Extended Data Fig. 9: Protection from myocardial injury by targeting Fgr.
(A) Micrographs of NETs (positive for citH3 and MPO; red) and vessels (blue) in cremasteric venules of wild-type subjected or not to I/R, and Fgr−/− mice subjected to I/R. Right, quantification of NETs per tissue volume; Data shown as mean ± SEM and n are number of mice per group. (B) Competitive recruitment of wild-type and Fgr−/− neutrophils to the peritoneal cavity after zymosan injection, or to the bronchoalveolar space of lungs after LPS instillation in mixed chimeric mice; n are numbers of mice analyzed. Selplg−/− neutrophils are shown for comparison of impaired migration. Values are normalized to reference wild-type competitors across the different groups and given as migration efficiencies. Data shown as mean ± SEM and n are number of mice analyzed. (C) Micrographs of Weibel-Palade bodies (WPB) and vacuoles in myocardial vessels after sham or ischemic challenge, which are quantified in (D). These measures of vascular damage are dependent on neutrophils, as shown after experimental depletion with 1A8 antibody (E). Data shown as mean ± SEM, and n is the number of micrographs analyzed, from 2 mice. (F) Effect of the Fgr antagonist TL02-59 in myocardial death upon ischemia-reperfusion, when given after ischemia at the time of reperfusion. Micrographs of heart sections at left illustrate the protective effect on myocardial death (outlined whitish regions). Data are normalized to the area at risk (AAR) and shown as mean from 4 mice per group. (G) Combination of neutrophil depletion with 1A8 antibody, and Fgr deficiency in transplanted mice. The infarcted areas are normalized with the areas at risk; n is number of mice per group. (H) Combination of neutrophil depletion with the Fgr agonist TL02-59. Data shown as mean ± SEM; n are mice per group. (I) Combination of the Fgr inhibitor in hematopoietic Fgr−/− mice, with no effect in further protecting from myocardial death; n are mice per group. (J) Myocardial fibrosis (left ventricle) determined by hematoxylin and eosin staining in control wild-type and Fgr−/− mice subjected to permanent ischemia and analyzed after 28 days. The fibrosis area is represented at right; n are mice per group. All data from (A, B) was analyzed by one-way ANOVA with Tukey’s multiple comparison test; (G–I) was analyzed by two-way ANOVA with Tukey’s multiple comparisons test. All other panels are compared by two-tailed unpaired-t test (C-F and J). The number of replicates (n) per group is indicated in each panel.
