Non-Invasive Macrophage Imaging for Detection of Vulnerable PlaqueApplications and LimitationsAmershamMorteza Naghavi, MD

*

*

AB

Which one is vulnerable, A or B?The one with inflammation (macrophage infiltration)

*

Morphology vs. Activity ImagingInactive and non-inflamed plaqueActive and inflamed plaqueSimilar in IVUSOCTMRI w/o CMMorphologyDifferent ActivityThermography, Spectroscopy, immunoscintigraphy, MRI with targeted contrast media

*

Plaque Morphology vs.

Plaque Activity We Need Both

*

Imaging Inflammation- by MRI- by CTBoth are available today!

Imaging Inflammation- by MRIPioneered in 1980sRummeny E, Weissleder R, Stark DD, Elizondo G, Ferrucci JT. Radiologe. 1988 Aug;28(8):380-6.

*

Superparamagnetic and Ultra-superparamagnetic Iron OxideBlood pool Magnetic resonance (MR) imaging contrast media with a central core of iron oxide generally coated by a polysaccharide layer

Shortening MR relaxation timeEngulfed by and accumulated in cells with phagocytic activity

Particle Core Size Particle Size Blood (nm) (nm) Half-life

Combidex 5-6 20-30 8hFeridex 4-6 35-50 2.40.2hDDM 43/34/102 6.4 20-30 6h ClariscanMION 4-6 17 variesFeruglose --- --- --- --- Examples of commercially available SPIOs

USPIOs Enter the Atherosclerotic Plaque ThroughMacrophages that engulfed themFissured or thin cap Extensive angiogenesis vasa vasorum leakage Intra plaque hemorrhage

In-vitro Study of Macrophage SPIO Uptake In a series of in-vitro studies we have tested

the rate of SPIO uptake by human activated monocytes in different conditions regardingincubation time and concentration of SPIO.All SPIO were labeled by a fluorescent dye(DCFA).

FL-labeled SPIO Incubated Macrophages 24hr

Double DAPI Staining with Fluorescence-labeled SPIO Macrophages after 24hr Incubation

Hypothesis

vasa vasorum

Over magnification is a major advantage of SPIO

Darkening property of SPIO in the white background of fat and water of plaque is another advantage

SPIO and T2 EffectIn-vitro study to show the effect of macrophage SPIO uptake on their T2 relaxation time

Macrophage Uptake of Feridex with Time and Concentration Shown by T2 ReductionConcentration mol/ml

Histopathologic Studies of ApoE KO Mice Injected with SPIO (Abdominal Aorta)H&E stainingIron Staining

CD 68 stainingIron particles

Histopathologic Study of Wild Type MiceInjected With SPIO (Thoracic Aorta) H&E stainingCD68 stainingIron staining

Comparison of the Number of the Iron Particles (per HPF) in ApoE KO Mice Plaque vs. Normal Wall

MR Image of Abdominal Aorta After SPIO Injection in ApoE and Control Mice

ApoE deficient mouse

C57B1 (control) mouse

Before Injection

After Injection (5 Days )

Dark (negatively enhanced) aortic wall, full of iron particles

Bright aortic lumen and wall without negative enhancement and no significant number of iron particles in pathology

Injection of Cytokine Increases SPIO-Loaded Macrophage Density in Aortic Plaques (Apo E Deficient Mice)Naghavi et al Circulation 2003

Naghavi et al Circulation 2003

Injection of Cytokine Increases SPIO-Loaded Macrophage Density in a Coronary Plaque (Apo E Deficient Mice)

Histopathologic studies of Thoracic aorta in WatanabeHereditary Hypercholesterolemic rabbit after SPIO injectionH&E stainingIron stainingIron staining

Histopathologic studies of Thoracic aorta in Watanabe Hereditary Hypercholesterolemic rabbit after SPIO injectionH&E stainingIron stainingIron stainingIron particles

R=0.956Correlation between Iron positive cells in Iron staining and cell density in H&E staining in rabbitatherosclerotic aorta.

Chart1

35

0

12

6

24

17

9

8

8

10

18

22

8

0

5

6

12

3

50

24

10

40

2

35

4

10

52

1

Cell Denity in H&E staining

SPIO positive cell -Iron staining

Plaque Cell Density vs SPIO

Sheet1

HPFsNumber of nucleated cells per HPF(H&E staining)0.956113827

13735

270

32012

4106

52824

62617

7129

8118

9128

101810

111818

123322

1388

1430

15115

16146

171412

1853

196050

201424

211410

226540

2332

245035

2554

261010

276452

2821

574431

574431

Sheet1

Cell Denity in H&E staining

SPIO positive cell -Iron staining

Plaque Cell Density vs SPIO

Sheet2

Sheet3

MR Angiography 3D with Gadolinium-DTPA in Watanabe RabbitBefore SPIO injectionAfter SPIO injection

MRI Identifies Plaque Inflammation by SPIO NanoparticlesWatanabe rabbitPost-SPIOWatanabe rabbitcontrolNZW rabbitcontrolNZW rabbitPost-SPIO

Ex-vivo MR study of the thoracic aorta in Watanabe and Wild type rabbit after SPIO injection compared to control.(Gradient echo)

Schmitz et al

Schmitz et al

Schmitz et al

Ruhem et al

Ruhem et al

Ruhem et al

Ruhem et al

Ruhem et al

Kooi et al

Kooi et al

Kooi et al

Kooi et al

Ho et al

Ho et al

MR signal changes after CsA treatment. A group with allotransplants was treated with CsA for either 7 days (n=5) or 4 days (n=5) after initial MRI experiments showed decrease of MR signal intensity according to degree of graft rejection. On POD 14, transplanted rats were reinjected with USPIO particles, and then MRI experiments were performed. Animals treated 7 days showed minimal changes in MR signal intensity 24 hours after reinjection (A and B), and animals treated 4 days showed MR signal intensity that was significantly decreased after USPIO injection (C and D). MR images of short-axis view of graft are shown. MR images before USPIO infusion are at A and C, and images taken 24 hours after infusion are at B and D. Ho et al

Ho et al

*

Imaging Macrophage Activity in the Brain Using Ultrasmall Particles of Iron Oxide

Jeff W.M. Bultea and Joseph A. Franka Laboratory of Diagnostic Radiology Research National Institutes of Health Bethesda, MD

*

SPIO MR imagine of macrophage is feasible for clinical applications.

It will be of great interest to investigate the cost-effectiveness of this technique in comparison with alternative diagnostic approaches.

Conclusion

*

Poor spatial and temporal resolution for coronary applicationsCostCumbersome (injection and delayed imaging)

Limitations

*

*

*

*

*

*

*

*

*

*

*