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Cited by 3Alzheimer's disease research and treatmentsDementia and Cognitive Impairment ResearchNeuroinflammation and Neurodegeneration Mechanisms

Tuberous sclerosis complex‐1 (TSC1) contributes to selective neuronal vulnerability in Alzheimer's disease

Open access
Adriaanse, Bryan A., Brady, Sinead, Wang, Minghui, Beard, Daniel J., Spencer, Jonathan I., Pansieri, Jonathan, Sutherland, Brad A., Esiri, Margaret M. +4 more·2023·Neuropathology and Applied Neurobiology
DOI 10.1111/nan.12904
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Abstract

Aims: Selective neuronal vulnerability of hippocampal Cornu Ammonis (CA)‐1 neurons is a pathological hallmark of Alzheimer's disease (AD) with an unknown underlying mechanism. We interrogated the expression of tuberous sclerosis complex‐1 (TSC1; hamartin) and mTOR‐related proteins in hippocampal CA1 and CA3 subfields. Methods: A human post‐mortem cohort of mild ( n = 7) and severe ( n = 10) AD and non‐neurological controls ( n = 9) was used for quantitative and semi‐quantitative analyses. We also developed an in vitro TSC1 knockdown model in rat hippocampal neurons, and transcriptomic analyses of TSC1 knockdown neuronal cultures were performed. Results: We found a selective increase of TSC1 cytoplasmic inclusions in human AD CA1 neurons with hyperactivation of one of TSC1's downstream targets, the mammalian target of rapamycin complex‐1 (mTORC1), suggesting that TSC1 is no longer active in AD. TSC1 knockdown experiments showed accelerated cell death independent of amyloid‐beta toxicity. Transcriptomic analyses of TSC1 knockdown neuronal cultures revealed signatures that were significantly enriched for AD‐related pathways. Conclusions: Our combined data point to TSC1 dysregulation as a key driver of selective neuronal vulnerability in the AD hippocampus. Future work aimed at identifying targets amenable to therapeutic manipulation is urgently needed to halt selective neurodegeneration, and by extension, debilitating cognitive impairment characteristic of AD.

AI snapshot

Written from the paper’s full text.

TL;DR

TSC1 forms cytoplasmic inclusions selectively in vulnerable CA1 hippocampal neurons in Alzheimer’s disease, where they correlate with mTOR activation, neuronal loss, and cognitive decline. In vitro TSC1 knockdown causes mTORC1-independent cell death and transcriptomic changes overlapping human AD signatures, implicating TSC1 loss-of-function in selective CA1 vulnerability.

Key findings

  • In human post-mortem hippocampus, TSC1 inclusions were increased in CA1 in severe AD compared with CA3 and controls (severe AD, p = 0.004), and phos-mTOR was increased in severe AD vs control only in CA1 (*p < 0.05).
  • CA1 TSC1 inclusions negatively correlated with neuronal density (r = −0.59, p = 0.0030) and MMSE (r = −0.62, p = 0.0045), and positively correlated with phos-mTOR (r = 0.61, p = 0.0025); in multiple linear regression, TSC1 inclusions were the only significant predictor of MMSE (F[3,15] = 8.927, p = 0.0012, adjusted R² = 0.5692; TSC1 p = 0.0097).
  • TSC1 knockdown in rat hippocampal neurons increased cell death and caspase-3 activation (caspase-3 at 48 h: 32.9 ± 5.9% vs 19.1 ± 5.2%, p = 0.0001); rapamycin did not reduce TSC1 knockdown-related cell death (21.6 ± 4.2% vs 24.3 ± 3.1%, p = 0.92).
  • RNA-seq of TSC1 knockdown cultures detected close to 4000 DEGs at each time point, with significant overlap between up-regulated (4.2-fold enrichment) and down-regulated (4.5-fold enrichment) signatures; five AD GWAS risk genes were modulated, and signatures overlapped human AD CA1/CA3 signatures up to 2.4-fold (FET adjusted p up to 5.0E-33).

Methods

  • Human post-mortem cohort from OPTIMA/Oxford Brain Bank: 10 severe AD, 7 mild AD, and 9 non-neurological controls, age- and sex-matched; formalin-fixed paraffin-embedded hippocampal tissue containing CA1 and CA3 was assessed by quantitative and semi-quantitative immunohistochemistry, with blinded measurements and non-parametric statistics plus multiple linear regression.
  • In vitro model used rat hippocampal primary cultures transduced with TSC1 shRNA or control, with hTSC1 overexpression rescue, Aβ 1-42, and rapamycin; cell death and caspase-3 activation were monitored by live-cell imaging, and Western blot assessed mTOR activation, tau phosphorylation, and TSC1 expression.
  • Transcriptomic analysis used RNA-seq of wild-type and TSC1 knockdown rat hippocampal neurons treated with Aβ or vehicle, aligned to Rat Rn6, analysed with Limma, and tested for enrichment against MSigDB pathways, human AD DEGs, and human AD co-expression modules.

Limitations

  • Stated by the paper: incomplete rescue of the TSC1 knockdown phenotype by overexpression and rapamycin raises the possibility of off-target effects; rapamycin may act on mTORC2, shRNA may affect other genes, irreversible cell death cascades may be triggered, and other TSC1 downstream targets such as Rheb may be involved.
  • Stated by the paper: it is unclear what role TSC1 loss-of-function plays outside the hippocampus or in neurological diseases other than AD; Aβ treatment had no additional effect in knockdown cultures, which was unexpected.
  • Evident from the text: the human cohort is small (n = 26; severe AD n = 10, mild AD n = 7, controls n = 9) and cross-sectional post-mortem, so causal direction cannot be established; the in vitro and transcriptomic findings are model-based and associative rather than direct proof of mechanism in human AD.

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

Integrity signals

Registry

Crossref · 10.1111/nan.12904

Cites

50 of 50 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 44 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    TSC1 knockdown accelerates cell death in rat hippocampal neurons.

    “TSC1 knockdown experiments showed accelerated cell death independent of amyloid-beta toxicity.”
    • Also stated in: § TSC1 knockdown in rat hippocampal neurons results in increased cell death · plain
      “TSC1 knockdown led to increased cell death throughout the culture period”
    Quote in the paperSubstantial claim
  • Findingown data

    TSC1 knockdown-induced cell death in rat hippocampal neurons is independent of amyloid-beta toxicity.

    “TSC1 knockdown experiments showed accelerated cell death independent of amyloid-beta toxicity.”
    • Also stated in: § TSC1 knockdown in rat hippocampal neurons results in increased cell death · plain
      “Unexpectedly, treatment with Aβ 1-42 in TSC1 knockdown or hTSC1 rescue experiments did not have additional effects on mTOR activation (Figures S11-S14), Tau burden (Figures S16 and S17), total cell death or caspase-3 activation (Figure S18).”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Aβ 1-42 at 5 μM is toxic to rat hippocampal neurons in this model.
  • Findingown data

    In human AD, TSC1 cytoplasmic inclusions are selectively increased in CA1 hippocampal neurons.

    “We found a selective increase of TSC1 cytoplasmic inclusions in human AD CA1 neurons”
    • Also stated in: § TSC1 cytoplasmic inclusions and phos-mTOR expression are selectively increased in CA1 hippocampal neurons in human AD · plain
      “A selective increase in the number of hippocampal neurons with TSC1 inclusions in the CA1 subfield was pronounced in severe AD”
    Quote in the paperSubstantial claim
  • Findingown data

    In human AD, mTORC1 is hyperactivated, as indicated by increased phos-mTOR, in CA1 hippocampal neurons.

    “with hyperactivation of one of TSC1's downstream targets, the mammalian target of rapamycin complex-1 (mTORC1)”
    • Also stated in: § TSC1 cytoplasmic inclusions and phos-mTOR expression are selectively increased in CA1 hippocampal neurons in human AD · plain
      “the intensity and proportion of neurons expressing phos-mTOR in CA1 hippocampal neurons were increased in severe AD compared with controls”
    Quote in the paperSubstantial claim
  • Findingown data

    Transcriptomic signatures of TSC1 knockdown neuronal cultures are significantly enriched for AD-related pathways.

    “Transcriptomic analyses of TSC1 knockdown neuronal cultures revealed signatures that were significantly enriched for AD-related pathways.”
    • Also stated in: § Transcriptomic analysis of TSC1 knockdown in cultured rat hippocampal neurons shows similarities to human AD · plain
      “The TSC1 DEG signatures significantly overlap previously published transcriptomic signatures of AD in different human brain regions [24][25][26][27][28][29][30][31][32][33] (Figure 4C).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [24]

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

    • cites [25]

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

    • cites [26]

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

  • May combine two claims. 39
  • The quote only partly states this claim. 2
  • Classified as probably not a substantial claim. 21

  • May combine two claims.

    Findingown data

    In AD, TSC1 aggregates are found in CA1 through CA4 hippocampal subfields and in the subiculum, but not in the dentate gyrus.

    “In AD, we found TSC1 aggregates in the CA1 through CA4 hippocampal subfields and in the subiculum with an absence of TSC1 aggregates noted in the dentate gyrus (Figure S2).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Five known AD GWAS risk genes are modulated by TSC1 knockdown, including down-regulated CELF1 and FERMT2 and up-regulated ETFA, PLD3 and PSEN2.

    “Five known AD GWAS risk genes were modulated by TSC1 knockdown (Figure 4B), including down-regulated genes CELF1 and FERMT2 and up-regulated genes ETFA, PLD3 and PSEN2”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    TSC1 inclusions co-localise to separate compartments with granules resembling those found in granulovacuolar degeneration and frustrated autophagy, the latter being most prevalent in the CA1 subfield in severe AD.

    “However, the TSC1 inclusions observed in our study co-localised to separate compartments with the granules seen resembling not only those found in granulovacuolar degeneration but also in frustrated autophagy, the latter being most prevalent in the CA1 subfield in severe AD [34].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [34]

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

  • May combine two claims.

    Findingown data

    hTSC1 overexpression reduces total cell death and reverses the effects of TSC1 knockdown on caspase-3 activation.

    “hTSC1 overexpression reduced total cell death and reversed the effects of TSC1 knockdown on caspase-3 activation (Figure 3C,D).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Phos-mTOR expression increases with disease severity, but does not differ between CA1 and CA3 subfields.

    “A concurrent increase in phos-mTOR expression was noted with disease severity, though this did not differ between the CA1 and CA3 subfields (Figure 1H).”
    Quote in the paperNot a substantial claim

  • May combine two claims. The quote only partly states this claim.

    Findingcitation

    The present transcriptomic signatures were significantly enriched in immune and nerve ensheathment-related human AD gene network modules that were strongly associated with AD pathology.

    “the present transcriptomic signatures were significantly enriched in the immune and nerve ensheathment-related human AD gene network modules”
    • Also stated in: § DISCUSSION · boosted
      “were strongly associated with AD pathology [33]”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [33]

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

  • May combine two claims.

    Findingown data

    TSC1 knockdown experiments identified AD-related pathway changes, including down-regulation of axon guidance and synaptic transmission and up-regulation of interferon-gamma signalling and lysosome biology.

    “In our TSC1 knockdown experiments, we identified AD-related pathway changes, including down-regulation of axon guidance and synaptic transmission and up-regulation of interferon-gamma signalling and lysosome biology.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In AD cases, immunofluorescence studies show reduced LAMP-1 inclusions and increased TSC1 inclusions in neurons with abundant NFTs, most markedly in the CA1 subfield.

    “Of note, our immunofluorescence studies highlighted reduced expression of LAMP-1 inclusions and increased TSC1 inclusions in neurons with abundant NFTs, most markedly noted in the CA1 subfield of AD cases (Figures S8-S9).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Hippocampal Aβ and tau burden increase with disease severity, with a predisposition for CA1 compared with CA3.

    “whereas hippocampal Aβ and tau burden increased with disease severity, with a predisposition for CA1 compared with CA3 (Figure S4).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The extent of TSC1 inclusions relates to decreased neuronal density in CA1, severity of cognitive impairment during life and markers of AD pathology after death.

    “Importantly, the extent of these inclusions related to decreased neuronal density in CA1, the severity of cognitive impairment during life and markers of AD pathology after death.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: TSC1 inclusions in post-mortem tissue reflect antemortem TSC1 dysregulation.
    • Rests on an assumption: MMSE score obtained in the last 2 years of life reflects cognitive decline.

  • May combine two claims.

    Findingown data

    In human post-mortem tissue, cytoplasmic TSC1 inclusions and evidence of mTOR activation are mainly restricted to CA1 neurons in AD.

    “In human post-mortem tissue, we found striking cytoplasmic TSC1 inclusions and evidence of mTOR activation mainly restricted to CA1 neurons in AD.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    GVs and TSC1 inclusions showed independent compartments, and TSC1 inclusions were partially located in GVs as shown by overlap.

    “GVs (red, arrows) and TSC1 inclusions (green asterisks) showed independent compartments, and TSC1 inclusions were partially located in GVs as shown by overlap (yellow, arrowheads).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Overexpression of hTSC1 combined with rat TSC1shRNA (30.9 ± 5.0%) significantly reduced cell death compared with TSC1shRNA + overexpression at 120 h (p = 0.0002), whereas cell death remained increased compared with the control group (p < 0.0001).

    “Overexpression of hTSC1 combined with rat TSC1shRNA (30.9 ± 5.0%) significantly reduced cell death compared with TSC1shRNA + overexpression at 120 h (p = 0.0002), whereas cell death remained increased compared with the control group (p < 0.0001).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    TSC1 inclusions were more abundant in neurons and GVs were abundant in neurons from both CA1 and CA3 regions.

    “TSC1 inclusions were more abundant in neurons and (C, G) GVs were abundant in neurons from both CA1 and CA3 regions.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    TSC1 knockdown results in increased cell death at 48 h, whereas no difference was observed between TSC1shRNA + rapamycin vs TSC1shRNA + vehicle (21.6 ± 4.2% vs 24.3 ± 3.1%, p = 0.92).

    “TSC1 knockdown results in increased cell death at 48 h, whereas no difference was observed between TSC1shRNA + rapamycin vs TSC1shRNA + vehicle (21.6 ± 4.2% vs 24.3 ± 3.1%, p = 0.92).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In the TSC1shRNA + hTSC1 overexpression group, this effect was reduced to the control level throughout the culture period, such that there was no difference between rescue and control conditions at any point (34.9 ± 5.7% at 120 h, p = 0.216).

    “In the TSC1shRNA + hTSC1 overexpression group, this effect was reduced to the control level throughout the culture period, such that there was no difference between rescue and control conditions at any point (34.9 ± 5.7% at 120 h, p = 0.216).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    There was a selective accumulation of TSC1 in the CA1 compared with the CA3 subfield in neuropathological analysis, and TSC1 inclusions were rarely found in entorhinal cortical neurons.

    “Although there was a selective accumulation of TSC1 in the CA1 compared with the CA3 subfield in our neuropathological analysis, TSC1 inclusions were rarely found in entorhinal cortical neurons.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Aβ treatment had no additional effect in TSC1 knockdown cultures and did not have any effect in the transcriptomic data.

    “Aβ treatment had no additional effect in our knockdown cultures and did not have any effect in our transcriptomic data.”
    Quote in the paperSubstantial claim

  • The quote only partly states this claim.

    Interpretationreasoning

    TSC1 is no longer active in AD CA1 neurons.

    “suggesting that TSC1 is no longer active in AD.”
    • Also stated in: § DISCUSSION · hedged
      “suggesting that TSC1 is inactive in these inclusions.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: TSC1 inclusions in post-mortem tissue reflect antemortem TSC1 dysregulation.

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

    Study design and conductnone given

    The specificity of reactions was confirmed using both positive and negative control sections, the latter where the primary antibody was omitted.

    “The specificity of reactions was confirmed using both positive and negative control sections, the latter where the primary antibody was omitted.”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    Pathological hallmarks of AD include cerebral atrophy, extracellular amyloid-beta plaques and intraneuronal neurofibrillary tangles composed of hyper-phosphorylated tau.

    “Pathological hallmarks of AD include cerebral atrophy, extracellular amyloid-beta (Aβ) plaques and intraneuronal neurofibrillary tangles (NFTs) composed of hyper-phosphorylated tau [1].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1]

      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

    Significant changes in ECM have been reported in the early stages of AD, and inhibiting hippocampal ECM changes restored long-term potentiation and contextual memory performance in APP/PS1 transgenic mice.

    “Significant changes in ECM have been reported in the early stages of AD [41] and inhibiting hippocampal ECM changes restored both long-term potentiation and contextual memory performance in APP/PS1 transgenic mice [42].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [41]

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

    • cites [42]

      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).

    Study design and conductown data

    Multicollinearity did not appear to affect the multiple linear regression model (all VIF scores <5).

    “Multicollinearity did not appear to have an effect (all VIF scores <5).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    The presence of TSC1 inclusions must not be confused with increased TSC1 expression, which has been shown by others to be neuroprotective.

    “Thus, the presence of TSC1 inclusions must not be confused with increased TSC1 expression, which has been shown by others to be neuroprotective [7].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [7]

      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

    Enhanced susceptibility of CA1 to neurodegeneration is not unique to AD, with similar patterns observed in cerebral ischaemia and epilepsy.

    “Interestingly, enhanced susceptibility of CA1 to neurodegeneration is not unique to AD, with similar patterns observed in cerebral ischaemia [5] and epilepsy [6].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [5]

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

    • cites [6]

      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).

    Study design and conductnone given

    Measurements and observations were taken by four experimenters blinded to disease category.

    “Measurements and observations were taken by four experimenters (BA, SB, JP and GD) blinded to disease category.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Rapamycin acts on mTORC2, another mTOR complex that when activated leads to PKC phosphorylation and cytoskeletal changes.

    “We provide some evidence that rapamycin acts on mTORC2, another mTOR complex that-when activated-leads to PKC phosphorylation and cytoskeletal changes [38].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [38]

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

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

    Premisecitation

    Dysregulation of mTOR signalling has been suggested as an early event in AD pathogenesis with implications for tau pathology through decreased autophagy.

    “Dysregulation of mTOR signalling has even been suggested as an early event in AD pathogenesis [11,12] with implications for tau pathology through decreased autophagy [9,[13][14][15].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [11]

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

    • cites [12]

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

    • cites [9]

      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

    Neuronal and volumetric losses in CA1 uniquely predict conversion from mild cognitive impairment to AD.

    “and conversion from mild cognitive impairment to AD [4].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [4]

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

  • May combine two claims.

    Premisecitation

    The activity of mTORC1 is negatively regulated by TSC1 physiologically and by rapamycin.

    “The activity of this complex is negatively regulated by a host of factors, including tuberous sclerosis complex-1 (TSC1, also known as hamartin) physiologically and the drug rapamycin [8].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [8]

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

  • Classified as probably not a substantial claim.

    Premisecitation

    Recent data suggest that mTORC1 signalling in CA1 may explain selective neuronal vulnerability to cerebral ischaemia.

    “Recent data suggest that the mammalian target of rapamycin complex 1 (mTORC1) signalling in CA1 may explain selective neuronal vulnerability to cerebral ischaemia [7].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [7]

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

  • May combine two claims.

    Interpretationown data

    TSC1 loss-of-function activates AD-associated immune responses and suppresses neuronal function.

    “Overall, our findings suggest that TSC1 loss-of-function activates AD-associated immune responses and suppresses neuronal function, thus supporting a role of TSC1 in selective neuronal vulnerability in AD.”
    Quote in the paperSubstantial claim

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

    Study design and conductnone given

    All groups were age- and sex-matched with minimal evidence of small vessel disease.

    “All groups were age-and sex-matched with minimal evidence of small vessel disease.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    In an experimental ischaemia model, hippocampal CA3 neurons increased expression of functional TSC1 in the setting of ischaemic injury, whereas CA1 hippocampal neurons did not.

    “In an experimental ischaemia model, hippocampal CA3 neurons, which are resistant to cell death, increased expression of functional TSC1 in the setting of ischaemic injury, whereas CA1 hippocampal neurons, which are vulnerable to death, did not.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [7]

      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.

    Study design and conductown data

    Data were not normally distributed so non-parametric tests were used.

    “Data were not normally distributed so non-parametric tests were used.”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    mTORC1 is a pleomorphic complex involved in protein synthesis, cellular growth, proliferation and differentiation and has an inhibitory effect on autophagy.

    “mTORC1 is a pleomorphic complex involved in protein synthesis, cellular growth, proliferation and differentiation and has an inhibitory effect on autophagy [8].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [8]

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    Premisecitation

    mTORC1 plays a role in memory and cognition through modulating synaptogenesis and synaptic plasticity.

    “mTORC1 plays a role in memory and cognition through modulating synaptogenesis and synaptic plasticity [10].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [10]

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

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

    Study design and conductown data

    There were no significant differences in age, sex or post-mortem interval between subject groups.

    “There were no significant differences in age, sex or post-mortem interval between subject groups.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    In vitro data show TSC2 down-regulation leads to the formation of TSC1 aggregates with resultant loss of function leading to mTORC1 hyperactivation.

    “Our finding of reduced TSC2 expression in the CA1 hippocampal subfield of AD cases is relevant here given in vitro data that show TSC2 down-regulation leads to the formation of TSC1 aggregates with resultant loss of function leading to mTORC1 hyperactivation [35][36][37].”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [35]

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

    • cites [36]

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    • cites [37]

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

    Premisecitation

    FTO has been identified as upstream of TSC1/TSC2 and the mTOR pathway in AD.

    “A gene associated with insulin signalling defects called FTO has been identified as upstream of TSC1/TSC2 and the mTOR pathway in AD [48]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [48]

      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

    The overall amount of amyloid and its correlation with the severity of dementia in AD remains elusive.

    “Indeed, the overall amount of amyloid and its correlation with the severity of dementia in AD remains elusive [45].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [45]

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

  • May combine two claims.

    Interpretationown data

    TSC1 loss-of-function contributes to driving cell death that is separate from amyloid toxicity, which may cast light onto the debated role of amyloids in cognitive decline in AD.

    “we highlight an important contribution of TSC1 loss-of-function in driving cell death that is separate from amyloid toxicity, which may cast light onto the debated role of amyloids in cognitive decline in AD.”
    Quote in the paperSubstantial claim

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

    Interpretationown data

    It is unclear what role TSC1 loss-of-function plays in neurodegeneration outside the hippocampus and in neurological diseases other than AD.

    “Therefore, it is unclear what role TSC1 loss-of-function plays in neurodegeneration outside the hippocampus and in neurological diseases other than AD.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    In vitro experiments provide evidence that TSC1 loss-of-function promotes mTOR hyperactivation and leads to cell death independent of amyloid exposure.

    “Our in vitro experiments provide evidence that TSC1 loss-offunction promotes mTOR hyperactivation and leads to cell death independent of amyloid exposure.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: TSC1 shRNA knockdown is specific and does not act on other genes to cause the observed effects.
    • Rests on an assumption: The rat hippocampal culture model is relevant to human AD hippocampal selective vulnerability.

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

    Premisecitation

    Amyloid increases mTOR activation and mTOR activation increases amyloid build-up.

    “Others have previously shown amyloid increases mTOR activation and that mTOR activation increases amyloid build-up [14,44]”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [14]

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

    • cites [44]

      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).

    Study design and conductnone given

    Ethical approval was obtained via the Oxford Brain Bank (REC 07/0606/85) in compliance with relevant guidelines, including the Human Tissue Act 2006.

    “Ethical approval was obtained via the Oxford Brain Bank (REC 07/0606/85) in compliance with relevant guidelines, including the Human Tissue Act 2006.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationown data

    Our findings fill the critical knowledge gap of no research into cell-specific mTOR changes in AD.

    “Our findings fill this critical knowledge gap.”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: There has been no research into cell-specific mTOR changes in AD.

  • May combine two claims.

    Interpretationown data

    Previous work has implicated mTOR hyperactivation globally in the AD brain, but our findings point to a more region-specific role of mTOR dysregulation in the pathogenesis of neuronal loss in AD.

    “Previous work has implicated mTOR hyperactivation globally in the AD brain but our findings point to a more region-specific role of mTOR dysregulation in the pathogenesis of neuronal loss in AD.”
    Quote in the paperSubstantial claim

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

    Premisecitation

    In a model of cerebral ischemia, TSC1 was neuroprotective independent of amyloid or tau pathology.

    “A previous study looking at TSC1 in a model of cerebral ischemia highlights this [7] -TSC1 was neuroprotective independent of amyloid or tau pathology.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [7]

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

  • May combine two claims.

    Interpretationown data

    mTOR activation was far more pronounced in vulnerable CA1 neurons compared with resistant CA3 hippocampal neurons, which lie only micrometres away, providing a biological clue to the specific substrate of selective neuronal vulnerability in AD.

    “The fact that mTOR activation was far more pronounced in vulnerable CA1 neurons compared with resistant CA3 hippocampal neurons, which lie only micrometres away, provides an important biological clue to the specific substrate of selective neuronal vulnerability in AD.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    It is plausible that there are no synergistic effects between Aβ and TSC1 loss-of-function in our culture system, with the impact of TSC1 knockdown overshadowing a potential cumulative Aβ toxic effect.

    “It remains then plausible that there are no synergistic effects between Aβ and TSC1 loss-of-function in our culture system, with the impact of TSC1 knockdown overshadowing a potential cumulative Aβ toxic effect.”
    Quote in the paperSubstantial claim
51 of 51

  • The paper relies on this without support.

    Rapamycin at 2 nM effectively inhibits mTORC1 in rat hippocampal neurons.

    “cells were treated with 5-μM Aβ 1-42 or a control vehicle in the absence or presence of 2-nM rapamycin.”
    “Surprisingly, rapamycin treatment did not reduce the extent of TSC1 knockdown-related cell death or caspase-3 activation at any time point”

  • The paper relies on this without support.

    Aβ 1-42 at 5 μM is toxic to rat hippocampal neurons in this model.

    “cells were treated with 5-μM Aβ 1-42 or a control vehicle in the absence or presence of 2-nM rapamycin.”
    “Unexpectedly, treatment with Aβ 1-42 in TSC1 knockdown or hTSC1 rescue experiments did not have additional effects on mTOR activation (Figures S11-S14), Tau burden (Figures S16 and S17), total cell death or caspase-3 activation (Figure S18).”

  • The paper relies on this without support.

    TSC1 exerts its functional activity when associated with the lysosome.

    “As TSC1 exerts its functional activity when associated with the lysosome”

  • The paper relies on this without support.

    Data derived from whole brain homogenate preparations support a role of mTOR hyperactivation in the mesial temporal lobe in AD.

    “Data derived from whole brain homogenate preparations support a role of mTOR hyperactivation in the mesial temporal lobe in AD”

  • The paper relies on this without support.

    TSC1 inclusions in post-mortem tissue reflect antemortem TSC1 dysregulation.

    “TSC1 inclusions in CA1 correlated with cognitive decline, measured by MMSE score, obtained in the last 2 years of life (Figure 1K).”
    “Importantly, the extent of these inclusions related to decreased neuronal density in CA1, the severity of cognitive impairment during life and markers of AD pathology after death.”

  • The paper relies on this without support.

    MMSE score obtained in the last 2 years of life reflects cognitive decline.

    “Indicators of cognition (Mini-Mental Status Examination [MMSE]) obtained in the last 2 years of life and post-mortem interval are listed in Table 1.”
    “Further, TSC1 inclusions in CA1 correlated with cognitive decline, measured by MMSE score, obtained in the last 2 years of life (Figure 1K).”

  • The paper relies on this without support.

    There has been no research into cell-specific mTOR changes in AD.

    “there has been no research into cellspecific mTOR changes in AD.”

  • The paper relies on this without support.

    TSC1 inclusions are indicative of loss-of-function.

    “TSC1 inclusions (indicative of loss-of-function)”

  • The paper relies on this without support.

    TSC1 shRNA knockdown is specific and does not act on other genes to cause the observed effects.

    “Lentiviral knockdown of TSC1 in rat hippocampal cultures was performed using pLKO.1 TSC1 shRNA (Sigma-Aldrich, targeting nucleotides 2262-2282 of the rat TSC1 mRNA) and pLKO.1 control shRNA (Sigma-Aldrich, control sequence that targets TurboGFP, a sequence that does not correspond to any known mammalian gene) at a multiplicity of infection (MOI) of 2.”

  • The paper relies on this without support.

    The rat hippocampal culture model is relevant to human AD hippocampal selective vulnerability.

    “Lentiviral knockdown of TSC1 in rat hippocampal cultures was performed using pLKO.1 TSC1 shRNA (Sigma-Aldrich, targeting nucleotides 2262-2282 of the rat TSC1 mRNA) and pLKO.1 control shRNA (Sigma-Aldrich, control sequence that targets TurboGFP, a sequence that does not correspond to any known mammalian gene) at a multiplicity of infection (MOI) of 2.”
10 of 10

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?

  • Neurons with heightened mTOR activity, particularly hippocampal CA1 neurons, are selectively predisposed to tau accumulation and degeneration.

    “Experimental data further indicate that neurons with heightened mTOR activityparticularly hippocampal CA1 neurons, a region highly vulnerable in AD-are selectively predisposed to tau accumulation and degeneration, implicating mTOR dysregulation as a central upstream driver (14,15).”

    From Amyloid-Independent Elevation of Plasma phosphorylated tau-217 Tracks Neurodegenerative Risk in Tuberous Sclerosis Complex: Implications for mTOR-Targeted Intervention (2026) · cites this paper as [14]

    The CA1 results (page not available in our copy) report increased phos-mTOR, greater tau burden than CA3, and reduced neuronal density in severe AD. The Discussion links CA1 mTOR activation to selective vulnerability, but the experiments do not establish that heightened mTOR activity predisposes neurons to tau accumulation or causes their degeneration; rapamycin did not reduce TSC1-knockdown-related cell death. (LLM, conf=0.86)

    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.

  • CA1-versus-CA3 mTOR activation claim conflicts with the reported comparison

    Major

    The discussion says mTOR activation was far more pronounced in CA1 than CA3, while the results state that phos-mTOR expression did not differ between those subfields.

  • Figure error bars are described as both SD and SEM

    Minor

    The general figure note says data in graphs are presented as mean ± SD, but the Figure 1 caption identifies its error bars as SEM.

2 of 2

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Authors
Adriaanse, B. A., Brady, S., Wang, M., Beard, D. J., Spencer, J. I., Pansieri, J., Sutherland, B. A., Esiri, M. M., Buchan, A. M., Cader, Z., Zhang, B., & DeLuca, G. C.
Affiliations
  • University of Oxford (GB)
  • Icahn School of Medicine at Mount Sinai (US)
  • University College Hospital (GB)
  • University of Tasmania (AU)
  • Nuffield Department of Clinical Neurosciences University of Oxford Oxford UK
Corresponding author
  • Zhang, Bin
  • DeLuca, Gabriele C.
ORCID
Venue
Neuropathology and Applied Neurobiology
Publisher
Wiley
Published
2023-05-11
Volume
49
Issue
3
Pages
e12904-e12904
ISSN
  • 0305-1846 (ISSN-L)
  • 1365-2990
DOI
10.1111/nan.12904
OpenAlex
W4362658033
Cited by
2
References
48
Citations by year
2025: 1 · 2024: 1
FWCI
0.29
Type
journal article
Language
en
Open access
Licence
Locations
  • Neuropathology and Applied Neurobiology (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • NOVA (University of Newcastle Australia) (submittedVersion)
Funders
  • National Institute on Aging
  • National Institutes of Health
  • NIHR Oxford Biomedical Research Centre
  • National Institute for Health and Care Research
Topics
  • Health Sciences › Medicine › Physiology › Alzheimer's disease research and treatments
  • Health Sciences › Medicine › Psychiatry and Mental health › Dementia and Cognitive Impairment Research
  • Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
Keywords
tuberous sclerosis complex 1, hamartin, Alzheimer's disease, selective neuronal vulnerability, CA1 pyramidal neurons, mTORC1 hyperactivation, neuronal cell death, gene expression profiling, postmortem brain, neurodegeneration
MeSH terms
Alzheimer Disease, Animals, Hippocampus, Humans, Mammals, Neurons, Tuberous Sclerosis, Rats, TOR Serine-Threonine Kinases

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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 2026-10-27.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-27.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

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