Computational protocol: Age-related functional brain changes in FMR1 premutation carriers

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[…] Statistical analysis on fMRI data was carried out using Statistical Parametric Mapping software (SPM12) (Wellcome Department of Clinical Neurology) using standard procedures.The functional images from all participants were preprocessed using the following steps: 1) images were realigned, estimated (for optimal transformation from individual images to the reference using SPM12 default quality, separation, smoothing, num passes, interpolation, wrapping and weighting parameters) and resliced 2) images were slice timed, adjusting for interleaved and bottom up slice order 3) subsequent images were then coregistered with the source structural image from the T1-weighted MPRAGE anatomical scan 4) coregistered images were segmented into grey matter, white matter and CSF outputs 5) segmented (forward deformation) images were normalised to MNI space and 6) finally, normalised images were smoothed with an 8 mm full-width at half maximum (FWHM) Gaussian smoothing kernel.Movement was controlled for by adding realignment parameters for each participant as a multiple regressor into the first level model. For the first-level analysis, the onsets and durations of the conditions were modelled, excluding the prompt screen stimuli. The contrast of interest here was between the sequential and random conditions, as we aimed to focus on brain regions associated with the changes in demand for movement cognition and planning. For the second-level analysis, an explicit mask was derived from an average binarised image created from the combined grey matter and white matter segmented images from all participants and was used to exclude voxel data out-with the brain tissue. Between group analyses were completed using a p threshold value of p < 0.005 and significant suprathreshold clusters were defined as FWE corrected p < 0.05. Firstly, between group analyses were performed using a two-sample t-test with age as a covariate. A full factorial design was used for a secondary analysis, carried out with age added into the model as a possible interacting variable.CGG repeat analysis revealed that some individuals in the sample were mosaic or borderline for the full mutation. To ensure this was not causing a bias in the results, a one-way ANOVA of 3 age-matched groups, followed by a Tukey post-hoc test was performed on extracted voxel data from the maximum voxel of the significantly different between group cluster and a univariate analysis of variance was performed on extracted voxel data from the age interaction significant cluster. [...] Imaging analyses are detailed above. Further statistical analyses were carried out in SPSS Statistics 22. A two-tailed t-test was used to discern differences between the premutation and control groups for the tremor, balance and coordination measurements, in addition to the FMR1 mRNA data. Simple linear regressions were also performed on extracted raw voxel values from significant clusters against the movement and molecular variables. All significance levels were assumed at p < 0.05. […]

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