre controls. N=6 mice in each group and P value is for Mann-Whitney test. (D) Western Blot of phospho-p38 and total p38 for fibroblasts isolated from Pdgfrb-Cre: P2rx4 fl/fl or Pdgfrb-Cre controls. Quantification shows the ratio of P-p38 to total p38 by densitometry. P value is shown for unpaired one-tailed t-test and n=3 mice. (E) Model of fibroblast activation by paracrine ATP derived from monocyte-derived macrophages in lung fibrosis.