PR attenuate PA SN-triggered lung cell death and limits PA SN-mediated lung injury in thrombocytopenic Mpl^−/−mice. (A) WT mice BAL platelet counts (10⁹/L) at baseline (0 hours, n = 5) and 20 hours after IT 10⁶ CFU PA (n = 8). (B) BAL thrombin–antithrombin complexes (nM) in WT and Mpl^−/− mice at 0 and 4 hours after IT PA SN (n = 4 each group). In separate experiments using MLE cells, fold change in MLE cytotoxicity 6 hours after PA SN in the presence or absence of pooled thrombin-stimulated PR from WT mice (C) (n = 4 trials, where 3 of the 4 trials included 10 µg group) or denatured PR (D). Cleaved caspase-3, total caspase-3, and β-actin expression in MLE cells treated with PA SN in the presence or absence of increasing PR (100 and 300 µg total protein) at 4 hours (E), fold change in MLE cytotoxicity 6 hours after PA SN in the presence or absence of apyrase (0.2 U) or PR from WT mice (F), or thrombin-stimulated platelets pooled from healthy volunteers (G) (n = 4). For panels C-D and F-G, each point represents the median cytotoxicity of 3 to 4 technical replicates from a single trial, relative to PA SN and vehicle treatment cytotoxicity, which is set at 1.0. In separate experiments with Mpl^−/− mice 20 hours after IT PA SN with either vehicle or 100 µg PR, lung tissue expression of cleaved PARP and β-actin is shown, n = 3 in each group (H). Fold change in BAL protein concentration. n = 14 (vehicle) and n = 17 (PR-treated group) from 3 separate experiments (I). Each point represents an individual mouse, and the group median is displayed. Statistical comparison by Mann-Whitney U test (*P < .05 and ***P < .001) (A-B,I) or Kruskal-Wallis test with Dunn’s test for multiple comparisons (lines represent post hoc analysis; *P < .05 (C-D,F).