Multimodality imaging reveals a gradual increase in matrix metalloproteinase activity at aneurysmal lesions in live fibulin-4 mice

Background-We imaged the protease activity of matrix metalloproteinases (MMPs) upregulated during aneurysm formation, using protease-activatable near-infrared fluorescence probes. We tested whether these protease-activatable sensors can directly report the in vivo activity of the key biomarkers in aneurysm, using our genetically modified fibulin-4 mouse models for aneurysm formation. Mice homozygous for the fibulin-4 reduced-expression allele (fibulin-4R/R) show dilatation of the ascending aorta and a tortuous, stiffened aorta resulting from disorganized elastic fiber networks. Strikingly, even a moderate reduction in expression of fibulin-4 in the heterozygous fibulin-4+/Rmice occasionally results in modest aneurysm formation. Methods and Results-Aorta transcriptome and protein expression analysis of fibulin-4+/Rand fibulin-4R/Ranimals identified excessive transforming growth factor-β signaling as the critical event in the pathogenesis of aneurysm formation. To determine whether a perturbed elastin lamellar structure arose from induction of transforming growth factor-β-regulated MMPs, we performed gelatin zymography and used a protease-activatable near-infrared fluorescence probe to monitor and quantify MMP upregulation in animals, using various in vivo optical imaging modules and coregistration of the fluorescence signal with CT images of the same animals. Gelatin zymography demonstrated a significant increase in the presence of the active form of MMP-9 in the aortic arch of fibulin-4R/Rmice. In vivo analysis of MMP upregulation using the near-infrared fluorescence probe and subsequent isosurface concentration mapping from reconstructed tomographic images from fibulin-4+/Rand fibulin-4R/Rmice revealed a graded increase in activation of MMPs within the aneurysmal lesions. Conclusions-We aimed to develop molecular imaging procedures for faster, earlier, and easier recognition of aortic aneurysms. We show that in vivo coregistration of MMP activity by noninvasive tomographic imaging methods allows the detection of increased MMP activity, even before the aneurysm has actually formed.

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