Anti-inflammatory drug evaluation in ApoE-/- mice by ultrasmall superparamagnetic iron oxide-enhanced magnetic resonance imaging.

OBJECTIVES: The renin-angiotensin system and local phagocytic activity play a major role in atherosclerotic plaque development. Treatment with irbesartan, an antagonist of angiotensin II receptor, can decrease atherosclerotic lesion formation. Iron oxide-enhanced magnetic resonance imaging (MRI) can be successfully used to evaluate the phagocytic activity in the atherosclerotic plaque in mice. In this study, we used 2 iron oxide-enhanced MRI strategies, in vivo labeling by injection of iron oxide particles and injection of in vitro labeled macrophages, to investigate the effect of irbesartan on both atherosclerotic plaque size and macrophage content in apolipoprotein (Apo) E-deficient mice. MATERIALS AND METHODS: ApoE-/- female mice (C57BL/6 background; Charles-River, France) were divided into 2 groups (irbesartan treated [TG] or not treated [NTG]) and started on a high-fat diet (Harlan TD88137 Western Diet, 21% fat, 0.2% cholesterol). Animals underwent magnetic resonance examinations on a 7-T scanner at baseline and at 14 and 28 weeks of treatment. At each time point, 2 MRI sessions were performed, before and 48 hours after administration of an iron oxide agent (P904; Guerbet, France) or magnetically labeled macrophages (MФΦ). At the end of the follow-up, blood samples were taken for plasma lipid dosing and aorta samples for histology. The study was approved by the animal experimentation ethic committee of our institution.Vessel wall area measurements were performed on high-resolution spin echo transverse images. Multiecho gradient echo images acquired with the same geometry were used to calculate T2* maps of the vessel wall using a pixel-by-pixel monoexponential fit. Irbesartan effect on vessel wall area over time was assessed using a factorial analysis of variance test. T2* values of the vessel wall at pre- and post-ultrasmall superparamagnetic iron oxide (USPIO) administration were analyzed with a 1-way analysis of variance test with Bonferroni post hoc. RESULTS: Irbesartan treatment resulted in significantly smaller vessel wall areas at 28 weeks of treatment (P = 0.04). Postinjection values varied significantly over time for both the NTG-P904 (P = 0.02) and the TG-P904 (P = 0.01) groups. Furthermore, when comparing the TG-P904 with the NTG-P904 group at 28 weeks of treatment, a significant difference was obtained for both pre- and post-USPIO administration values (P = 0.01). In the labeled-macrophage group, postinjection T2* values were smaller than the preinjection ones for the NTG animals at 14 weeks of treatment. No T2* changes were observed in the TG-MΦ group.The difference between pre- and post-USPIO administration T2* values (ΔT2*) was significantly smaller in the TG-P904 group compared with the NTG-P904 group at 28 weeks of treatment. At this point, a good correlation (R = 0.7, P = 0.03) was found between the ΔT2* values in the P904 imaging group and the macrophage-covered area by immunohistological analysis. CONCLUSIONS: The present study illustrates an MRI follow-up of intraplaque macrophages using in vivo labeling by iron oxide particle injection and macrophage injection after in vitro USPIO labeling in the assessment of a therapeutic effect in a mouse model of atherosclerosis. Even though in vivo labeling is not fully specific of macrophage uptake, it enabled the detection of a treatment-related reduction in the macrophage content of atherosclerotic plaques in ApoE-/- mice.

Assessment of age modulated vascular inflammation in ApoE-/- mice by USPIO-enhanced magnetic resonance imaging.

OBJECTIVE: Inflammation within atherosclerotic lesions increases the risk for plaque rupture and thrombosis. A functional approach to plaque analysis is the intravenous administration of ultrasmall superparamagnetic particles of iron oxide (USPIO) that enables visualization of macrophages residing in the plaques. In this study, we sought to characterize the age-related inflammatory status associated with atherosclerosis lesion progression in ApoE mice using USPIO-enhanced magnetic resonance imaging (MRI). MATERIALS AND METHODS: A total of 24 ApoE mice were divided in 4 groups (N = 6) and were given a high cholesterol diet from 6 weeks of age to the end of the protocol. One group per MR time point was investigated at 10, 16, 24, and 34 weeks of age. Each MR examination was performed on a 4.7 T scanner and consisted of baseline and 48 hours post-USPIO administration imaging sessions. P904, a USPIO contrast agent (Guerbet, Paris, France) with a potential for plaque macrophage targeting, was used.Vessel wall area measurements were performed on high resolution spin echo transverse images. Multi-echo gradient-echo images acquired with the same geometry were used to calculate T2* maps of the vessel wall using a pixel-by-pixel monoexponential fit. A one-way analysis of variance was performed to characterize the temporal variation of vessel wall area, susceptibility artifact area, baseline, and post-USPIO T2* values. MR measurements were correlated with the histologic findings. RESULTS: A significant increase was found in the aortic wall area from 1.4 ± 0.2 at 10 weeks to 2.0 ± 0.3 mm at 34 weeks of age (P < 0.05). Concerning the post-USPIO MRI, signal loss regions, with patterns spanning from focal to the complete disappearance of the vessel wall, were observed on all postcontrast images. A significant increase in the size of the susceptibility artifact was observed from 0.5 ± 0.2 to 2.4 ± 1.0 at 24 weeks (P < 0.05) and to 2.0 ± 0.9 mm at 34 weeks (P < 0.05).The T2* values calculated on the 48 hours post-USPIO images were shorter compared with baseline. The decrease was 34% ± 16% at 10 weeks, 57% ± 11% at 16 weeks, 57% ± 16% at 24 weeks, and 48% ± 13% at 34 weeks.The Pearson's correlation test between measurement of aortic wall area performed on both MR images and histologic analysis showed a statistically significant correlation (r = 0.695 and P < 0.05). A correlation was also obtained between the signal loss area and the macrophages covered area (r = 0.68 and P < 0.05). CONCLUSIONS: This study demonstrated the feasibility of USPIO-enhanced MRI in assessing the inflammatory status related to the temporal progression of the atherosclerosis plaque in ApoE transgenic mice model of atherosclerosis. In our experimental conditions, the vascular inflammation peak, for the ApoE mice feeding high-fat/high-cholesterol diet is measured between 16 and 24 weeks of age.