The Application of Plant Viral Nanoparticles in Tissue-Specific Imaging - Nanotechnology for Biomedical Imaging and Diagnostics - page 398

Confocal microscopy revealed that the level of CpMV uptake by heLa, hT-29,
and MDA-MB-231 cancer cells correlated with the level of surface vimentin
expression in vitro [54]. heLa and hT-29 cells that are positive for surface vimentin
staining showed detectable levels of CpMV internalization, while MDA-MB-231
cells with very low levels of surface vimentin displayed no detectable CpMV uptake
(Fig. 14.3a). Similar CpMV targeting was observed in surface vimentin-positive
prostate cancer pC-3 and LNCap cell lines [73]. Surface vimentin is upregulated in
many cancers and inflamed cells and has also been shown to mediate internalization
of CpMV by endothelial cells. immunohistochemical staining of rat aortic endothe-
lium ex vivo revealed strong colocalization between surface vimentin expression
and CpMV binding (Fig. 14.3b) [71]. Thus, CpMV is a particularly useful tool for
imaging tumor neovasculature and vascular disease. The binding of CpMV to surface
vimentin facilitates targeting of CpMV to areas of disease, which can be applied for
tissue-specific drug delivery [64] and imaging [53].
Detection of atherosclerosis is one such application for CpMV. Compared to
normal, healthy vasculature, there is an elevation in the expression of surface vimen-
tin in atherosclerotic lesions. Using a LDL-receptor knockout mouse model of ath-
erosclerosis, it was found that CpMV uptake increased at sites of inflammation
within the lesion. Surface vimentin was detected via immunostaining of aorta tissue
sections followed by confocal imaging. An increase of uptake was detected in lesion
areas within 12 weeks of placing the mice on a high-fat diet. Correspondingly, fluo-
rescence levels from dye-labeled CpMV injected intraorbitally increased by more
than two-fold in these areas of lesion compared to nonlesion tissue. The confocal
images were analyzed, and the areas within the endothelium that are positive for
CpMV were determined. A 27-fold increase in the percent area where CpMV was
detected was observed after 12 weeks, and a 32-fold increase was observed at 20
weeks. This data suggests that the interaction of CpMV with surface vimentin can
allow it to be used as an imaging probe for early detection of atherosclerotic lesions
where surface vimentin is upregulated [76].
The natural interaction of CpMV with vimentin can also be utilized for intra-
vital imaging of vascular development in tumors. Fluorescently labeled CpMV
nanoparticles have been used to image the vasculature of a chicken chorioallantoic
membrane (CAM) tumor model and mouse embryos. in the CAM model, hT1080
fibrosarcoma tumor xenografts were implanted in the CAM of shell-less chicken
embryos. Dyes were conjugated onto the surface of CpMV particles via their lysine
residues, and the CpMV was then injected intravenously and imaged live under a
fluorescence microscope. The particles were rapidly taken up by endothelial cells
in vivo in both models (Fig. 14.3c-h). CpMV was observed to line the endothelial
wall, leading to high-resolution intravital imaging of the vasculature for at least
72 h, enabling high-resolution vascular mapping studies to be conducted (Fig. 14.3d).
Tumor-mediated angiogenesis can be examined by injecting CpMV labeled with a
different dye after a period of 24 h, allowing one to image and distinguish the newly
formed blood vessels. Visualization could be performed up to a depth of 500 µm for
both microvasculature and larger vessels. Attempts to obtain the same resolution
with QD nanoparticles were unsuccessful, as they tended to aggregate and obstruct