Loss of epigenetic information as a cause of mammalian aging

May combine two claims.

Reduced H3K122ac extends lifespan of budding yeast, and H3K27ac and H3K56ac decrease in many human immune cell types.

“Reduced levels of H3K122ac extend the lifespan of budding yeast (Sen et al., 2015) and levels of H3K27ac and H3K56ac decrease in many human immune cell types (Cheung et al., 2018; Dang et al., 2009).”

• cites Sen et al., 2015 sourceCould not verify

  Sen et al. is cited for the budding-yeast H3K122ac lifespan claim, not the separate human immune-cell claim. The Results section on histone mutations and the Discussion report lifespan effects involving H3K36 methylation, but the passages found by searching for H3K122ac and H3K122 do not address whether reduced H3K122ac extends lifespan. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites Cheung et al., 2018 sourceDisputed

  Cheung et al. report the opposite overall age trend: the Results (EpiTOF Reveals Aging-Associated Changes; page unavailable) say the vast majority of chromatin-mark measurements were elevated in older subjects, and the Discussion says most marks increased across a broad array of immune cell subtypes. The Results identify reduced marks in central memory CD8+ T cells as an exception, not a decrease across many immune cell types. The budding-yeast claim is attributed to Sen et al., not Cheung et al. (LLM, conf=0.82)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured
• cites Dang et al., 2009 sourcePartly supported

  The “Histone modifications change in old cells” section (page unavailable) reports that H3K56ac decreased with age in yeast mother cells. It does not establish a decrease in H3K56ac or H3K27ac across human immune cell types. The H3K122ac lifespan claim is attached to Sen et al., not to Dang et al. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

May combine two claims.

Old mice develop opaque lenses and lose retinal ganglion cells in the innermost retinal layer.

“Old mice develop opaque lenses and lose retinal ganglion cells (RGCs) in the innermost retinal layer (Calkins, 2013; Downs, 2015; Samuel et al., 2011; Wolf et al., 2000) (Figure 7J).”

• cites Calkins, 2013 sourceNot checked — no full text of the source
• cites Downs, 2015 sourcePartly supported

  Section 3.A (Aging; page unavailable) discusses retinal ganglion cell loss in an experimental mouse model of glaucoma. It does not say that aging itself causes this loss in mice, and the passages found on lens opacity do not support the claim about opaque lenses. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites Samuel et al., 2011 sourceDisputed

  The Discussion mentions lens opacification as an age-related optical factor, but does not establish that the study's old mice developed opaque lenses. More decisively, the Results section, “Neurons are preserved in old retina,” reports little change in retinal neuron numbers and no detectable age-related increase in cell death, contrary to the claim that old mice lose RGCs. (LLM, conf=0.90)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured

May combine two claims.

Double-strand DNA breaks are the damage type most consistently linked to aging, occurring at 10 to 50 per cell per day.

“The type of DNA damage that is most consistently linked to aging is the double-stranded DNA break (DSB), occurring at a rate of 10 to 50 per cell per day (Tian et al., 2019; Vilenchik and Knudson, 2003).”

• cites Tian et al., 2019 sourceNot checked — no full text of the source
• cites Vilenchik and Knudson, 2003 sourcePartly supported

  The Background Occurrence of DSBs section estimates about 50 endogenous double-strand breaks per human cell per cell cycle, supporting the claim that these breaks occur frequently. It does not give a rate of 10–50 per cell per day or identify double-strand breaks as the damage type most consistently linked to aging; the paper discusses their implications for cancer instead. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

May combine two claims.

Overexpression of SIR2 or histones, or deletion of SET2, extends yeast lifespan, indicating epigenetic changes are a cause, not merely a biomarker, of yeast aging.

“Overexpression of SIR2, histones, or deletion of the histone methyltransferase gene SET2, extends yeast lifespan, indicating that epigenetic changes are not merely a biomarker, but a cause of yeast aging (Dang et al., 2009; Feser et al., 2010; Hu et al., 2014; Kaeberlein et al., 1999; Ryu et al., 2014).”

• cites Dang et al., 2009 sourcePartly supported

  The section ‘Sas2 has opposing effects on H4K16ac and lifespan’ (page not supplied) reports that SIR2 overexpression extends yeast lifespan. The Discussion (page not supplied) links histone acetylation and telomeric chromatin to longevity, supporting the point that epigenetic changes can affect aging. This paper does not establish lifespan extension from histone overexpression or SET2 deletion; it reports lifespan extension from SAS2 deletion instead. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Feser et al., 2010 sourceSupported
• cites Hu et al., 2014Not matched

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

May combine two claims.

Many old cells have few mutations, mouse or human strains with higher mutation rates show little premature aging, and mammals can be cloned from old somatic cells to produce normal-lifespan individuals.

“Many types of old cells have a paucity of mutations (De Majo et al., 2021; Kaya et al., 2015), strains of mice or people with higher mutation rates show little to no evidence of premature aging (Narayanan et al., 1997; Robinson et al., 2021), and mammals can be cloned from old somatic cells to produce new individuals with normal lifespans (Burgstaller and Brem, 2017).”

• cites De Majo et al., 2021 sourcePartly supported

  De Majo et al. supports the first clause for naturally aged mouse hearts: the Discussion reports negligible genomic instability in physiologically aging myocardium. It does not establish that many types of old cells have few mutations; the Results report increased mutational burden with age in mouse intestine, lung, and liver. The later clauses are attributed to other citations, not De Majo et al. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Kaya et al., 2015 sourceSupported
• cites Narayanan et al., 1997 sourceNot checked — no full text of the source

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

Mammalian aging is associated with decline in central nervous system function.

“Mammalian aging is associated with a decline in central nervous system function (Figure 3A) (Johnson et al., 2018; Ungvari et al., 2017).”

• cites Johnson et al., 2018 sourceSupported
• cites Ungvari et al., 2017 sourceNot checked — no full text of the source

May combine two claims.

At 10 months post-treatment, ICE mice had 6-fold fewer COX-positive myofibers, about half the capillary-to-fiber ratio, and loss of silencing at repetitive elements and transcription of retrotransposons, paralleling normal aging.

“At 10-months post-treatment, the ICE mice had 6-fold fewer COX-positive myofibers (Figure S5G and S5H), about half the capillary to fiber ratio (Figure 4G and 4H), a loss of silencing at repetitive elements and the transcription of retrotransposons, paralleling normal aging (Das et al., 2019; De Cecco et al., 2019; Oberdoerffer et al., 2008; Wenz et al., 2009) (Figure S5I).”

• cites Das et al., 2019Not matched

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

• cites De Cecco et al., 2019 sourceSupported
• cites Oberdoerffer et al., 2008 sourceNot checked — no full text of the source

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

Hippocampal function declines with age and is often measured by fear conditioning, which measures short-term memory.

“Hippocampal function, critical for spatial and memory consolidation, declines with age (Gallagher et al., 2010; Miller and O’Callaghan, 2005; Park and Reuter-Lorenz, 2009) and is often measured by fear-conditioning, which measures short-term memory.”

• cites Gallagher et al., 2010 sourceSupported
• cites Miller and O’Callaghan, 2005 sourceNot checked — no full text of the source
• cites Park and Reuter-Lorenz, 2009 sourceSupported

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

Molecular hallmarks of muscle aging include reduced ATP, mitochondrial DNA, and alterations in subsarcolemmal and intermyofibrillar morphology.

“Molecular hallmarks of muscle aging include reduced ATP, mitochondrial DNA, and alterations in subsarcolemmal and intermyofibrillar morphology (Demontis et al., 2013; Leduc-Gaudet et al., 2015).”

• cites Demontis et al., 2013 sourcePartly supported

  The section “Decline in mitochondrial function and metabolic homeostasis with age” (page unavailable) reports lower ATP levels in aged muscle, but describes increased mitochondrial DNA mutations rather than reduced mitochondrial DNA. The Fig. 2 caption in “Intrinsic defects leading to loss of muscle function during aging” (page unavailable) describes abnormal mitochondrial size and distribution, but does not specify subsarcolemmal and intermyofibrillar morphology. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Leduc-Gaudet et al., 2015 sourcePartly supported

  The Discussion reports altered subsarcolemmal and intermyofibrillar mitochondrial morphology in aged mouse muscle. The copy does not establish reduced ATP or mitochondrial DNA: the Introduction mentions impaired energetics, while the Results report no significant change in ATP synthase subunit content, which is not a measure of ATP levels. Searches for mitochondrial DNA and reduced ATP found no supporting passage. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

May combine two claims.

As mice age, they undergo characteristic physical and physiological changes, including alopecia, hair greying, kyphosis, decreased body weight, motion in dark phase, and reduced respiratory exchange ratio.

“As mice age, they undergo characteristic physical and physiological changes, including alopecia, hair greying, kyphosis, decreased body weight, motion in dark phase, and a reduced respiratory exchange ratio (RER) (Ackert-Bicknell et al., 2015; Harkema et al., 2016; Houtkooper et al., 2011; Koks et al., 2016).”

• cites Ackert-Bicknell et al., 2015 sourcePartly supported

  The Introduction reports age-related declines in physical function, and Commentary > Critical Parameters identifies sudden weight loss and hunched posture as signs of terminal decline in aging mice. Our copy does not substantiate alopecia, hair greying, kyphosis, a change in dark-phase motion, or reduced respiratory exchange ratio; it also does not present weight loss as a universal consequence of aging. The cited passages have no page numbers in our copy. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites Harkema et al., 2016 sourceNot checked — no full text of the source
• cites Houtkooper et al., 2011 sourcePartly supported

  The Results section (“Metabolic characterization and physical performance”; page not provided) reports lower respiratory exchange ratio and less running during the night phase in old mice. It reports that old mice were significantly heavier, not lighter, and does not report alopecia, hair greying, or kyphosis. (LLM, conf=0.92)

  Experimental — accuracy not yet measured

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

Astrocytes and microglia become hyper-activated with age.

“Astrocytes and microglia, mediators of the innate immune response of the central nervous system, become hyper-activated with age (Baruch et al., 2014; Norden and Godbout, 2013).”

• cites Baruch et al., 2014 sourcePartly supported

  The passage discussing Fig. 4F–G reports that blocking IFN-I in aged mice markedly decreased astrogliosis and microgliosis, which the authors describe as age-related neuroinflammation. This supports glial activation in aged mice, but the cited passage does not directly show that astrocyte and microglial activation increases from young to old age. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites Norden and Godbout, 2013 sourcePartly supported

  The section “Astrocytes also have a more inflammatory profile with age” reports age-related changes resembling an activated astrocytic profile. The Abstract and “Innate immune challenge leads to prolonged and exaggerated neuroinflammation” describe aged microglia as primed, with hyper-activation occurring following immune challenge. The claim omits that condition and overstates what the review says about activation with age alone. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

Exercise endurance, strength, muscle mass, vascularization, and mitochondrial function decrease in skeletal muscle with age.

“Exercise endurance, strength, muscle mass, vascularization, and mitochondrial function decrease in skeletal muscle with age (Das et al., 2019; Demontis et al., 2013) (Figure 4A).”

• cites Das et al., 2019Not matched

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

• cites Demontis et al., 2013 sourcePartly supported

  The Introduction reports age-related losses of muscle mass and strength, and its Fig. 1 caption describes decreased capillarization in aging mammalian muscle. The section ‘Decline in mitochondrial function and metabolic homeostasis with age’ reports mitochondrial defects but notes conflicting findings about respiratory function in muscle fibers. The paper does not clearly state that exercise endurance declines with age, so it does not support the full list as written. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

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

H3K27ac is enriched in accessible regions, consistent with its known association with active promoters and enhancers.

“Based on ChIP-seq and Assay for Transposase Accessible Chromatin using sequencing (ATAC-seq), H3K27ac was enriched in accessible regions, consistent with the known association of H3K27ac with active promoters and enhancers (Heinz et al., 2015; Klemm et al., 2019).”

• cites Heinz et al., 2015 sourcePartly supported

  The Enhancer characteristics section (page unavailable) identifies H3K27ac as a feature of active enhancers and describes its correlation with cell-type-specific enhancer activity at an accessible locus. The passages do not establish its association with active promoters. The ChIP-seq and ATAC-seq enrichment result is presented as the citing authors’ own finding, not a result attributed to this review. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites Klemm et al., 2019 sourceNot checked — no full text of the source

May combine two claims.

These data imply that muscle of post-treated ICE mice exdifferentiate towards an immune signature, consistent with data from aged mouse tissues.

“These data imply that the muscle of post-treated ICE mice exdifferentiate towards an immune signature, consistent with data from aged mouse tissues (Benayoun et al., 2019).”

• cites Benayoun et al., 2019 sourceSupported

May combine two claims.

Repeated triggering of the RCM response disrupts youthful gene expression and releases retrotransposons, including those normally silenced by SIRT1 and SIRT6.

“Studies of these proteins indicate that repeated triggering of the RCM response disrupts youthful gene expression patterns and liberates retrotransposons, such as those silenced by SIRT1/SIRT6 (De Cecco et al., 2019; Oberdoerffer et al., 2008; Simon et al., 2019).”

• cites De Cecco et al., 2019 sourcePartly supported

  The Abstract (page unavailable) reports that LINE-1 retrotransposons become derepressed during cellular senescence, supporting the retrotransposon portion of the claim. The paper does not establish that repeated triggering of an RCM response causes this or disrupts youthful gene-expression patterns. Its References list names a study of SIRT6-mediated LINE-1 repression, but this paper does not show that SIRT1 or SIRT6 silences the elements it examined. (LLM, conf=0.87)

  Experimental — accuracy not yet measured
• cites Oberdoerffer et al., 2008 sourceNot checked — no full text of the source
• cites Simon et al., 2019 sourcePartly supported

  The Discussion reports elevated LINE1 expression in aged wild-type and SIRT6-deficient mice and suggests that loss of silencing or redistribution of chromatin modifiers such as SIRT1 and SIRT6 may explain its activation. It does not establish that repeated triggering of an RCM response disrupts youthful gene-expression patterns or causes retrotransposon release. The indexed passages provide section labels but no page numbers. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

May combine two claims.

In mammals, cellular identity is established prenatally and maintained by H3K27ac-enriched super-enhancers.

“In mammals, cellular identity is established prenatally and maintained by H3K27ac-enriched super-enhancers (Hnisz et al., 2013; Whyte et al., 2013).”

• cites Hnisz et al., 2013 sourcePartly supported

  The Results section, “Super-enhancers in many cell types” (page unavailable), says super-enhancers drive genes that control and define cell identity and identifies H3K27ac as a useful mark for finding them. The paper does not say that mammalian cellular identity is established prenatally or demonstrate that H3K27ac-enriched super-enhancers maintain it thereafter. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Whyte et al., 2013 sourcePartly supported

  The Results report that super-enhancers have substantially higher H3K27ac levels than typical enhancers, and the Discussion links super-enhancers to genes that establish and maintain cellular identity. The paper does not say that mammalian cellular identity is established prenatally; its statement about establishment and maintenance is conditional. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

May combine two claims.

While certain types of DNA damage mimic aging, including chemotherapy, radiation, smoking, and progeria, evidence to the contrary includes cloned animals or mice engineered to accumulate mutations having normal health and lifespans.

“While it is incontrovertible that certain types of DNA damage mimic aging, including chemotherapy, radiation, smoking, and progeria”

• Also stated in: § DISCUSSION · plain

  “evidence to the contrary includes the fact that cloned animals or mice engineered to accumulate mutations have normal health and lifespans (Narayanan et al., 1997; Wakayama et al., 2013).”

• cites Carrero et al., 2016 sourcePartly supported

  The sections “Defects in DNA repair, and premature ageing” and “Increased DNA damage and defective DNA repair” link DNA damage and impaired repair to ageing and progeroid syndromes, and mention susceptibility to ionizing radiation. The copy does not establish the claim’s specific examples of chemotherapy or smoking. Its “Mouse models of progeroid syndromes” section reports some mutant mice without accelerated senescence or altered lifespan, but does not support the broader assertion that mutation-accumulating or cloned animals have normal health and lifespans. Page numbers are unavailable in the indexed copy. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites Hofstatter et al., 2018 sourcePartly supported

  The Results and Discussion (pages unavailable) report an association between smoking and accelerated epigenetic age in normal breast tissue, supporting the smoking-related part of the claim. They do not establish that DNA damage causes aging-like changes, or support the claims about chemotherapy, radiation, progeria, cloned animals, or mutation-accumulating mice. The Methods mention chemotherapy and radiation only to exclude patients who had received them before surgery. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Horvath and Levine, 2015 sourcePartly supported

  The Discussion links HIV-induced DNA damage to epigenetic age acceleration only as a possible explanation, conditional on viral integration causing epigenomic instability. It does not establish that DNA damage mimics aging or address the listed examples of chemotherapy, radiation, smoking, and progeria. The cited copy also does not address the cloned-animal or mutation-accumulating-mouse claim in the supplied quote. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

May combine two claims.

Cyclic expression of Yamanaka factors Oct4, Sox2, Klf4, and Myc alleviates symptoms and extends lifespan of progeroid mice and other aspects of aging.

“The cyclic expression of Yamanaka factors Oct4, Sox2, Klf4 and Myc (OSKM) (Takahashi and Yamanaka, 2006) alleviates the symptoms and extends the lifespan of progeroid mice and other aspects of aging (Ocampo et al., 2016; Sarkar et al., 2020).”

• cites Takahashi and Yamanaka, 2006 sourceNot checked — no full text of the source
• cites Ocampo et al., 2016 sourceSupported
• cites Sarkar et al., 2020 sourcePartly supported

  The Abstract reports prior evidence that transient reprogramming improves aging hallmarks and extends lifespan in progeroid mice, and identifies Ocampo et al. as the source of the mouse finding. Sarkar et al.’s own Results concern other aspects of aging in cells treated with transient OSKMLN expression, not cyclic OSKM treatment or lifespan in mice. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

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

Studies of simple organisms indicate that loss of epigenomic information may be an underlying cause of aging in those species.

“Studies of simple organisms, such as budding yeast, worms and flies, indicate that a loss of epigenomic information, may be an underlying cause of aging in those species (Imai and Kitano, 1998; Jiang et al., 2013; Lu et al., 2014; Mills et al., 1999; Oberdoerffer et al., 2008).”

• cites Imai and Kitano, 1998Not matched

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

• cites Jiang et al., 2013 sourceSupported
• cites Lu et al., 2014Not matched

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

May combine two claims.

During normal aging, H3K9me3 levels in kidneys decrease and H3K36me2 levels in muscle increase.

“During normal aging, levels of H3K9me3 in kidneys decrease and H3K36me2 in muscle increase (Tvardovskiy et al., 2017).”

• cites Tvardovskiy et al., 2017 sourcePartly supported

  The Discussion table (page unavailable) lists H3K9me3 as decreasing with age in kidney, supporting the first part. The Results section on global H3 methylation (page unavailable) reports an increasing trend for H3K36me2 in heart, but the paper does not report this finding for muscle generally or for skeletal muscle. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Classified as probably not a substantial claim.

Spatial organization of chromatin, including long-range DNA interactions, plays a key role in controlling gene expression and maintaining cellular identity.

“The spatial organization of chromatin, including a variety of long-range DNA interactions, plays a key role in controlling gene expression and maintaining cellular identity (Gorkin et al., 2014; Kundu et al., 2017; Narendra et al., 2015; Schoenfelder et al., 2015; Zirkel et al., 2018).”

• cites Gorkin et al., 2014 sourceSupported
• cites Kundu et al., 2017 sourcePartly supported

  The Results (page unavailable) link changes in PRC1-associated chromatin domains to Hox gene derepression during differentiation. The Discussion (page unavailable) hypothesizes that these domains might help silence genes and maintain their silenced state; it does not establish that spatial organization plays a key role in maintaining cellular identity. (LLM, conf=0.84)

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

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims. The quote only partly states this claim.

ChIP-seq gene ontology analysis used GREAT and ChIP-Enrich, and only GO terms with five or more genes were kept.

“GO biological processes were ranked by HyperFdrQ and only GO terms made up of at least 5 genes were included. ChIP-seq data were also analyzed using ChIP-Enrich (Welch et al., 2014). GO terms with at least 5 genes were ranked by FDR.”

• cites Welch et al., 2014 sourceSupported

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

Variant calling from whole-genome sequencing adhered to Broad Institute best practices.

“The best practices recommended by the Broad Institute for variant calling were then followed (Van der Auwera et al., 2013).”

• cites Van der Auwera et al., 2013 sourceSupported

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

Peaks failing a signal-to-noise cutoff of p > 10^-5 and peaks in ENCODE blacklist regions were discarded.

“We removed peaks with significance (signal relative to noise) of p > 10−5. Peaks that fell into the ENCODE blacklist regions were removed (Landt et al., 2012).”

• cites Landt et al., 2012 sourceCould not verify

  The citation is attached to the sentence about removing peaks in ENCODE blacklist regions. The Peak calling section discusses significance thresholds and false positives, but the passages found in our copy do not mention blacklist regions or their removal (searched: “blacklist,” “blacklist regions,” and filtering problematic genomic regions). (LLM, conf=0.76)

  Experimental — accuracy not yet measured

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

Equal amounts of Drosophila melanogaster DNA were added to ChIP-seq samples, and dm6-to-mouse read ratios were compared to estimate total ChIP-seq signal.

“Equal amounts of D. Melanogaster DNA were spiked-into ChIP-seq samples. In addition to aligning to the mouse genome, we aligned reads to the D. Melanogaster dm6 genome (Cunningham et al., 2019). To provide a sense of total ChIP-seq signal strength, the proportion of reads aligning to the dm6 genome were compared to the proportion aligning to the mouse genome (Orlando et al., 2014).”

• cites Cunningham et al., 2019 sourceCould not verify

  Cunningham et al. is cited specifically for the D. melanogaster dm6 genome used in read alignment, not for the spike-in or read-ratio method. The Introduction and “Supporting annotation of all species” discuss Ensembl genome resources, but searches for “dm6” and “Drosophila melanogaster” found no passage identifying that assembly in our copy. (LLM, conf=0.74)

  Experimental — accuracy not yet measured
• cites Orlando et al., 2014 sourcePartly supported

  The Results describe using the proportion of reads mapping to a fixed Drosophila reference to assess changes in ChIP-seq signal relative to the test genome. But Orlando et al. tested human cells and aligned reads to human hg19 and Drosophila dm3, not mouse and dm6 (Discussion, “Library Construction, Sequencing, and Data Collection”). The cited paper supports the comparison’s rationale, not those specific genomes. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

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

Processing of ChIP-seq and ATAC-seq reads adhered to ENCODE/Roadmap guidelines, with small changes.

“The techniques described for processing the ChIP-seq and ATAC-seq reads are based on ENCODE/Roadmap guidelines with a few modifications (Gjoneska et al., 2015; Landt et al., 2012; Roadmap Epigenomics et al., 2015).”

• cites Gjoneska et al., 2015 sourcePartly supported

  The Materials and Methods sections on ChIP-seq read mapping and differential analysis say the procedures were consistent with ENCODE standards and describe an adjustment for limited sequencing depth. The paper does not describe ATAC-seq read processing or say that its read-processing procedures followed Roadmap guidelines. Page numbers are not available in our copy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Landt et al., 2012 sourceSupported
• cites Roadmap Epigenomics et al., 2015 sourcePartly supported

  Online Methods §1.2 describes the Roadmap consortium’s ChIP-seq read-mapping and processing procedures, supporting its use as a reference for ChIP-seq processing. That section covers ChIP-seq and DNase-seq, not ATAC-seq; the paper also cannot establish whether the citing authors followed those procedures with only a few modifications. No page number is available in our copy. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

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

Primer sequences were analyzed with an in silico PCR tool to determine how many genomic elements contribute to each amplification product.

“Primer sequences were analyzed using the UCSC genome browser in silico PCR tool (genome.ucsc.edu/cgi-bin/hgPcr) to determine the number of genomic elements that contribute to the amplification products (De Cecco et al., 2013).”

• cites De Cecco et al., 2013 sourceCould not verify

  The Experimental procedures section on PCR primer design refers readers to supplemental methods, while the Results say that some primers were empirically verified to amplify individual elements with single-copy kinetics. Neither passage says primer sequences were analyzed with the UCSC in silico PCR tool to count the genomic elements contributing to amplification products; searches for that tool and purpose found no such passage in our copy. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

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

rDNA instability and nucleolar size affect lifespan in lower organisms.

“Indeed, rDNA instability and nucleolar size influence the lifespan of lower organisms (Sinclair et al., 1997; Tiku et al., 2017).”

• cites Sinclair et al., 1997Not matched

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

• cites Tiku et al., 2017 sourceSupported

May combine two claims.

In the ICE system, accurate repair of DNA breaks drives aging-related changes at physiological, cognitive, and molecular levels, including epigenetic landscape erosion, cell exdifferentiation, senescence, and a faster DNA methylation clock.

“Using a system called “ICE” (Inducible Changes to the Epigenome), we find the act of faithful DNA repair advances aging at physiological, cognitive, and molecular levels, including erosion of the epigenetic landscape, cellular exdifferentiation, senescence, and advancement of the DNA methylation clock”

May combine two claims.

At 16 months of age, ICE mice had significantly less muscle mass, reduced endurance, greater post-exercise lactate, reduced grip strength, and other molecular hallmarks of muscle aging.

“At 16 months of age, ICE mice had significantly less muscle mass (Figure 4B), reduced endurance (Figure 4C), greater lactate levels post-exercise (Figure S5A), reduced grip strength (Figure S5B) and other molecular hallmarks of muscle aging (Figure 4D–4F and S5C–S5F).”

May combine two claims.

In post-treated ICE cells, regions with higher accessibility lost H3K27ac and those with lower accessibility gained it.

“In post-treated ICE cells, regions with higher accessibility lost H3K27ac and those with lower accessibility gained it (Figure S6H).”

May combine two claims.

Based on the two mouse clocks, the rate of epigenetic aging was about 50% faster in ICE mice than Cre controls (p < 0.0001), closely paralleling treated ICE fibroblasts.

“Based on the two mouse clocks, the rate of epigenetic aging was ~50% faster in the ICE mice than Cre controls (p < 0.0001), closely paralleling treated ICE fibroblasts (Figure 4K–M).”

May combine two claims.

ICE mice moved about 50% less in the dark phase and had impaired gait.

“ICE mice moved ~50% less in the dark phase (Figure 3B) with an impaired gait (Figure S4N–S4P).”

May combine two claims.

Erosion was seen across H3K27ac peaks genome-wide and in the top or bottom 40% quantile of H3K27ac signals.

“Erosion was also seen across H3K27ac peaks genome-wide and in the top or bottom 40% quantile of H3K27ac signals.”

The quote only partly states this claim.

In skeletal muscle, genes significantly dysregulated in ICE mice correlated with changes in old mice, including Cdkn1a, Myl4, Nlrc5, and Mrpl55.

“In skeletal muscle, genes that were significantly dysregulated in ICE mice correlated with changes in old mice (Figure 4I, 4J, S5M–S5O and Table S2), including Cdkn1a (Cyclin Dependent Kinase Inhibitor 1A or p21), a mediator of p53-mediated cellular senescence”

• cites Beggs et al., 2004 sourceNot checked — no full text of the source
• cites Choudhury et al., 2007 sourceNot checked — no full text of the source
• cites Welle et al., 2004 sourceNot checked — no full text of the source

May combine two claims.

During and after I-PpoI induction, there was no detectable change in cell cycle profile, apoptosis, or senescence in ICE MEFs.

“During and after I-PpoI induction, there was no detectable change in cell cycle profile, apoptosis, or senescence (Figure S1E–S1G).”

May combine two claims.

At 10 months post-treatment, ICE mice exhibited classic features of old age, including reduced body weight and fat mass independent of food intake, lower daytime RER, and decreased motion in the dark phase.

“At 10-months post-treatment, the ICE mice exhibited classic features of old age, including reduced body weight and fat mass, independent of food intake (Figure 2B–2E and S4A–S4E), a lower respiratory exchange ratio (RER) during the day (Figure 2F and S4F) and decreased motion in the dark phase (Figure 2G).”

May combine two claims.

At the rDNA, there was no evidence of residual cutting, changes in copy number, 28S expression levels, or protein synthesis.

“At the rDNA in particular, there was no evidence of residual cutting (Figure S2J), changes in copy number (Figure S2O), 28S expression levels (Figure S2P) or protein synthesis (S2M and S2N).”

May combine two claims.

GO analysis indicated that cell identity and developmental processes were disrupted in ICE cells; of the top 20 processes, half were involved in development.

“analysis of genes with significant increases in H3K27ac and H3K56ac and decreases in H3K27me3 by Gene Ontology (GO) analysis indicated that cell identity and developmental processes were disrupted in ICE cells. Of the top 20 processes, half of them were involved in development (Figure 5C, Table S5 and S6).”

May combine two claims.

The ICE:Cre ratio of H3K27ac signals was inversely correlated with basal H3K27ac signals, demonstrating that ICE treatment corrupted epigenetic information and increased Shannon entropy.

“The ICE:Cre ratio of H3K27ac signals was inversely correlated with basal H3K27ac signals (Figure 5B and S6J), demonstrating that the ICE treatment corrupted epigenetic information and increased Shannon entropy (Hannum et al., 2013).”

• cites Hannum et al., 2013 sourceNot checked — no full text of the source

May combine two claims.

There was no difference in mutation frequency at canonical or non-canonical I-PpoI recognition sites or across the genome.

“There was also no difference in mutation frequency at canonical or non-canonical I-PpoI recognition sites (Wittmayer et al., 1998) or across the genome (Figure S3G–S3I).”

• cites Wittmayer et al., 1998 sourceNot checked — no full text of the source

May combine two claims.

In the Barnes maze test, recall of ICE mice was about half that of Cre controls, similar to 24-month-old WT mice.

“In the Barnes maze test, a measure of long-term memory, the recall of ICE mice was about half that of Cre controls, similar to that of 24-month-old WT mice (Figure 3F and 3G).”

May combine two claims.

In skeletal muscle, liver, and kidney, the Tmem56 intron and the 28S I-PpoI site were cut during TAM treatment, and no cutting was detectable 1 and 10 months later.

“In skeletal muscle, liver and kidney, the Tmem56 intron and the 28S I-PpoI site were cut during TAM treatment and, as expected, no cutting was detectable 1- and 10-month later (Figure 1N and S3F).”

May combine two claims.

Validation sets had R2 = 0.915 for muscle and 0.944 for blood, demonstrating accurate age estimations using independent datasets.

“Validation sets had R2 = 0.915 and 0.944 for muscle and blood, respectively, demonstrating accurate age estimations using independent datasets.”

May combine two claims.

The authors could not detect changes in mutation frequency at the 28S rDNA, RNA levels, or overall translation efficiency in ICE MEFs.

“We could not detect changes in mutation frequency at the 28S rDNA (Figure S2A–S2C and 1K), RNA levels (Figure S2D and S2F), or overall translation efficiency (Figure S2K and S2L).”

May combine two claims.

ICE mice had thinner left ventricular posterior walls, implying possible dilated cardiomyopathy.

“The ICE mice had thinner left ventricular (LV) posterior walls, implying possible dilated cardiomyopathy (Figure S5K and S5L).”

May combine two claims.

At 120-168 hours post-treatment, ICE cells had lower Lamin B1 levels and increased indicators of cellular senescence.

“At later times post-treatment (120–168 hours), ICE cells had lower Lamin B1 levels (Figure S1J and S1K) and increased indicators of cellular senescence (Figure 1F–1H).”

May combine two claims.

After TAM addition, HA-I-PpoI was detected in the nucleus of ICE MEFs but not controls, and γH2AX foci reached a maximum of 4-fold above background after 24 hours with locus-dependent cutting.

“After the addition of TAM, HA-I-PpoI was detected in the nucleus of ICE cells, but not controls (Figure 1B), and the number of serine 139-phosphorylated H2AX (γH2AX) foci, a marker of DSBs, reached a maximum of 4-fold above background after 24-hours, with locus-dependent cutting (Figure 1C and 1D).”

May combine two claims.

Similar to hippocampi of aged mice, ICE mice had greater numbers of activated astrocytes (1.6x) and microglia (3.5x).

“Similar to the hippocampi of aged mice, ICE mice had greater numbers of activated astrocytes (1.6x) and microglia (3.5x) (Figure 3H–3K).”

May combine two claims.

Loci with higher H3K27ac peak intensities tended to lose the most signal and vice versa, consistent with genome-wide erosion of the H3K27ac landscape, with no apparent changes specifically at I-PpoI recognition sites.

“Loci with higher H3K27ac peak intensities tended to lose the most signal and vice versa, consistent with a genome-wide erosion of the H3K27ac landscape, with no apparent changes specifically at I-PpoI recognition sites (Figure S6I).”

May combine two claims.

Key features of skin aging were apparent in ICE mice, including subepidermal thinning and hair greying due to loss of KIT/CD117-positive melanocyte stem cells.

“Key features of skin aging were apparent, including subepidermal thinning and hair greying due to a loss of KIT/CD117-positive melanocyte stem cells (Matsumura et al., 2016; Nishimura et al., 2005) (Figure S4J–S4M).”

• cites Matsumura et al., 2016 sourceNot checked — no full text of the source
• cites Nishimura et al., 2005 sourceNot checked — no full text of the source

May combine two claims.

On the second day of fear conditioning, about 75% of young mice and about 40% of old mice froze, indicating reduced contextual recall, with a similar difference between Cre and ICE mice.

“On the second day, ~75% of the young mice and ~40% of old mice froze, indicating reduced contextual recall, with a similar difference between Cre and ICE mice (Figure 3C–3E).”

May combine two claims.

At 1 month post-treatment, there was no significant difference in frailty index between ICE and controls, but at 10 and 12 months ICE mice had higher frailty index resembling 24-month-old WT mice (p=0.0006 and <0.0001), along with kyphosis and loss of cortical bone thickness and trabecular bone density.

“At 1-month post-treatment there was no significant difference in FI between ICE mice and controls, but at 10- and 12-months the ICE mice had a higher FI resembling that of 24-month-old WT mice (p=0.0006 and < 0.0001, respectively) (Figure 2H), along with other features of aging, including kyphosis, loss of cortical bone thickness and trabecular bone density in the inner layer”

May combine two claims.

In ICE mice, lens opacity was greater and the number of RGC axons in the myelinated region was fewer than controls.

“In the ICE mice, lens opacity was greater (Figure S7N–S7P) and the number of RGC axons in the myelinated region were fewer than controls (Figure 7K–7N).”

May combine two claims.

H3K27ac ChIP-seq on skeletal muscle of 10-month post-treated ICE mice showed regions with lower H3K27ac in Cre vs ICE had strongest enrichment for muscle tissue signatures (p=9.0×10−8), while regions with higher H3K27ac were enriched for immune cell enhancers (p=9.3×10−28).

“Regions with lower H3K27ac (p < 0.01) in Cre vs. ICE showed the strongest enrichment for muscle tissue signatures (p=9.0×10−8) while regions with higher H3K27ac showed an enrichment for immune cell enhancers (p=9.3×10−28) (Figure 6H and S6M).”

May combine two claims.

From Hoxa1 to Hoxa6, H3K27ac and H3K56ac decreased, and from Hoxa9 to Hoxa13 they increased, with concomitant changes in H3K27me3 and mRNA levels.

“From Hoxa1 to Hoxa6, levels of H3K27ac and H3K56ac decreased and from Hoxa9 to Hoxa13 they increased, with concomitant changes in H3K27me3 and mRNA levels (Figure 5F and S6K).”

May combine two claims.

After ICE treatment, spleen super-enhancers in muscle were derepressed, gaining H3K27ac, as were the Class II MHC cluster and Nfkbid gene.

“After ICE treatment, spleen super-enhancers in muscle were derepressed, gaining H3K27ac (Figure 6J), as were the Class II major histocompatibility complex (MHC) cluster and the Nfkbid gene, a regulator of humoral immunity (Figure 6K).”

May combine two claims.

Consistent with in vitro aging forward and reversal data, 7/10 of top upregulated processes in RGCs of 12-month-old mice were involved in development, and 86% of nervous system developmental genes were restored by OSK.

“Consistent with the in vitro aging forward and reversal data, 7/10 of the top upregulated processes in the RGCs of 12-month-old mice were involved in development (Figure 7P), the majority of which (86% of nervous system developmental genes) were restored by OSK (Figure 7Q).”

May combine two claims.

Of the top 20 processes increased in ICE muscle, all were elevated in old WT mice, consistent with WT and ICE mice undergoing similar epigenetic changes.

“Of the top 20 processes that were increased in ICE muscle, all were elevated in old WT mice, consistent with WT and ICE mice undergoing similar epigenetic changes (Figure 6I).”

May combine two claims.

After chemical reprogramming of post-treated Cre and ICE cells into neurons, Neurod1 and Nefh were 8-15-fold more easily derepressed in ICE MEFs, coincident with increases in neuron count and neuronal marker TUJ1.

“To test this, we reprogrammed the post-treated Cre and ICE cells into neurons by chemical means (Li et al., 2015) and found that Neurod1 and Nefh were 8–15-fold more easily derepressed in the ICE MEFs (Figure 6D and 6E), coincident with increases in neuron count and the neuronal cell marker TUJ1 (Figure 6F and 6G).”

• cites Li et al., 2015Not matched

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

May combine two claims.

Expression of OSK in treated ICE cells reversed age-associated mRNA changes, including those for Hmgb, Chaf1, Hoxa, and canonical histone genes, similar to aged fibroblasts, and did not erase cell identity or induce Nanog.

“Expression of OSK in treated ICE cells reversed age-associated mRNA changes, including those for Hmgb, Chaf1, Hoxa and canonical histone genes (Figure 7A–7C and S7E), similar to the effect of OSK expression in aged fibroblasts (Figure 7D–7F and S7F), and did not go so far back in age as to begin to erase cell identity or induce the pluripotency gene Nanog (Figure S7A–S7E).”

May combine two claims.

H3K27ac-positive posterior Hoxa gene promoters (Hoxa 9-13) formed new contacts with active enhancers in an adjacent TAD, with concomitant increases in mRNA levels, consistent with weakened insulation and disordered promoter-enhancer communication.

“H3K27ac-positive posterior Hoxa gene promoters (Hoxa 9–13) formed new contacts with active enhancers in an adjacent TAD, with concomitant increases in mRNA levels consistent with weakened insulation and disordered promoter-enhancer (P-E) communication.”

May combine two claims.

Of genes with decreased H3K27me3, 4/10 of top GO processes and 6/10 tissue-specific transcriptional profiles were related to neuronal processes or neuronal tissue types.

“Of the genes with decreased H3K27me3, 4/10 of the top GO processes and 6/10 tissue-specific transcriptional profiles were related to neuronal processes or neuronal tissue types (Figure 6A and 6B).”

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

Each RRBS sample covered over 3.6 million CpG sites, with 2.7 million covered across all samples.

“More than 3.6 M CpG sites were covered in each sample and 2.7M were covered in all samples.”

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

A higher CpG coverage cutoff lowered how many clock sites entered the analysis.

“Increase of the threshold for the CpG sites coverage decreased the number of the clock sites included in the analysis.”

May combine two claims.

The authors observed no effects on rDNA mutations, rRNA levels, or protein translation.

“That said, we saw no effects on rDNA mutations, rRNA levels, or protein translation, and I-SceI, which does not cut the rDNA, generated a I-PpoI-like gene expression pattern.”

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

The RCM Hypothesis and Information Theory of Aging propose that eukaryotic aging results from loss of transcriptional networks and epigenetic information over time, driven by a conserved mechanism that evolved to co-regulate responses to cellular damage such as DSBs or crush injury.

“Our Relocalization of Chromatin Modifiers or “RCM” Hypothesis and subsequent Information Theory of Aging propose that aging in eukaryotes is due to the loss of transcriptional networks and epigenetic information over time, driven by a conserved mechanism that evolved to co-regulate responses to cellular damage, such as a DSB or a crush injury (Mills et al., 1999; Oberdoerffer et al., 2008; Oberdoe”

• cites Mills et al., 1999 sourceNot checked — no full text of the source
• cites Oberdoerffer et al., 2008 sourceNot checked — no full text of the source
• cites Sinclair and LaPlante, 2019Not matched

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

May combine two claims.

The data support the Information Theory of Aging, which holds that loss of epigenetic information is a reversible cause of aging.

“These data are consistent with the Information Theory of Aging, which states that a loss of epigenetic information is a reversible cause of aging.”

• AssumptionRests on an assumption: The I-PpoI-induced DNA breaks at the 20 genomic sites are a valid model of the DNA breaks that occur during normal mammalian aging.

May combine two claims.

In yeast, DSBs are a major driver of aging, and their repair requires epigenetic regulators Sir2, Hst1, Rpd3, Gcn5, and Esa1.

“A major driver in yeast is the DSB (Park et al., 1999), the repair of which requires epigenetic regulators Sir2, Hst1, Rpd3, Gcn5, and Esa1 (Martin et al., 1999; McAinsh et al., 1999; Mills et al., 1999; Tamburini and Tyler, 2005).”

• cites Park et al., 1999 sourceNot checked — no full text of the source
• cites Martin et al., 1999 sourceNot checked — no full text of the source
• cites McAinsh et al., 1999 sourceNot checked — no full text of the source

May combine two claims.

In yeast, chromatin-modifying proteins relocalize to DNA breaks over time, causing loss of epigenetic information and cell identity, which is a hallmark of yeast aging.

“In yeast, epigenetic information is lost over time due to the relocalization of chromatin modifying proteins to DNA breaks, causing cells to lose their identity, a hallmark of yeast aging.”

• Also stated in: § INTRODUCTION · boosted

  “We and others showed that relocalization of the silent information regulator complex (Sir2/3/4) away from silent mating-type loci to the unstable rDNA is the cause of sterility, a hallmark of yeast aging”

• cites Smeal et al., 1996 sourceNot checked — no full text of the source

May combine two claims.

The authors' data are consistent with mammalian aging being a software problem: corrupted epigenetic information that can be restored from an existing back-up copy.

“Our data are consistent with aging in mammals being the equivalent of a software problem, the result of corrupted epigenetic information that can be restored from an existing back-up copy.”

• AssumptionRests on an assumption: The I-PpoI-induced DNA breaks at the 20 genomic sites are a valid model of the DNA breaks that occur during normal mammalian aging.

May combine two claims.

The ICE system induces specific DNA breaks with no evidence of mutations or immediate deleterious effects.

“Thus, the ICE system induces specific DNA breaks with no evidence of mutations or immediate deleterious effects.”

May combine two claims.

I-PpoI recognizes the sequence CTCTCTTAA▼GGTAGC, which occurs at 20 targets in the mouse genome, 19 non-coding including rDNA, and none in mitochondrial DNA.

“I-PpoI recognizes the DNA sequence CTCTCTTAA▼GGTAGC (Monnat et al., 1999), which occurs at 20 targets in the mouse genome, 19 of which are non-coding, including the rDNA, and none of which occur in mitochondrial DNA (Berkovich et al., 2007).”

• cites Monnat et al., 1999 sourceNot checked — no full text of the source
• cites Berkovich et al., 2007 sourceNot checked — no full text of the source

May combine two claims.

By introducing DSBs without causing mutations, the authors provide evidence that the cell's reaction to damage and the resulting loss of epigenetic information drives mammalian aging forward.

“By introducing DSBs without causing mutations, we provide evidence that it is the cell’s reaction to damage and the resulting loss of epigenetic information that drives mammalian aging forward.”

• AssumptionRests on an assumption: The I-PpoI-induced DNA breaks at the 20 genomic sites are a valid model of the DNA breaks that occur during normal mammalian aging.

May combine two claims.

Based on yeast work, the authors proposed the RCM and Information Theories of Aging, which state that loss of epigenetic information driven in part by cellular repair events is a cause of aging.

“Based on yeast work performed in our lab and others, we proposed the RCM and Information Theories of Aging, which state that a loss of epigenetic information driven in part by cellular repair events, are a cause of aging (Oberdoerffer et al., 2008; Sinclair and Guarente, 1997; Sinclair and LaPlante, 2019; Sinclair et al., 1997).”

• cites Oberdoerffer et al., 2008 sourceNot checked — no full text of the source
• cites Sinclair and Guarente, 1997Not matched

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

• cites Sinclair and LaPlante, 2019Not matched

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

May combine two claims.

In a parallel study, the authors found it possible to safely reverse DNA methylation age and gene expression patterns of old and damaged neurons to cure blindness in mice, a process requiring DNA demethylation.

“In a parallel study to this one, we found it possible to safely reverse DNAme age and gene expression patterns of old and damaged neurons to cure blindness in mice, a process requiring DNA demethylation (Lu et al., 2020).”

• cites Lu et al., 2020Not matched

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

May combine two claims.

In mammals, the efficiency of double-strand break repair tracks with lifespan, while nucleotide excision repair and base excision repair do not.

“The fact that the DSB repair efficiency of mammals correlates with lifespan, but not other types of repair, such as NER and BER, indicates that DSB are a special case, perhaps because only severe threats to cell survival disrupt the epigenome sufficiently to cause aging (Brown and Stuart, 2007; Tian et al., 2019).”

• cites Brown and Stuart, 2007 sourceNot checked — no full text of the source
• cites Tian et al., 2019 sourceNot checked — no full text of the source

May combine two claims.

Together with neuronal priming of ICE cells and increased EMT in ICE kidney, the authors conclude that induction of non-mutagenic DSBs accelerates the DNA methylation clock and age-related changes to chromatin, gene expression, and cellular identity.

“Together with the neuronal priming of ICE cells and increased EMT in the ICE kidney, we conclude that the induction of non-mutagenic DSBs accelerates the DNAme clock and age-related changes to chromatin, gene expression, and cellular identity.”

May combine two claims.

In this paper, the authors show that non-mutagenic DSB repair causes the epigenetic landscape to erode and cells to drift into adjacent Waddington valleys, where they malfunction and cause aging, consistent with the Information Theory of Aging.

“In this paper, we show that non-mutagenic DSB repair causes the epigenetic landscape to erode and cells to drift into adjacent Waddington valleys, where they malfunction and cause aging, consistent with the Information Theory of Aging.”

• AssumptionRests on an assumption: The I-PpoI-induced DNA breaks at the 20 genomic sites are a valid model of the DNA breaks that occur during normal mammalian aging.

May combine two claims.

Wnt and Hox genes direct embryonic development and also promote double-strand break repair.

“Indeed, Wnt and Hox genes direct embryonic development and also facilitate DSB repair (Feltes, 2019; Liu et al., 2007; Rubin et al., 2007; Ye et al., 2007; Zhao et al., 2018).”

• cites Feltes, 2019 sourceNot checked — no full text of the source
• cites Liu et al., 2007 sourceNot checked — no full text of the source
• cites Rubin et al., 2007 sourceNot checked — no full text of the source

May combine two claims.

Repeated activation over time causes chromatin factors to disperse and long-range DNA interactions to change.

“Over time, however, repeated activation causes chromatin factors to scatter and long-range DNA interactions to change over time, thereby aging cells and tissues.”

Classified as probably not a substantial claim.

DNase digestion of RNA was monitored with reverse-transcriptase-omitted controls and repeated until those controls showed no γ-satellite signal.

“The effectiveness of the digestion was assessed using controls that omitted reverse transcriptase (RT). Digestion with DNase was repeated until the control lacking RT was negative for γ-satellite sequences.”

May combine two claims.

The study did not perform single-cell epigenomic analyses and induced the ICE system throughout the body rather than in a tissue-specific way, so it cannot draw conclusions about cell autonomy.

“We did not perform epigenomic analyses at the single cell level and induced the ICE system in the whole body but not in a tissue-specific manner, so cannot make conclusions about cell autonomy.”

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

ChIP-seq analyses omitted sex chromosomes because the MEF sexes were inconsistent.

“Sex chromosomes were excluded from analyses due to the inconsistency of the sexes of the MEFs.”

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

Mitochondrial network, size, and number were quantified blindly using FUJI ImageJ.

“Mitochondrial network, size and number were quantified blindly using FUJI ImageJ.”

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

To reduce non-specific binding to beads, diluted chromatin was pre-cleared with Dynabeads protein A/G for 1 h at 4°C before antibody incubation.

“To reduce non-specific binging to beads, the diluted chromatin was mixed with Dynabeads protein A/G for 1 h at 4 °C and the beads were removed prior to incubating chromatin with 2 ug of the appropriate antibodies with Dynabeads protein A/G.”

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

ComBat from the SVA package in R was used to normalize the RRBS data.

“Data were normalized using ComBat from the SVA package in R.”

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

DNA methylation clock analysis used only CpG sites present in every sample; this left 89 of 90 blood clock sites covered.

“Only CpG sites covered in all samples were considered for DNA methylation clocks application. This resulted in 89 out of 90 sites being covered for the blood DNA methylation clock.”

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

Pearson correlations of Hi-C matrices were used for principal component analysis, and the first eigenvector's sign was flipped to match GC content.

“Matrix pearson’s correlations were calculated in R v 3.6.1 (Bunn, 2008, 2010) and used to perform a principal component analysis. The sign of the first eigenvector for each chromosome was adjusted to correlate with GC content, and were smoothed with loess smoothing using 1 megabase windows.”

• cites Bunn, 2008, 2010Not matched

  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).

Repetitive element qPCR used tissue from six animals, each analyzed in triplicate.

“Tissue from 6 individual animals was analyzed in triplicate.”

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

Picard's MarkDuplicates was used to remove duplicate reads prior to germline variant calling.

“The sorted binary alignments underwent post-processing to remove duplicates via Picard’s MarkDuplicates (v.2.01; http://broadinstitute.github.io/picard) before germline variants were identified using Genome Analysis Toolkit (GATK; v. 3.7) HaplotypeCaller (McKenna et al., 2010).”

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

MEF ChIP-seq differential peaks were called using an adjusted p-value cutoff below 0.01.

“For MEF experiments, we used a stringent threshold of adjusted p < 0.01.”

Probably not a substantial claim: may not pass S3 (load-bearing). The quote only partly states this claim.

Reads of low quality or with mapping quality under 20 were discarded from ChIP-seq/ATAC-seq data.

“Low quality reads and reads (q < 20) that did not map to the genome were removed.”

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

The study did not identify which chromatin factors are relocalized and did not examine chromatin contacts in vivo.

“In this study, we did not determine which chromatin factors are relocalized, nor did we study chromatin contacts in vivo.”

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

All primers were validated by serial dilution of cDNA to confirm quantitative amplification of their targets.

“All primers were tested with serial dilutions of cDNA to ensure they amplified their target sequences quantitatively.”

Probably not a substantial claim: may not pass S3 (load-bearing). The quote only partly states this claim.

Scorers were blinded to experimental groups when quantifying optic nerve axons.

“Scorers were blinded to experimental groups.”

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

For Hi-C, intra-chromosome contact frequencies were divided by the mean number of contacts seen at the same distance.

“Pairwise interaction frequencies were binned in 50-kb nonoverlapping windows and intra-chromosome interaction frequencies were normalized by dividing each interaction by the average number of interactions observed with that distance.”

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

Because RRBS libraries have low complexity, 10-20% phiX was added as a spike-in.

“To compensate for the low complexity of RRBS libraries 10–20% of phiX was spiked in.”

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

For MEF ChIP, Drosophila S2R+ cells were added as a spike-in control at one-quarter the number of mouse cells and processed together.

“1/4 number of Drosophila S2R+ cells relative to mouse cells were added as a spiked-in control and combined cells were treated as a single sample during the rest of the procedures.”

Probably not a substantial claim: may not pass S4 (based).

Mammals possess a backup copy of youthful epigenetic information that can safely restore old tissue function.

“Fortunately, it is now apparent that mammals retain a backup copy of youthful epigenetic information that can safely restore the function of old tissues, akin to reinstalling software.”

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

The quality of RNA was checked with an Agilent Bioanalyzer 2100 and an RNA-nano chip.

“RNA integrity was determined using an Agilent Bioanalyzer 2100 and an RNA-nano chip.”

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

Since few muscle ChIP-seq peaks met the stricter significance cutoff, analysis was limited to peaks enriched at p < 0.01.

“For muscle ChIP-Seq experiments, very few peaks attained significance levels at that cutoff. Therefore, we restricted our analysis to looking at the group of peaks enriched at p < 0.01.”