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RGD targeting of human ferritin iron oxide nanoparticles can enhance in vivo carotid MRI of experimental atherosclerosis


Human ferritin (HFn) is a promising nanoscale protein cage platform for molecular/cellular imaging, and we have developed engineered HFn nanoparticles as MRI agents. Inflammation and angiogenesis contribute to atherosclerosis, and RGD is a well-studied ligand of the αvβ3 integrin expressed by activated macrophages and angiogenic endothelial cells.


To evaluate RGD-conjugated HFn-iron oxide nanoparticles for enhanced in vivo MRI detection of murine carotid atherosclerosis.


1) Mice

Fourteen FVB mice underwent left carotid ligation after 4 weeks of high-fat diet and diabetes induction by streptozotocin.

2) RGD-conjugated HFn-iron oxide nanoparticles

Using the recombinant human heavy-chain ferritin protein cage, HFn was genetically engineered to display 24 copies of an RGD-4C peptide (CDCRGDCFC) on the exterior surface of the protein cage. Magnetite (Fe3O4) was encapsulated in interior cavities of RGD-conjugated HFn (RGD+ HFn) and non-targeted HFn (RGD- HFn) at loading factors of 5000Fe per cage, giving R2 values of 93 mM-1s-1 (magnetite diameter: 5-7nm, exterior diameter: 12nm). The injected dose was adjusted to 25mgFe/kg.

3) MRI

Two weeks post ligation, mice were imaged on a whole-body 3T MRI scanner (Signa HDx, GE Healthcare) with a phased array mouse coil (RAPID MR International), using a gradient echo sequence (TR/TE=100ms/10ms, slice thickness=1.0mm, FOV=3cm, matrix=256x256, FA=60, NEX=10). Mice were then injected with either RGD+ (n=7) or RGD- (n=7) HFn nanoparticles, followed by MRI at 24 and 48 hours post injection. The nanoparticle accumulation was assessed by measuring the extent of T2*-induced reduction in carotid lumen size (% reduction of carotid lumen area).

4) Histology

Perl’s iron staining was performed to identify accumulation of the nanoparticles in the carotid lesions.


Both RGD+ and RGD- HFn nanoparticles caused a reduction in lumen size of the ligated left carotid arteries at 48 hrs due to T2* signal loss (p<0.001 vs. preinjection, Figures 1, 2), but the effect was significantly greater with RGD+ HFn (p=0.01 vs. RGD- HFn). There was no significant lumen reduction in the non-ligated (control) right carotid arteries. Perl’s iron staining confirmed greater accumulation of RGD+ HFn in the lesion compared to RGD- HFn, primarily in neointimal macrophages (Figure 3).

figure 1

Figure 1

figure 2

Figure 2

figure 3

Figure 3


Human ferritin protein cage is a versatile nanoparticle imaging platform for in vivo cellular/molecular MRI, with enhanced atherosclerosis imaging through multivalent RGD targeting.

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This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Kitagawa, T., Kosuge, H., Uchida, M. et al. RGD targeting of human ferritin iron oxide nanoparticles can enhance in vivo carotid MRI of experimental atherosclerosis. J Cardiovasc Magn Reson 13 (Suppl 1), P373 (2011).

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