Cryogenic contrast-enhanced microCT enables nondestructive 3D quantitative histopathology of soft biological tissues

Biological tissues comprise a spatiallycomplex structure, composition andorganization at the microscale, named the microstructure. Given the closestructure-function relationships in tissues, structural characterization isessential to fully understand the functioning of healthy and pathological tis-sues, as well as the impact of possible treatments. Here, we present a non-destructive imaging approach to perform quantitative 3D histo(patho)logy ofbiological tissues, termed Cryogenic Contrast-Enhanced MicroCT (cryo-CECT). By combining sample staining,using an X-ray contrast-enhancingstaining agent, with freezing the sample at the optimal freezing rate, cryo-CECT enables 3D visualization and structural analysis of individual tissueconstituents, such as muscle and collagenfibers. We applied cryo-CECT onmurine hearts subjected to pressure overload following transverse aorticconstriction surgery. Cryo-CECT allowed to analyze, in an unprecedentedmanner, the orientation and diameter of the individual musclefibers in theentire heart, as well as the 3D localization offibrotic regions within the myo-cardial layers. We foresee further applications of cryo-CECT in the optimiza-tion of tissue/food preservation and donor banking, showing that cryo-CECTalso has clinical and industrial potential.

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