Abstract 17275: Induction of Arteriogenesis in Mouse Hindlimb by Human Peripheral Blood B Cells as Determined by Micro-CT Imaging

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For patients with critical limb ischemia, restoration of blood flow through mechanical or pharmacological revascularization is an important therapeutic goal. Clinical studies have been unsuccessful in enhancement of neovascularization by treatment with angiogenic growth factors. We have shown that bone marrow derived B cells delivered into rat ischemic myocardium preserves cardiac function (Goodchild et al, 2009) and we sought to determine if B cells could be therapeutically beneficial in a mouse model of critical limb ischemia. Since commonly used histological determination of angiogenesis only provides semi-quantitative measure of vessels in limited tissue area without confirming perfusion functionality, this study measured the angiogenic and arteriogenic activities of B cell therapy using high resolution computed tomography. Mice (n=16, Balb/c) had iliac and femoral arteries and veins ligated. Mouse bone marrow B cells (CD19+) isolated by antibody-magnetic bead selection or saline control was injected directly into ischemic muscles. A post-ligation laser Doppler scan was performed to ensure ligation consistency then scanned weekly. On Day 28, mice were euthanized, vasculature flushed, fixed and perfused with contrast agent. Limbs were imaged by micro-CT with reconstruction at 25um. Data represented as vascular segment number and area counted in 500 slices in 3D micro-CT images. At Days 7, 14, 21 and 28, blood flow increased in B cell treated ischemic hindlimbs compared to controls. At Day 28, B cell treatment increased number of vessels in treated-ligated legs compared to unligated legs (8033±439 vs. 5297±710 vessels, respectively; P=0.010) and vessel area (175.9±45.4 vs. 99.0±28.6 mm2, respectively; P=0.012). Vessels ranging from 25-175 and 225-350um were increased in B cell treated-ligated legs compared to unligated legs (P<0.05). Vessel number ratio in treated-ligated legs to unligated legs was higher in B cell treated compared to control treated mice (1.66 vs. 1.05; P=0.025). These data show that unlike angiogenic response to treatment with growth factors, B cells enhanced development of vessels with a wide range of diameters suggesting that B cells may offer therapeutic potential for treatment of ischemic peripheral artery disease.