Localization and Quantification of Platelet-Rich Thrombi in Large Blood Vessels With Near-Infrared Fluorescence Imaging

Abstract

Background-Imaging of thrombus formation in vivo has been limited by the inability to directly visualize and measure thrombi in large blood vessels in real time. Near-infrared light, with its superior tissue penetration and reduced scatter, could potentially solve this problem. Methods and Results-Platelets were labeled with the near-infrared fluorophore IR-786. Optimal total fluorescence yield occurred at 6 attomoles of IR-786 per platelet. IR-786 -labeled platelets were tested for their ability to detect thrombus formation in large animal model systems relevant to common human vascular procedures. Invisible near-infrared light did not distort the surgical field in any way, and even after optimization of per-platelet fluorescent yield, platelets remained fully functional. Intravenous infusion of just 3.6ϫ10 10 labeled platelets into a 35-kg Yorkshire pig permitted thrombus visualization, with a signal-to-background ratio Ն2, for at least 2 hours in coronary, carotid, and femoral vessels. Platelet-rich, actively growing clots were monitored in real time and quantified with respect to size and kinetics after injury to vessels, cutaneous incisions, intravascular stent insertion, or introduction of embolic coils. Similarly, formed clots were monitored in real time during thrombolysis with streptokinase and heparin. Vessel patency was assessed independently with a second near-infrared fluorescent blood pool agent. Conclusions-IR-786 -labeled platelets provide sensitive, specific, and real-time visualization of thrombi in thick-walled blood vessels. In addition to immediate application in cardiac, transplant, and vascular surgery, the mechanisms that underlie thrombus formation in large blood vessels can now be investigated.

Magnetic Resonance Imaging Contrast Agent Targeted Toward Activated Platelets Allows In Vivo Detection of Thrombosis and Monitoring of Thrombolysis

“…Recently, localization and quantification of platelet-rich thrombi in large blood vessels were performed with fluorescence-labeled platelets, which allowed imaging of clot formation and real-time imaging of thrombolysis. 24 In this approach, 3.6ϫ10 10 in vitro-labeled platelets were used to achieve good contrast properties. Conversely, the in vivo MR approach reported here uses only an exogenous reagent and does not require subject-specific preparation, which would not be practicable in the context of acute thrombotic syndromes.…”

Prevention of Ventricular Arrhythmias With Sarcoplasmic Reticulum Ca 2+ ATPase Pump Overexpression in a Porcine Model of Ischemia Reperfusion

“…The areas of gene distribution identified by IRDYE 786 16 and fluorescent microspheres exhibited homogeneous distribution of the particles in the left ventricle ( Figure 3). The expression efficiency was demonstrated by immunohistochemistry, immunoblotting, and qRT-PCR.…”

“…102185-03-5, Sigma-Aldrich, St Louis, Mo) 16 and fluorescent microspheres (Molecular Probes, Carlsbad, Calif) instead of adenoviral solution ( Figure 3). …”

“…To assess the area at risk (AAR), a solution with red fluorescent microspheres (Molecular Probes) mixed to near-infrared fluorescent dye (IRDYE 786) 16 was injected immediately before euthanization into the proximal coronary arteries while a balloon was inflated at the same location used during ischemia in all reperfused pigs and with no balloons in all pigs with PO or the sham procedure. To delineate infarct size, the hearts were sliced into 1-cm-thick slices and stained with triphenyltetrazolium chloride (TTC; Sigma) as previously described.…”

Imaging Biomarkers in Atherosclerosis Trials

“…14 Molecular imaging techniques are available for most imaging modalities, including CT, 15 integrated nuclear CT imaging, 16 -21 optical imaging, [22][23][24][25][26][27] and MRI. 28 -34 Although major advances have been made in molecular imaging techniques, the clinical emergence of targeted molecular probes is slower.…”