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Cited by 107Alzheimer's disease research and treatmentsNeuroinflammation and Neurodegeneration MechanismsAmyotrophic Lateral Sclerosis Research

CREB-activity and nmnat2 transcription are down-regulated prior to neurodegeneration, while NMNAT2 over-expression is neuroprotective, in a mouse model of human tauopathy

Open access
Ljungberg, M. Cecilia, Ali, Yousuf O., Zhu, Jie, Wu, Chia-Shan, Oka, Kazuhiro, Zhai, R. Grace, Lu, Hui-Chen·2012·Human Molecular Genetics
DOI 10.1093/hmg/ddr492PMC PMC3276285
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Abstract

Tauopathies, characterized by neurofibrillary tangles (NFTs) of phosphorylated tau proteins, are a group of neurodegenerative diseases, including frontotemporal dementia and both sporadic and familial Alzheimer's disease. Forebrain-specific over-expression of human tau(P301L), a mutation associated with frontotemporal dementia with parkinsonism linked to chromosome 17, in rTg4510 mice results in the formation of NFTs, learning and memory impairment and massive neuronal death. Here, we show that the mRNA and protein levels of NMNAT2 (nicotinamide mononucleotide adenylyltransferase 2), a recently identified survival factor for maintaining neuronal health in peripheral nerves, are reduced in rTg4510 mice prior to the onset of neurodegeneration or cognitive deficits. Two functional cAMP-response elements (CREs) were identified in the nmnat2 promoter region. Both the total amount of phospho-CRE binding protein (CREB) and the pCREB bound to nmnat2 CRE sites in the cortex and the hippocampus of rTg4510 mice are significantly reduced, suggesting that NMNAT2 is a direct target of CREB under physiological conditions and that tau(P301L) overexpression down-regulates CREB-mediated transcription. We found that over-expressing NMNAT2 or its homolog NMNAT1, but not NMNAT3, in rTg4510 hippocampi from 6 weeks of age using recombinant adeno-associated viral vectors significantly reduced neurodegeneration caused by tau(P301L) over-expression at 5 months of age. In summary, our studies strongly support a protective role of NMNAT2 in the mammalian central nervous system. Decreased endogenous NMNAT2 function caused by reduced CREB signaling during pathological insults may be one of underlying mechanisms for neuronal death in tauopathies.

AI snapshot

Written from the paper’s full text.

TL;DR

In rTg4510 tau P301L mice, NMNAT2 mRNA/protein and CREB activity decline in forebrain before neurodegeneration, and nmnat2 is a direct CREB target via two promoter CREs. Hippocampal overexpression of NMNAT1 or NMNAT2, but not NMNAT3, reduces tau-induced CA1 degeneration.

Key findings

  • NMNAT2 protein in rTg4510 cortex and hippocampus was often reduced to <60% of age-matched controls at 1, 2, and 7 months; nmnat2 mRNA was significantly down-regulated in cortex at 1, 2, and 7 months and in hippocampus at 2 and 7 months, with no change in cerebellum.
  • pCREB(Ser133), but not total CREB, was significantly reduced in 2-month-old rTg4510 cortex and hippocampus; pCREB/CREB ratios were ~50% of control, with no cerebellar difference. ChIP showed reduced pCREB occupancy at nmnat2 promoter regions containing CRE1+CRE2 and CRE2 alone in cortex and hippocampus, but not cerebellum.
  • Two functional CREs were identified in the nmnat2 promoter. Forskolin increased Pro-2Kb luciferase activity almost 4-fold (2.9 ± 0.57% to 11.44 ± 1.62% normalized Renilla activity; P = 0.001), and mutating either CRE abolished induction; constitutively active CREB DIEDML increased Pro-2Kb activity (18.99 ± 1.9%; P < 0.001).
  • rAAV overexpression of NMNAT1 or NMNAT2, but not NMNAT3, in rTg4510 hippocampi from 6 weeks reduced CA1 degeneration at 5 months: NMNAT1 stratum radiatum 156 ± 19% (P = 0.027) and stratum pyramidale 203 ± 36% (P = 0.029); NMNAT2 stratum radiatum 131 ± 13% (P = 0.047) and stratum pyramidale 154 ± 22% (P = 0.049). NMNAT2 also increased NF-M (207 ± 32%; P = 0.02), reduced cleaved caspase 3 (17 ± 12%; P = 0.01), GFAP (65 ± 13%; P < 0.05), CP-13/hTau (39 ± 7%; P < 0.01), and MC-1/hTau (42 ± 10%; P = 0.01).

Methods

  • Used rTg4510 mice overexpressing human tau P301L and littermate controls; analyzed cortex, hippocampus, and cerebellum at 2 weeks, 1, 2, and 7 months by western blot and real-time qPCR, with ChIP for pCREB occupancy at the nmnat2 promoter.
  • Tested nmnat2 promoter CRE function by luciferase reporter assays in 293T cells with wild-type or mutated CREs, pcDNA-CREB, dominant-negative CREB S133A, or constitutively active CREB DIEDML, with DMSO or 10 μM forskolin.
  • Injected serotype 6 rAAV carrying human NMNAT1, NMNAT2, NMNAT3, or EGFP into rTg4510 and control hippocampi at 6 weeks; assessed neuroprotection at 5 months by EGFP/synapsin immunofluorescence, CA1 layer thickness, and western blot. Statistical analysis used Student’s t-test unless otherwise indicated, with P < 0.05 considered significant.

Limitations

  • Stated: the authors note it remains to be determined whether NMNAT1 or NMNAT2 rescue of neurodegeneration can alleviate synaptic and cognitive deficits in rTg4510 mice; the mechanism of NMNAT neuroprotection is also unresolved, since NAD+ and NADH levels were unchanged and possible chaperone/clearance effects are discussed.
  • Evident: neuroprotection was assessed mainly by CA1 layer thickness and selected protein markers rather than direct stereological neuron counts or behavioral outcomes; some analyses used small samples (e.g., n = 3 per genotype/age for qPCR, n = 3 per group for ChIP, and three animals for NMNAT2 western quantification).

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Integrity signals

Registry

Crossref · 10.1093/hmg/ddr492

Cites

109 of 109 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

51 of 89 cited sources checked · 38 supported · 0 flagged · accuracy not yet measured

  • Premisecitation

    From 2.5 months onward, phosphorylated tau protein and neuronal loss can be detected in the forebrain of rTg4510 mice.

    “From 2.5 months onward, phosphorylated tau protein and neuronal loss can be detected in the forebrain of these mice (14,34,35).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [14] Spires, 2006, Region-specific dissociation of neuronal loss and neurofibrillary pathology in a mouse model of tauopathy, Am. J. Pathol. source
    • cites [34] Ramsden, 2005, Age-dependent neurofibrillary tangle formation, neuron loss, and memory impairment in a mouse model of human tauopathy (P301L), J. Neurosci. source

      The Figure 2 caption reports phosphorylated-tau labeling in hippocampal neurons at 2.5 months. The age-dependent neurodegeneration section reports marked neuronal loss in 10-month-old mice, while the Discussion cites neuron counts at 5.5 months; neither establishes detectable neuronal loss from 2.5 months onward. Page numbers are unavailable in our copy. (LLM, conf=0.89)

      Experimental — accuracy not yet measured
    • cites [35] Santacruz, 2005, Tau suppression in a neurodegenerative mouse model improves memory function, Science source
  • Findingown data

    The reduced abundance of two well-characterized CREB targets TrkB and cFos provides further confirmation that CREB-mediated transcription is reduced in rTg4510 mice.

    “The reduced abundance of two well-characterized CREB targets TrkB and cFos (52,53) provide further confirmation that CREBmediated transcription is reduced in rTg4510 mice (Supple- mentary Material, Fig. S3).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [52] Flammer, 2006, Cyclic AMP response element-binding protein (CREB) and CAAT/enhancer-binding protein beta (C/EBPbeta) bind chimeric DNA sites with high affinity, Biochemistry source
    • cites [53] Zhang, 2005, Genome-wide analysis of cAMP-response element binding protein occupancy, phosphorylation, and target gene activation in human tissues, Proc. Natl Acad. Sci. USA source

      The citation is attached to the description of TrkB and cFos as CREB targets; the reduction in rTg4510 mice is attributed to the citing paper’s supplementary figure. The Results and Promoter passages discuss CREB target genes generally, but searches for TrkB, cFos, and FOS found no passage identifying either named gene as a target. (LLM, conf=0.72)

      Experimental — accuracy not yet measured
  • Premisecitation

    In rTg4510 mice, age-dependent loss of neurons and progressive cognitive dysfunction closely mimic the clinical features of tauopathy.

    “In rTg4510 mice (34,35), the age-dependent loss of neurons and progressive cognitive dysfunction closely mimic the clinical features of tauopathy.”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [34] Ramsden, 2005, Age-dependent neurofibrillary tangle formation, neuron loss, and memory impairment in a mouse model of human tauopathy (P301L), J. Neurosci. source
    • cites [35] Santacruz, 2005, Tau suppression in a neurodegenerative mouse model improves memory function, Science source
  • Findingown data

    No difference was found in the levels of P300, cREL or NFATc in rTg4510 mice.

    “No difference was found in the levels of P300, cREL or NFATc (data not shown), transcription factors that have been implicated in different neurodegeneration models (73 -76).”
    Quote in the paperNot a substantial claim2 of 4 sources checked
    • cites [73] Martin-Loeches, 2001, P300 amplitude as a possible correlate of frontal degeneration in schizophrenia, Schizophr. Res. source
    • cites [74] Rouaux, 2003, Critical loss of CBP/p300 histone acetylase activity by caspase-6 during neurodegeneration, EMBO J. source
    • cites [75] Wu, 2010, Amyloid beta induces the morphological neurodegenerative triad of spine loss, dendritic simplification, and neuritic dystrophies through calcineurin activation, J. Neurosci. source
    • cites [76] Kogel, 2004, S100B potently activates p65/c-Rel transcriptional complexes in hippocampal neurons: Clinical implications for the role of S100B in excitotoxic brain injury, Neuroscience source
  • Premisecitation

    Mitochondria dysfunction has been demonstrated in tau P301L transgenic mice.

    “Mitochondria dysfunction has been demonstrated in tau P301L transgenic mice (92 -94)”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites [92] David, 2005, Proteomic and functional analyses reveal a mitochondrial dysfunction in P301L tau transgenic mice, J. Biol. Chem. source
    • cites [93] Rhein, 2009, Amyloid-beta and tau synergistically impair the oxidative phosphorylation system in triple transgenic Alzheimer's disease mice, Proc. Natl Acad. Sci. USA source
    • cites [94] Perreault, 2009, Increased association between rough endoplasmic reticulum membranes and mitochondria in transgenic mice that express P301L tau, J. Neuropathol. Exp. Neurol. source
  • May combine two claims. 26
  • Classified as probably not a substantial claim. 37
  • The quote only partly states this claim. 1

  • May combine two claims.

    Premisecitation

    Substantial neurodegeneration is observed after 5 months of age in rTg4510 mice, being most apparent in the hippocampal CA1 area.

    “Substantial neurodegeneration is observed after 5 months of age, being most apparent in the hippocampal CA1 area (14,34,35).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [14] Spires, 2006, Region-specific dissociation of neuronal loss and neurofibrillary pathology in a mouse model of tauopathy, Am. J. Pathol. source

      The section “rTg4510 Brain Suffers Dramatic Neuron Loss with Regional Vulnerability” reports that CA1 neuron loss begins by 5.5 months and reaches 82% by 8.5 months. But it does not identify CA1 as the most affected area: dentate gyrus loss begins earlier, at 2.5 months, and reaches 85% by 8.5 months. The supplied copy does not identify a page for these findings. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [34] Ramsden, 2005, Age-dependent neurofibrillary tangle formation, neuron loss, and memory impairment in a mouse model of human tauopathy (P301L), J. Neurosci. source
    • cites [35] Santacruz, 2005, Tau suppression in a neurodegenerative mouse model improves memory function, Science source

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

    Premisecitation

    Three mammalian homologs NMNAT1-3 have been identified, with NMNAT2 being the most labile of the three isoforms with a half-life of approximately 4 h.

    “Three mammalian homologs, NMNAT1 -3, have been identified (24)(25)(26), with NMNAT2 (nicotinamide mononucleotide adenylyltransferase 2) being the most labile of the three isoforms with a half-life of ,4 h (27).”
    Quote in the paperSubstantial claim3 of 4 sources checked
    • cites [24] Emanuelli, 2001, Molecular cloning, chromosomal localization, tissue mRNA levels, bacterial expression, and enzymatic properties of human NMN adenylyltransferase, J. Biol. Chem. source
    • cites [25] Raffaelli, 2002, Identification of a novel human nicotinamide mononucleotide adenylyltransferase, Biochem. Biophys. Res. Commun. source
    • cites [26] Zhang, 2003, Structural characterization of a human cytosolic NMN/NaMN adenylyltransferase and implication in human NAD biosynthesis, J. Biol. Chem. source
    • cites [27] Gilley, 2010, Endogenous Nmnat2 is an essential survival factor for maintenance of healthy axons, PLoS Biol. source

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

    Premisecitation

    Transgenic mice expressing human mutated tau showed NFT formation and neuronal loss, implying a causal relationship between tau over-expression and neurodegeneration.

    “transgenic mice expressing human mutated tau showed NFT formation and neuronal loss, implying a causal relationship between tau over-expression and neurodegeneration (8 -11).”
    Quote in the paperSubstantial claim1 of 4 sources checked
    • cites [8] Gotz, 2001, Tau filament formation in transgenic mice expressing P301L tau, J. Biol. Chem. source

      The Abstract reports neurofibrillary tangles in mice expressing human P301L tau, and the Results report cortical neurons undergoing apoptosis. The paper does not demonstrate neuronal loss in the mice; its Discussion mentions severe neuronal loss in human patients instead. It links the mouse pathology to the P301L mutation in combination with high expression, not to tau overexpression alone. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
    • cites [9] Lewis, 2000, Neurofibrillary tangles, amyotrophy and progressive motor disturbance in mice expressing mutant (P301L) tau protein, Nat. Genet. source
    • cites [10] Andorfer, 2003, Hyperphosphorylation and aggregation of tau in mice expressing normal human tau isoforms, J. Neurochem. source
    • cites [11] Yoshiyama, 2007, Synapse loss and microglial activation precede tangles in a P301S tauopathy mouse model, Neuron source

  • Classified as probably not a substantial claim.

    Premisecitation

    In rTg4510 mice, tau with the P301L mutation is over-expressed in the forebrain driven by the CaMKII promoter.

    “In rTg4510 mice (34,35), tau with the mis-sense mutation P301L, found in some FTDP-17 cases (36), is over-expressed in the forebrain driven by the calcium-calmodulin kinase II (CaMKII) promoter.”
    Quote in the paperSubstantial claim1 of 3 sources checked
    • cites [34] Ramsden, 2005, Age-dependent neurofibrillary tangle formation, neuron loss, and memory impairment in a mouse model of human tauopathy (P301L), J. Neurosci. source
    • cites [35] Santacruz, 2005, Tau suppression in a neurodegenerative mouse model improves memory function, Science source
    • cites [36] Hutton, 1998, Association of missense and 5′-splice-site mutations in tau with the inherited dementia FTDP-17, Nature source

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

    Premisecitation

    Neuronal loss appears to be independent of tangle formation in some transgenic mouse models.

    “However, neuronal loss appears to be independent of tangle formation in some transgenic mouse models (12)(13)(14).”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites [12] Wittmann, 2001, Tauopathy in Drosophila: neurodegeneration without neurofibrillary tangles, Science source
    • cites [13] Andorfer, 2005, Cell-cycle reentry and cell death in transgenic mice expressing nonmutant human tau isoforms, J. Neurosci. source
    • cites [14] Spires, 2006, Region-specific dissociation of neuronal loss and neurofibrillary pathology in a mouse model of tauopathy, Am. J. Pathol. source

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

    Premisecitation

    Reducing NMNAT2 levels in cultured mouse superior cervical ganglia neurons induced Wallerian-like degeneration, suggesting NMNAT2 is required to maintain axonal health in the peripheral nervous system.

    “Reducing NMNAT2 levels in cultured mouse superior cervical ganglia neurons induced Wallerian-like degeneration, suggesting that NMNAT2 is required to maintain axonal health in the peripheral nervous system (27).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [27] Gilley, 2010, Endogenous Nmnat2 is an essential survival factor for maintenance of healthy axons, PLoS Biol. source

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

    Premisecitation

    Mutant tau expression in rTg4510 mice starts around postnatal day 7 and peaks at 1 month of age.

    “Mutant tau expression starts around postnatal day 7 and peaks at 1 month of age (34,35).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [34] Ramsden, 2005, Age-dependent neurofibrillary tangle formation, neuron loss, and memory impairment in a mouse model of human tauopathy (P301L), J. Neurosci. source

      The ‘Generation of transgenic mice expressing mutant human tau P301L’ section reports that transgenic tau was first detected at postnatal day 7, supporting the onset part of the claim. Neither that passage nor the Figure 1 caption states that expression peaks at one month; the caption identifies P7 as the earliest expression shown by the postnatal forebrain immunoblot. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [35] Santacruz, 2005, Tau suppression in a neurodegenerative mouse model improves memory function, Science source

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

    Premisecitation

    Several tau mutations linked to FTDP-17 have been identified.

    “Several tau mutations linked to FTDP-17 have been identified (2 -7)”
    Quote in the paperNot a substantial claim1 of 6 sources checked
    • cites [2] Hutton, 2000, Molecular genetics of chromosome 17 tauopathies, Ann. N. Y. Acad. Sci. source
    • cites [3] Spillantini, 1998, Tau protein pathology in neurodegenerative diseases, Trends Neurosci. source
    • cites [4] Sperfeld, 1999, FTDP-17: an early-onset phenotype with parkinsonism and epileptic seizures caused by a novel mutation, Ann. Neurol. source
    • cites [5] Bugiani, 1999, Frontotemporal dementia and corticobasal degeneration in a family with a P301S mutation in tau, J. Neuropathol. Exp. Neurol. source
    • cites [6] Poorkaj, 2001, Frequency of tau gene mutations in familial and sporadic cases of non-Alzheimer dementia, Arch. Neurol. source
    • cites [7] Goedert, 2005, Mutations causing neurodegenerative tauopathies, Biochim. Biophys. Acta source

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

    Premisecitation

    Over-expression of NMNAT provides neuroprotection against several degenerative conditions in Drosophila.

    “Over-expression of NMNAT, on the other hand, provides neuroprotection against several degenerative conditions in Drosophila (21,23).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [21] Zhai, 2008, NAD synthase NMNAT acts as a chaperone to protect against neurodegeneration, Nature source
    • cites [23] Zhai, 2006, Drosophila NMNAT maintains neural integrity independent of its NAD synthesis activity, PLoS Biol. source

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

    Premisecitation

    Loss of nmnat in the Drosophila eye causes rapid neuronal degeneration.

    “Loss of nmnat in the Drosophila eye causes rapid neuronal degeneration (23).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [23] Zhai, 2006, Drosophila NMNAT maintains neural integrity independent of its NAD synthesis activity, PLoS Biol. source

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

    Premisecitation

    In mammals, over-expressing NMNATs significantly delays Wallerian degeneration of peripheral nerves.

    “In mammals, over-expressing NMNATs significantly delays Wallerian degeneration of peripheral nerves (28)(29)(30)(31)(32)(33).”
    Quote in the paperNot a substantial claim5 of 6 sources checked
    • cites [28] Press, 2008, Nmnat delays axonal degeneration caused by mitochondrial and oxidative stress, J. Neurosci. source

      The Introduction (page unavailable) reports delayed Wallerian degeneration after nerve injury in Wldˢ mutant mice and explains that the protective chimeric protein contains Nmnat1. It also says increased Nmnat expression delayed axonal degeneration in cultured dorsal root ganglion neurons. The paper does not establish that overexpressing NMNATs themselves significantly delays Wallerian degeneration of peripheral nerves in mammals. (LLM, conf=0.85)

      Experimental — accuracy not yet measured
    • cites [29] Araki, 2004, Increased nuclear NAD biosynthesis and SIRT1 activation prevent axonal degeneration, Science source
    • cites [30] Wang, 2005, A local mechanism mediates NAD-dependent protection of axon degeneration, J. Cell Biol. source

      Figure 2 in Results and discussion reports that overexpressing Nmnat1 significantly delayed Wallerian degeneration (P < 0.001). Materials and methods identifies the model as cultured rat dorsal root ganglion explants, not intact peripheral nerves. The paper tests Nmnat1 and a fusion protein containing it, rather than NMNATs generally. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [31] Sasaki, 2010, Axonal degeneration is blocked by nicotinamide mononucleotide adenylyltransferase (NMNAT) protein transduction into transected axons, J. Biol. Chem. source

      The opening section says Nmnat1 overexpression can delay axonal degeneration, and Results and Discussion reports axonal protection in transgenic mice expressing Nmnat proteins. The paper’s own experiments use cultured dorsal-root-ganglion neurons; our copy does not establish the more specific claim that overexpression significantly delays Wallerian degeneration of peripheral nerves in vivo. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
    • cites [32] Sasaki, 2009, Transgenic mice expressing the Nmnat1 protein manifest robust delay in axonal degeneration in vivo, J. Neurosci. source
    • cites [33] Yahata, 2009, Nicotinamide mononucleotide adenylyltransferase expression in mitochondrial matrix delays Wallerian degeneration, J. Neurosci. source

      In the Results section on transgenic overexpression (Figs. 2–3), NMNAT3-overexpressing mice showed a significant delay in degeneration of transected sciatic nerves. The claim generalizes this effect to NMNATs, but the same section reports no detectable delay in NMNAT1-overexpressing mice. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    Cognitive impairment in spatial reference memory and synaptic dysfunction in hippocampal CA1 pyramidal neurons begin to manifest at 4.5 months of age in rTg4510 mice.

    “Cognitive impairment in spatial reference memory and synaptic dysfunction in hippocampal CA1 pyramidal neurons begin to manifest at 4.5 months of age (37).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [37] Hoover, 2010, Tau mislocalization to dendritic spines mediates synaptic dysfunction independently of neurodegeneration, Neuron source

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

    Premisecitation

    NAD+ is used to synthesize NADP+, whose reduced form NADPH is involved in defense against oxidative stress.

    “NAD + is used to synthesize NADP + , whose reduced form NADPH is involved in defense against oxidative stress (91).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [91] Pollak, 2007, The power to reduce: pyridine nucleotides—small molecules with a multitude of functions, Biochem. J. source

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

    Premisecitation

    The protective effects in WldS mice are attributed to a chimeric gene Ube4b/NMNAT1, which encodes an in-frame fusion protein of the amino-terminal of UBE4B and the complete coding region of NMNAT1.

    “The protective effects occurring in these mice are attributed to a chimeric gene Ube4b/NMNAT1, which encodes an in-frame fusion protein of the amino-terminal of ubiquitination factor E4B (UBE4B) and the complete coding region of NMNAT1 (83,84).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [83] Conforti, 2000, A Ufd2/D4Cole1e chimeric protein and overexpression of Rbp7 in the slow Wallerian degeneration (WldS) mouse, Proc. Natl Acad. Sci. USA source

      The Results identify a WldS-specific in-frame fusion of the N-terminal 70 amino acids of Ufd2 and the C-terminal 302 amino acids of D4Cole1e. The Abstract and Discussion call it a promising candidate for slowing Wallerian degeneration, not an established cause of protection. Our copy does not identify D4Cole1e as NMNAT1 or say that the fusion contains NMNAT1’s complete coding region. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [84] Mack, 2001, Wallerian degeneration of injured axons and synapses is delayed by a Ube4b/Nmnat chimeric gene, Nat. Neurosci. source

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

    Premisecitation

    NMNATs are essential enzymes for NAD biosynthesis and several studies have found the enzymatic activity to be important for their neuroprotective effect.

    “NMNATs are essential enzymes for NAD biosynthesis and several studies have found the enzymatic activity to be important for their neuroprotective effect (reviewed in 81,89).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [81] Coleman, 2010, Wallerian degeneration, wld(s), and nmnat, Annu. Rev. Neurosci. source
    • cites [89] Zhai, 2009, Nicotinamide/nicotinic acid mononucleotide adenylyltransferase, new insights into an ancient enzyme, Cell Mol. Life Sci. source

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

    Premisecitation

    Ab oligomers cause tau mislocalization.

    “Ab oligomers also cause tau mislocalization (77,78).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [77] Ittner, 2010, Dendritic function of tau mediates amyloid-beta toxicity in Alzheimer's disease mouse models, Cell source
    • cites [78] Zempel, 2010, Abeta oligomers cause localized Ca(2+) elevation, missorting of endogenous Tau into dendrites, Tau phosphorylation, and destruction of microtubules and spines, J. Neurosci. source

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

    Premisecitation

    NMDAR-mediated calcium influx increases pCREB levels through several signaling pathways.

    “NMDAR (N-methyl-D-aspartate receptors)-mediated calcium influx is known to increase pCREB levels through several signaling pathways (65).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [65] Lonze, 2002, Function and regulation of CREB family transcription factors in the nervous system, Neuron source

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

    Premisecitation

    Genetic manipulations that decrease CREB-mediated transcription lead to deficits in both synaptic plasticity and long-term memory, whereas overexpression of CREB results in memory enhancements.

    “Genetic manipulations that decrease CREB-mediated transcription lead to deficits in both synaptic plasticity and long-term memory, whereas overexpression of CREB results in memory enhancements (38).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [38] Lee, 2009, The molecular and cellular biology of enhanced cognition, Nat. Rev. Neurosci. source

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

    Premisecitation

    Removing the creb1 gene in adult mouse forebrain or chronic inhibition of CREB function leads to extensive neurodegeneration in the hippocampus.

    “Removing the creb1 gene in adult mouse forebrain or chronic inhibition of CREB function leads to extensive neurodegeneration in the hippocampus (67,68).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [67] Mantamadiotis, 2002, Disruption of CREB function in brain leads to neurodegeneration, Nat. Genet. source
    • cites [68] Valor, 2010, Ultrastructural and transcriptional profiling of neuropathological misregulation of CREB function, Cell Death Differ. source

      The section “Chronic inhibition of CREB function causes dark neuron degeneration” reports CA1 neuron loss in mice expressing the CREB inhibitor A-CREB, supporting the chronic-inhibition part of the sentence. The passages do not report removal of the creb1 gene in adult mouse forebrain, so this source does not support that part. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

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

    Premisecitation

    In the CNS, WldS protects nigrostriatal axons from degeneration in a model of Parkinson's disease, reduces the number of axonal spheroids in mice with gracile axonal dystrophy, and protects axons from neuronal degeneration induced by toxic insults.

    “In the CNS, Wld S has been shown to protect nigrostriatal axons from degeneration in a model of Parkinson's disease, to reduce the number of axonal spheroids in mice with gracile axonal dystrophy, and to protect axons from neuronal degeneration induced by toxic insults (85)(86)(87).”
    Quote in the paperNot a substantial claim1 of 3 sources checked
    • cites [85] Gillingwater, 2004, Neuroprotection after transient global cerebral ischemia in Wld(s) mutant mice, J. Cereb. Blood Flow Metab. source
    • cites [86] Sajadi, 2004, Wlds-mediated protection of dopaminergic fibers in an animal model of Parkinson disease, Curr. Biol. source
    • cites [87] Mi, 2005, The slow Wallerian degeneration gene, WldS, inhibits axonal spheroid pathology in gracile axonal dystrophy mice, Brain source

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

    Premisecitation

    CREB is important for cell survival.

    “CREB is also important for cell survival (65,66).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [65] Lonze, 2002, Function and regulation of CREB family transcription factors in the nervous system, Neuron source
    • cites [66] Bito, 2003, Ca(2+)/CREB/CBP-dependent gene regulation: a shared mechanism critical in long-term synaptic plasticity and neuronal survival, Cell Calcium source

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

    Premisecitation

    NMNAT over-expression can prevent axonal damage resulting from exogenous application of oxidants.

    “Interestingly, NMNAT over-expression can prevent axonal damage resulting from exogenous application of oxidants (28).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Press, 2008, Nmnat delays axonal degeneration caused by mitochondrial and oxidative stress, J. Neurosci. source

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

    Premisecitation

    The role of NMNATs in neuroprotection was first implicated in studies using Wallerian Slow mice (WldS).

    “The role of NMNATs in neuroprotection was first implicated in studies using Wallerian Slow mice (Wld S ; reviewed in 81).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [81] Coleman, 2010, Wallerian degeneration, wld(s), and nmnat, Annu. Rev. Neurosci. source

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

    Premisecitation

    NMNAT3-transgenic mice show delayed Wallerian degeneration similar to WldS mice.

    “NMNAT3-transgenic mice show delayed Wallerian degeneration similar to Wld S mice (33).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [33] Yahata, 2009, Nicotinamide mononucleotide adenylyltransferase expression in mitochondrial matrix delays Wallerian degeneration, J. Neurosci. source

  • Classified as probably not a substantial claim.

    Premisecitation

    rTg4510 neurons have fewer NMDARs on their dendritic spines due to the mislocalization of tau protein from axons to dendritic spines.

    “A recent study by Hoover et al. (37) found that rTg4510 neurons have fewer NMDARs on their dendritic spines due to the mislocalization of tau protein from axons to dendritic spines.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [37] Hoover, 2010, Tau mislocalization to dendritic spines mediates synaptic dysfunction independently of neurodegeneration, Neuron source

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

    Premisecitation

    The differential subcellular distribution of NMNAT1, NMNAT2 and NMNAT3 is consistent with previous observations that found NMNAT1 in the nucleus, NMNAT2 in the Golgi complex and neurites and NMNAT3 in the mitochondria.

    “This differential subcellular distribution is consistent with previous observations that found NMNAT1 in the nucleus, NMNAT2 in the Golgi complex and neurites and NMNAT3 in the mitochondria (25,26,55,56).”
    Quote in the paperNot a substantial claim1 of 3 sources checked
    • cites [25] Raffaelli, 2002, Identification of a novel human nicotinamide mononucleotide adenylyltransferase, Biochem. Biophys. Res. Commun. source
    • cites [26] Zhang, 2003, Structural characterization of a human cytosolic NMN/NaMN adenylyltransferase and implication in human NAD biosynthesis, J. Biol. Chem. source

      In Results and Discussion (page not supplied), the paper reports nuclear hsPNAT-1 and mitochondrial hsPNAT-3. It does not place hsPNAT-2 in the Golgi complex or neurites; it says its exact cellular location remains to be determined. (LLM, conf=0.91)

      Experimental — accuracy not yet measured
    • cites [55] Berger, 2005, Subcellular compartmentation and differential catalytic properties of the three human nicotinamide mononucleotide adenylyltransferase isoforms, J. Biol. Chem. source

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

    Premisecitation

    It has been proposed that WldS or NMNATs protect neurons at a step common to many degenerative processes.

    “Together with the broad protective effects of NMNAT found in Drosophila (21,23,88), it has been proposed that Wld S or NMNATs protect neurons at a step common to many degenerative processes.”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites [21] Zhai, 2008, NAD synthase NMNAT acts as a chaperone to protect against neurodegeneration, Nature source
    • cites [23] Zhai, 2006, Drosophila NMNAT maintains neural integrity independent of its NAD synthesis activity, PLoS Biol. source
    • cites [88] MacDonald, 2006, The Drosophila cell corpse engulfment receptor Draper mediates glial clearance of severed axons, Neuron source

      The Results report that dNmnat protects severed Drosophila axons from Wallerian degeneration, and the Discussion identifies Nmnat activity as part of WldS neuroprotection. The paper does not propose that WldS or NMNATs act at a step common to many different degenerative processes; its findings here concern injury-induced axon degeneration. Page numbers are unavailable in our copy. (LLM, conf=0.84)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The ratios of pCREB (Ser133) to CREB in the cortex or hippocampus of rTg4510 mice were less than 50% of control values, while no difference was found in the cerebellum.

    “The ratios of pCREB (Ser133) to CREB in the cortex or hippocampus of rTg4510 mice were ,50% of control values, while no difference was found in the cerebellum (Fig. 2B).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The initial reduction in nmnat2 expression coincided with peak tau P301L expression and remained low throughout the life.

    “The initial reduction in nmnat2 expression coincided with peak tau P301L expression and remained low throughout the life.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    rAAVs generated in this study support efficient and long-lasting expression of NMNAT and EGFP in neurons located near the injection site.

    “Clearly, the rAAVs generated here support efficient and longlasting expression of NMNAT and EGFP in neurons located near the injection site.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Analysis in 293T cells found pCREB binding to these regions and an increase in the amount of pCREB bound to the specific CRE-containing regions upon forskolin treatment.

    “Analysis in 293T cells also found pCREB binding to these regions and an increase in the amount of pCREB bound to the specific CRE-containing regions upon forskolin treatment (Fig. 3B).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In mice with NMNAT3 over-expression, the widths of the stratum radiatum and stratum pyramidale were similar to the EGFP expressing side (stratum radiatum, 89 + 8%, P = 0.657; stratum pyramidale, 98 + 12%, P = 0.883).

    “However, in mice with NMNAT3 over-expression, the widths of the stratum radiatum and stratum pyramidale were similar to the EGFP expressing side (stratum radiatum, 89 + 8%, P ¼ 0.657; stratum pyramidale, 98 + 12%, P ¼ 0.883).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both the CREs are required for nmnat2 transcription as mutating either one abolishes transcription of nmnat2, suggesting transcriptional regulation in a cooperative manner.

    “Furthermore, our luciferase studies indicated that both the CREs are required for nmnat2 transcription as mutating either one abolishes transcription of nmnat2, suggesting transcriptional regulation in a cooperative manner.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    EGFP was expressed at high levels in dendrites, and transgene expression had no detectable impact on the hippocampal structure.

    “EGFP was expressed at high levels in dendrites, and transgene expression had no detectable impact on the hippocampal structure (Fig. 6A).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Reduced levels of activated cleaved caspase 3 (ratios of activated cleaved caspase 3 to un-cleaved caspase 3: ctrl, 100 + 26%; NMNAT2, 17 + 12%; P = 0.01) and GFAP (ctrl, 100 + 10%; NMNAT2, 65 + 13%; P < 0.05) were observed in the NMNAT2-injected rTg4510 hippocampi compared with the EGFP injected rTg4510 hippocampi.

    “Interestingly, we also observed reduced levels of activated cleaved caspase 3 (Fig. 7A and D; ratios of activated cleaved caspase 3 to un-cleaved caspase 3: ctrl, 100 + 26%; NMNAT2, 17 + 12%; P ¼ 0.01) and GFAP (Fig. 7A and E; ctrl, 100 + 10%; NMNAT2, 65 + 13%; P , 0.05) in the NMNAT2-injected rTg4510 hippocampi compared with the EGFP injected rTg4510 hippocampi.”
    Quote in the paperSubstantial claim

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

    Findingown data

    Ectopic expression of EGFP or NMNAT1 is strong at 5 months of age in the CA1 area of Tau mice injected with rAAV.

    “Dual EGFP (green) and NMNAT1 (N1, red) immunostaining shows strong ectopic expression of EGFP or NMNAT1 at 5 months of age (C).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    pCREB occupancy was significantly reduced at both the promoter with CRE1 + CRE2 and the one with CRE2 alone in the rTg4510 cortex (P < 0.05 for CRE1 + CRE2; P < 0.001 for CRE2) and hippocampus (P < 0.001 for CRE1 + CRE2 and P < 0.05 for CRE2).

    “We found that pCREB occupancy was significantly reduced at both the promoter with CRE1 + CRE2 and the one with CRE2 alone in the rTg4510 cortex (P , 0.05 for CRE1 + CRE2; P , 0.001 for CRE2) and hippocampus (P , 0.001 for CRE1 + CRE2 and P , 0.05 for CRE2) (Fig. 4; Supplementary Ma- terial, Table S3).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In cultured cortical neurons, NMNAT1 immunoreactivity was found in the nucleus, NMNAT2 immunoreactivity was found in neurites but not in the nucleus, and NMNAT3 was mainly found in the cytoplasm.

    “As expected, NMNAT1 immunoreactivity was found in the nucleus (Fig. 5E), while NMNAT2 immunoreactivity was found in neurites (Fig. 5F), but not in the nucleus.NMNAT3 was mainly found in the cytoplasm (Fig. 5G).”
    Quote in the paperNot a substantial claim

  • The quote only partly states this claim.

    Findingown data

    NMNAT3 over-expression did not have an impact on hippocampal size in rTg4510 mice.

    “NMNAT3 over-expression did not have an impact on hippocampal size.”
    Quote in the paperNot a substantial claim

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

    Findingown data

    Ectopic NMNAT1-3 expression has a distinct subcellular distribution in cultured cortical neurons.

    “The distinct subcellular distribution of ectopic NMNAT1-3 expression in cultured cortical neurons.”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    Several gene-array studies found that NMNAT2 levels were reduced in brain specimens from patients with Alzheimer's disease.

    “Interestingly, several gene-array studies found that NMNAT2 levels were reduced in brain specimens from patients with Alzheimer's disease (AD; https://www.nextbio.com).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites https://www.nextbio.com

      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).

    Premisecitation

    Tauopathies include several neurodegenerative diseases such as Pick's disease and frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17).

    “Tauopathies include several neurodegenerative diseases such as Pick's disease and frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17; reviewed in 1).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1] Lee, 2001, Neurodegenerative tauopathies, Annu. Rev. Neurosci. source

  • May combine two claims.

    Interpretationown data

    The decrease in cortical NMNAT2 coincides with tau P301L over-expression and precedes significant neuronal loss.

    “Thus, the decrease in cortical NMNAT2 coincides with tau P301L over-expression and precedes significant neuronal loss.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Immunoprecipitation was performed with anti-pCREB (Ser133), Rabbit IgG as negative control, and anti-RNAPolII as positive control.

    “Immunoprecipitation was performed with the following antibodies: anti-pCREB (Ser133) (Millipore), Rabbit IgG (negative control; Millipore) or anti-RNAPolII (positive control; Millipore).”
    Quote in the paperSubstantial claim

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

    Study design and conductnone given

    Only brains with similar amounts of EGFP in the CA1 area of hippocampus in both hemispheres were chosen for further immunofluorescence analysis.

    “Only the brains with similar amounts of EGFP in the CA1 area of hippocampus in both hemispheres were chosen for further immunofluorescence analysis.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    A housekeeping gene, gapdh, was used as an internal control to normalize mRNA expression.

    “A housekeeping gene, gapdh, was used as an internal control to normalize mRNA expression.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    nmnat2 is likely to be a CREB target necessary for maintaining neuronal health at physiological conditions.

    “nmnat2 is likely to be a CREB target necessary for maintaining neuronal health at physiological conditions.”
    Quote in the paperSubstantial claim

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

    Premisecitation

    In WldS mice, the Wallerian degeneration process is significantly delayed.

    “In this spontaneous mutant, the Wallerian degeneration process is significantly delayed (82).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [82] Lunn, 1989, Absence of Wallerian degeneration does not hinder regeneration in peripheral nerve, Eur. J. Neurosci. source

  • May combine two claims.

    Interpretationown data

    Our studies demonstrate down-regulation of NMNAT2 and CREB activity in the central nervous system of a tauopathy animal model.

    “Taken together, our studies demonstrate down-regulation of NMNAT2 and CREB activity in the central nervous system (CNS) of a tauopathy animal model.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Overexpression of NMNAT2 protects against tau-induced degeneration by reducing the toxic hyperphosphorylated tau load and ameliorates tau-induced cytotoxicity.

    “These results suggest that overexpression of NMNAT2 protects against tau-induced degeneration by reducing the toxic hyperphosphorylated tau load and ameliorates tau-induced cytotoxicity.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: The EGFP-rAAV-injected hemisphere provides a valid internal control for the effects of NMNAT-rAAV in rTg4510 mice.

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

    Study design and conductnone given

    ChIP experiments were replicated three times incorporating all the appropriate controls.

    “ChIP experiments were replicated three times incorporating all the appropriate controls.”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    Various transgenic and pharmacological approaches that augment NMDAR signaling increase learning and memory in mice.

    “Various transgenic and pharmacological approaches that augment NMDAR signaling have been found to increase learning and memory in mice, for example, NR2B overexpression and application of NMDAR partial agonist, D-cycloserine (79,80).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [79] Thompson, 1992, Hippocampus-dependent learning facilitated by a monoclonal antibody or D-cycloserine, Nature source
    • cites [80] Flood, 1992, Effect on memory processing by D-cycloserine, an agonist of the NMDA/glycine receptor, Eur. J. Pharmacol. source

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

    Premisecitation

    CREB binds to CREs in the promoter region of its target genes, but only activates gene transcription upon phosphorylation at Ser133.

    “CREB binds to CREs in the promoter region of their target genes (69), but will only activate gene transcription upon phosphorylation at Ser133 (40).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [69] Impey, 2004, Defining the CREB regulon: a genome-wide analysis of transcription factor regulatory regions, Cell

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

    • cites [40] Mayr, 2001, Transcriptional regulation by the phosphorylation-dependent factor CREB, Nat. Rev. Mol. Cell Biol. source

  • Classified as probably not a substantial claim.

    Study design and conductown data

    The extensive EGFP-positive area in the CA1 region indicates successful rAAV injections.

    “The extensive EGFP-positive area in the CA1 region indicates successful rAAV injections.”
    Quote in the paperSubstantial claim
54 of 54

  • The paper relies on this without support.

    The precise mechanism of cell death in tauopathies remains unknown.

    “Although there is evidence for the presence of activated caspase and morphological features of necrosis, autophagy and abnormal cell cycle events (10,13,15 -18), the precise mechanism of cell death in tauopathies remains unknown.”

  • The paper relies on this without support.

    The EGFP-rAAV-injected hemisphere provides a valid internal control for the effects of NMNAT-rAAV in rTg4510 mice.

    “NMNAT1-, NMNAT2-or NMNAT3-rAAV was injected into the CA1/ Thicknesses were analyzed by taking six measurements per section and three sections per animal.The three 50 mm sections were separated by dentate gyrus area of the hippocampus in the right hemisphere, while EGFP-rAAV was injected into the left hemisphere as an internal control.”

  • The paper relies on this without support.

    The reduction in NMNAT2 protein levels in rTg4510 mice is primarily due to reduced nmnat2 transcription rather than altered protein stability.

    “Thus, the decrease in the NMNAT2 protein in rTg4510 mice was accompanied by a down-regulation of nmnat2 transcripts in the cortex at 1 month of age and older, and in the hippocampus at 2 months of age and older.”

  • The paper relies on this without support.

    The thickness of the stratum radiatum and stratum pyramidale reflects hippocampal neuronal loss in rTg4510 mice.

    “To quantify the impact of NMNAT over-expression on CA1 structures, we measured the thickness of the stratum radiatum and the stratum pyramidale guided by synapsin immunoreactivity and DAPI staining (see Materials and Methods).”

  • The paper relies on this without support.

    NMDAR-dependent long-term potentiation is impaired in the Schaffer collateral pathway of rTg4510 mice at 4.5 months of age.

    “NMDAR-dependent long-term potentiation was impaired in the Schaffer collateral pathway of rTg4510 mice at 4.5 months of age.”

  • The paper relies on this without support.

    Over-expression of NMNAT substantially reduces the formation of protein aggregates such as hAtx-1[82Q] protein in Drosophila.

    “Over-expression of NMNAT substantially reduced the formation of protein aggregates such as hAtx-1[82Q] protein, the cause of spinocerebellar ataxia 1, in Drosophila.”

  • The paper relies on this without support.

    Synapsin immunoreactivity is a valid estimate of neuronal morphology and anatomical integrity in rTg4510 mice.

    “In this study, we used the immunoreactivity of synapsin, a synaptic marker, to estimate neuronal morphology and anatomical integrity in rTg4510 mice.”
7 of 7

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • The Benjamini-Hochberg procedure was applied to control the false discovery rate at 0.1.

    “To control false discovery rate, Benjamini-Hochberg procedure was applied with a false discovery rate of 0.1 35, 36”

    From Amyloid-beta induced retrograde axonal degeneration in a mouse tauopathy model (2019) · cites this paper as [36]

    The Protein extraction and western blotting section (page unavailable) says statistical analysis used Student’s t-test unless otherwise indicated and treated P < 0.05 as significant. Searches for Benjamini–Hochberg, false discovery rate, and 0.1 found no passage stating that the procedure or threshold was used; the reported t-test does not itself rule out a separate correction. (LLM, conf=0.76)

    Experimental — accuracy not yet measured

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.

  • One-month western-blot sample size differs between methods and figure

    Major

    The methods specify three mice per genotype and age for protein analysis, but the Figure 1 western-blot caption specifies ten per genotype at one month.

  • Cultured-neuron staining is reported at both 6 and 8 DIV

    Minor

    The results say NMNAT expression was examined at 6 DIV, whereas the Figure 5 caption says the neurons were stained at 8 DIV. The methods also describe transduction at 2 DIV followed by another 6 days of growth.

  • Seven-month samples are reported but absent from the stated collection ages

    Minor

    The tissue-processing methods list only 2 weeks, 1 month and 2 months as collection ages for qPCR and protein analysis, while the results report both analyses at 7 months.

3 of 3

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Related

Authors
Ljungberg, M. C., Ali, Y. O., Zhu, J., Wu, C.-S., Oka, K., Zhai, R. G., & Lu, H.-C.
Affiliations
  • Baylor College of Medicine (US)
  • University of Miami (US)
  • Neurological Research Institute (US)
Corresponding author
Lu, Hui-Chen
ORCID
Venue
Human Molecular Genetics
Publisher
Oxford University Press (OUP)
Published
2011-10-25
Volume
21
Issue
2
Pages
251-267
ISSN
  • 0964-6906 (ISSN-L)
  • 1460-2083
DOI
10.1093/hmg/ddr492
PMC
PMC3276285
OpenAlex
W2127395697
Cited by
107
References
101
Citations by year
2026: 5 · 2025: 4 · 2024: 3 · 2023: 4 · 2022: 9 · 2021: 9 · 2020: 6 · 2019: 13 · 2018: 12 · 2017: 11 · 2016: 6 · 2015: 1 · 2014: 8 · 2013: 11 · 2012: 5
FWCI
5.22
Type
journal article
Language
en
Open access
Locations
  • Human Molecular Genetics (publishedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • American Heart Association
  • National Heart, Lung, and Blood Institute (HL73144, R01HL073144)
  • National Institute on Drug Abuse (R21DA029381, DA029381)
  • National Institute of Diabetes and Digestive and Kidney Diseases (DK-79638, P30DK079638)
  • National Institute of Neurological Disorders and Stroke (R01NS048884, R01NS064269, R56NS064269, NS048884, NS64269)
  • Eunice Kennedy Shriver National Institute of Child Health and Human Development (HD065561, R21HD065561)
Topics
  • Health Sciences › Medicine › Physiology › Alzheimer's disease research and treatments
  • Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
  • Health Sciences › Medicine › Neurology › Amyotrophic Lateral Sclerosis Research
Keywords
tauopathy, NMNAT2, CREB, cAMP response element, P301L tau, rTg4510 mice, neurodegeneration, neuroprotection, neurofibrillary tangles, frontotemporal dementia, Alzheimer's disease, hippocampus, adeno-associated virus vectors, neuronal survival
MeSH terms
Animals, Base Sequence, Disease Models, Animal, Fluorescent Antibody Technique, Hippocampus, Humans, Mice, Transgenic, Nicotinamide-Nucleotide Adenylyltransferase, Promoter Regions, Genetic, Transcription, Genetic, Blotting, Western, Down-Regulation, DNA Primers, Tauopathies, CREB-Binding Protein, Mice, Real-Time Polymerase Chain Reaction

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-30.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-30.

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