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CEST imaging of ApoE –/– mouse brain during atherosclerosis development1City University of Hong Kong, Hong Kong, Hong Kong, 2Hong Kong Centre for Cerebro-Cardiovascular Health Engineering (COCHE), Hong Kong, Hong Kong, 3Russell H. Morgan Department of Radiology and Radiological Science, , The Johns Hopkins University School of Medicine, Baltimore, MD, United States, 4City University of Hong Kong Shenzhen Research Institute, Shenzhen, China, 5Tung Biomedical Sciences Centre, City University of Hong Kong, Hong Kong, Hong Kong
Synopsis
Keywords: Atherosclerosis, Atherosclerosis
Motivation: The molecular imaging and relationship between the brain and atherosclerosis are poorly understood.
Goal(s): To study the brain during atherosclerosis development and the association of atherosclerosis and brain using CEST MRI.
Approach: 3 months old ApoE –/– and C57BL/6 mouse brains were imaged using 3T preclinical MRI and CEST was applied, then three offsets, 3.5, –1.6 and –3.5 ppm, were extracted for post-processing.
Results: ApoE –/– mouse showed higher APT and rNOE signals than WT in thalamus, hippocampus and cortex, which are associated with neuroinflammation and cholesterol deposition during atherosclerosis development.
Impact: The underlying molecular changes of the brain during atherosclerosis development could enhance the identification of diseases at early stage using CEST MRI.
Introduction
Atherosclerosis is developed when plaques composed of cholesterol, fat or cells build up inside the artery, followed by immune-inflammatory process1. It is a chronic development that vulnerable plaque might ultimately lead to myocardial infarction or stroke2. Therefore, early detection and intervention of atherosclerosis are essential. The mechanism of early stages of atherosclerosis is well established, which could lead to molecular alterations in the brain globally or specifically in certain regions, due to immunological process activation3. This prompts us to image molecular changes during the plaque development, especially those related to atherosclerosis. Here, we have applied CEST-MRI onto the Apolipoprotein E (ApoE) deficient mouse brain to observe the brain during atherosclerosis development at a molecular level.Methods
C57BL/6 (wildtype) and ApoE –/– mouse (female, 3 months) were used for mouse brain imaging using 3T Bruker Biospec system (Bruker, Germany) with a 82 mm volume transmitter and a single surface coil as a receiver. For CEST acquisition, the parameters follow as below: TR/TE = 5000/ 56.96 ms, B1=0.8 µT , tsat= 3000 ms. For post-processing, three offsets, 3.5, –1.6 and –3.5 ppm, were extracted using Lorentzian difference analysis using custom-written Matlab code. For MISL and MT, –10 ppm was used for the labeled MT signal and 300 ppm for the control image. Images were acquired by subtracting labeled MT signal from the control, followed by normalizing Z-spectrum images to M0 images. Weight of the mice was measured after the scanning.Results and Discussion
Given that the foam cell lesion is detectable when ApoE –/– mouse reaches at 10 weeks old4, the corresponding age-matched mice with wildtype were compared. From the whole brain, ApoE –/– mouse showed higher signals than WT at all offsets, especially at 3.5 and –3.5 ppm (P=0.0140, 0.0055, Fig 1). At 3.5 ppm, 17.10% higher signals were observed in ApoE –/– mouse, which could be attributed to neuroinflammation in the brain5, where vascular dysfunction caused by atherosclerosis results in inflammation and BBB breakdown6 and increased inflammatory cells could lead to an increase in APT signal5. Similarly, offsets at –1.6 and –3.5 ppm were higher than WT (16.67 and 14.83%), which could be associated with the cholesterol deposition in the plaque7, 8. As the serum cholesterol level elevates along with the progression of atherosclerosis, it might influence the brain through interaction with brain lipoproteins and regulation of cholesterol homeostasis9, where increased aliphatic protons in cholesterol have led to rNOE signal increase7.In order to further study other contributions to the brain caused by atherosclerosis, certain brain regions were studied, such as thalamus, hippocampus, and cortex. Interestingly, all regions showed constant trends as the whole brain that all signals in ApoE –/– mouse were higher than those in WT (Fig 2B-D). At –3.5 ppm, three regions of ApoE –/– mouse had significantly higher signals than WT, indicating that the cholesterol accumulation in atherosclerotic plaque affects cerebral vasculature, leading to higher rNOE signals at those regions9. Especially at hippocampus region, ApoE –/– mouse showed significantly higher CEST signals at all offsets than WT mouse (3.5 ppm: 18.17%; –1.6 ppm: 24.59%; –3.5 ppm: 16.36%). Notably, hippocampus is the only region, which showed significant differences at both APT and rNOE signals. In ApoE –/– mouse, many studies have reported strong neuroinflammatory responses, such as GFAP -positive astrocytes and Iba 1-positive microglia in hippocampus10, 11, which aligns with our results at 3.5 ppm (Fig 2C).
Further neuropathological analysis and studies, such as longitudinal observations of ApoE –/– mouse, are underway to monitor the continuous molecular changes when the atherosclerotic plaques become more evident. At the same time, histological validations are under investigation to confirm different stages of the disease. Our study might reveal the relationship between the brain and atherosclerosis with the molecular alterations.
Conclusion
Atherosclerosis is a major cause of cardiovascular disease and its current understandings related to the brain are not clear, especially at a molecular level. Here, with ApoE –/– mouse, which is widely used animal model for atherosclerosis, we have imaged and analyzed different regions of the brain using CEST-MRI to see molecular differences compared to WT mouse. Our results indicate that APT and rNOE signals were higher than WT due to neuroinflammation and cholesterol elevations, respectively. This hypothesis is currently under investigation and further studies are underway.Acknowledgements
Research Grants Council (11102218, 11200422, RFS2223-1S02, C1134-20G), City University of Hong Kong (7005433, 7005626, 9609307, 9610560 and 9610616), National Natural Science Foundation of China (81871409), Tung Biomedical Sciences Centre and Hong Kong Centre for Cerebro-cardiovascular Health Engineering (COCHE).References
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