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1 Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA
2 Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA; Veterans Affairs Medical Center, Atlanta, GA, USA
3 Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA
4 Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA, USA
* To whom correspondence should be addressed. E-mail: robert.guldberg{at}me.gatech.edu.
Transgenic mouse models are increasingly being used to investigate the functions of specific growth factors or matrix proteins to design therapeutic strategies for controlling blood vessel growth. However, the available methodologies for evaluating angiogenesis and arteriogenesis in these models are limited by animal size, user subjectivity, the power to visualize the threedimensional vessel networks, or the capability to employ a vigorous quantitative analysis. In this study, we employed contrast enhanced microcomputed tomography imaging to assess collateral development following induction of hind limb ischemia in the mouse. The morphological parameters vessel volume, connectivity, number, thickness, thickness distribution, separation, and degree of anisotropy were evaluated in control and surgery limbs 0, 3, and 14 days post surgery. Results indicate that the vascular volume of the surgically manipulated limb was reconstituted as early as 3 days following femoral artery excision through development of a series of highly connected, small caliber, closely spaced, and isotropically oriented collateral vessels. Parametric analyses were completed to assess the sensitivity of the calculated morphological parameters to variations in image binarization threshold and voxel size. Images taken at 36 micron voxel size were found to be optimal for evaluating collateral vessel formation, while 8-16 micron voxel sizes were needed to resolve smaller vascular structures. This study demonstrates the utility of microcomputed tomography as a robust method for quantitative, three-dimensional analysis of blood vessel networks. While these initial efforts focused on the mouse hind limb ischemia model, the developed techniques may be applied to a variety of model systems to investigate mechanisms of angiogenesis and arteriogenesis.
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