Micro-computed tomography in murine models of cerebral cavernous malformations as a paradigm for brain disease

Abstract

Background Cerebral cavernous malformations (CCMs) are hemorrhagic brain lesions, where murine models allow major mechanistic discoveries, ushering genetic manipulations and preclinical assessment of therapies. Histology for lesion counting and morphometry is essential yet tedious and time consuming. We herein describe the application and validations of X-ray micro-computed tomography (micro-CT), a nondestructive technique allowing three-dimensional CCM lesion count and volumetric measurements, in transgenic murine brains. New Method We hereby describe a new contrast soaking technique not previously applied to murine models of CCM disease. Volumetric segmentation and image processing paradigm allowed for histologic correlations and quantitative validations not previously reported with the micro-CT technique in brain vascular disease. Results Twenty-two hyper-dense areas on micro-CT images, identified as CCM lesions, were matched by histology. The inter-rater reliability analysis showed strong consistency in the CCM lesion identification and staging (K=0.89, p<0.0001) between the two techniques. Micro-CT revealed a 29% greater CCM lesion detection efficiency, and 80% improved time efficiency. Comparison with Existing Method Serial integrated lesional area by histology showed a strong positive correlation with micro-CT estimated volume (r2= 0.84, p<0.0001). Conclusions Micro-CT allows high throughput assessment of lesion count and volume in pre-clinical murine models of CCM. This approach complements histology with improved accuracy and efficiency, and can be applied for lesion burden assessment in other brain diseases.

RhoA Kinase Inhibition With Fasudil Versus Simvastatin in Murine Models of Cerebral Cavernous Malformations

“…The combined effect of therapy on lesional count and cross sectional area promises to be more easily and accurately assessed in ongoing experiments using novel micro-computed tomography technique recently introduced and validated by our team, with volumetric quantification of CCM lesion burden in murine brains. 50 …”

Endothelial TLR4 and the microbiome drive cerebral cavernous malformations

“…For all experiments utilizing microCT quantification of CCM lesion volume, brains were harvested and immediately placed in 4% PFA/PBS fixative. Brains remained in fixative until staining with non-destructive, iodine contrast and subsequent microCT imaging performed as previously described 47 . Importantly, all tissue processing, imaging, and volume quantification were done in a blinded manner by investigators at The University of Chicago without any knowledge of experimental details.…”

Phenotypic characterization of murine models of cerebral cavernous malformations

“…Micro-CT imaging and volumetric assessments of lesion burden were performed using a published protocol. 40 Fixed extracted brains from the respective models were shipped to the University of Chicago, where they were processed by soaking in iodine to increase the differential contrast between CCMs and brain tissue on micro-CT images. Brains were then scanned using a transmission 180 kV nanofocus X-ray of a Phoenix v|tome|x s micro-computed tomography system (General Electric, Fairfield, CT, USA).…”