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Objective: Current management of aortic aneurysms (AAs) relies primarily on size criteria to determine whether invasive repair is indicated to preempt rupture. We hypothesized that emerging molecular imaging tools could be used to more sensitively gauge local inflammation. Because macrophages are key effector cells that destabilize the extracellular matrix in the arterial wall, it seemed likely that they would represent suitable imaging targets. We here aimed to develop and validate macrophage-targeted nanoparticles labeled with fluorine-18 ((18)F) for positron emission tomography-computed tomography (PET-CT) detection of inflammation in AAs., Methods and Results: Aneurysms were induced in apolipoprotein E-/- mice via systemic administration of angiotensin II. Mice were imaged using PET-CT and a monocyte/macrophage-targeted nanoparticle. AAs were detected by contrast-enhanced micro-CT and had a mean diameter of 1.85 ± 0.08 mm, whereas normal aortas measured 1.07 ± 0.03 (P < 0.05). The in vivo PET signal was significantly higher in aneurysms (standard uptake value, 2.46 ± 0.48) compared with wild-type aorta (0.82 ± 0.05, P < 0.05). Validation with scintillation counting, autoradiography, fluorescence, and immunoreactive histology and flow cytometry demonstrated that nanoparticles localized predominantly to monocytes and macrophages within the aneurysmatic wall., Conclusions: PET-CT imaging with (18)F-labeled nanoparticles allows quantitation of macrophage content in a mouse model of AA.

Unlabelled: The contribution of (18)F-FDG uptake by endothelial cells to uptake values measured by PET in various tissues is as yet unclear. We therefore sought to characterize (18)F-FDG uptake in an in vitro model of human endothelial cells., Methods: Commercially obtained human umbilical vein endothelial cells (HUVECs) were seeded in 6-multiwell plates 48-96 h before incubation with 1-2 MBq (18)F-FDG per well. Radioactivity measurements were performed after washing and mechanical dissolvation of the cellular monolayers. Cellular (18)F-FDG uptake was referred to protein concentration. This experimental protocol was subsequently varied to study the effect of different parameters of interest. Furthermore, radio-thin-layer chromatography was used to identify intracellular (18)F-FDG metabolites. (18)F-FDG uptake in HUVECs was compared with that by a human monocyte-macrophage (HMM) preparation and by glioblastoma cells (GLIOs) under identical experimental conditions., Results: (18)F-FDG accumulated in HUVECs in a time-dependent manner and was trapped mainly as (18)F-FDG-6-phosphate and (18)F-FDG-1,6-diphosphate. Unlabeled glucose and cytochalasin B competitively inhibited (18)F-FDG uptake, whereas phlorizin had no significant effect. Glucose deprivation significantly enhanced (18)F-FDG uptake by a factor of 2.7, whereas sodium depletion had no significant influence. HUVECs treated with vascular endothelial growth factor (VEGF) showed a significant 82% increase in (18)F-FDG accumulation after a 2-h exposure to 50 ng/mL VEGF. (18)F-FDG uptake in HUVECs was significantly higher than that in HMMs and in the range of the uptake values measured in GLIOs., Conclusion: (18)F-FDG accumulates in HUVECs by mechanisms analogous to those in neoplastic cells or neurons. VEGF significantly stimulates endothelial (18)F-FDG uptake. The observed differences in (18)F-FDG uptake between HUVECs, HMMs, and GLIOs are difficult to extrapolate to in vivo conditions but stimulate further studies on the contribution of endothelial (18)F-FDG uptake to the overall uptake of that tracer in neoplastic or vascular lesions.

Unlabelled: Activated monocytes with a high (18)F-FDG accumulation can affect the results of clinical PET studies. To better understand the mechanisms regulating monocytic (18)F-FDG uptake, we investigated the effect of priming and respiratory-burst generation and further evaluated the role of potential protein kinase pathways., Methods: Purified human monocytes were primed with interferon-gamma (IFN-gamma), and respiratory burst was generated by stimulation of primed cells with phorbol-12-myristate-13-acetate (PMA). Oxygen-intermediate generation was assessed by luminescence measurements after the addition of lucigenin. (18)F-FDG uptake after 30 min of incubation was measured for unprimed control cells, primed cells, and PMA-stimulated cells. The role of protein kinases was investigated using respective inhibitors., Results: PMA stimulation of primed monocytes dramatically increased oxygen-intermediate generation, leading to a 42.2 +/- 1.1 fold higher level of cumulative luminescence compared with unprimed control cells, whereas IFN-gamma priming alone resulted in low luminescence levels (13.9% +/- 4.6% of PMA-stimulated cells). In contrast, priming alone was sufficient to augment monocytic (18)F-FDG uptake to 273.3% +/- 16.7% of control levels (P < 0.001), and it was not further increased by PMA stimulation. The tyrosine kinase inhibitor, genistein, and the specific protein kinase C inhibitor, staurosporine, completely abolished the priming-induced enhancement of (18)F-FDG uptake and lowered uptake to control levels. Under the same conditions, wortmannin, a phosphatidylinositol 3 kinase (PI3 kinase)-specific inhibitor, and cycloheximide, a protein synthesis inhibitor, were associated with only minor reductions in the enhanced-uptake effect of priming., Conclusion: IFN-gamma priming alone, without stimulation of respiratory-burst activity, is sufficient to induce maximal augmentation of (18)F-FDG uptake in monocytes. Furthermore, this metabolic effect appears to involve tyrosine kinases and the protein kinase C pathway but is independent of the PI3 kinase pathway.

Therapeutic angiogenesis is the controlled induction or stimulation of new blood vessel formation to reduce unfavourable tissue effects caused by local hypoxia and to enhance tissue repair. The effects of ultrasound on wound healing, chronic ulcers, fracture healing and osteoradionecrosis may be explained by the enhancement of angiogenesis. The aim of this study was to identify which cytokines and angiogenesis factors are induced by ultrasound in vitro. Two ultrasound machines were evaluated, a "traditional" (1 MHz, pulsed 1:4, tested at four intensities), and a "long wave" machine (45 kHz, continuous, also tested at four intensities). The ultrasound was applied to human mandibular osteoblasts, gingival fibroblasts and peripheral blood mononuclear cells (monocytes). The following cytokines and angiogenesis factors were assayed by ELISA techniques: interleukin-1beta(IL-1beta), IL-6, tumour necrosis factor alpha (TNF-alpha), IL-8, fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF).A slight stimulation of IL-1beta was noted in all cell types. There was no difference in the IL-6 and TNF-alpha levels. The angiogenesis-related cytokines, IL-8 and bFGF, were significantly stimulated in osteoblasts, and VEGF was significantly stimulated in all cell types. Both ultrasound machines produced similar results, and the optimum intensities were 0.1 and 0. 4 W/cm2 (SATA) with 1 MHz ultrasound, and 15 and 30 mW/cm2 (SATA) with 45 kHz ultrasound.The results show that therapeutic ultrasound stimulates the production of angiogenic factors such as IL-8, bFGF and VEGF. This may be one of the mechanisms through which therapeutic ultrasound induces angiogenesis and healing., (Copyright 1999 Academic Press.)