Da Shi1, Jiachen Zhuo1, Su Xu1, Mary C. McKenna2, and Rao P. Gullapalli1
1Diagnostic Radiology, University of Maryland Baltimore, Baltimore, MD, United States, 2Department of Pediatrics, University of Maryland Baltimore, Baltimore, MD, United States
Synopsis
Fragile X
syndrome is the most common genetic cause of autism and is modeled
with the Fmr1 knockout mouse.
To
investigate recent report of myelination delay in Fragile X,
this study used
translational imaging techniques including T2 mapping and
magnetization transfer imaging to determine myelination changes in
the developing Fmr1 knockout mouse. Age-related trajectory changes in
regional white matter development were observed between the genotypes
and may provide insights into the pathophysiology of Fragile X.Introduction
Fragile X
syndrome (FXS) patients are afflicted with intellectual disability and
autistic behavior due to abnormal neuronal wiring
1.
Traditionally, the study of FXS focused on neuronal pathology but
recent evidence points towards myelination and white matter
alterations in both FXS and the Fmr1 knockout mouse (KO) model
2,3.
Myelin disruption has been shown to occur during the early postnatal
period in developing Fmr1 KO mice, including reduction of myelin proteins
and myelin thickness
3. We hypothesized that the reduction
in myelination could be best assessed in-vivo using T
2-mapping
techniques and magnetization transfer (MT) imaging.
Methods
Fmr1
KO (JAX-B6.129P2-fmr1tmICgr) and wildtype mice (JAX-C57Bl/6J; WT)
were bred at the University of Maryland, Baltimore. All experiments
were performed on a Bruker Biospec 7.0 Tesla 30cm horizontal bore
scanner. At postnatal-day 12 (P12), P20, P30, and P60 male mice were
anesthetized with isoflourane. Animals were grouped as: P12 Fmr1 KO
n=12, WT n=5; P20 Fmr1 KO n=11, WT n=4; P30 Fmr1 KO n=5, WT n=11; and
P60 Fmr1 KO n=12, WT n=8.
MT
images were acquired with FLASH T1 sequences with and
without off-resonance saturation pulse, TE/TR=3.6/3700ms, NA=1, slice
thickness=0.5mm, slice=14, and 150μm×150μm resolution. The
Gaussian off-resonance saturation pulse had a duration of 12.6ms and
flip-angle 540.0 degrees, off-resonance frequency of 1000Hz. T2
relaxation was measured using a multi-echo spin echo sequence with
TEs=8, 16, 24, 32, 40, 48, 56, 64ms, TR=10000ms, NA=1, slice
thickness=1mm, slice=2, and 100μm×100μm resolution.
Regions
of interests were drawn manually including the corpus callosum (CC),
external capsule (EC), internal capsule (IC), cerebral peduncle (CP)
and fimbria (FI). Medical Image Processing, Analysis and
Visualization tool (MIPAV v5.3.1, CIT; NIH) was used for MT images.
MT ratio (MTR) was calculated by
((M0-Ms)/M0)×100, where M0
is the signal without saturation and Ms is the signal
after saturation. T2 relaxation was
measured from averaged voxels from each of the regions using an
in-house program. The first echo at 8 ms was ignored to minimize
potential contamination from the stimulated echo.
Two-way
analysis of variance (ANOVA) was performed on MTR and T2
relaxation time, comparing the effect of genotype, age and
genotype×age interaction. One-way ANOVA
was used to determine age-related changes followed by Tukey's
post-hoc test. Genotype difference was determined with an independent
student's t-test.
Results
T2
relaxation time (Figure 1): The effect of age was significant for all
regions (p<0.001). One-way ANOVA and post-hoc revealed the T2
decreased with age in both Fmr1 KO and WT mice in all regions
studied. No effect of genotype or interaction were observed.
MTR: All
regions showed the effect of age with MTR (p<0.001) and the
developmental increase of MTR was revealed in both Fmr1 KO and WT
mice (p<0.05). Significant genotype effect was only observed in
the CP (p=0.046). The trajectories of age-related
changes in Fmr1 KO and WT mice were different in the CC, EC, and FI with age×genotype interaction observed in the CC
(p=0.12), EC (p=0.26) and FI (p=0.20).
Discussion
Our
results suggest a difference in the trajectory of regional white
matter development between the Fmr1 KO and WT mice. Patterns of
age-related T
2 reduction and MTR increase suggest enhanced
development of specific regions at the expense of other regions in
Fmr1 KO mice. The disparity in
age-related MTR increase may further indicate myelination delay in
the CC, EC, and FI in Fmr1 KO mice. The MTR change may be attributed to
the pervasive misregulation of protein synthesis in Fmr1 KO brain
4.
Significant genotype×age
interactions in the CC, EC and FI with MTR further solidify the
differences in the developmental trajectory of these regions between
Fmr1 KO and WT mice. The lack of genotype differences may
speak to the temporary nature of delayed myelination in Fmr1 KO mice,
which was observed only in early postnatal development and
subsequently normalized with age
3. These findings suggest
a difference in the temporal pattern of neurodevelopment in the Fmr1
KO mice compared to the WT mice and needs to be further substantiated
with further analysis of myelin.
Acknowledgements
Funding
partially supported by: FRAXA research
foundation
and NIH P01-HD016596-27.References
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