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Cheung, Sau Ching Honor

University of British Columbia

Neurovascular impairments such as blood brain barrier (BBB) dysfunction, decreased cerebral blood flow, and small vessel disease have been associated with traumatic brain injury (TBI), dementia and other neurological disorders, and contribute to cognitive impairment. Due to the complex structure of the cerebral vasculature, it is technically challenging to routinely analyse the murine vasculature at high resolution using traditional histopathological or magnetic resonance imaging (MRI) approaches. The objective of this thesis is to develop an experimental pipeline to map the cerebral vasculature using tissue clearing, light sheet microscopy (LSM) and 3D image analysis to provide high-resolution characterization of the structure and integrity of the entire murine vasculature after TBI. Additionally, Apolipoprotein E (ApoE), which carries lipids in the brain in the form of lipoproteins, is the major genetic risk factor for sporadic Alzheimer’s Disease (AD). Recent studies have shown that APOE can exacerbate AD pathology via vascular pathways, and even in the absence of AD diagnosis, the APOE4 allele has emerged as a risk factor for small vessel disease and vascular cognitive impairment in comparison to the more common isoform APOE3. As proof-of-concept of the utility of our 3D analysis pipeline, we used the CHIMERA (Closed-Head Impact Model of Engineered Rotational Acceleration) TBI model to subject both ApoE4 targeted replacement (ApoE4-TR) and ApoE3 targeted replacement (ApoE3-TR) mice to TBI or sham procedures followed by transcardial perfusion of florescent wheat germ agglutinin (WGA) to label the whole brain vasculature. Brains were fixed using a technique called stabilization under harsh conditions via intramolecular epoxide linkages to prevent degradation (SHIELD) and passively cleared for imaging of WGA-labelled vessels using LSM. Volumetric images were stitched, 3D rendered and analyzed using Imaris image analysis software, to achieve a resolution of 1250x1250 pixels with an image resolution at pixel size of 1.5 x 1.5 x 5 µm (xyz plane). In summary, the work presented here provides proof of concept data of the establishment of an efficient imaging and analysis pipeline, which has great flexibility to be applied to a myriad of research projects examining different pathologies of the murine brain.

Text; Moving Image

2024-04-29

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/88081

Pathology and Laboratory Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/