Direct Thrombus Imaging as a Means to Control the Variability of Mouse Embolic Infarct Models

Abstract

Background and Purpose-High experimental variability in mouse embolic stroke models could mask the effects of experimental treatments. We hypothesized that imaging thrombus directly would allow this variability to be controlled. Methods-We optically labeled thrombi with a near-infrared fluorescent (NIRF) probe C15 that is covalently linked to fibrin by factor-XIIIa. Labeled thrombus was injected into the left distal internal carotid artery (ICA) of C57/BL6 mice (nϭ47), near its bifurcation, and laser-Doppler cerebral-blood-flow (CBF) was assessed for 30 minutes. NIRF thrombus imaging was done ex vivo at 24 hours. Results-CBF variably decreased to 43.9Ϯ17.3% at 5 minutes (rCBF; 11.2ϳ80.4%). NIRF thrombus imaging at 24 hours showed variability in distribution (ICA bifurcation, adjacent and/or remote areas) and burden (2279Ϯ1270 pixels; 0ϳ5940 pixels). Final infarct size was also variable (21.0Ϯ10.3%; 4.7ϳ60.3% of the bihemispheric volume). Despite this heterogeneity, a strong thrombus-infarct correlation was maintained. The left hemispheric target infarct size (% of the hemisphere) correlated with thrombus burden, as a stronger predictor of infarct volume (PϽ0.001, rϭ0.50) than rCBF (Pϭ0.02, rϭϪ0.34). The infarct size was best predicted by a combination of thrombus imaging and CBF: left-hemispheric big-thrombi (Ͼ1865 pixels)/low-rCBF (Յ42%) had an infarct volume of 56.9Ϯ10.4% (nϭ12), big-thrombi/high-rCBF had 45.9Ϯ23.5% (nϭ11), small-thrombi/low-rCBF 35.7Ϯ17.3% (nϭ11) and small-thrombi/ high-rCBF 27.3Ϯ16.4% (nϭ12). Conclusions-This is the first study to demonstrate that the highly heterogeneous nature of the mouse embolic stroke model can be characterized and managed by using near-infrared fluorescent thrombus imaging combined with CBF monitoring to stratify animals into useful subgroups. Key Words: thrombus imaging Ⅲ embolic cerebral infarction Ⅲ molecular imaging Ⅲ optical imaging I schemic stroke caused by cerebral thromboembolism is a leading cause of death and disability. Much effort has been put into developing effective neuroprotective treatments, but with limited success. 1,2 A major difficulty in stroke research has been experimental variability in animal stroke models, [3][4][5] leading to failures in identifying neuroprotective drugs. Embolic stroke models, in which preformed clots are injected into the middle cerebral artery-anterior cerebral artery (MCA-ACA) bifurcation area, mimic human stroke more closely than do other models of cerebral ischemia. 6 -9 However, they provide less control over the location and extent of the resulting cerebral infarction. 3 Factors, such as spontaneous lysis 8 or distal embolization of thrombi after injection into the MCA, as well as anatomic variations in the circle of Willis, could have major and variable impacts on the resulting infarct. 10 This variability could mask and confound the potential therapeutic effects 3 of neuroprotective drugs, and by adding to experimental noise, might mask or heighten the effects of experimental treatments, leading to hard-to-interp...

Role of suture diameter and vessel insertion position in the establishment of the middle cerebral artery occlusion rat model

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Structured evaluation of rodent behavioral tests used in drug discovery research

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A New Micro–Computed Tomography–Based High-Resolution Blood–Brain Barrier Imaging Technique to Study Ischemic Stroke

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