Fig 1.
Ldlr-/- mice treated with MPO inhibitor PF-06282999 had reduced plasma MPO activity but not atherosclerotic lesion area.
Ldlr-/- mice were fed Western diet and treated with 15 mg/kg MPO inhibitor PF-06282999 PO, BID for 14 weeks. A-C. MPO levels and activity were assessed in plasma one hour after the final compound dose. A. MPO activity, B. MPO levels. C. MPO activity normalized to MPO levels. D-E. Aortas were extracted, pinned en face, and stained with Oil Red O. D. Representative images of aortas after vehicle or PF-06282999 treatment. E. Quantitation of Oil Red O staining as a percentage of aorta area. (N = 8–11, **; p<0.01, ****; p<0.0001).
Fig 2.
Reduced necrotic core area and enhanced percentage of collagen in aortic roots of Ldlr-/- mice treated with MPO inhibitor PF-06282999.
Ldlr-/- mice were fed Western diet and treated with 15 mg/kg MPO inhibitor PF-06282999 PO, BID for 14 weeks. Hearts were isolated, and aortic roots were sectioned and stained with H&E (A-C), Masson’s Trichrome (D-E), or Oil Red-O (F-G). A. Representative images of H&E stained aortic roots from vehicle or PF-06282999-treated animals. B. Quantitation of lesion area. C. Left panel, quantitation of necrotic core area. Right panel, quantitation of necrotic core area as a percentage of lesion area. D. Representative images of Masson’s trichrome stained aortic roots from vehicle or PF-06282999-treated animals. E. Left panel, quantitation of collagen area. Right panel, quantitation of collagen area as a percentage of lesion area. F. Representative images of Oil Red O stained aortic roots from vehicle or PF-06282999-treated animals. G. Left panel, quantitation of Oil Red O area. Right panel, quantitation of Oil Red O area as a percentage of lesion area. (N = 9–15, *; p<0.05, **; p<0.01). Aortic root images were taken at 5x magnification.
Fig 3.
Unaltered macrophage content in and immune cell trafficking to plaques after MPO inhibition.
Ldlr-/- mice were fed Western diet and treated with 5 or 15 mg/kg MPO inhibitor PF-06282999 PO, BID for 7 or 14 weeks. A-B. Hearts were isolated and aortic roots were sectioned and stained with CD68 antibodies. A. Representative images from vehicle or PF-6282999-treated animals (images were taken at 5x magnification). B. Left panel, quantitation of CD68-positive area. Right panel, quantitation of CD68-positive stained area as a percentage of lesion area. C-G. Fluorescently labeled peritoneal exudate cells were injected intravenously into mice that had been treated with vehicle or PF-06282999 as described above. C. Protocol for leukocyte homing experiment. D-E. Aortas were extracted, and red fluorescent cells were counted in Z stacks using high content imaging. D. Left panel, representative aorta image demonstrating red fluorescent cells throughout the aorta, concentrating in the aortic arch. Right panel, close up of image in the white box. E. Quantitation of red cells per aorta normalized to aorta area as measured after pinning en face. Left panel, after 7 weeks of Western diet. Right panel, after 14 weeks of Western diet. F-G. Aortic roots were sectioned and stained for RFP and DAPI (images were taken at 5x magnification). F. Left panel, representative image of aortic root containing red fluorescent cells in the atherosclerotic lesion. Right panel, close up of image in white box. White arrowhead denotes RFP+ cell in aortic plaque. G. Quantitation of RFP positive stained area as a percentage of lesion area (N = 6–8, **; p<0.01). Aortic root images were taken at 5x magnification.
Fig 4.
Reduced MPO activity and 18F-FDG in aortas after 4 weeks of MPO inhibitor treatment.
Ldlr-/- mice were fed Western diet for 12 weeks, at which time, treatment with 5 or 15 mg/kg MPO inhibitor PF-06282999 or vehicle PO, BID was performed for an additional 4 weeks. A. Study design. B. Aortas were extracted at the end of the study, and aortic MPO activity was measured one hour post dose. C. Animals were injected with 200–300μCi, <100μL 18F-FDG for 60 minutes, and aortas were extracted and counted (N = 5–18, **; p<0.01, ****; p<0.0001).