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Cited by 398Mitochondrial Function and PathologyMuscle Physiology and DisordersAutophagy in Disease and Therapy

DRP1-mediated mitochondrial shape controls calcium homeostasis and muscle mass

Open access
Favaro, Giulia, Romanello, Vanina, Varanita, Tatiana, Andrea Desbats, Maria, Morbidoni, Valeria, Tezze, Caterina, Albiero, Mattia, Canato, Marta +10 more·2019·Nature Communications
DOI 10.1038/s41467-019-10226-9PMC PMC6561930
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Abstract

Mitochondrial quality control is essential in highly structured cells such as neurons and muscles. In skeletal muscle the mitochondrial fission proteins are reduced in different physiopathological conditions including ageing sarcopenia, cancer cachexia and chemotherapy-induced muscle wasting. However, whether mitochondrial fission is essential for muscle homeostasis is still unclear. Here we show that muscle-specific loss of the pro-fission dynamin related protein (DRP) 1 induces muscle wasting and weakness. Constitutive Drp1 ablation in muscles reduces growth and causes animal death while inducible deletion results in atrophy and degeneration. Drp1 deficient mitochondria are morphologically bigger and functionally abnormal. The dysfunctional mitochondria signals to the nucleus to induce the ubiquitin-proteasome system and an Unfolded Protein Response while the change of mitochondrial volume results in an increase of mitochondrial Ca 2+ uptake and myofiber death. Our findings reveal that morphology of mitochondrial network is critical for several biological processes that control nuclear programs and Ca 2+ handling.

AI snapshot

Written from the paper’s full text.

TL;DR

Muscle-specific loss of DRP1 in mice causes a lethal mitochondrial myopathy with atrophy, weakness, and altered calcium homeostasis. DRP1-deficient mitochondria are larger and dysfunctional, triggering UPR/UPS activation, blocked autophagy/mitophagy, and increased MCU-mediated Ca²⁺ uptake that contributes to myofiber death.

Key findings

  • Constitutive muscle-specific Drp1 deletion caused postnatal lethality within 30 days (WT n = 43; KO n = 14), reduced body growth, fewer glycolytic 2B/2X fibers, smaller cross-sectional area, and a 30% reduction in protein synthesis.
  • Inducible adult Drp1 deletion reduced fiber size by 25% at 70 days and 50% at 180 days, decreased absolute and specific force, and produced center-nucleated fibers in 10% of tibialis anterior and 25% of gastrocnemius myofibers at 180 days.
  • Drp1-deficient mitochondria were larger and fewer with unchanged mtDNA content and mitochondrial mass, but showed reduced respiration (complex-I-driven respiration most affected), decreased respiratory chain supercomplexes, increased OPA1 cleavage, and in acute deletion about 10% damaged mitochondria versus 2% in WT.
  • DRP1 loss activated UPR (Bip/Grp78, p-eIF2α, ATF4, FGF21; FGF21 blood increased 20-fold in acute KO), induced FoxO3/UPS, blocked autophagy/mitophagy, and increased mitochondrial Ca²⁺ uptake via MCU upregulation and miR1 downregulation; MCU knockdown normalized cytosolic Ca²⁺ and reduced center-nucleated fibers.

Methods

  • Two muscle-specific Drp1 knockout models: constitutive Drp1fl/fl × MLC1f-Cre mice analyzed at postnatal day 12, and tamoxifen-inducible Drp1fl/fl × HSA-Cre-ER mice treated at 5 months of age; Cre-negative littermates served as controls.
  • Readouts included electron microscopy/stereology, histology and SDH staining, RT-qPCR, immunoblotting, mitochondrial respiration and Blue Native PAGE, SUnSET protein synthesis, TMRM membrane potential, Fura-2 and cameleon Ca²⁺ probes, in vivo/ex vivo force and fatigue measurements, mito-Keima mitophagy assay, MCU knockdown, and plasma FGF21/IGF1 measurements.
  • Data are means ± SEM; statistical analysis used two-tailed unpaired Student’s t test or two-way ANOVA with Bonferroni’s multiple comparison test, with significance at p < 0.05.

Limitations

  • Stated: MCU knockdown did not completely block degeneration/regeneration because transfection efficiency was low, so MCU was reduced only in some myofibers.
  • Evident: all findings are from mouse models; no human patient validation is reported.
  • Evident: some analyses used very small n (e.g., WT n = 2 for succinate/rotenone respiration in Fig. 5d), and the causal role of Ca²⁺ dysregulation is supported by rescue/knockdown experiments but not fully isolated from other DRP1-dependent defects.

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

Integrity signals

Registry

Crossref · 10.1038/s41467-019-10226-9

Cites

62 of 62 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 27 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    FGF21 blood levels are increased 20-fold in DRP1 knockout mice compared with controls.

    “FGF21 blood levels were also increased of 20-fold in DRP1 knockout mice when compared with controls”
    Quote in the paperNot a substantial claim
  • Findingown data

    Inducible deletion of Drp1 results in atrophy and degeneration.

    “inducible deletion results in atrophy and degeneration”
    Quote in the paperSubstantial claim
  • Findingown data

    In DRP1-deficient muscles, atrogin1 and MuRF1 do not change.

    “while atrogin1 and MuRF1 did not change”
    Quote in the paperNot a substantial claim
  • Findingown data

    Most FoxO-dependent autophagy-related genes are upregulated in DRP1-deficient muscles.

    “we found an upregulation of most of these genes”
    Quote in the paperNot a substantial claim
  • Findingown data

    Protein synthesis was reduced by 30% in Drp1−/− mice.

    “protein synthesis was reduced by 30% in Drp1−/− mice”
    Quote in the paperSubstantial claim
  • May combine two claims. 36
  • Classified as probably not a substantial claim. 14

  • May combine two claims.

    Findingown data

    UPR pathway is activated in DRP1-null muscles, with induction of Bip/Grp78, p-eIF2α, ATF4 and downstream genes including FGF21.

    “we found an activation of the UPR pathway. Bip/Grp78 and p-eIF2α (Fig. 6c; Supplementary Fig. 7a) and the transcripts of ATF4 and downstream genes, including FGF21, were induced in DRP1-null muscles”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In DRP1-deficient muscles, there is no increase of lipidated LC3II, but accumulation of LC3I.

    “we did not detect an increase of lipidated LC3II, but instead we found an accumulation of LC3I”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Ablation of DRP1 in adulthood reduces RCS assembly and Complex-I-, Complex-II-, and complex-IV-dependent respiration.

    “Ablation of DRP1 in adulthood modifies mitochondrial shape and reduces RCS assembly, Complex-I-, Complex-II-, and complex-IV-dependent respiration.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Muscle-specific loss of DRP1 induces muscle wasting and weakness.

    “Here we show that muscle-specific loss of the pro-fission dynamin related protein (DRP) 1 induces muscle wasting and weakness.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Drp1 deficient mitochondria are morphologically bigger and functionally abnormal.

    “Drp1 deficient mitochondria are morphologically bigger and functionally abnormal.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Ablation of DRP1 in adulthood modifies mitochondrial shape, producing bigger mitochondria in KO muscles compared with control.

    “SDH staining indicating the presence of bigger mitochondria in KO muscles compared with control.”
    • Also stated in: § Acute deletion of Drp1 causes mitochondrial dysfunction · plain
      “Ablation of DRP1 in adulthood modifies mitochondrial shape and reduces RCS assembly, Complex-I-, Complex-II-, and complex-IV-dependent respiration.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Mitochondrial matrix-free calcium concentration is not affected at rest but is higher in Drp1−/− during 60 Hz electrical stimulation.

    “Mitochondrial matrix-free calcium concentration is not affected at rest but is higher in Drp1−/− compared with control during a train of electrical stimulation at 60 Hz, 2 s duration”
    • Also stated in: § Inhibition of DRP1 alters normal calcium homeostasis · plain
      “the peak of Ca2+ concentration reached during high-frequency electrical stimulation was higher in DRP1-deficient myofibers”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Constitutive Drp1 ablation in muscles reduces growth and causes animal death.

    “Constitutive Drp1 ablation in muscles reduces growth and causes animal death”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In acute Drp1-deficient mice, 4EBP1 is hyper-phosphorylated at two mTOR-dependent sites and S6 is unchanged.

    “the Akt-mTOR axis appeared inconsistent with the atrophic phenotype being 4EBP1 robustly hyper-phosphorylated in two mTOR-dependent sites and S6 unchanged”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    These changes occurred early at 40 days after tamoxifen, before muscle mass loss, and progressively increased at 70 days.

    “These changes occurred early after 40 days from tamoxifen treatment when animals have not yet lost muscle mass and progressively increased at 70 days.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Inflammatory response is not present and oxidative stress is not increased in DRP1 knockout mice.

    “the inflammatory response is not present and oxidative stress is not increased in DRP1 knockout mice (Supplementary Figs. 9f, 6e).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    DRP1 overexpression reverts UPR in DRP1 KO and induces UPR in control mice.

    “DRP1 overexpression reverted or induced UPR in DRP1 knockout and control mice, respectively”
    Quote in the paperSubstantial claim

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

    Findingreasoning

    Under the stated calibration assumptions, the resting cytosolic calcium concentration will be 71 nM at R = 0.87 and 87 nM at R = 0.92.

    “Under these assumptions, the resting cytosolic calcium concentration will be 71 nM at R = 0.87 and 87 nM at R = 0.92.”
    Quote in the paperNot a substantial claim
    • The authors state this as an assumption: The calibration equation and constants (KD = 145 nM, Rmin, Rmax, β) used to convert Fura-2 fluorescence ratios to free cytosolic calcium concentration are valid.

  • May combine two claims.

    Findingown data

    CRUs are modified in DRP1 KO compared with WT fibers, with increased frequency of oblique/longitudinal triads and decreased number/area of CRUs.

    “we performed EM analyses and found some modification in CRUs compared with WT fibers”
    • Also stated in: § Inhibition of DRP1 alters normal calcium homeostasis · boosted
      “we showed an increased frequency of oblique/longitudinal triads (inserts D and E) and a decrease in the number/area of CRUs compared with WT fibers”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Reducing MCU expression for two months reduces center-nucleated fibers and suppresses UPR in DRP1 KO.

    “By reducing MCU expression for two months, we reduced the amount of center-nucleated fibers (Fig. 7j) and suppressed UPR (Supplementary Fig. 9e).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    Loss-of-function approach shows decrease of mitofusin2 and downregulation of calnexin and upregulation of Bip/Grp78.

    “Another peculiarity of the loss-of-function approach is the decrease of the ER-mitochondria tethering protein mitofusin2 and the down- and upregulation of the ER chaperones calnexin and Bip/Grp78, respectively11.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [11] Touvier, T. et al. Muscle-specific Drp1 overexpression impairs skeletal muscle growth via translational attenuation. Cell Death Dis. 6, e1663 (2015). source

      Not checked in the free run.

  • May combine two claims.

    Findingown data

    Mitofusin2 and calnexin expression are downregulated in DRP1 KO.

    “we found a downregulation in their expression”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    Change of mitochondrial volume results in an increase of mitochondrial Ca2+ uptake and myofiber death.

    “the change of mitochondrial volume results in an increase of mitochondrial Ca2+ uptake and myofiber death”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    Acute inhibition of DRP1 causes muscle loss and important weakness, but ameliorates fatigue.

    “Thus, acute inhibition of DRP1 causes muscle loss and important weakness, but ameliorates fatigue.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Loss of DRP1 in adult muscle is sufficient to induce a catabolic condition involving activation of UPS and inhibition of autophagy and mitophagy.

    “loss of DRP1 in adult muscle is sufficient to induce a catabolic condition that involves activation of the UPS system and inhibition of autophagy and mitophagy pathways34.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [34] Masiero, E. et al. Autophagy is required to maintain muscle mass. Cell Metab. 10, 507–515 (2009). source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    DRP1 ablation alters mitochondrial morphology and function.

    “Therefore, DRP1 ablation alters mitochondrial morphology and function.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    UPR is strongly activated in Drp1−/− muscles and can account for the translation impairment.

    “supporting the concept that UPR is strongly activated in Drp1−/− muscles and can account for the translation impairment”
    Quote in the paperSubstantial claim

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

    Study design and conductcitation

    Autophagic flux was monitored in basal condition using colchicine.

    “We monitored autophagic flux in basal condition using colchicine62.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [62] Ju, J. S., Varadhachary, A. S., Miller, S. E. & Weihl, C. C. Quantitation of “autophagic flux” in mature skeletal muscle. Autophagy 6, 929–935 (2010). source

      Not checked in the free run.

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

    Premisecitation

    Inhibition of PTP opening is beneficial in dystrophic muscles.

    “it has been shown that inhibition of PTP opening is beneficial in dystrophic muscles45–47.”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [45] Millay, D. P. et al. Genetic and pharmacologic inhibition of mitochondrial-dependent necrosis attenuates muscular dystrophy. Nat. Med. 14, 442–447 (2008). source

      Not checked in the free run.

    • cites [46] Irwin, W. A. et al. Mitochondrial dysfunction and apoptosis in myopathic mice with collagen VI deficiency. Nat. Genet. 35, 367–371 (2003). source

      Not checked in the free run.

    • cites [47] Angelin, A. et al. Mitochondrial dysfunction in the pathogenesis of Ullrich congenital muscular dystrophy and prospective therapy with cyclosporins. Proc. Natl Acad. Sci. USA 104, 991–996 (2007). source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Both fusion and fission machinery inhibitions induce mitochondrial dysfunction (decreased respiration, supercomplex formation, membrane potential), but downstream signaling pathways and biological effects are peculiar for fusion or fission deficiency.

    “In conclusion, even though both fusion and fission machinery inhibitions induce mitochondrial dysfunction, revealed by decreased respiration, supercomplex formation, and mitochondrial membrane potential, the signaling pathways and the biological effects downstream mitochondria are peculiar for fusion or fission deficiency.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Two different knockout mice (muscle-specific and inducible muscle-specific) result in striking phenotypes, underlining critical role of DRP1 in mitochondrial function and muscle homeostasis.

    “The finding that two different knockouts mice, muscle-specific and inducible muscle-specific, result in striking phenotypes (Supplementary Table 3) underlines the critical role of DRP1 in mitochondria function and muscle homeostasis.”
    Quote in the paperSubstantial claim

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

    Study design and conductnone given

    Force was normalized to the muscle mass as an estimate of specific force.

    “Force was normalized to the muscle mass as an estimate of specific force.”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    ER stress and UPR activation are well-known triggers of FGF21 production in different organs.

    “ER stress and UPR activation are well-known triggers of FGF21 production in different organs, such as the liver, pancreas, or adipose tissue39.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [39] Gimeno, R. E. & Moller, D. E. FGF21-based pharmacotherapy-potential utility for metabolic disorders. Trends Endocrinol. Metab. 25, 303–311 (2014). source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Shape of mitochondria is critical for mitochondrial function, but triggers different signaling pathways and biological processes peculiar to fusion or fission impairment.

    “Therefore, the shape of the mitochondria is critical for mitochondrial function, but triggers different signaling pathways and biological processes that are peculiar of fusion or fission impairment.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Impairment of SR-mitochondria tethering and ER chaperone proteins explains the abnormalities of CRUs and the presence of ER stress.

    “Thus, the impairment of the SR–mitochondria tethering and ER chaperon proteins would explain the abnormalities of CRUs and the presence of ER stress36,37.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [36] Gutierrez, T. & Simmen, T. Endoplasmic reticulum chaperones tweak the mitochondrial calcium rheostat to control metabolism and cell death. Cell Calcium 70, 64–75 (2018). source

      Not checked in the free run.

    • cites [37] Debattisti, V., Pendin, D., Ziviani, E., Daga, A. & Scorrano, L. Reduction of endoplasmic reticulum stress attenuates the defects caused by Drosophila mitofusin depletion. J. Cell Biol. 204, 303–312 (2014). source

      Not checked in the free run.

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

    Study design and conductnone given

    The mtDNA copy number was calculated using 18S amplification as a reference for nuclear DNA content.

    “The mtDNA copy number was calculated using 18S amplification as a reference for nuclear DNA content.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Study design and conductown data

    The degeneration/regeneration process was not completely blocked because transfection efficiency was low and MCU was reduced only in some myofibers.

    “We did not completely block the degeneration/regeneration process because the efficiency of transfection was low, and consequently, MCU was reduced only in some myofibers.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Both chronic inhibition and acute hyperactivation of mitochondrial fission result in detrimental effects on muscle mass and function, but with different phenotypes and mechanisms.

    “Like other systems, such as autophagy, both chronic inhibition or acute hyperactivation of mitochondrial fission result in detrimental effects on muscle mass and function, but with different phenotypes and mechanisms.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    DRP1 inhibition causes increased MCU expression via miR1 downregulation and enhanced Ca2+ uptake of the mitochondria.

    “Therefore, DRP1 inhibition causes increased MCU expression via miR1 downregulation and enhanced Ca2+ uptake of the mitochondria.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Initial outcome of genetic DRP1 ablation is identical to its overexpression (muscle atrophy via FoxO); loss-of-function also reveals center-nucleated fibers and involvement of different signaling pathways including MCU upregulation, ER stress, UPR activation, and FGF21 induction.

    “However, while the initial outcome of genetic DRP1 ablation is identical to its overexpression, i.e., muscle atrophy via FoxO, our loss-of-function study also reveals the appearance of center-nucleated fibers, a sign of myopathy, and the involvement of different signaling pathways, including MCU upregulation, ER-stress, UPR activation, and great FGF21 induction (Fig. 8).”
    Quote in the paperSubstantial claim

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

    Study design and conductown data

    No evidence of necrosis or inflammation was observed after the transfection procedure.

    “No evidence of necrosis or inflammation were observed after the transfection procedure.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    Inhibition of fission machinery alters SR-mitochondria tethering, decreases CRU units, increases mitochondrial Ca2+ uptake, reduces Ca2+ availability for contraction, and induces myofiber death due to mitochondrial Ca2+ overload.

    “Altogether, these findings confirm that inhibition of the fission machinery alters the SR–mitochondria tethering, decreases CRU units, increases mitochondrial Ca2+ uptake that, altogether, reduces Ca2+ availability for contraction, and simultaneously induces myofiber death, due to mitochondrial Ca2+ overload.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    Increased MCU expression could induce mitochondrial Ca2+ overload and possibly explains swollen/damaged mitochondria.

    “The increased expression of MCU in Drp1−/− fibers could induce mitochondrial Ca2+ overload, and possibly explains the increased presence of the swollen and damaged mitochondria (Fig. 5 b; Supplementary Table 1).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S4 (based). May combine two claims.

    Premisenone given

    ATP-synthase can reversely transport protons across the inner mitochondrial membrane, so maintaining the potential also in dysfunctional mitochondria.

    “ATP-synthase can reversely transport protons across the inner mitochondrial membrane, so maintaining the potential also in dysfunctional mitochondria.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductcitation

    Free Ca2+ concentrations in the mitochondrial matrix and SR lumen were determined using cameleon Ca2+ probes, 4mtD3cpv for mitochondria and D1-ER for sarcoplasmic reticulum.

    “Free Ca2+ concentrations in the mitochondrial matrix and SR lumen were determined using cameleon Ca2+ probes, 4mtD3cpv54 for mitochondria and D1-ER 55 for sarcoplasmic reticulum, kindly donated by Dr. R.Y. Tsien (University of California, San Diego).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [54] Giacomello, M. et al. Ca2+ hot spots on the mitochondrial surface are generated by Ca2+ mobilization from stores, but not by activation of store-operated Ca2+ channels. Mol. Cell 38, 280–290 (2010). source

      Not checked in the free run.

    • cites [55] Palmer, A. E., Jin, C., Reed, J. C. & Tsien, R. Y. Bcl-2-mediated alterations in endoplasmic reticulum Ca2+ analyzed with an improved genetically encoded fluorescent sensor. Proc. Natl Acad. Sci. USA 101, 17404–17409 (2004). source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Ca2+ overload triggers PTP opening and mitochondrial swelling.

    “it is well known that Ca2+ overload triggers permeability transition pore (PTP) opening and mitochondrial swelling44.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [44] Giorgio, V., Guo, L., Bassot, C., Petronilli, V. & Bernardi, P. Calcium and regulation of the mitochondrial permeability transition. Cell Calcium 70, 56–63 (2017). source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    The improved fatigue can be explained by lower force, less contractile proteins, reduced cytosolic Ca2+, decreased ATP consumption, delaying ATP drop.

    “The apparent paradox could be explained by the lower force developed by DRP1-deficient muscles compared with control muscles. The reduction of force due to less contractile proteins and of cytosolic Ca2+ level leads to a decreased ATP consumption during contraction by myosin and SERCA ATPase pump, delaying ATP drop and consequently, improving fatigue.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductcitation

    Mitochondrial volume was determined using the well-established stereology point-counting technique.

    “Mitochondrial volume (Supplementary Table 1, column A and Supplementary Fig. 2a) was determined using the well-established stereology point-counting technique49,50 in micrographs taken from transversal sections at ×7.100 magnifications.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [49] Loud, A. V., Barany, W. C. & Pack, B. A. Quantitative evaluation of cytoplasmic structures in electron micrographs. Lab. Invest. 14, 996–1008 (1965).

      Not checked in the free run.

    • cites [50] Mobley, B. A. & Eisenberg, B. R. Sizes of components in frog skeletal muscle measured by methods of stereology. J. Gen. Physiol. 66, 31–45 (1975). source

      Not checked in the free run.

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

    Study design and conductnone given

    Only mitochondria which were entirely visualized in the micrograph were measured.

    “Only mitochondria which were entirely visualized in the micrograph were measured.”
    Quote in the paperNot a substantial claim

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

    Study design and conductcitation

    In vivo protein synthesis was measured by using the SUnSET technique.

    “In vivo protein synthesis was measured by using the SUnSET technique25,60.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [25] Schmidt, E. K., Clavarino, G., Ceppi, M. & Pierre, P. SUnSET, a nonradioactive method to monitor protein synthesis. Nat. Methods 6, 275–277 (2009). source

      Not checked in the free run.

    • cites [60] Goodman, C. A. et al. Novel insights into the regulation of skeletal muscle protein synthesis as revealed by a new nonradioactive in vivo technique. FASEB J. 25, 1028–1039 (2011). source

      Not checked in the free run.

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

    Study design and conductnone given

    The results of mitochondrial respiratory chain enzymatic activities were normalized to citrate synthase enzymatic activity.

    “The results were normalized to citrate synthase enzymatic activity.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Micrographs of non-overlapping regions were randomly collected from transversal and longitudinal sections of internal fiber areas of EDL muscles.

    “Micrographs of non-overlapping regions were randomly collected from transversal and longitudinal sections of internal fiber areas of EDL muscles.”
    Quote in the paperSubstantial claim
47 of 47

  • The paper relies on this without support.

    Mitochondrial quality control is essential in highly structured cells such as neurons and muscles.

    “Mitochondrial quality control is essential in highly structured cells such as neurons and muscles.”

  • The paper relies on this without support.

    Insulin signaling through mTOR plays a critical role in controlling protein synthesis.

    “Insulin signaling, through mTOR, plays a critical role in controlling protein synthesis.”

  • The paper relies on this without support.

    FGF21 causes GH resistance.

    “Because FGF21 causes GH resistance”

  • The authors state this as an assumption.

    The calibration equation and constants (KD = 145 nM, Rmin, Rmax, β) used to convert Fura-2 fluorescence ratios to free cytosolic calcium concentration are valid.

    “Under these assumptions, the resting cytosolic calcium concentration will be 71 nM at R = 0.87 and 87 nM at R = 0.92.”
4 of 4

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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.

  • Neonatal fiber loss is described as occurring without hypoplasia

    Major

    The Results state that Drp1-deficient muscles have fewer fibers, while the Fig. 1 caption describes the fiber-count analysis as showing no hypoplasia.

  • Fura-2 excitation wavelengths disagree within the calcium-measurement method

    Minor

    The cytosolic calcium method first specifies a 360/380-nm fluorescence ratio, but its calibration defines that ratio using 340/380 nm.

2 of 2

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Authors
Favaro, G., Romanello, V., Varanita, T., Andrea Desbats, M., Morbidoni, V., Tezze, C., Albiero, M., Canato, M., Gherardi, G., De Stefani, D., Mammucari, C., Blaauw, B., Boncompagni, S., Protasi, F., Reggiani, C., Scorrano, L., Salviati, L., & Sandri, M.
Affiliations
  • University of Padua (IT)
  • University of Chieti-Pescara (IT)
Corresponding author
  • Salviati, Leonardo
  • Sandri, Marco
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2019-06-12
Volume
10
Issue
1
Pages
2576-2576
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-019-10226-9
PMC
PMC6561930
OpenAlex
W2950169360
Semantic Scholar
f4a2921dcb6e0705b3925fc5a9a47319adb62545
Cited by
398
References
62
Citations by year
2026: 30 · 2025: 44 · 2024: 53 · 2023: 62 · 2022: 53 · 2021: 78 · 2020: 67 · 2019: 10
FWCI
17.20
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
  • Padua Research Archive (University of Padova) (submittedVersion, cc-by)
Funders
  • Fondazione Cassa di Risparmio di Padova e Rovigo
  • European Commission (MSCA-RISE-2014, H2020, 645648, H2020‐MSCA‐RISE‐2014, H2020-MSCA, H2020 MSCA Rise)
  • Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (133130)
  • Ministero della Salute (GR-2011-02352681)
  • Associazione Italiana per la Ricerca sul Cancro (PRIN 2015, 17388, PRIN 2010)
  • Horizon 2020 Framework Programme (H2020-MSCA-RISE-2014, H2020-MSCA, H2020-MSCA-RISE, 645648)
  • FP7 Ideas: European Research Council (282310)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Muscle Physiology and Disorders
  • Health Sciences › Medicine › Epidemiology › Autophagy in Disease and Therapy
Keywords
Drp1, mitochondrial fission, mitochondrial morphology, calcium homeostasis, muscle atrophy, skeletal muscle, mitochondrial quality control, ubiquitin-proteasome system, unfolded protein response, myofiber degeneration, mitochondrial calcium uptake, sarcopenia, cancer cachexia
MeSH terms
Animals, Calcium, Cell Nucleus, Disease Models, Animal, Homeostasis, Humans, Mitochondria, Muscle, Ubiquitins, Mitochondrial Myopathies, Mice, Knockout, Muscle, Skeletal, Dynamins, Proteasome Endopeptidase Complex, Mice, Unfolded Protein Response, Proteolysis, Mitochondrial Dynamics

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 2026-10-06.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-06.

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.

What counts as a claim ›How we measure confidence ›How we check this ›

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