Figure 1.
(A) Cartoon depicting the model used for inducing atherothrombosis. A tiny curved needle introduced through a branch of the external carotid was advanced up to the plaque surface under persistent blood flow. After positioning the tip of the needle in front of the plaque, it was applied to the plaque and moved forth and back twice in order to scratch the fibrous cap. (B) The experimental timeline. “I” stands for injection. (C) Transversal histological section of the carotid artery after the scratch injury. Immunohistochemistry depicts platelets and therefore thrombus in red. The scratch with superimposed thrombosis is clearly visible and induced here a small dissection (arrow). A corresponding section of the site of the plaque rupture stained with Masson-Trichrome is depicted in (D).
Figure 2.
Axial MRI images of the external carotid arteries from two representative animals.
Depicted are four consecutive slices (H1-H4) at three time points: t(−1) (before injection), t2 (16 minutes after injection) and t3 (24 minutes after injection). Red circles indicate the left external carotid artery which underwent surgery and scratching of the plaque, and green circles mark the contra lateral vessel. The animals orientation is indicated by labeling its posterior (P) and left (L) direction. Signal attenuation can be observed in the animal A which received LIBS-MPIO contrast agent whereas the blood signal remains hyperintense in animal B receiving the control-MPIO.
Figure 3.
Quantitative analysis of MRI data.
Subgroup analysis of animals with thrombus size >2% is shown in Fig. 3A for animals with LIBS-MPIO and control-MPIO injection. At each time point after the injection of LIBS-MPIO, the MR signal detected at the level of the scratched plaque was significantly lower than the signal detected at the contralateral and non-injured plaque level (A, red line). Conversely, the ratio of signals slightly increased after the injection of control-MPIO (A, blue line). In animals with thrombus size <2%, the signal ratio had not been altered by the injection of LIBS-MPIO in comparison to the injection of control-MPIO (3B).
Figure 4.
Histological identification of MPIOs.
(A): Representative anti-CD41 immunostaining of a carotid artery section from a mouse receiving LIBS-MPIOs. (B) Enlargement of the area from (A) shows MPIOs inside the thrombus (circles). MPIOs can be recognized as regular round structures with a blue halo and a brownish inner colour.
Figure 5.
Quantification of MPIOs on histological sections.
MPIOs were about four-fold more abundant in animals with LIBS-MPIO-injection as compared to the animals receiving control-MPIO. A similar difference was observed between injured and non injured vessels in animals receiving LIBS-MPIO. Correlation of thrombus size with the number of bound MPIOs in animals with LIBS-MPIO injection is not significant; however the number of MPIOs is in average within a similar range of 10–20 MPIOs per section as expected from the observed signal attenuation.