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Cited by 186Advanced Neuroimaging Techniques and ApplicationsFunctional Brain Connectivity StudiesAdvanced MRI Techniques and Applications

The role of myelination in measures of white matter integrity: Combination of diffusion tensor imaging and two-photon microscopy of CLARITY intact brains

Open access
Chang, Eric H., Argyelan, Miklos, Aggarwal, Manisha, Chandon, Toni-Shay S., Karlsgodt, Katherine H., Mori, Susumu, Malhotra, Anil K.·2017·NeuroImage
DOI 10.1016/j.neuroimage.2016.11.068PMC PMC5560594
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Abstract

Diffusion tensor imaging (DTI) is used extensively in neuroscience to noninvasively estimate white matter (WM) microarchitecture. However, the diffusion signal is inherently ambiguous because it infers WM structure from the orientation of water diffusion and cannot identify the biological sources of diffusion changes. To compare inferred WM estimates to directly labeled axonal elements, we performed a novel within-subjects combination of high-resolution ex vivo DTI with two-photon laser microscopy of intact mouse brains rendered optically transparent by Clear Lipid-exchanged, Anatomically Rigid, Imaging/immunostaining compatible, Tissue hYdrogel (CLARITY). We found that myelin basic protein (MBP) immunofluorescence significantly correlated with fractional anisotropy (FA), especially in WM regions with coherent fiber orientations and low fiber dispersion. Our results provide evidence that FA is particularly sensitive to myelination in WM regions with these characteristics. Furthermore, we found that radial diffusivity (RD) was only sensitive to myelination in a subset of WM tracts, suggesting that the association of RD with myelin should be used cautiously. This combined DTI-CLARITY approach illustrates, for the first time, a framework for using brain-wide immunolabeling of WM targets to elucidate the relationship between the diffusion signal and its biological underpinnings. This study also demonstrates the feasibility of a within-subject combination of noninvasive neuroimaging and tissue clearing techniques that has broader implications for neuroscience research.

AI snapshot

Written from the paper’s full text.

TL;DR

Combining ex vivo DTI with CLARITY whole-brain myelin basic protein (MBP) immunofluorescence in the same mouse brains, the authors found that FA correlates with MBP, especially in coherent, highly myelinated white matter regions. RD correlated with MBP only in a subset of tracts, indicating that RD–myelin associations should be interpreted cautiously.

Key findings

  • CLARITY MBP-labeled ROI volumes showed excellent structural correspondence with DTI ROIs (Spearman’s ρ=0.876, P<0.00001).
  • Across all WM ROIs, FA significantly correlated with MBP immunofluorescence (Χ²(1)=7.74, P<0.01), whereas AD (Χ²(1)=0.27, P=0.61) and RD (Χ²(1)=0.05, P=0.83) did not.
  • In a WM subset including corpus callosum, fimbria, and anterior commissure, MBP positively correlated with FA (Χ²(1)=12.56, P<0.0005) and negatively with RD (Χ²(1)=10.54, P<0.005); in remaining tracts, FA still correlated with MBP (Χ²(1)=10.62, P<0.005) but RD did not (P=0.37).
  • Within the stria medullaris, FA–MBP correlation was significant in high-FA ROIs >0.5 (ρ=0.666, P<0.00001, n=44 regions) but not in low-FA ROIs (P=0.118); high-FA regions had higher linear anisotropy (Cl=0.51±0.021) than diverging-fascicle regions (Cl=0.35±0.024).

Methods

  • Within-subjects design combining ex vivo DTI at 11.7 T with CLARITY/PACT tissue clearing and two-photon microscopy of MBP immunofluorescence in intact mouse brains.
  • Main experiments used four male C57BL/6 J mice, 3–6 months old; three male thy1-eYFP-H mice were used for initial pilot compatibility testing.
  • DTI used a 3D DW-GRASE sequence, 15 directions, b=1500 s/mm², 100 μm isotropic resolution, ~16.5 h scan time; 14 discrete 3D WM ROIs were traced in DTI and CLARITY spaces, and mixed-effect models tested DTI metrics as predictors of MBP with ROI as fixed effect and subject as random effect.

Limitations

  • The paper states that incomplete antibody penetration or uneven labeling may leave some myelinated areas uncaptured; fixation and the liquid hydrogel may affect diffusivity in ways that cannot be fully ruled out; and MBP immunofluorescence quantification in fixed tissue must be interpreted cautiously, with electron microscopy needed for further validation.
  • The paper also notes that the FA–myelin relationship is complex and cannot be resolved by this correlational approach alone, and that MBP expression does not necessarily index myelin sheath thickness or the percentage of myelinated axons.
  • Evident from the text: the main sample is small (four mice), imaging is ex vivo at 28 °C, and even at 100 μm resolution each voxel may contain 1,000–13,000 axons, over 80 oligodendrocytes, and more than 20 astrocytes, limiting biological specificity.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1016/j.neuroimage.2016.11.068

Cites

65 of 66 references verified1 citation errors · 0 not checkable

Statistics

Claims the paper makes

12 of 36 cited sources checked · 4 supported · 0 flagged · accuracy not yet measured

  • Interpretationown data

    CLARITY could be successfully performed despite the presence of a contrast agent in the hydrogel and minor deviations from the original published protocol.

    “This demonstrated that CLARITY could be successfully performed despite the presence of a contrast agent in the hydrogel and minor deviations from the original published protocol (Chung et al., 2013).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Chung et al., 2013 source
  • Premisecitation

    When coupled with two-photon or light-sheet laser microscopy, whole mouse brains can be imaged entirely intact with sub-cellular resolution.

    “When coupled with two-photon or light-sheet laser microscopy, whole mouse brains can be imaged entirely intact with sub-cellular resolution (Tomer et al., 2014).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Tomer et al., 2014 source

      The Overview of procedure and Anticipated Results (page unavailable) report high-resolution imaging through an intact mouse brain and whole-brain imaging with CLARITY-optimized light-sheet microscopy. The paper does not demonstrate whole-brain imaging with two-photon microscopy or establish sub-cellular resolution throughout the whole brain. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
  • Premisecitation

    WM integrity can be affected by many different parameters including axon packing density, axon caliber, myelination, microglia, inflammation, and tissue architecture.

    “This has led to considerable debate, since the inception of DTI, over the biological significance of these DTI-derived measures since WM integrity can be affected by many different parameters including axon packing density, axon caliber, myelination, microglia, inflammation, and tissue architecture (Mori and Zhang, 2006; Le Bihan and Johansen-Berg, 2012; Jones et al., 2013).”
    Quote in the paperSubstantial claim1 of 3 sources checked
    • cites Mori and Zhang, 2006 source
    • cites Le Bihan and Johansen-Berg, 2012 source

      The section on diffusion anisotropy in white matter links the measure to fiber packing, membrane organization, myelin, and tissue orientation; the section on brain development also discusses axon density and diameter. The cited paper supports the broader point that DTI measures have multiple biological influences, but our copy does not identify microglia or inflammation as influences on white-matter integrity or these measures (searched: “microglia” and “inflammation”). No page numbers were provided for these passages. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites Jones et al., 2013 source
  • Premisecitation

    Up to 90% of WM voxels in a DTI map contain complex fiber architecture and cannot be modeled accurately using a single tensor model.

    “It has been estimated that up to 90% of WM voxels in a DTI map contain complex fiber architecture and cannot be modeled accurately using a single tensor model (Jeurissen et al., 2013).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Jeurissen et al., 2013 source
  • Findingown data

    There is a discrepancy between the principal eigenvectors and the physical fiber bundles present at those locations when moving from macroscopic to microscopic scales.

    “Moving from macroscopic to microscopic scales further highlights the discrepancy between the principal eigenvectors and the physical fiber bundles that are actually present at those locations in the brain (Fig. 5A and B).”
    Quote in the paperSubstantial claim
  • May combine two claims. 19
  • Classified as probably not a substantial claim. 9

  • May combine two claims.

    Premisecitation

    CLARITY achieves tissue transparency to enable deep laser penetration in normally highly light-scattering mouse brains, while preserving proteins in situ and the native 3D structure of brain tissue.

    “CLARITY achieves tissue transparency to enable deep laser penetration in normally highly light-scattering mouse brains, while preserving proteins in situ and the native 3D structure of brain tissue.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Chung et al., 2013 source

  • Classified as probably not a substantial claim.

    Premisecitation

    CLARITY is a clearing technique that transforms intact brain tissue into nanoporous, lipid-free, hydrogel-tissue hybrids that are optically transparent but macromolecule permeable.

    “Clear Lipid-exchanged, Anatomically Rigid, Imaging/immunostaining compatible, Tissue hYdrogel (CLARITY; Chung et al., 2013) is a clearing technique that transforms intact brain tissue into nanoporous, lipid-free, hydrogel-tissue hydrids that are optically transparent but macromolecule permeable.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Chung et al., 2013 source

      The Abstract describes CLARITY as transforming intact tissue into a nanoporous, optically transparent, macromolecule-permeable hydrogel hybrid; the ‘Hydrogel-electrophoretic tissue transmutation’ section describes lipid extraction. However, the citing sentence expands the acronym differently from the paper’s Methods > CLARITY section, which specifies ‘Acrylamide-hybridized Rigid’ rather than ‘Anatomically Rigid.’ (LLM, conf=0.84)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    Even at 100 μm resolution and ∼17 h scan times, each imaging voxel can contain anywhere from 1,000 to 13,000 axons, over 80 oligodendrocytes, and more than 20 astrocytes.

    “However, even at this 100 μm resolution and ∼17 h scan times, each imaging voxel can contain anywhere from 1,000 to 13,000 axons, over 80 oligodendrocytes, and more than 20 astrocytes (Walhovd et al., 2014).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Walhovd et al., 2014 source

      In “Toward a multiscale understanding: From a myelin biology point of view, what does a voxel contain?” (page unavailable), the paper reports 1,000 to over 13,000 axons and averages of 86 oligodendrocytes and 22 astrocytes in a 100 μm-cubed volume of rodent white matter. These support the cited counts, though the glial figures are averages rather than counts guaranteed for each voxel. The passages found do not substantiate the ∼17-hour scan time. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductcitation

    Multiple variables can influence the quantitation of fluorophores in fixed tissue samples, so the interpretation of MBP immunofluorescence as a marker of myelination must be considered cautiously.

    “Lastly, we have used MBP immunofluorescence as a marker of myelination, however, multiple variables can influence the quantitation of fluorophores in fixed tissue samples (Straight, 2007; Waters, 2009) and this interpretation must be considered cautiously.”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites Straight, 2007 source
    • cites Waters, 2009 source

  • May combine two claims.

    Study design and conductcitation

    We used an ultra high field 11.7 T scanner with a 3D DW-GRASE sequence that minimized distortion while allowing for a 12-fold acceleration compared to spin echo imaging.

    “we used an ultra high field 11.7 T scanner with a 3D DW-GRASE sequence that minimized distortion while allowing for a 12-fold acceleration compared to spin echo imaging (Aggarwal et al., 2010).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Aggarwal et al., 2010 source

      Materials and Methods (DTI acquisition and Pulse sequence details; page unavailable) describe an 11.7 T instrument and a 3D DW-GRASE sequence. The Results (Comparison of GRASE and spin echo DTI; page unavailable) report no discernible geometric distortion relative to multiple spin echo imaging, but a threefold—not 12-fold—reduction in scan time. (LLM, conf=0.93)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    While temperature and fixation affect diffusivity values, and ADC in fixed tissue is significantly lower than in vivo, anisotropy values are comparable between in vivo and ex vivo samples.

    “While temperature and fixation affect diffusivity values, and ADC in fixed tissue is significantly lower than in vivo (Richardson et al., 2014), anisotropy values are comparable between in vivo and ex vivo samples (Sun et al., 2001, 2003; Zhang et al., 2012).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites Richardson et al., 2014 source

      The Introduction states that temperature differences greatly affect measured diffusion coefficients, and DTI (Experiment A) reports lower axial diffusivity in fixed than viable isolated tissue. But the study compares fixed tissue with viable isolated tissue, not directly with in vivo samples, and the cited passages do not establish a significantly lower overall ADC. The anisotropy clause is attributed to other sources and is not part of this verdict. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
    • cites Sun et al., 2001, 2003

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Zhang et al., 2012 source

      The Discussions report no significant difference between in vivo and ex vivo fractional anisotropy in the normal mouse corpus callosum. But they also report significantly different values in the demyelinated corpus callosum, so the unqualified claim that anisotropy is comparable across samples overextends the finding. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductcitation

    While it appears that we have labeled the major myelinated WM tracts of the mouse brain, there may be areas that are not fully captured due to incomplete antibody penetration or uneven labeling.

    “First, while it appears that we have labeled the major myelinated WM tracts of the mouse brain, there may be areas that are not fully captured due to incomplete antibody penetration or uneven labeling, which have been reported by other groups using intact brain approaches (Epp et al., 2015; Li et al., 2015).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites Epp et al., 2015 source

      The citing authors’ statement about which tracts they labeled is their own; the point attributed to Epp et al. is the reported risk of incomplete antibody penetration or uneven labeling in intact-brain approaches. The Materials and Methods section describes antibody incubation, but neither it nor the passages found by searching for penetration and uneven labeling report that problem (page unavailable). (LLM, conf=0.72)

      Experimental — accuracy not yet measured
    • cites Li et al., 2015

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Findingown data

    Regions where clearly diverging fascicles could be observed in the CLARITY sample had lower linear anisotropy (Cl=0.35 ± 0.024) and lower FA-MBP correlations.

    “In contrast, regions where clearly diverging fascicles could be observed in the CLARITY sample had lower linear anisotropy (Cl=0.35 ± 0.024) and also lower FA-MBP correlations (ρ=0.185, P=0.388, n=24 regions).”
    Quote in the paperSubstantial claim
    • Rests on an assumption: MBP immunofluorescence intensity is linearly proportional to myelin density across WM regions.
    • Rests on an assumption: The CLARITY clearing and immunolabeling procedures do not differentially affect MBP detection across different WM regions.
    • Rests on an assumption: The ex vivo DTI scanning and CLARITY hydrogel perfusion do not alter the relationship between MBP and DTI metrics.

  • May combine two claims.

    Findingown data

    Mean MBP in the genu and splenium were both higher than in the body of the corpus callosum.

    “Specifically, mean MBP in the genu and splenium were both higher than in the body”
    Quote in the paperSubstantial claim
    • Rests on an assumption: MBP immunofluorescence intensity is linearly proportional to myelin density across WM regions.
    • Rests on an assumption: The CLARITY clearing and immunolabeling procedures do not differentially affect MBP detection across different WM regions.

  • May combine two claims.

    Findingown data

    There were significant positive FA-MBP correlations for the genu and splenium, but not for the body of the corpus callosum.

    “we found that there were significant positive correlations for the genu (P < 0.0005) and splenium (P < 0.0005), but not for the body (P=0.138; Fig. 5D).”
    Quote in the paperSubstantial claim
    • Rests on an assumption: MBP immunofluorescence intensity is linearly proportional to myelin density across WM regions.
    • Rests on an assumption: The CLARITY clearing and immunolabeling procedures do not differentially affect MBP detection across different WM regions.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The stria medullaris is a WM tract within the thalamus that carries axons from the septum to the habenula.

    “The stria medullaris is a WM tract within the thalamus that carries axons from the septum to the habenula (Watson et al., 2012).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Watson et al., 2012 source

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    RD is considered to be sensitive to the degree of myelination and thus was expected to provide a significant negative correlation with MBP.

    “This was unexpected for the scalar metric RD, which is considered to be sensitive to the degree of myelination and thus was expected to provide a significant negative correlation (Song et al., 2002; Sun et al., 2006a, 2006b).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Song et al., 2002 source
    • cites Sun et al., 2006a, 2006b

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    DTI is the most commonly employed diffusion model and utilizes the magnitude and orientation of anisotropic water diffusion to estimate the WM structure of brain tissue, producing maps of macroscopic axonal organization.

    “Diffusion tensor imaging (DTI) is the most commonly employed diffusion model and utilizes the magnitude and orientation of anisotropic water diffusion to estimate the WM structure of brain tissue, thereby producing maps of macroscopic axonal organization in the brain (Basser and Pierpaoli, 1996; Le Bihan et al., 2001; Mori and Zhang, 2006).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Basser and Pierpaoli, 1996 source
    • cites Le Bihan et al., 2001 source
    • cites Mori and Zhang, 2006 source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Diffusion-weighted MRI is the only noninvasive method available for mapping fiber architecture of tissue in vivo.

    “Diffusion-weighted MRI is the only noninvasive method available for mapping fiber architecture of tissue in vivo (Le Bihan et al., 1986; Basser et al., 1994)”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Le Bihan et al., 1986 source
    • cites Basser et al., 1994 source

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductreasoning

    Common sources of noise for in vivo neuroimaging include head motion, cardiac pulses, and arousal, which we have excluded by imaging ex vivo.

    “Common sources of noise for in vivo neuroimaging include head motion, cardiac pulses, and arousal, which we have excluded by imaging ex vivo.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    MBP is the predominant protein within CNS myelin, representing as much as 30% of the protein in myelin, and is essential to myelin formation.

    “we chose to label MBP as a putative biological index for myelination because it is the predominant protein within CNS myelin, representing as much as 30% of the protein in myelin (Baumann and Pham-Dinh, 2001) and is essential to myelin formation (Boggs, 2006).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Baumann and Pham-Dinh, 2001 source
    • cites Boggs, 2006 source

  • May combine two claims.

    Premisecitation

    Diffusion anisotropy is present even when compact myelin is completely absent, although it is significantly reduced.

    “Indeed, diffusion anisotropy is present even when compact myelin is completely absent, although it is significantly reduced (Roach et al., 1985; Watanabe et al., 2010).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Roach et al., 1985 source
    • cites Watanabe et al., 2010 source

  • May combine two claims.

    Interpretationreasoning

    Compared to the other WM ROIs, the subset of tracts with RD-MBP association tended to have larger volumes, which may minimize a potential partial volume effect that may have confounded the RD-MBP association in smaller WM tracts.

    “Compared to the other WM ROIs, this subset tended to have larger volumes, which may minimize a potential partial volume effect that may have confounded the RD-MBP association in smaller WM tracts.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Some of the RD-myelination association evidence is based on studies involving myelin damage to the mouse optic nerve, which may differ from WM tracts within brain tissue.

    “Additionally, some of the RD-myelination association evidence is based on studies involving myelin damage to the mouse optic nerve (Song et al., 2003, 2006), which may differ from WM tracts within brain tissue.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Song et al., 2003, 2006

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • May combine two claims.

    Premisecitation

    The association of RD to myelination has been previously studied in some of these same WM structures, such as the corpus collosum and anterior commissure, but many other WM structures analyzed here have not been examined previously.

    “The association of RD to myelination has been previously studied in some of these same WM structures, such as the corpus collosum and anterior commissure (Song et al., 2002), however, many other WM structures analyzed here have not been examined previously.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Song et al., 2002 source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given

    The liquid hydrogel may have changed diffusivity during DTI in some unpredictable way, and we cannot rule out uneven labeling or altered diffusivity as possibilities.

    “Another point of consideration is that the liquid hydrogel may have changed diffusivity during DTI in some unpredictable way. While we have not seen evidence for uneven labeling or altered diffusivity in our samples, we cannot rule these out as possibilities.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    The relationship between FA and myelin is complex and cannot be resolved using this correlational approach alone.

    “The relationship between FA and myelin is complex and cannot be resolved using this correlational approach alone.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    The percentage of myelinated axons and myelin sheath thickness are not necessarily indexed by MBP expression.

    “Also, the percentage of myelinated axons and myelin sheath thickness are not necessarily indexed by MBP expression (Shine et al., 1992).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Shine et al., 1992 source
23 of 23

  • The paper relies on this without support.

    MBP immunofluorescence intensity is linearly proportional to myelin density across WM regions.

    “MBP fluorescence intensity values ranged from 0–4095 and mean intensity values within ROIs were used for comparisons.”

  • The paper relies on this without support.

    The CLARITY clearing and immunolabeling procedures do not differentially affect MBP detection across different WM regions.

    “We performed the CLARITY procedure as previously described (Chung et al., 2013; Tomer et al., 2014; Yang et al., 2014) with custom modifications in order to maximize antibody penetration and minimize tissue expansion .”

  • The paper relies on this without support.

    The ex vivo DTI scanning and CLARITY hydrogel perfusion do not alter the relationship between MBP and DTI metrics.

    “In order to test the feasibility of CLARITY following DTI, we first tested three thy1-eYFP-H mice that underwent ∼18 h of ex vivo DTI in a high-field scanner.”

  • The paper relies on this without support.

    Additional factors such as fiber architecture, axonal diameter, glial cells, and inflammation also contribute to diffusion anisotropy.

    “However, additional factors such as fiber architecture, axonal diameter, glial cells, and inflammation also contribute to diffusion anisotropy.”

  • The paper relies on this without support.

    The spatial resolution of diffusion MRI is low, typically on the order of 1–2 mm in the human brain in vivo, while myelinated axons are characterized on a micron scale.

    “The spatial resolution of diffusion MRI is low, typically on the order of 1–2 mm in the human brain in vivo, while myelinated axons are characterized on a micron scale.”

  • The paper relies on this without support.

    Diffusion anisotropy carries microscopic anatomical information, but this information is averaged over the large volume of an imaging voxel.

    “While diffusion anisotropy does carry microscopic anatomical information, this information is averaged over the large volume of an imaging voxel.”

  • The paper relies on this without support.

    None of the prior studies utilized a 3D-IHC approach for making comparisons to diffusion measures.

    “None of the prior studies utilized a 3D-IHC approach for making comparisons to diffusion measures.”

  • The paper relies on this without support.

    DTI remains a powerful method for noninvasively estimating WM structure in the brain with high sensitivity, but low specificity.

    “Nonetheless, DTI remains a powerful method for noninvasively estimating WM structure in the brain with high sensitivity, but low specificity.”

  • The paper relies on this without support.

    WM in the brain has a very complex structure that cannot be accurately estimated using a single diffusion tensor model, such as DTI.

    “WM in the brain has a very complex structure that cannot be accurately estimated using a single diffusion tensor model, such as DTI.”
9 of 9

Statistics recomputed

Every reported test result, recomputed from its test statistic and degrees of freedom. A result shows as not adding up only once an independent check has confirmed it.

  • Consistentχ²(1) = 7.74reported p < .01 · recomputed p = .005

    “Χ2(1)=7.74, p < .01”

  • Consistentχ²(1) = 0.27reported p = .61 · recomputed p = .603

    “Χ2(1)=0.27, p = .61”

  • Consistentχ²(1) = 0.05reported p = .83 · recomputed p = .823

    “Χ2(1)=0.05, p = .83”

  • Consistentχ²(1) = 12.56reported p < .0005 · recomputed p < .001

    “Χ2(1)=12.56, p < .0005”

  • Consistentχ²(1) = 10.54reported p < .005 · recomputed p = .001

    “Χ2(1)=10.54, p < .005”

  • Consistentχ²(1) = 10.62reported p < .005 · recomputed p = .001

    “Χ2(1)=10.62, p < .005”

  • Consistentχ²(1) = 0.804reported p = .37 · recomputed p = .370

    “Χ2(1)=0.804, p = .37”

7 of 7

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • 3DISCO reference has conflicting publication years

    Minor

    The text cites the Ertürk et al. 3DISCO paper as published in 2012, but its reference entry lists 1983.

1 of 1

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Related

Authors
Chang, E. H., Argyelan, M., Aggarwal, M., Chandon, T.-S. S., Karlsgodt, K. H., Mori, S., & Malhotra, A. K.
Affiliations
  • Northwell Health (US)
  • Johns Hopkins University (US)
  • Kennedy Krieger Institute (US)
Corresponding author
Chang, Eric H.
ORCID
Venue
NeuroImage
Publisher
Elsevier BV
Published
2017-02
Volume
147
Pages
253-261
ISSN
1053-8119 (ISSN-L)
DOI
10.1016/j.neuroimage.2016.11.068
PMC
PMC5560594
OpenAlex
W2563000655
Semantic Scholar
aa30e8c1e8b1d8d5eeafb64fca929a49ee935650
Cited by
186
References
66
Citations by year
2026: 16 · 2025: 27 · 2024: 13 · 2023: 14 · 2022: 17 · 2021: 27 · 2020: 19 · 2019: 17 · 2018: 21 · 2017: 13 · 2016: 1
FWCI
13.66
Type
journal article
Language
en
Open access
Licence
www.elsevier.com/tdm/userlicense/1.0/ (tdm)
Locations
  • NeuroImage (publishedVersion)
  • PubMed (publishedVersion)
  • eScholarship (California Digital Library) (submittedVersion)
  • PubMed Central (submittedVersion)
Funders
Center for Intervention Development and Applied Research (P50MH080173)
Topics
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Advanced Neuroimaging Techniques and Applications
  • Life Sciences › Neuroscience › Cognitive Neuroscience › Functional Brain Connectivity Studies
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Advanced MRI Techniques and Applications
Keywords
myelination, white matter integrity, diffusion tensor imaging, myelin basic protein, fractional anisotropy, radial diffusivity, fiber dispersion, mouse brain, ex vivo brain imaging
MeSH terms
Animals, Fluorescent Antibody Technique, Male, Mice, Inbred C57BL, Myelin Sheath, Anisotropy, Microscopy, Fluorescence, Multiphoton, Mice, Diffusion Tensor Imaging, White Matter

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