Brain virtual histology with X-ray phase-contrast tomography Part II: 3D morphologies of amyloid-β plaques in Alzheimer’s disease models

Abstract

While numerous transgenic mouse strains have been produced to model the formation of amyloid-β (Aβ) plaques in the brain, efficient methods for whole-brain 3D analysis of Aβ deposits have to be validated and standardized. Moreover, routine immunohistochemistry performed on brain slices precludes any shape analysis of Aβ plaques, or require complex procedures for serial acquisition and reconstruction. The present study shows how in-line (propagation-based) X-ray phase-contrast tomography (XPCT) combined with ethanol-induced brain sample dehydration enables hippocampus-wide detection and morphometric analysis of Aβ plaques. Performed in three distinct Alzheimer mouse strains, the proposed workflow identified differences in signal intensity and 3D shape parameters: 3xTg displayed a different type of Aβ plaques, with a larger volume and area, greater elongation, flatness and mean breadth, and more intense average signal than J20 and APP/PS1. As a label-free non-destructive technique, XPCT can be combined with standard immunohistochemistry. XPCT virtual histology could thus become instrumental in quantifying the 3D spreading and the morphological impact of seeding when studying prion-like properties of Aβ aggregates in animal models of Alzheimer’s disease. This is Part II of a series of two articles reporting the value of in-line XPCT for virtual histology of the brain; Part I shows how in-line XPCT enables 3D myelin mapping in the whole rodent brain and in human autopsy brain tissue.

Why are amyloid-β plaques detected by X-ray phase-contrast imaging? Role of metals revealed through combined synchrotron infrared and X-ray fluorescence microscopies

“…Using propagation-based imaging, now the most widely employed XPCT method for virtual histology of label-free samples, we made original observations. In animals, Aβ plaques from the J20 mouse strain and the TgF344 rat strain appeared as small bright (hyperintense) spots, as already reported for these and for other transgenic models such as 3xTg [10], APP/PS1 [8,10], TauPS2APP [5] and B6C3-Tg [13]. But surprisingly, Aβ plaques in the newly imaged APPPS1 and ArcAβ mouse strains appeared as dark (hypointense) spots, with the largest plaques frequently exhibiting an hyperintense core.…”

“…X-ray phase-contrast tomography (XPCT) refers to a family of techniques which uses the phase shift rather than the attenuation of X-rays to probe the microstructure in soft biological tissues. X-ray 3D imaging of Aβ plaques has been described in transgenic rodent models that develop amyloidosis for the past decade, using propagation-based phase contrast (either with synchrotron radiation [5, 6, 7, 8, 9, 10] or with a laboratory source [11]) or using gratingbased phase contrast (with synchrotron radiation [12, 13, 14, 5]). These techniques generate 3D images with an isotropic resolution ranging from 1 µm to 10 µm, thus providing a so-called “virtual histology” of the excised brain [6, 15, 16, 17, 18, 19] with minimal preparation (fixation, dehydration or paraffin embedding).…”

“…Imaging AD samples with XPCT is particularly interesting because plaques stand out in the images without the need to add any staining agents. Previous studies highlighted the great potential of XPCT for the quantitative analysis of amyloid-β plaques (overall burden [14, 5], spatial distribution [8], 3D morphometry [10]). These advances (which bring original information not available to standard immunohistochemistry), combined with increasingly faster acquisitions (less than 5 min per sample [10]), position XPCT as a potential screening tool for amyloid pathology.…”

“…Previous studies highlighted the great potential of XPCT for the quantitative analysis of amyloid-β plaques (overall burden [14, 5], spatial distribution [8], 3D morphometry [10]). These advances (which bring original information not available to standard immunohistochemistry), combined with increasingly faster acquisitions (less than 5 min per sample [10]), position XPCT as a potential screening tool for amyloid pathology. However, up to now, the substrate underlying contrast of Aβ plaques in XPCT images remains unexplored.…”

Revealing the three-dimensional murine brain microstructure by contrast-enhanced computed tomography

“…However, to obtain a high discriminative power, methods for enhancing the contrast between different tissue constituents are required. At present, the most promising contrast-enhancing methods include phase-contrast microCT (PCCT) ( Takeda et al, 2013 ; Barbone et al, 2021 ; Rodgers et al, 2021 , 2022 ; Chourrout et al, 2022a , b ; Palermo et al, 2022 ) and contrast-enhanced microCT (CECT) ( Rodrigues et al, 2021 ). While PCCT makes use of the refractive properties of the X-rays, CECT aims to increase the X-ray attenuating properties of different tissue constituents (e.g., white matter versus gray matter) by incubating or infiltrating the tissue with contrast-enhancing staining agents (CESAs).…”