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Maranzano, J.

Publications and source records attributed to Maranzano, J..

At least 19 recordsLinked to original sources

T2 relaxometry for myelin water fraction: an ex vivo brain imaging study

Abstract Introduction: Myelin water imaging provides a non-invasive approach for indirectly investigating myelin content of the brain by magnetic resonance Imaging (MRI). Myelin water fraction (MWF) reflects the fraction of water signal associated with water trapped between myelin layers. Postmortem imaging provides a unique opportunity to validate MWF metrics against gold-standard postmortem neuropathology. However, there is a need for validated sequences that would be applicable in postmortem settings. In this study, we validated and compared the applicability of turbo spin echo (TSE) and gradient-and-spin-echo (GRASE) techniques for myelin water imaging in formaldehyde fixed postmortem human brains. Methods: 40 postmortem human brain hemispheres were scanned with TSE and GRASE sequences. Acquired data were reconstructed using a non-parametric multicomponent T2 relaxometry approach using a consistent framework to support comparison between GRASE- and TSE-derived measures. Agreement between derived measures was quantified at both voxel and regional level. Maps derived from the full 32-echo GRASE reconstruction were compared against a truncated 14-echo reconstruction to examine the contribution of the longer echoes to the derived maps. Finally, a random forest regression model was trained based on TSE data to predict the GRASE-derived MWF maps. Results: Both sequences provided robust multicomponent T2 characterization across the brain, and the derived measures showed anatomical patterns consistent with expected differences across tissue types. TSE and GRASE derived maps had moderate to strong agreement at voxel (0.41< {rho} < 0.82, all pFDR<0.001) and regional levels (0.87< {rho} < 0.98, all pFDR<0.001), with consistently stronger agreement observed for regional metrics. Echo reduction comparisons between the full 32-echo GRASE reconstruction and a truncated 14-echo reconstruction showed minimal impact of echo truncation (all {rho} > 0.98, pFDR < 0.001). The nonlinear random forest regression model trained on TSE maps was able to accurately reproduce the MWF maps derived from GRASE (r = 0.91, RMSE = 8.87). Discussion: TSE and GRASE32 capture strongly related multicomponent T2 information in fixed postmortem human brain tissue, but with systematic and tissue-dependent quantitative differences across derived measures. These differences suggest that direct interchangeability should not be assumed and that cross-sequence mapping or calibration is required when quantitative equivalence is desired.

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Neuronal quantification in the primary motor cortex of mouse brains fixed with solutions from human gross anatomy laboratories

Background: Brain banks usually provide small tissue blocks fixed by immersion in neutral-buffered formalin (NBF). While still underexploited for research, gross anatomy laboratories could provide full brains fixed by perfusion with solutions better suited for gross anatomy dissection. However, the chemicals in these solutions might have a different impact on histology protocols for cell quantification than in NBF-fixed brains. The main goal of this study is to compare the effects on the number and size of labeled neurons of the primary motor cortex (PMC) of mouse brains fixed with three different solutions: (1) NBF, typical of brain banks, (2) a saturated salt solution (SSS), and (3) an alcohol-formaldehyde solution (AFS), both used in human anatomy laboratories. Methods: 27 C57BL/6J mouse brains were perfused with the NBF (N=9), SSS (N=9) or AFS (N=9), then cut in 40-m slices and processed with immunohistochemistry to target neurons. Various quantitative variables were assessed manually and automatically on photomicrographs of 3 regions of interest (ROIs) of the PMC per specimen, namely the total and individual neuronal profile areas, number and diameters. The effects of the three fixatives on these variables were compared using ANOVA or Kruskal-Wallis, depending on the distribution. For measures on individual cells, a generalized linear mixed model was applied. Dice coefficients and correlations were applied to evaluate the agreement of the manual and automatic methods. Results: There was no significant difference between the brains fixed by the three fixatives for the total and individual cell areas, the total cell count and the cell diameters. The values obtained from manual and automatic measures had an overall good agreement (Dice coefficients > 0.79). Conclusion: It was found that the SSS and AFS had similar impacts on the quantitative variables in the tissue as the NBF. These results are promising for neuroscientists interested in using brains from anatomy laboratories for quantitative research on neurons from the PMC.

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Spatiotemporal trajectories of formaldehyde fixation effects on quantitative MRI in postmortem human brains

Introduction: Postmortem human brain magnetic resonance imaging (MRI) offers a unique opportunity to study finer neuroanatomical details and enables direct correlations with gold standard histological and immunohistochemical assessments. However, to prevent tissue decay, postmortem brains are preserved in fixative solutions which can alter tissue properties and exert substantial impacts on the MRI signals. The present study investigates the impact of formalin fixation, the most commonly used solution for postmortem human brain preservation, on different quantitative MRI contrasts. Methods: 142 intact human brain hemispheres immersed in 10% formalin for a range of fixation durations (between 0 days and 20 years) were imaged in a 3T MRI scanner. A subset of 10 brains were further scanned repeatedly at days 0, 3, 10, 20, 30, 60, 90, and 120 to allow for better characterization of the initial transient effects of fixation. Voxel-wise T1 and T2* relaxation, T1/T2 ratio, and myelin water fraction (MWF) maps were generated for each specimen and timepoint, and linear and nonlinear models were used to examine the spatiotemporal changes associated with progressive fixation. Results: All investigated metrics were significantly impacted by formalin fixation, albeit at different rates and with differing regional patterns. T1 and T2* relaxation time decreased as a result of progressive fixation, whereas T1/T2 ratio and MWF measures increased. T1 relaxation and T1/T2 ratio showed nonlinear patterns with initially accelerated changes that decelerate in the first few months, whereas T2* relaxation and MWF changes followed a more linear trend. Conclusion: Formaldehyde fixation exerts systematic changes on quantitative MRI signals that can be modeled and adjusted for to allow for harmonized comparisons of MRI metrics across brains fixed for differing durations. The distinct temporal trajectories observed across metrics highlight the need to account for fixation duration in study design and downstream analyses, particularly when integrating datasets acquired under heterogeneous conditions. Our findings provide a quantitative framework for correcting fixation-induced biases, thereby improving the interpretability and reproducibility of postmortem MRI studies.

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Immunofluorescence quality of human brain tissue fixed with solutions used in gross anatomy laboratories

Brain banks provide small tissue samples fixed in neutral-buffered-formalin (NBF), but human anatomy teaching laboratories could provide full brains fixed with solutions that are more appropriate for gross anatomy such as a saturated salt solution (SSS) or an alcohol-formaldehyde solution (AFS). Advanced aging and prolonged exposure to aldehydes are known to enhance brain tissue autofluorescence (AF), limiting the efficacy of immunofluorescence (IF) procedures. We have previously shown by IF staining the antigenicity preservation in mouse brains fixed with the three solutions. We now aimed to compare the quality of IF staining in human brains fixed with SSS, AFS and NBF. In addition, we compared the efficiency of AF quenching methods, namely the application of SudanBlackB (SBB) and the treatment of sections with sodium borohydride (NaBH4). Blocks of neocortex were extracted from 18 brains (NBF=6, SSS=6, AFS=6) and cut into 40{micro}m sections. Neurons (anti-NeuN, AlexaFluor-488) and astrocytes (anti-GFAP, AlexaFluor-555) were revealed with IF after an antigen retrieval protocol, while two treatments (SBB or NaBH4) were used to quench AF. We then assessed the degree of AF (criteria: background or cell AF) and the immunostaining quality with excitation wavelengths of 488nm, 555nm and 647nm. Brains fixed with all three solutions showed well-labeled astrocytes, whereas neurons werent always stained, but this was not associated to the fixative solution. The overall AF intensity was similar in sections from brains fixed with all three solutions. Finally, the SBB treatment was the most effective at reducing AF in all specimens. Given the similarity in AF and antigenicity assessment across the three solutions, we conclude that brains fixed with SSS and AFS could be good alternatives for NBF-fixed specimens in the context of IF experiments processed with a SBB protocol. Highlights- Immunofluorescence staining is feasible in brains fixed with anatomy labs solutions - GFAP is less affected by fixation than NeuN - Autofluorescence can be reduced by Sudan Black treatment

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Effects of prolonged post-fixation on vascular biomarkers in postmortem human brains

Postmortem human brains stored in brain banks are important research resources to study the mechanisms underlying normal brain functions as well as various neurodegenerative disorders. Immunohistochemical (IHC) and histochemical (HC) staining have been used to examine human brains fixed in neutral-buffered formalin (NBF) for months, years, and even decades. As such, it is essential to establish the effects of prolonged fixation in NBF on both IHC and HC stains. Previously, we found that prolonged NBF fixation resulted in differential effects on IHC and HC staining on postmortem brains. In this study, we further examined the effects of prolonged fixation on IHC stains targeting 6 antigens and 2 HC stains of known biomarkers of cerebrovascular diseases in prefrontal cortex of human brains fixed for 1, 5, 10, 15, and 20 years. The IHC targets included microvasculature markers of the blood brain barrier (Collagen-IV and Claudin-5), a type III intermediate filament marker (Vimentin), an activated microglia marker (CD68), a biomarker for oligodendrocytic myelin proteolipid protein (PLP) and a marker for iron accumulation (Ferritin). The HC included Massons Trichrome Stain (MTS) and Bielschowsky silver stain (BSS). We found that staining intensities of Ferritin, Vimentin, Collagen-IV and BSS decreased with prolonged fixation, while no significant differences were observed in the staining intensity of other markers. Hence, these differential alterations should be taken into consideration when interpreting the results from processed tissues with prolonged fixation. We recommend performing IHC and HC staining for human brains with the same fixation times to offset any impact on downstream neuropathological analyses, as well as adding the fixation duration as a covariate in the analysis.

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Comparison of T1- and T2-weighted MRI contrasts of ex vivo ex situ brains fixed with solutions used in gross anatomy laboratories

Post-mortem magnetic resonance imaging (MRI) offers high resolution and histological correlation, so protocols have been developed by brain banks using hemispheres fixed by immersion in Neutral-buffered formalin (NBF), but they provide limited tissue samples. Conversely, anatomy laboratories could supply complete brains perfused either with a salt-saturated (SSS) or an alcohol-formaldehyde (AFS) solution. These fixation methods alter the brains molecular properties, potentially affecting MRI quality and structural characteristics. T1- and T2-weighted (T1w, T2w) contrasts change with NBF fixation, but the effects of SSS or AFS remain unknown. We compared T1w and T2w intensities of different regions of interest (ROIs), including subcortical white matter (WM), cortical and deep gray matter (GM), in brains fixed with NBF, SSS and AFS. We scanned 20 ex situ hemispheres (NBF-immersed=7; SSS-perfused=7; AFS-perfused=6) in a 3T MRI scanner using T1w (0.7mm3) and T2w (0.64mm3) sequences overnight. Mean intensities of 29 ROIs in T1w and T2w MRIs and GM-WM ratios were calculated and compared in brains fixed with the three solutions. We found that T1w images were more affected by the fixation process, inverting the contrast of in vivo T1w and reducing the GM-WM contrast in AFS-fixed brains. T2w images resembled in vivo scans and maintained a sharp contrast in brains fixed with the three solutions, although the GM-WM intensity ratios were lowered in SSS-fixed brains. In conclusion, brains fixed with SSS and AFS from anatomy laboratories could be used for MRI studies, especially with the T2w sequence that seems more appropriate for structural analyses in different ROIs.

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Histology to MRI registration quality of ex vivo human brain blocks fixed with solutions used in anatomy laboratories

IntroductionPost-mortem brain tissue is obtained from brain banks that provide small tissue samples, while gross anatomy laboratories could become a source of complete brains for neuroscientists. These are preserved with solutions whose chemical composition differs from the classic neutral-buffered formalin (NBF) used in brain banks, such as a saturated-salt-solution (SSS) or an alcohol-formaldehyde solution (AFS) that preserve antigenicity of the main brain cell populations. Since histology remains the gold standard in neuroscientific research, while MRI is the most common imaging modality, MRI-histology registration quality needs to be assessed to ensure the suitability using brains fixed with innovative solutions for research procedures. Hence, our goal was to compare the registration quality of human brains fixed with NBF, SSS and AFS, as well as the histological characteristics that could affect the registration. MethodsWe used 12 human brain blocks of 3x3x3 cm3 fixed in our anatomy laboratory using SSS (N=4), AFS (N=4), or NBF (N=4). The blocks were scanned using a 7Tesla Bruker animal MRI scanner with a T2-TurboRARE sequence at 0.13x0.13x0.5mm3. Blocks were then cut into 40m thick sections (parallel to the 0.13x0.13 plane) using a vibratome. Sections were stained with histochemistry (HC) (Cresyl violet, Prussian blue, Luxol fast blue, H&E, and Bielschowsky) and immunohistochemistry (IHC) of the 4 main cell populations: neurons (NeuN), astrocytes (GFAP), microglia (Iba1) and myelin (PLP) either with or without an antigen retrieval (AR) protocol. Stained sections were imaged using a slide scanner microscope and segmented with masks using Display, and the sections were manually registered to the T2-TurboRare images using landmarks in Register (MincToolKit). ResultsMore landmarks were needed to achieve proper alignment of the histology to MRI images for the SSS-fixed blocks, due to the lower GM-WM contrast in these brains. However, there was no significant difference in the staining intensity of the histology sections of blocks fixed with the three solutions, while SSS-fixed blocks showed a lower percentage of overlap between the good histology quality masks and the MRI masks. This resulted in a sufficient registration quality of all blocks, although more challenging when fixed with SSS. ConclusionWe have developed histology, MRI and registration protocols that are of good quality in brain blocks fixed with solutions used in gross anatomy laboratories. These results are promising for neuroscientists interested in using full brains from anatomy laboratories, either using MRI, histology or registration of both modalities to study normal aging and neurodegenerative conditions.

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Ex vivo human brain volumetry: validation of magnetic resonance imaging measurements

BackgroundNeurodegenerative diseases are associated with brain atrophy. The volume of in vivo human brains is determined with various magnetic resonance imaging (MRI) measurement tools of which the validity has not been assessed against a gold standard. Here, we propose to validate the MRI brain volumes by scanning ex vivo-in situ specimens (i.e., anatomical heads), which allows the extraction of the brain after the scan to compare its volume with the gold standard water displacement method (WDM). MethodsWe acquired 3T MRI T2-weighted, T1-weighted, and MP2RAGE images of seven anatomical heads fixed with an alcohol-formaldehyde solution routinely used in anatomy laboratories and segmented the gray and white matter of the brain using two methods: 1) a manual intensity-based threshold segmentation using Display (MINC-ToolKit, McConnell BIC), and 2) an automatic Deep-Learning-based segmentation tool (SynthSeg). The brains were then extracted, and their volumes were measured with the WDM after the removal of their meninges and a midsagittal cut (to allow water penetration into the ventricles). Volumes from all methods were compared to the ground truth (WDM volumes) using a repeated-measures ANOVA. ResultsMean brain volumes, in cubic centimeters, were 1111.14{+/-}121.78 for WDM, 1020.29{+/-}70.01 for manual T2-weighted, 1056.29{+/-}90.54 for automatic T2-weighted, 1094.69{+/-}100.51 for automatic T1-weighted, 1066.56{+/-}96.52 for automatic MP2RAGE INV1, and 1156.18{+/-}121.87 for MP2RAGE INV2. All volumetry methods were significantly different (F=17.874; p<0.001) from the WDM volumes, except the automatic T1-weighted volumes. ConclusionWe demonstrate that SynthSeg accurately determines the brain volume in ex vivo-in situ T1-weighted MRI scans. Our results also suggest that given the contrast similarity between our ex vivo and in vivo sequences, the brain volumes of clinical studies are most probably sufficiently accurate, with some degree of underestimation depending on the sequence used.

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The Douglas Bell Canada Brain Bank Post-mortem Brain Imaging Protocol

Magnetic resonance imaging (MRI) is a valuable non-invasive tool that has been widely used for in vivo investigations of brain morphometry and microstructural characteristics. Postmortem MRIs can provide complementary anatomical and microstructural information to in vivo imaging and ex vivo neuropathological assessments without compromising the sample for future investigations. We have developed a postmortem MRI protocol for the brain specimens of the Douglas-Bell Canada Brain Bank (DBCBB), the largest brain bank in Canada housing over 3000 neurotypical and diseased brain specimens, that allows for acquisition of high-resolution 3T and 7T MRIs. Our protocol can be used to scan DBCBB specimens with minimal tissue manipulation, allowing for feasibly scanning large numbers of postmortem specimens while retaining the quality of the tissue for downstream histology and immunohistochemistry assessments. We demonstrate the robustness of this protocol in spite of the dependency of image quality on fixation by acquiring data on the first day of extraction and fixation, to over twenty years post fixation. The acquired images can be used to perform volumetric segmentations, cortical thickness measurements, and quantitative analyses which can be potentially used to link MRI-derived and ex vivo histological measures, assaying both the normative organization of the brain and ex vivo measures of pathology.

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Comparison of histological procedures and antigenicity of human post-mortem brains fixed with solutions used in gross anatomy laboratories

BackgroundBrain banks provide small tissue samples to researchers, while gross anatomy laboratories could provide larger samples, including complete brains to neuroscientists. However, they are preserved with solutions appropriate for gross-dissection, different from the classic neutral-buffered formalin (NBF) used in brain banks. Our previous work in mice showed that two gross-anatomy laboratory solutions, a saturated-salt-solution (SSS) and an alcohol-formaldehyde-solution (AFS), preserve antigenicity of the main cellular markers. Our goal is now to compare the quality of histology and antigenicity preservation of human brains fixed with NBF by immersion (practice of brain banks) vs. those fixed with a SSS and an AFS by whole body perfusion, practice of gross-anatomy laboratories. MethodsWe used a convenience sample of 42 brains (31 males, 11 females; 25-90 years old) fixed with NBF (N=12), SSS (N=13), and AFS (N=17). One cm3 tissue blocks were cut, cryoprotected, frozen and sliced into 40 m sections. The four cell populations were labeled using immunohistochemistry (neuronal-nuclei (NeuN), glial-fibrillary-acidic-protein (GFAP), ionized-calcium-binding-adaptor-molecule1 (Iba1) and myelin-proteolipid-protein (PLP)). We qualitatively assessed antigenicity and cell distribution, and compared the ease of manipulation of the sections, the microscopic tissue quality, and the quality of common histochemical stains (e.g., Cresyl violet, Luxol fast blue, etc.) across solutions. ResultsSections of SSS-fixed brains were more difficult to manipulate and showed poorer tissue quality than those from brains fixed with the other solutions. The four antigens were preserved, and cell labeling was more often homogenous in AFS-fixed specimens. NeuN and GFAP were not always present in the NBF and SSS samples. Some antigens were heterogeneously distributed in some specimens, independently of the fixative, but an antigen retrieval protocol successfully recovered them. Finally, the histochemical stains were of sufficient quality regardless of the fixative, although neurons were more often paler in SSS-fixed specimens. ConclusionAntigenicity was preserved in human brains fixed with solutions used in human gross-anatomy (albeit the poorer quality of SSS-fixed specimens). For some specific variables, histology quality was superior in AFS-fixed brains. Furthermore, we show the feasibility of frequently used histochemical stains. These results are promising for neuroscientists interested in using brain specimens from anatomy laboratories.

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Antigenicity is preserved with fixative solutions used in human gross anatomy: A mice brain immunohistochemistry study.

BackgroundHistology remains the gold standard to assess human brain biology, so ex vivo studies using tissue from brain banks are standard practice in neuroscientific research. However, a larger number of specimens could be obtained from gross anatomy laboratories. These specimens are fixed with solutions appropriate for dissections, but whether they also preserve brain tissue antigenicity is unclear. Therefore, we perfused mice brains with solutions used for human body preservation to assess and compare the tissue quality and antigenicity of the main cell populations. Methods28 C57BL/6J mice were perfused with: 4% formaldehyde (FAS, N=9), salt-saturated solution (SSS, N=9), and alcohol-solution (AS, N=10). The brains were cut into 40m sections for antigenicity analysis and were assessed by immunohistochemistry of four antigens: neuronal nuclei (NeuN), glial fibrillary acidic protein (GFAP-astrocytes), ionized calcium binding adaptor molecule1 (Iba1-microglia), and myelin proteolipid protein (PLP). We compared the fixatives according to multiple variables: perfusion quality, ease of manipulation, tissue quality, immunohistochemistry quality, and antigenicity preservation. ResultsThe perfusion quality was better using FAS and worse using AS. The manipulation was very poor in SSS brains. FAS and AS fixed brains showed higher tissue and immunohistochemistry quality than the SSS brains. All antigens were readily observed in every specimen, regardless of the fixative solution. ConclusionSolutions designed to preserve specimens for human gross anatomy dissections also preserve tissue antigenicity in different brain cells. This offers opportunities for the use of human brains fixed in gross anatomy laboratories to assess normal or pathological conditions. SIGNIFICANCE STATEMENTNeuroscientists currently obtain tissue samples from brain banks. Alternatively, a much larger amount of tissue may be obtained from bodies donated to gross anatomy laboratories. However, they are preserved with different fixative solutions that are used for dissection purposes. We ignore if these solutions also preserve antigenicity of the main cell populations, essential in neuroscientific research. This work is the first to show that two solutions currently used in human gross anatomy laboratories preserve sufficient histological quality, in addition to preserve antigenicity of the main cell populations of the mice brain. This work opens the door to the use of human brain tissue obtained from anatomy laboratories in neuroscientific research.

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Using Transfer Learning for Automated Microbleed Segmentation

IntroductionCerebral microbleeds are small perivascular haemorrhages that can occur in both grey and white matter brain regions. Microbleeds are a marker of cerebrovascular pathology, and are associated with an increased risk of cognitive decline and dementia. Microbleeds can be identified and manually segmented by expert radiologists and neurologists, usually from susceptibility-contrast MRI. The latter is hard to harmonize across scanners, while manual segmentation is laborious, time-consuming, and subject to inter- and intra-rater variabiltiy. Automated techniques so far have shown high accuracy at a neighborhood ("patch") level at the expense of a high number of false positives voxel-wise lesions. We aimed to develop an automated, more precise microbleeds segmentation tool able to use standardizable MRI contrasts. MethodsWe first trained a ResNet50 network on another MRI segmentations task (cerberospinal fluid versus background segmentation) using T1-weighted, T2-weighted, and T2* MRI. We then used transfer learning to train the network for the detection of microbleeds with the same contrasts. As a final step, we employed a combination of morphological operators and rules at the local lesion level to remove false positives. Manual segmentations of microbleeds from 78 participants were used to train and validate the system. We assessed the impact of patch size, freezing weights of the initial layers, mini-batch size, learning rate, as well as data augmentation on the performance of the Microbleed ResNet50 network. ResultsThe proposed method achieved a high performance, with a patch-level sensitivity, specificity, and accuracy of 99.57%, 99.16%, and 99.93%, respectively. At a per lesion level, sensitivity, precision, and Dice similarity index values were 89.1%, 20.1%, and 0.28 for cortical GM; 100%, 100%, and 1.0 for deep GM; and 91.1%, 44.3%, and 0.58 for WM, respectively. DiscussionThe proposed microbleed segmentation method is more suitable for the automated detection of microbleeds with high sensitivity.

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White Matter Hyperintensity Distribution Differences in Aging and Neurodegenerative Disease Cohorts

IntroductionWhite matter hyperintensities (WMHs) are common magnetic resonance imaging (MRI) findings in the aging population in general, as well as in patients with neurodegenerative diseases. They are known to exacerbate the cognitive deficits and worsen the clinical outcomes in the patients. However, it is not well-understood whether there are disease-specific differences in prevalence and distribution of WMHs in different neurodegenerative disorders. MethodsData included 976 participants with cross-sectional T1-weighted and fluid attenuated inversion recovery (FLAIR) MRIs from the Comprehensive Assessment of Neurodegeneration and Dementia (COMPASS-ND) cohort of the Canadian Consortium on Neurodegeneration in Aging (CCNA) with eleven distinct diagnostic groups: cognitively intact elderly (CIE), subjective cognitive impairment (SCI), mild cognitive impairment (MCI), vascular MCI (V-MCI), Alzheimers dementia (AD), vascular AD (V-AD), frontotemporal dementia (FTD), Lewy body dementia (LBD), cognitively intact elderly with Parkinsons disease (PD-CIE), cognitively impaired Parkinsons disease (PD-CI), and mixed dementias. WMHs were segmented using a previously validated automated technique. WMH volumes in each lobe and hemisphere were compared against matched CIE individuals, as well as each other, and between men and women. ResultsAll cognitively impaired diagnostic groups had significantly greater overall WMH volumes than the CIE group. Vascular groups (i.e. V-MCI, V-AD, and mixed dementia) had significantly greater WMH volumes than all other groups, except for FTD, which also had significantly greater WMH volumes than all non-vascular groups. Women tended to have lower WMH burden than men in most groups and regions, controlling for age. The left frontal lobe tended to have a lower WMH burden than the right in all groups. In contrast, the right occipital lobe tended to have greater WMH loads than the left. ConclusionsThere were distinct differences in WMH prevalence and distribution across diagnostic groups, sexes, and in terms of asymmetry. WMH burden was significantly greater in all neurodegenerative dementia groups, likely encompassing areas exclusively impacted by neurodegeneration as well as areas related to cerebrovascular disease pathology.

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Diffusely abnormal white matter converts to T2 lesion volume in the absence of acute inflammation

Diffusely abnormal white matter (DAWM), characterised by biochemical changes of myelin in the absence of frank demyelination, has been associated with clinical progression in secondary progressive MS (SPMS). However, little is known about changes of DAWM over time and their relation to focal white matter lesions (FWML). The objectives of this work were: 1) To characterize the longitudinal evolution of FWML, DAWM, and DAWM that transforms into FWML, and 2) To determine whether gadolinium enhancement, known to be associated with the development of new FWML, is also related to DAWM voxels that transform into FWML. Our data included 4220 MRI scans of 689 SPMS participants, followed for 156 weeks and 2677 scans of 686 RRMS participants, followed for 96 weeks. FWML and DAWM were segmented using a previously validated, automatic thresholding technique based on normalized T2 intensity values. Using longitudinally registered images, DAWM voxels at each visit that transformed into FWML on the last MRI scan as well as their overlap with gadolinium enhancing lesion masks were identified. Our results showed that the average yearly rate of conversion of DAWM-to-FWML was 1.27cc for SPMS and 0.80cc for RRMS. FWML in SPMS participants significantly increased (t=3.9; p=0.0001) while DAWM significantly decreased (t=-4.3 p<0.0001) and the ratio FWML:DAWM increased (t=12.7; p<0.00001). RRMS participants also showed an increase in the FWML:DAWM Ratio (t=6.9; p<0.00001) but without a significant change of the individual volumes. Gadolinium enhancement was associated with 7.3% and 18.7% of focal New T2 lesion formation in the infrequent scans of the RRMS and SPMS cohorts, respectively. In comparison, only 0.1% and 0.0% of DAWM-to-FWML voxels overlapped with gadolinium enhancement. We conclude that DAWM transforms into FWML over time, in both RRMS and SPMS. DAWM appears to represent a form of pre-lesional pathology that contributes to T2 lesion volume increase over time, independent of new focal inflammation and gadolinium enhancement.

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PhyloBrain atlas: a cortical brain MRI atlas following a phylogenetic approach

Cortical atlases constitute a consistent division of the human cortex into areas that have common structural as well as meaningful and distinctive functional characteristics. The most widely used atlases follow the cytoarchitectonic and myeloarchitectonic characteristics of the cortex and have been combined to the standard anatomical nomenclature of gyri and sulci. More recently, common functional features depicted by resting state functional MRI have also guided the division of the cortical brain in functional regions of interest. However, to date, there are no atlases that divide the cortex considering the common evolutionary changes experienced by the mammalian cortex. Hence, the present study proposes the division of cortical areas into five main regions of interest (ROIs) following a phylogenetic approach: 1- archicortex, 2- paleocortex, 3- peri-archicortex, 4- proisocortex, 5-neocortex, and twelve neocortical sub-ROIs: 5.1.temporopolar, 5.2.post-central, 5.3.pre-central, 5.4.pericalcarine, 5.5.superior temporal, 5.6.middle temporal, 5.7.precuneus, 5.8.insular, 5.9.inferior parietal, 5.10.caudal anterior, 5.11.posterior cingulate, and 5.12.lingual gyrus. The segmentations were done using the T1-weighted MNI-ICBM152 non-linear 6th generation symmetric average brain MRI model.

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Proof of concept of a novel ex vivo, in situ method for MRI and histological brain assessment

MRI-histology correlation studies of the ex vivo brain mostly employ fresh, extracted (ex situ) specimens, aldehyde fixed by immersion. This method entails manipulation of the fresh brain during extraction, introducing several disadvantages: deformation of the specimen prior to MRI acquisition; introduction of air bubbles in the sulci, creating artifacts; and uneven or poor fixation of the deeper regions of the brain. We propose a new paradigm to scan the ex vivo brain, exploiting a technique used by anatomists: fixation by whole body perfusion, which implies fixation of the brain in situ. This allows scanning the brain surrounded by fluids, meninges, and skull, thus preserving the structural relationships of the brain in vivo and avoiding the disadvantages of ex situ scanning. Our aims were: 1) to assess whether months of in situ fixation resulted in a loss of fluid around the brain; 2) to evaluate whether in situ fixation modified antigenicity for myelin and neuron specific marker; 3) to assess whether in situ fixation improved the register of ex vivo brain images to standard neuroanatomical templates in pseudo-Talairach space for morphometry studies. Five head specimens fixed with a saturated sodium chloride solution (a non-standard fixative used in our anatomy laboratory for neurosurgical simulation) were employed. We acquired 3D T1-weighted (MPRAGE), 2D fluid-attenuated inversion recovery T2-weighted turbo spin echo (T2w-FLAIR), and 3D gradient-echo (3D-GRE) pulse sequences of all brains on a 1.5T MRI. After brain extraction, sections were processed for binding with myelin basic protein (MBP) and neuronal nuclei (NeuN) primary antibodies by immunofluorescence. This study showed that all but one specimen retained fluids in the subarachnoid and ventricular spaces. The specimen that lost fluid was the oldest one, with the longest interval between the time of death and the MRI scanning day being 403 days. All T1-weighted images were successfully processed through a validated pipeline used with in vivo MRIs. The pipeline did not require any modification to run on the ex vivo-in situ scans. All scans were successfully registered to the brain template, more accurately than an ex vivo-ex situ scan and exhibited positive antigenicity for MBP and NeuN. MRI and histology study of the ex vivo-in situ brain fixed by perfusion is feasible and allows for in situ MRI imaging for of at least 10 months post-mortem prior to histology analyses. Fluids around and inside the brain specimens and antigenicity for myelin and neurons were all well preserved.

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Conversion of Diffusely Abnormal White Matter to Focal Lesions is Linked to Progression in Secondary Progressive Multiple Sclerosis

Objectives1) To automatically segment focal white matter lesions (FWML) and Diffusely abnormal white matter (DAWM), i.e. regions of diffuse abnormality observed on conventional (T2-weighted) MRI and characterize their longitudinal volumetric and normalized T1-weighted (T1w) intensity evolution, 2) To assess associations of FWML and DAWM with Expanded Disability Status Scale (EDSS) and confirmed disability progression (CDP).\n\nMethodsData includes 3951 timepoints of 589 SPMS participants followed for three years. FWML and DAWM were automatically segmented using a 2-weighted-intensity thresholding technique. Screening DAWM volumes that transformed into FWML at the last visit (DAWM-to-FWML) and normalized T1w intensities (as a marker of severity of damage) in those voxels were calculated.\n\nResultsFWML volume significantly increased and DAWM volume significantly decreased as disease duration increased (p<0.001). Global EDSS scores were positively associated with FWML volumes (p=0.002), but not with DAWM volumes. Median volume of DAWM-to-FWML was significantly higher in patients who progressed (2.75 vs 1.70 cc; p<0.0001), and represented 14% of the total DAWM volume at screening, compared to 10% in patients who did not progress (p=0.001). Normalized T1w intensity values of DAWM-to-FWML were negatively associated with CDP status (p<0.00001).\n\nConclusionDAWM transformed into FWML over time, and this transformation was significantly associated with clinical progression. DAWM voxels that transformed had greater normalized T1w intensity decrease over time, in keeping with relatively greater tissue damage evolution. Evaluation of DAWM in progressive MS provides a useful measure to evaluate therapies that aim to protect this at-risk tissue with the potential to slow progression.

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Automated Separation of Diffusely Abnormal White Matter from Focal White Matter Lesions on MRI in Multiple Sclerosis

BackgroundPrevious histopathology and MRI studies have addressed the differences between focal white matter lesions (FWML) and diffusely abnormal white matter (DAWM) in multiple sclerosis (MS). These two categories of white matter T2-weighted (T2w) hyperintensity show different degrees of demyelination, axonal loss and immune cell density on histopathology, potentially offering distinct correlations with symptoms.\n\nObjectives1) To automate the separation of FWML and DAWM using T2w MRI intensity thresholds and to investigate their differences in magnetization transfer ratios (MTR), which are sensitive to myelin content; 2) to correlate MTR values in FWML and DAWM with normalized signal intensity values on fluid attenuated inversion recovery (FLAIR), T2w, and T1-weighted (T1w) contrasts, as well as with the ratio of T2w/T1w normalized values, in order to determine whether these normalized intensities can be used as myelin-sensitive markers when MTR is not available.\n\nMethodsUsing similar 3T MRI protocols, 2 MS cohorts of 20 participants each were scanned in 2 centers, including: 3D T1w (MP2RAGE), 3D FLAIR, 2D T2w, and 3D magnetization-transfer (MT) contrasts. We used the first dataset to develop an automated technique to separate FWML from DAWM and the second one to validate the automation of the technique. We applied the automatic thresholds to both datasets and assessed the overlap of the manual and the automated masks using Dice kappa. We also assessed differences in mean MTR values between NAWM, DAWM and FWML, using manually and automatically derived masks in both datasets. Finally, we used the mean intensity of manually-traced areas of NAWM on T2w images as the normalization factor for each MRI contrast, and compared these with the normalized-intensity values obtained using automated NAWM (A-NAWM) masks as the normalization factor. Paired t-tests assessed the MTR differences across tissue types. Wilcoxon Signed Rank test and paired t-tests assessed FWML and DAWM differences in manual and automated derived volumes. Pearson correlations assessed the relationship between MTR and normalized intensity values in the manual and automated derived masks.\n\nResultsThe mean Dice-kappa values for dataset 1 were: 0.8 for DAWM masks and 0.7 for FWML masks. In dataset 2, mean dice-kappa values were: 0.8 for DAWM and 0.8 for FWML. Also, manual and automated DAWM and FWML volumes were not significantly different in both datasets. MTR values (expressed as mean and standard deviation, arbitrary units) were significantly lower in manually derived FWML compared with DAWM in both datasets: 1) FWML: 37.1 {+/-} 3.2 vs DAWM: 43.3 {+/-} 2.1; t=13.2; p<0.0001, and 2) FWML: 32.5 {+/-} 3.9 vs DAWM: 38.8 {+/-} 1.7; t=9.8; p<0.0001. Similar results were obtained using the automatic derived masks in both datasets: 1) FWML: 35.8 {+/-} 2.8 vs DAWM: 43.1 {+/-} 1.6; t=15.3; p<0.0001, and 2) FWML: 31.3 {+/-} 3.1 vs DAWM: 38.1 {+/-} 1.5; t=12.8; p<0.0001. MTR was also significantly lower in manually derived DAWM masks compared with normal appearing white matter (NAWM) in both datasets: 1) NAWM: 46.7 {+/-} 1.3; t=10.1; p<0.0001, and 2) NAWM: 39.3 {+/-} 0.8; t=3.1; p=0.003. Similar results were obtained using the automatic derived masks in both datasets: 1) NAWM: 46.3 {+/-} 1.1; t=13.7; p<0.0001, and 2) NAWM: 39.9 {+/-} 1.1; t=9.6; p<0.0001.\n\nIn both datasets, manually derived FWML and DAWM MTR values showed significant correlations with normalized FLAIR (r=-0.35 to -0.67; p=0.06 to <0.0001), T2w (r=-0.60 to -0.72; p=0.003 to <0.0001), T1w/T2w (r=0.63 to 0.77; p=0.002 to <0.0001), and T1w (r=0.77 to 0.92; p<0.0001) intensities. Finally, normalized intensity values obtained using automatic derived masks were significantly correlated with the manually derived values in both datasets.\n\nConclusionsThe separation of FWML and DAWM on MRI scans of MS patients using automated intensity thresholds on T2w images is feasible. MTR values are significantly lower in FWML than DAWM, and DAWM values are significantly lower than NAWM, reflecting potentially greater demyelination within focal lesions. Normalized intensity values of different MRI contrasts exhibit a significant correlation with MTR values in both tissues of interest and could be used as a proxy to assess demyelination when MTR images are not available.

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