Jean-Baptiste Perot1, Marina Célestine1, Marc Dhenain1, Sandrine Humbert2, Emmanuel Brouillet1, and Julien Flament1
1Université Paris-Saclay, Commissariat à l’Energie Atomique et aux Energies Alternatives (CEA), Centre National de la Recherche Scientifique (CNRS), Molecular Imaging Research Center (MIRCen), Laboratoire des Maladies Neurodégénératives, Fontenay-aux-Roses, France, 2Université Grenoble-Alpes, Grenoble Institute of Neurosciences (GIN), INSERM U1216, Grenoble, France
Synopsis
Huntington’s
Disease (HD) is a neurodegenerative disorder caused by the expansion of CAG
repeats on the exon 1 of the HTT gene. Although genetic origin of HD is well-established,
early and predictive biomarkers of disease onset and progression are still
lacking. In the present study, we performed a multiparametric longitudinal MRI
study on a mouse model of HD. Our results in gluCEST, Magnetization Transfer, morphometry
and Diffusion Tensor Imaging revealed early modifications of white matter followed
by progressive functional and anatomical changes in
HD mice. Such network seems to point out the central role of white matter in HD
pathogenesis.
Purpose
Huntington’s
Disease (HD) is a neurodegenerative disorder caused by the expansion of CAG
repeats on the exon 1 of the HTT gene1. The mutation causes
progressive neurodegeneration with preferential vulnerability of the striatum.
Atrophy of this structure is currently the main biomarker of the disease2
but there is a need to find earlier and more functional biomarkers.
In
a previous work, we showed that gluCEST imaging3 was able to
highlight defects in the white matter of a slowly progressive mouse model of HD4.
In the present study, we combined gluCEST with other state-of-the-art MRI
techniques to investigate the pivotal role of white matter alteration in HD
pathogenesis in this mouse model. In addition, we evaluated the potential of
different biomarkers that could help refining HD diagnosis and disease
monitoring.Material & Methods
Mouse model:
Knock-in mice expressing mouse/human exon 1 containing 140 CAG repeats inserted
in the murine huntingtin (Htt) gene were used5. Heterozygous mice
for the Htt gene (Ki140, n=11 males) were compared to their relative
age-matched littermates (WT, n=12 males).
MRI protocol:
Animals were scanned longitudinally (2.5, 5, 8, 12 and 18 months of age) on a
horizontal 11.7T Bruker magnet using a Cryoprobe. The MRI protocol included
anatomical (TSE sequence, 100 slices, 0.1 x 0.1 mm², 0.2 mm slice thickness),
gluCEST (Magnetization Transfer Ratio (MTRasym) at ±3 ppm calculated from a Zspectrum
acquired between -5 and 5 ppm, B1=5 µT, Tsat=1 s, WASSR
correction for B0 inhomogeneity6), magnetization transfer
(MT) (MTRasym at ±16 ppm, B1=10 µT, Tsat=800 ms) and
Diffusion Tensor Image sequences (EPI, 10 slices, 0.1125 x 0.1125 mm², 0.5 mm slice
thickness, b-value=1000 s/mm², 30 directions). Resting-state fMRI sequence was
added at 18 months (TE/TR=10/1000 ms, 0.2 x 0.2 mm2, 0.7 mm slice thickness, 12
slices, 450 repetitions).
Image Analysis: Images
were co-registered and automatically segmented using an atlas composed of a
high-resolution template based on Allen mouse brain atlas7. The
registration pipeline used an in-house python library (Sammba-MRI8).
DTI images were analyzed using the Tract-Based Spatial Statistics (TBSS9)
pipeline (FSL10).
Statistical analysis: After
Shapiro-Wilk normality test, one-way ANOVA with repeated measures was used for
statistical analysis and was followed by Fisher LSD post-hoc test.Results
Morphometry: Based
on volume measurements, variation maps between WT and Ki140 mice were
calculated (Fig.1a). Ki140 mice
exhibited progressive global atrophy of the brain, leading to significant
atrophy of the striatum (-4.3%, p<0.05), frontal cortex (-5.3%, p<0.05)
and motor cortex (-3.3%, p<0.05) at 18 months.
GluCEST and MT imaging: Variation
maps of gluCEST and MT signal were generated using the same process (Fig.1b-c).
GluCEST signal was similar in Ki140 and WT at 2 and 5 months of age. At 8
months, Ki140 mice displayed reduced gluCEST in the corpus callosum (CC,
-10.8%, p<0.05). Significant decrease of gluCEST signal was also measured in
the CC (-19%, p<0.01), in the frontal (-7.3%, p<0.05) and piriform
(-16.7%, p<0.05) cortices and in the pallidum (-21.0%, p<0.05) of Ki140
mice at 12 months. At the same timepoint, a downward trend was also visible in
the striatum of HD mice (-12%, p=0.12). MT contrast displayed the same behavior
with no alteration in young animals until 12 months, when a significant decrease
was measured in the septum (-21.7%, p<0.05) and a trend in the striatum
(-12.5%, p=0.12). In the latest timepoint, gluCEST and MT signals exhibited no significant variation.
DTI:
TBSS analysis showed decreased Fractional Anisotropy (FA) in anterior CC of Ki140
mice at 5 months (Fig.2). Clusters of voxels with altered diffusivity
also appeared at 8 and 12 months in CC and fimbria.
RS-fMRI:
Correlation matrix analysis showed reduced connectivity of the striatum to
other regions of interest in Ki140 compared with WT (n.s., Fig. 3)
Discussion
Atrophy
observed at 18 months is consistent with clinical HD according to literature11-13, as well as reduced gluCEST contrast measured in the CC at 12 months4.
However, our results showed that gluCEST and MT imaging could give earlier and
more functional information than morphometric analysis.
Using
axonal projection data of the mouse brain14 (Fig.4), we discovered that striatal and cortical regions displaying
defects in MT, gluCEST or atrophy were strongly connected. FA alterations at 5
months showed evidence of microstructural modifications in the anterior CC,
which drives most of these connections. CC also exhibited earliest and firmest
metabolic impairments. This highlights the central role that white matter
alteration may play in HD pathogenesis.
Finally,
a resting-state fMRI sequence acquired during the last timepoint showed a reduced
functional connectivity of the striatum to other regions of interest,
especially motor, retrosplenial and piriform cortices (Fig.3). This tends to
confirm that white matter, and more precisely cortico-striatal connections, was
strongly involved in HD pathogenesis in this mouse model.Conclusion
Our
study emphasizes the interest of DTI, CEST and MT imaging combined with
automated segmentation as biomarkers of neurodegenerative diseases. We showed that, in the Ki140 mouse model, white matter alteration precedes striatal
impairments and may induce the vulnerability of this structure by affecting
cortico-striatal connectivity. In the future, it would be of great interest to
explore further early alterations of white matter and to transfer our protocol
to clinical HD.Acknowledgements
Acknowledgment
Project
was supported by eRARE ERA-Net (“TreatPolyQ” ANR-17-RAR3-0008-01) and NeurATRIS,
(“Investissements d'Avenir”, ANR-11-INBS-0011). The 11.7T scanner was funded by
NeurATRIS (“Investissements d'Avenir”, ANR-11-INBS-0011).References
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