Quiescent Hematopoietic Stem Cells Accumulate DNA Damage during Aging that Is Repaired upon Entry into Cell Cycle

May combine two claims.

Young HSCs are not free of DNA damage; about 33% show evidence of single or double strand breaks, defined as greater than 10% DNA in comet tails.

“young HSCs are not impervious to DNA damage, with 33% showing evidence of single or double strand breaks, defined here as having greater than 10% DNA in their comet tails”

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

Most aged progenitor populations show significantly more DNA breaks than their young counterparts, although less than aged HSCs.

“the majority of these aged progenitors nonetheless showed evidence of significantly greater levels of DNA breaks than their young counterparts”

May combine two claims.

Significant DNA damage (>30% tail DNA) is more prevalent in old HSCs (42%) than young HSCs (9%), while modest damage (10-30% tail DNA) occurs at comparable frequencies.

“cells showing evidence of significant DNA damage (>30% Tail DNA) were much more prevalent in HSCs purified from old (42%) compared to young (9%) mice”

May combine two claims.

Old HSCs cultured in cytokine-rich media for 24 hours show significantly fewer damaged cells than old HSCs at steady state, and young and old HSCs have similar comet profiles after stimulation.

“young and old HSCs assayed 24-hours post culture stimulation showed very similar comet profiles, with the old cells showing significantly reduced numbers of damaged cells compared to steady state”

May combine two claims.

Donor-derived reconstitution from old HSCs is significantly lower than from young HSCs and shows a marked myeloid bias.

“examination of donor-derived reconstitution from old HSCs was significantly lower compared to the young with a marked myeloid bias lineage output”

• cites Beerman et al., 2013 sourceNot checked yet

  Not checked in the free run.

• cites Cho et al., 2008 sourceNot checked yet

  Not checked in the free run.

• cites Rossi et al., 2005 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Young downstream progenitor populations and young HSCs have comparable levels of DNA damage, with no significant differences in Olive tail moment.

“Analysis of the Olive tail moment of each downstream progenitor populations compared to HSCs isolated from young mice showed no significant differences”

May combine two claims.

Most DNA damage repair and response pathways are significantly up-regulated (p<0.001) in the majority of downstream progenitor populations compared with HSCs, and this attenuation in HSCs is largely age-independent.

“most pathways were significantly up-regulated (p<0.001) in the majority of downstream progenitors populations examined, and further that attenuation of these pathways in HSCs was largely age-independent”

May combine two claims.

Trp53 expression is significantly increased downstream of HSCs in young mice but significantly diminished downstream of HSCs in aged progenitors.

“Trp53 (which encodes p53) that showed statistically significant increased expression downstream of HSCs in young mice, whereas aged progenitors had significantly diminished expression downstream of HSCs”

May combine two claims.

The majority of DNA damage repair and response genes are not significantly age-regulated in HSCs, although some genes including Trp53 and Wrn are age-regulated.

“Although we found a number of age-regulated genes, which included Trp53 and Wrn, the majority of genes involved in DNA damage repair and response were not significantly age-regulated in HSCs”

May combine two claims.

Wrn expression is significantly decreased in most young progenitors compared with HSCs, but significantly increased in most old progenitors compared with HSCs.

“Wrn, which encodes the Werner syndrome homolog, showed an opposite pattern where the majority of progenitors from young mice showed significantly decreased expression compared to HSCs, whereas most progenitors from old mice exhibited significantly increased expression”

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

Old HSCs generally give rise to smaller colonies than young HSCs at day 12 (p<0.001), although cell types in colonies are comparable.

“the old HSCs generally gave rise to smaller colonies than the young HSCs (p<0.001”

May combine two claims.

Many DNA damage response and repair genes show reduced expression at early time points after stimulation, leading to significant down-regulation of many pathways at 3 hours, with old cells showing attenuation of almost all DNA response and repair pathways.

“many of the genes we examined exhibited reduced expression at the early time points post-stimulation, which led to many of the pathways displaying significant down-regulation at 3 hours, with the old cells showing attenuation of almost all DNA response and repair pathways”

May combine two claims.

All but one DNA damage response and repair pathway are not significantly different between young and aged HSCs; the NER pathway is significantly down-regulated in aged HSCs (p<0.01).

“all but one of the DNA damage response and repair pathways were not significantly different between young and aged HSCs (Figure S4B). The sole exception was the NER pathway (p<0.01), which was significantly down-regulated in aged HSCs.”

May combine two claims.

NHEJ is the only pathway not significantly differentially regulated between HSCs and progenitor cells, remaining mostly unchanged in both young and old mice.

“The only pathway that was not significantly differentially regulated in HSCs and progenitor cells was NHEJ, which remained mostly unchanged between HSCs and downstream progenitors in both young and old mice”

• cites Mohrin et al., 2010 sourceNot checked yet

  Not checked in the free run.

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

The skewed expression pattern between fetal and adult HSCs is specific to DNA damage response and repair genes, as global gene expression shows comparable numbers of up- and down-regulated genes.

“global analysis of all genes significantly differentially regulated between FL-HSC and young or old HSCs showed comparable numbers of genes up- and down-regulated”

May combine two claims.

Many of the 190 DNA damage response and repair genes show robust up-regulation starting at 12 hours and further increasing by 24 hours after stimulation into cycle, with most pathways significantly up-regulated by 24 hours.

“many of the 190 genes showed robust up-regulation starting at 12 hours and further increasing by 24-hours post-stimulation (Figure 7A-B) with most pathways being significantly up-regulated by 24 hours”

May combine two claims.

The HSC population as a whole does not comprehensively accrue strand breaks during aging, because over 30% of old HSCs show no evidence of strand breaks.

“the fact that over 30% of old HSCs showed no evidence of strand breaks indicates that the HSC population as a whole does not comprehensively accrue strand breaks during aging”

May combine two claims.

HSCs driven into cycle by transplantation repair accumulated DNA damage, resetting both young and old HSCs to a non-damaged status, and both young and old HSCs acquire similar DNA damage over the experiment.

“This suggests that HSCs driven into cycle by transplantation repaired their accumulated DNA damage resetting both the young and old HSCs to a non-damaged status, and further that both young and old HSCs acquired similar levels of DNA damage over the time course of the experiment”

May combine two claims.

DNA repair and response pathways are broadly attenuated in quiescent but not cycling HSCs, and exit from G0 and progression into cell cycle leads to induction of DNA damage repair and response genes.

“DNA repair and response pathways are broadly attenuated in quiescent but not cycling HSCs. They further suggest that HSCs exit from G0, and progression into the cell cycle, leads to induction of DNA damage repair and response genes.”

May combine two claims.

Downstream progenitors are more actively cycling, with a progressive increase in steady-state cycling rate from MPPs to oligo-potent progenitors.

“downstream progenitors are more actively cycling with a progressive increase in the steady-state cycling rate from MPPs (Figure S5) to oligo-potent progenitors (Passegue et al., 2005).”

• cites Passegue et al., 2005 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

HSCs reside largely in a dormant state, with >90% in quiescent G0 phase in both young and old mice, and enter cycle infrequently throughout adult life.

“HSCs reside largely in a state of dormancy, with >90% in the quiescent G0 phase of the cell cycle in both young and old mice (Figure S5) (Rossi et al., 2007b; Sudo et al., 2000), entering into cycle infrequently throughout the adult lifespan (Cheshier et al., 1999; Foudi et al., 2008; Wilson et al., 2008).”

• cites Rossi et al., 2007bNot checked yet

  Not checked in the free run.

• cites Sudo et al., 2000 sourceNot checked yet

  Not checked in the free run.

• cites Cheshier et al., 1999 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Old HSCs repair accrued strand breaks upon entry into cycle rather than undergoing cell death during the first 24 hours after stimulation.

“This data supports the idea that old HSCs repair accrued strand breaks upon entry into cycle, and do not undergo cell death during the first 24 hours post stimulation”

May combine two claims.

HSCs exhibit attenuated expression of most DNA damage repair and response pathways compared with downstream progenitors, largely independent of age, and HSC aging itself is not associated with significant differences in regulation of most of these pathways.

“HSCs exhibit attenuated expression of most DNA damage repair and response pathways in comparison to their downstream progenitors that is largely age-independent. Moreover, HSC aging itself is not associated with significant differences in the regulation of the majority of these pathways.”

May combine two claims.

Aged HSCs repair accrued DNA strand breaks upon entry into cycle but still display diminished proliferative potential.

“aged HSCs repair accrued DNA strand breaks upon entry into cycle, however even after strand break repair, aged HSCs displayed diminished proliferative potential”

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

Both myelodysplastic syndrome and acute and chronic myelogenous leukemias begin with nonlethal mutations in the HSC pool, often leading to expansion of mutant HSC clones and progression to leukemia.

“Both myelodysplastic syndrome (Pang et al., 2013) and acute and chronic myelogenous leukemias begin with nonlethal mutations in the HSC pool, often leading to successful expansion of mutant HSC clones at the expense of normal HSC, and which progress eventually to leukemia (Corces-Zimmerman et al., 2014; Jamieson et al., 2004; Jan et al., 2012)”

• cites Pang et al., 2013 sourceNot checked yet

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• cites Corces-Zimmerman et al., 2014 sourceNot checked yet

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• cites Jamieson et al., 2004 sourceNot checked yet

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

In two additional experiments, comets of over two thousand young and old HSCs purified using an alternative HSC marker combination (LKSCD150+CD48−) were assayed.

“In two additional experiments, we assayed comets of over two thousand young and old HSCs purified using an alternative cell surface marker combination for HSCs-LKSCD150+CD48−”

• cites Kiel et al., 2005 sourceNot checked yet

  Not checked in the free run.

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

HSCs subjected to gamma irradiation were assayed in parallel as a positive control.

“HSCs subjected to gamma irradiation were assayed in parallel as a positive control.”

Classified as probably not a substantial claim.

The study used alkaline comet assays to measure single and double strand breaks.

“we used alkaline comet assays (Figure S1) (Olive and Banath, 2006; Olive et al., 2001; Singh et al., 1988) to measure single and double strand breaks”

• cites Olive and Banath, 2006Not checked yet

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• cites Olive et al., 2001 sourceNot checked yet

  Not checked in the free run.

• cites Singh et al., 1988 sourceNot checked yet

  Not checked in the free run.

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

The two most reliable DNA damage measurements used were Olive moment and percent tail DNA.

“Analysis of the two most reliable DNA damage measurements, Olive moment and percent tail DNA (Kumaravel and Jha, 2006)”

• cites Kumaravel and Jha, 2006Not checked yet

  Not checked in the free run.

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

Comet assay analysis was performed on blinded files.

“Analysis was performed on blinded files using CometScore (TriTek Corp)”

Classified as probably not a substantial claim.

HSCs are the primary reservoir for mutation accrual that underlies diverse age-associated hematopoietic diseases such as MDS and AML.

“the idea that HSCs serve as the primary reservoir for mutation accrual underlying the development of diverse age-associated hematopoietic diseases such a MDS and AML is now widely accepted”

• cites Jan et al., 2012 sourceNot checked yet

  Not checked in the free run.

• cites Krivtsov et al., 2013 sourceNot checked yet

  Not checked in the free run.

• cites Rossi et al., 2007aNot checked yet

  Not checked in the free run.

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

For each experiment, all cells used in comparison analyses were assayed on a single high-throughput slide, so that all subsequent steps were consistent for those comparisons.

“For each experiment, all cells used in comparison analyses were assayed on a single high throughput slide, such that all of the following steps were consistent for those comparisons.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Other adult stem cell systems, such as those in brain, gastrointestinal tract, and skin, very likely follow the same energy-saving paradigm: they accumulate DNA damage while quiescent and repair it upon entry into the cell cycle.

“other adult stem cell tissue systems such as brain, gastrointestinal tract, skin, etc., will very likely follow the same energy saving paradigm- and accumulate DNA damage in quiescent stem cells, and repair these insults when it is necessary to enter cell cycle.”

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

The study did not assess the fidelity of repair or examine other types of DNA damage.

“we did not address the fidelity of repair or examine other types of DNA damage in this study”