Figure 2.
Eosinophil recruitment and platelet-induced activation in thrombosis. (A) Representative images of thrombus formation (marked by dotted line) in the carotid artery (marked by solid white lines) of an ApoE−/−;ΔdblGATA1−/− and an ApoE−/−;ΔdblGATA1+/+ mouse. In ApoE−/−;ΔdblGATA1−/− mice (n = 17), thrombus stability was significantly reduced compared with that of ApoE−/−;ΔdblGATA1+/+ littermate controls (n = 17), as shown by occlusion time. (B) Eosinophils (red, arrows) in EoCre;PC-G5-tdTfl/fl mice adhere immediately after initial endothelial injury at the lesion site of the carotid artery after FeCl3 application. Platelets labeled with fluorescent 2′,7′-dichlorofluorescein (green); thrombus shown by dotted line. (C) Quantification of eosinophil recruitment in vivo after FeCl3 application on the carotid artery (n = 3). (D) Representative images of thrombus formation 20 minutes after endothelial injury by FeCl3 in the carotid artery of EoCre;Kindlin-3fl/fl and EoCre;Kindlin-3wt/wt mice (thrombus area marked by dotted white line; carotid artery marked by solid white lines). Scale bar, 100 µm. (E) EoCre;Kindlin-3fl/fl mice (n = 5) had drastically reduced thrombus formation compared with that of wild-type littermates (n = 6). (F) During thrombus formation in the carotid artery, platelets (green) interacted with an eosinophil (red), which already adhered to the endothelium. Interacting cells are marked with arrows. (G) Eosinophils (red) in EoCre;PC-G5-tdTflox mice collect platelets (white) around themselves and form elongated processes (arrow in higher magnification, top right inset). Thrombosis induced by FeCl3 in the mesenteric microcirculation; images acquired by confocal microscopy. Scale bar, 20 µm. Images representative of 3 experiments. (H) Calcium signal (pseudocolored; blue/green, low signal; yellow/red, high signal) in an eosinophil interacting with several platelet aggregates (white) in the mesenteric microcirculation. Cropped images from intravital confocal microscopy in EoCre;PC-G5-tdTflox mice. Image representative of 3 experiments. (I) Calcium signal (pseudocolored) in an eosinophil (red) adhering to the endothelium before and during platelet contact (white). Image representative of 4 experiments. Cropped images from intravital epifluorescence microscopy in the carotid artery of EoCre;PC-G5-tdTflox mice after FeCl3 application. Scale bar, 20 µm. Representative of 4 experiments. For panels A-C and E, data are mean ± SD. *P < .05; **P < .01.

Eosinophil recruitment and platelet-induced activation in thrombosis. (A) Representative images of thrombus formation (marked by dotted line) in the carotid artery (marked by solid white lines) of an ApoE−/−;ΔdblGATA1−/− and an ApoE−/−;ΔdblGATA1+/+ mouse. In ApoE−/−;ΔdblGATA1−/− mice (n = 17), thrombus stability was significantly reduced compared with that of ApoE−/−;ΔdblGATA1+/+ littermate controls (n = 17), as shown by occlusion time. (B) Eosinophils (red, arrows) in EoCre;PC-G5-tdTfl/fl mice adhere immediately after initial endothelial injury at the lesion site of the carotid artery after FeCl3 application. Platelets labeled with fluorescent 2′,7′-dichlorofluorescein (green); thrombus shown by dotted line. (C) Quantification of eosinophil recruitment in vivo after FeCl3 application on the carotid artery (n = 3). (D) Representative images of thrombus formation 20 minutes after endothelial injury by FeCl3 in the carotid artery of EoCre;Kindlin-3fl/fl and EoCre;Kindlin-3wt/wt mice (thrombus area marked by dotted white line; carotid artery marked by solid white lines). Scale bar, 100 µm. (E) EoCre;Kindlin-3fl/fl mice (n = 5) had drastically reduced thrombus formation compared with that of wild-type littermates (n = 6). (F) During thrombus formation in the carotid artery, platelets (green) interacted with an eosinophil (red), which already adhered to the endothelium. Interacting cells are marked with arrows. (G) Eosinophils (red) in EoCre;PC-G5-tdTflox mice collect platelets (white) around themselves and form elongated processes (arrow in higher magnification, top right inset). Thrombosis induced by FeCl3 in the mesenteric microcirculation; images acquired by confocal microscopy. Scale bar, 20 µm. Images representative of 3 experiments. (H) Calcium signal (pseudocolored; blue/green, low signal; yellow/red, high signal) in an eosinophil interacting with several platelet aggregates (white) in the mesenteric microcirculation. Cropped images from intravital confocal microscopy in EoCre;PC-G5-tdTflox mice. Image representative of 3 experiments. (I) Calcium signal (pseudocolored) in an eosinophil (red) adhering to the endothelium before and during platelet contact (white). Image representative of 4 experiments. Cropped images from intravital epifluorescence microscopy in the carotid artery of EoCre;PC-G5-tdTflox mice after FeCl3 application. Scale bar, 20 µm. Representative of 4 experiments. For panels A-C and E, data are mean ± SD. *P < .05; **P < .01.

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