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Relationship: 2811

Title

A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Oxidative Stress leads to Increase, DNA strand breaks

Upstream event

The causing Key Event (KE) in a Key Event Relationship (KER). More help

Oxidative Stress

Downstream event

The responding Key Event (KE) in a Key Event Relationship (KER). More help

Increase, DNA strand breaks

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes.Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name	Adjacency	Weight of Evidence	Quantitative Understanding	Point of Contact	Author Status	OECD Status
Deposition of energy leading to occurrence of cataracts	adjacent	Moderate	Low	Vinita Chauhan (send email)	Open for citation & comment	Under Review
Deposition of Energy Leading to Learning and Memory Impairment	adjacent	Moderate	Moderate	Vinita Chauhan (send email)	Open for citation & comment	Under Review
Deposition of energy leads to abnormal vascular remodeling	adjacent	High	Moderate	Vinita Chauhan (send email)	Open for citation & comment

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER. More help

Term	Scientific Term	Evidence	Link
human	Homo sapiens	Low	NCBI
rat	Rattus norvegicus	Low	NCBI
rabbit	Oryctolagus cuniculus	Low	NCBI
bovine	Bos taurus	Low	NCBI
mouse	Mus musculus	Low	NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help

Sex	Evidence
Unspecific	Low
Male	Low

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER. More help

Term	Evidence
Adult	Low
Not Otherwise Specified	Low

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

Oxidative stress is an event that involves both a reduction in free radical scavengers and enzymes, and an increase in free radicals (Brennan et al., 2012). Oxidative stress needs to be maintained within an organism to avoid an excess of damage to biological structures, such as DNA. A redox homeostasis between the radicals and the scavengers is necessary. Between reactive oxygen species (ROS) and reactive nitrogen species (RNS), collectively known as RONS, ROS is particularly significant to oxidative damage and disease states. Radicals such as singlet oxygen and hydroxyl radical are highly unstable and will react with molecules near their generation point, while radicals such as H2O2 are more stable and membrane permeable, meaning they can travel further to find electrons (Spector, 1990). Since DNA is mainly found in nucleus, ROS needs to reach the nucleus to induce breaks. Hydroxyl radicals, in addition to being highly reactive, are capable of causing DNA damage (Halliwell et al., 2021; Engwa et al., 2020). The regulation of these radicals is achieved by the antioxidant defense response (ADR), which includes enzymatic and non-enzymatic processes. The ADR is recruited to manage RONS levels, with antioxidants such as superoxide dismutase (SOD) functioning as the first line of defense (Engwa et al., 2020). These antioxidants act as scavengers to oxidants, reacting with them before reaching other structures within the cell such as DNA strands (Cabrera et al., 2011; Engwa et al., 2020). The backbone of DNA can fragment upon sustained exposure to ROS (Uwineza et al., 2019; Cannan et al., 2016). Due to low oxidation potentials, adenine and guanine are the DNA bases more prone to oxidation, with oxidation potentials (normal hydrogen electrode) at pH 7 of 1.3 eV and 1.42 eV compared to the 1.6 eV and 1.7 eV of cytosine and thymine (Fong, 2016; Halliwell et al., 2021; Poetsch, 2020). In fact, certain radicals even target guanine in a selective fashion, including carbonate anion radical (CO3•-) and singlet oxygen (1O2) (Halliwell et al., 2021).

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER. For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings. Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

The strategy for collating the evidence to support the relationship is described in Kozbenko et al 2022. Briefly, a scoping review methodology was used to prioritize studies based on a population, exposure, outcome, endpoint statement.

Evidence Supporting this KER

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help

Overall Weight of Evidence: Moderate

Biological Plausibility

Addresses the biological rationale for a connection between KEupstream and KEdownstream. This field can also incorporate additional mechanistic details that help inform the relationship between KEs, this is useful when it is not practical/pragmatic to represent these details as separate KEs due to the difficulty or relative infrequency with which it is likely to be measured. More help

The biological plausibility of the relationship between increased oxidative stress leading to increased DNA double strand breaks (DSBs) is highly supported by the literature. Evidence was collected from studies conducted using in vitro lens epithelial cell models and derived from humans, bovine and germ line cells (Spector, 1990; Stohs, 1995; Aitken et al., 2001; Spector, 1995). As this evidence is derived from studies using a human cell model it limits the ability to compare between different taxonomies (Ahmadi et al., 2022; Cencer et al., 2018; Liu et al., 2013; Meng et al., 2021; Smith et al., 2015; Zhou et al., 2016). Other evidence comes from human-derived and rodent models of neuronal and endothelial cells (Cervelli et al., 2014; El-Missiry et al., 2018; Huang et al., 2021; Sakai et al., 2017; Ungvari et al., 2013; Zhang et al., 2017).

ROS that are generated specifically as a result of radiation are highly localized, increasing the likelihood of clustered regions of damage. Naturally generated ROS are more widespread and as a result less capable of generating clusters of damage. ROS will act on DNA bases to oxidize or delete them from the sequence, which create nicks on the strand (Cannan et al., 2016). This damage can occur to any DNA base but bases such as guanine and adenine are most vulnerable due to their low oxidation potentials (Fong, 2016). The mechanism through which the strand break occurs is a result of base excision repair (BER) happening at multiple sites that are too close together, resulting in the spontaneous conversion to DSBs prior to completion of repair. ROS damage to bases clustered together means that multiple sites of BER are happening very close together and while the strand may be able to support the damaged area for one repair, concurrent repairs make surrounding areas more fragile and the strand breaks at the nick sites are under added strain (Cannan et al., 2016). Endogenous damage to DNA as a result of radicals appears over time and mainly as isolated lesions, a pattern understood to be due to the diffusion of the radicals resulting in homogenous distribution patterns. This differs from the specific situations where radiation acts as the stressor to increase oxidative stress, as the radiation track will be highly localized and form radicals within that hit space. This leads to non-homologous lesions and clustered damage to the DNA (Ward et al., 1985).

Empirical Evidence

Provides specific (citable) evidence that supports the idea of a change in the upstream KE (KEupstream) leading to, or being associated with, a subsequent change in the downstream KE (KEdownstream), assuming the perturbation of KEupstream is sufficient. More help

This relationship is well supported through empirical evidence from studies using stressors such as H2O2, photons, γ- and X-ray, which cause an increase in markers of oxidative stress such as ROS-generating enzymes (lactate dehydrogenase, LDH), and a decrease in free radical scavengers, resulting in DNA strand fragmentation. These studies include both in vivo and in vitro human lens epithelial cells (LECs), mouse, rat and rabbit models, including neuronal cells lines and endothelial cells (Ahmadi et al., 2022; Cencer et al., 2018; Cervelli et al., 2014; El-Missiry et al., 2018; Huang et al., 2021; Liu et al., 2013; Meng et al., 2021; Spector et al., 1997; Ungvari et al., 2013; Zhang et al., 2017; Zhou et al., 2016; Sakai et al., 2017).

Dose/Incidence Concordance 

There is high evidence to support a dose concordance between oxidative stress and DNA strand breaks. One in vitro study demonstrated that when ROS levels in LECs are 10% above control following 0.5 Gy gamma ray exposure, DNA strand breaks increased 15-20% above control (Ahmadi et al., 2021). Another study with ultraviolet (UV)B radiation demonstrated higher ROS levels after exposure to 0.14 J/cm2 on in vitro LECs as compared to a lower dose exposure (0.014 J/cm2) for the same time. This corresponded to DNA strand break levels also increasing following high dose rate exposure, but not with the low dose exposure (Cencer et al., 2018).

A 30 µM of H2O2 treatment of in vitro LECs is associated with a 1.4x increase in lactate dehydrogenase (LDH) and 55% more DNA strand breaks (Liu et al., 2013; Smith et al., 2015). Following exposure of in vitro LECs to 50 µM H2O2, increased ROS levels, 4x for LDH, and decreased antioxidant levels, 2x control for GSH-Px and SOD, are associated with a 3x increase in γ-H2AX, a marker of DNA strand breaks (Meng et al., 2021). SOD and GSH decreased by 2-fold following 100 µM H2O2 exposure on LECs with an in vitro model (Zhou et al., 2016). At 125 µM H2O2 intact DNA can be reduced to near 1% of pre-treatment levels for in vitro LECs (Spector et al., 1997). Following 400 µM H2O2 LDH increased to 1200% of control in neuroblastoma cells (Feng et al., 2016) and DNA strand breaks increased to over 150% of control in in vitro LECs (Li et al., 1998).

Exposure of in vitro mouse hippocampal neuronal cells (HT22 cell line) to 10 Gy of X-irradiation resulted in a 5x increase in ROS generation and 3x increase in γ-H2AX (Huang et al., 2021). Another study exposed the same cell line to 8 and 12 Gy of X-irradiation and found a ~2x increase in ROS at 8 Gy and a 4.4x and 3.2x increase in phosphorylation of ataxia telangiectasia mutated (ATM) and γ-H2AX, respectively, 30 minutes after 12 Gy (Zhang et al., 2017). A separate study exposed adult male rats to 4 Gy of γ-irradiation and found 2x increase in 4-hydroxy-2-nonenal (4-HNE) (lipid peroxidation marker) and 3x increase in protein carbonylation. Glutathione reductase decreased by approximately 5x, whereas glutathione and glutathione peroxidase levels decreased by approximately 3x each. Tail DNA %, tail length and tail moment (DNA strand break parameters) increased by approximately 2x, 3x and 6x, respectively (El-Missiry et al., 2018).

Endothelial cells exposed to irradiation also demonstrated the relation between oxidative stress and DNA strand breaks. Rat cerebromicrovascular endothelial cells (CMVECs) exposed to 8 Gy 137Cs gamma rays showed increased cellular peroxide production and mitochondrial oxidative stress. Tail DNA content indicating DNA damage was also increased from 0 to 45% (Ungvari et al., 2013). Human umbilical vein endothelial cells (HUVECs) were irradiated with single (0.125, 0.25, 0.5 Gy), or fractionated (2 × 0.125 Gy, 2 × 0.250 Gy) doses of X-rays. Intracellular ROS production increased in a dose-dependent manner following 0.125, 0.25, 0.5 Gy, and γ-H2AX foci positive cells were observed at all doses (Cervelli et al., 2014). Human aortic endothelial cells (HAECs) exposed to 100µM H2O2 showed 3.7-fold increase in intracellular ROS and a 3.4- and 4.7-fold increase in γ-H2AX and p-ATM, respectively (Sakai et al., 2017).

Time Concordance

There is low evidence to support a time concordance between oxidative stress to strand breaks on DNA. Non-protein-thiol levels, an antioxidant, in in vitro LECs decreased to near zero by 30 minutes post-exposure to 300 µM H2O2, before recovering to 70% of control by 120 minutes. At 60 minutes post-exposure to 125 µM H2O2 there was a start to a divergence from control level DNA fragmentation, one that increased logarithmically, with the treated group having a 14~18% reduction in intact DNA by 9 h post-exposure (Yang et al., 1998). Time response information is difficult to monitor for DNA strand breaks because repair will occur, reducing the number of breaks over time. At 0 minutes post in vitro exposure to 40 µM H2O2 LECs had ~145% of control level DNA strand breaks but that number dropped to ~105% by 30 minutes post-exposure (Li et al., 1998).

Essentiality 

Oxidative stress has been found to increase levels of DNA strand breaks above background levels (Li et al., 1998; Liu et al., 2013; Cencer et al., 2018; Ahmadi et al., 2022; El-Missiry et al., 2018; Huang et al., 2021; Cervelli et al., 2017; Sakai et al., 2017). It has been shown that inhibition of oxidative stress leads to a reduction in DNA strand breaks. Sulforaphane (SFN) is an isothiocyanate, which provides chemical protection against ROS by activating the release of enzymatic scavengers. When SFN was added to in vitro LECs exposed to 30 µM H2O2, LDH decreased to near unexposed cell levels from the 1.4x control level without SFN. This LDH drop was associated with reducing the levels of DNA strand breaks induced by oxidative stress almost 3-fold as compared to cells without SFN (Liu et al., 2013). In another study, intact DNA levels were returned to control when treated with µPx-11 (peroxidase that breaks down H2O2), following exposure to 125 µM H2O2. This was a near 100% recovery compared to the drop seen in LECs that did not contain µPx-11 (Spector et al., 1997).

Within the brain of Wistar rats, epigallocatechin-3-gallate (EGCG) ameliorated radiation-induced increases in lipid peroxidation and protein carbonylation, as well as decreases in glutathione (GSH), glutathione peroxidase (GPx) and glutathione reductase (GR) and reverted the levels back to those similar to controls. DNA strand break parameters also returned to those similar to controls after treatment with EGCG (El-Missiry et al., 2018). Similar effects were also shown in another study using treatment mesenchymal stem cell-conditioned medium in mouse hippocampal cells exposed to 10 Gy of X-irradiation (Huang et al., 2021).

HUVECs pretreated with the antioxidant mixture RiduROS blunted ROS generation in a concentration-dependent manner by 65% ± 5.6% and 98% ± 2%, at 0.1 and 1 μg/mL, respectively, compared with cells irradiated without pretreatment. Low-dose irradiation also increased DSB-induced γ-H2AX foci compared with control cells and 24 h of RiduROS pretreatment reduced the γ-H2AX foci number by 41% (Cervelli et al., 2017). Additionally, HAECs treated with eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) found significantly reduced intracellular ROS at 100µM, as well as reduced γ-H2AX foci formation by 47% and 48% following EPA and DHA treatment respectively. (Sakai et al., 2017).

Uncertainties and Inconsistencies

Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

N/A.

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references. More help

There is limited evidence demonstrating this relationship across different life stages/ages or sexes (Cencer et al., 2018; Li et al., 1998).

Modulating Factors	MF Details	Effects on the KER	References
Age	Reduced antioxidant capacities have been linked to aged lenses (in humans >30 years old). The development of a chemical barrier between the cortex and the nucleus is partially responsible, as it prevents GSH from protecting aged lens cells from ROS.	Prevention of RONS-mediated damage is primarily achieved by antioxidants, so a lowered capacity would likely lead to reduced damage mitigation abilities. 78% of lens over 30 had a low level of GSH in the center compared to 14% of lens under 30. Lens epithelial cells have an associated 3-fold increase in γ-H2AX (marker of DNA damage) when GSH-PX decreases by 2-fold.	Taylor & Davies, 1987; Cabrera & Chihuailaf, 2011; Quinlan & Hogg, 2018; Sweeney & Truscott, 1998; Meng & Fang, 2021
Free radical scavengers	ROS-scavengers are essential components of the body’s natural defense against oxidative damage. Increased ROS production leads to increased incidence of electron donation by scavengers, thus reducing the overall level of free radical scavengers available to deal with ROS.	Isothiocyanates, such as sulforaphane (SFN), activate the release of more enzymatic scavengers. When SFN was added to in vitro LECs, LDH decreased to near unexposed cell levels and was associated with 3.3x less DNA strand breaks compared to the non-SFN cells following stressor exposure. Epigallocatechin-3-gallate (EGCG) also has antioxidant properties and was shown to alleviate radiation-induced increases in oxidative stress and DNA strand breaks within rat hippocampi.	Taylor et al., 1987; Cabrera et al., 2011; Liu et al., 2013; El-Missiry et al., 2018
Media	Mesenchymal stem cell-conditioned medium (MSC-CM), which has self-renewal, differential and proliferation capacities.	MSC-CM treatment has also been shown to improve ROS levels and decrease radiation-induced DNA strand breaks within mouse hippocampal neuronal cells.	Huang et al., 2021

Quantitative Understanding of the Linkage

Captures information that helps to define how much change in the upstream KE, and/or for how long, is needed to elicit a detectable and defined change in the downstream KE. More help

The following tables provide representative examples of the relationship, unless otherwise indicated, all data is significantly significant.

Dose Concordance

Reference	Experiment Description	Result
Cencer et al., 2018	In vitro, human LECs exposed to UVB and tested for 120 min post exposure with fluorescent probes to detect ROS production and mitochondrial superoxide, and tetramethylrhodamine-dUTP (TMR) red assay to detect strand breaks.	Both ROS and DNA strand breaks were increased by both 0.014 J/cm2 and 0.14 J/cm2 UVB radiation. At 0.014 J/cm2, cellular ROS increased a maximum of 15 fluorescence units above the control at 5 min post-UVB, while DNA strand breaks increased about 115 fluorescence units above the control at this time. At 0.14 J/cm2, cellular ROS increased a maximum of about 35 fluorescence units above the control at 90 min post-UVB, while mitochondrial superoxide increased about 30 fluorescence units above the control and DNA strand breaks increased about 125 fluorescence units above the control at this time.
Ahmadi et al., 2021	In vitro, human LECs exposed to 0.065-0.3 Gy/min gamma radiation, with dihydroethdium (DHE) fluorescent probes to measure ROS levels and comet assay to measure strand breaks.	Human LECs exposed in vitro to 0.1 - 0.5 Gy gamma rays showed a gradual increase in ROS levels and a corresponding gradual increase in DNA in the tail from the comet assay (indicative of increased DNA strand breaks) with the maximum dose displaying a 10% increase in ROS levels and a 17% increase in DNA strand damage.
Li et al., 1998	In vitro, bovine LECs were exposed to 40 and 400 µM H2O2 with an alkaline unwinding assay to determine strand break levels.	Immediately after LECs were exposed to 40 µM and 400 µM H2O2, there were ~145% and ~150% DNA strand breaks compared to the unexposed control level, respectively. The amounts of DNA strand breaks in cells exposed to both concentrations were reduced to ~105% of the unexposed control level after 30 min. After 400 µM H2O2, oxidative stress as measured by LDH was 1200% of control in neuroblastoma cells.
Spector et al., 1997	In vitro, rat LECs exposed to 100 and 125 µM H2O2 with alkaline elution assay to determine single strand break level.	Exposure to 125 µM of H2O2 to lens epithelial cells resulted in reduction of intact DNA to near 1% by 9 hr post-exposure. Exposure to 100 µM H2O2 reduced SOD and GSH levels by 2-fold.
El-Missiry et al., 2018	In vivo, albino Wistar rats were exposed to 4 Gy of γ radiation (137Cs source) at 0.695 rad/s. Kits were used to measure 4-HNE (secondary product of lipid peroxidation) and protein carbonyl group levels as markers of oxidative stress. Antioxidants including GSH, GPx and GR were also assessed. The comet assay was used to analyze DNA strand breaks by visualizing DNA tail %, tail length and tail moment.	4-HNE and protein carbonyl levels increased by approximately 2- and 3-fold after radiation exposure. GSH and GPx levels decreased by approximately 3-fold each, whereas GR levels decreased by approximately 5-fold. Tail DNA %, tail length and tail moment increased by approximately 2-, 3- and 6-fold after exposure to 4 Gy.
Ungvari et al., 2013	In vitro. CMVECs and rat hippocampal neurons were irradiated with 2-8 Gy 137Cs gamma rays. 5(and 6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate acetyl ester (CM-H2DCFDA) staining, and flow cytometry were used to measure ROS production. DNA damage was quantified by measuring the tail DNA content (as a percentage of total DNA) using the Comet Assay-IV software.	Day 1 post-irradiation showed increased cellular peroxide production and increased mitochondrial oxidative stress in CMVECs in a dose-dependent manner, increasing a maximum of ~3-fold at 8 Gy. Tail DNA content also increased in a dose-dependent manner with an approximate increase from 0 to 45% at 8 Gy.
Huang et al., 2021	In vitro, HT22 cells (mouse hippocampal neuronal cell line) were exposed to 10 Gy of X-irradiation at 6 Gy/min. ROS levels were measured using H2-DCFDA staining and fluorescence microscope analysis, whereas western blotting was used to detect γ-H2AX.	At 10 Gy, intracellular ROS generation increased by 5-fold and γ-H2AX increased by 3-fold.
Zhang et al., 2017	In vitro. HT22 cells were exposed to 8 and 12 Gy X-rays. Relative intracellular ROS levels were determined by DCFDA. p-ATM, γ-H2AX were measured with Western blot.	Following 8 Gy irradiation, intracellular ROS levels increased ~1.8-fold. Phosphorylation of ATM and γ-H2AX were increased 4.4-fold and 3.2-fold, respectively, 30 min after 12 Gy.
Cervelli et al., 2014	In vitro. HUVECs were irradiated with single doses (0.125, 0.25, 0.5 Gy), or fractionated doses (2 × 0.125 Gy, 2 × 0.250 Gy) of X-rays. Intracellular ROS generation was measured with a fluorescent dye, C-DCFDA, using a spectrofluorometer. Immunofluorescence microscopy was used to measure γ-H2AX foci.	Intracellular ROS production was significantly increased in a dose-dependent manner (1.6-, 2- and 2.8-fold at 0.125, 0.25, 0.5 Gy, respectively). When HUVECs were exposed to fractionated doses, no increase in ROS generation was observed, compared with respective single doses. 24h post-irradiation the percentage of foci-positive cells exposed to 0.125 Gy, 2 × 0.125 Gy, 0.250 Gy, 2 × 0.250 Gy and 0.5 Gy, was 1.68, 1.48, 3.53, 2.59, 8.74-fold over the control, respectively.
Sakai et al., 2017	In vitro. HAECs were exposed to 100uM H2O2. Intracellular ROS was measured by CM-H2DCFDA. DNA DSBs were detected by immunofluorescent analysis with γ-H2AX as a marker.	Intracellular ROS increased by ~3.7-fold p-ATM increased by ~4.7-fold. γ-H2AX increased by ~3.4-fold.

Incidence Concordance

Reference	Experiment Description	Result
Meng et al., 2021	In vitro, human LECs exposed to 50 µM H2O2 with DCFH-DA fluorescent probe to detect ROS levels and immunofluorescence and western blot assay to detect γ-H2AX.	50 µM H2O2 exposure to lens epithelial cells increased oxidative stress, with ROS measured by LDH, by 4-fold and decreased the level of antioxidants by 2-fold as measured by SOD and GSH-PX. This resulted in 3-fold increase in γ-H2AX.
Smith et al., 2015	In vitro, human LECs exposed to 30 µM H2O2 with alkaline comet assay to determine amount of strand breaks.	Treatment of lens epithelial cells to 30 µM H2O2 induced DNA strand breaks by 55% at 0.5 hr after exposure and increased the level of LDH by ~1.4 fold at 24 hr post-exposure.
Liu et al. 2013	In vitro, human LECs exposed to 30 µM H2O2 with alkaline comet assay determination of strand breaks.	LDH increased by ~1.4 fold at 24 hr post-exposure, with a 5x increase from control levels in DNA strand breaks.

Time Concordance

Reference	Experiment Description	Result
Yang et al., 1998	In vitro, rabbit LECs exposed to H2O2 with TCA addition and thiol assay to determine non-protein thiol (NP-SH) level and alkaline elusion assay to determine strand breaks.	In rabbit LECs exposed in vitro to 125 µM H2O2, non-protein thiol levels decreased to <5% control (indicates oxidative stress) 30 min post-irradiation, and % DNA retained using alkaline elution decreased by 1.6 log (indicates increased DNA fragmentation) within the next 8.5 h.

Response-response Relationship

Provides sources of data that define the response-response relationships between the KEs. More help

Time-scale

Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help

Known Feedforward/Feedback loops influencing this KER

Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

Not identified. 

Domain of Applicability

A free-text section of the KER description that the developers can use to explain their rationale for the taxonomic, life stage, or sex applicability structured terms. More help

This KER is plausible in all life stages, sexes, and organisms with DNA. The evidence is from human, rodent, rabbit and bovine in vitro studies that do not specify the sex, as well as an adult rat in vivo study.

References

List of the literature that was cited for this KER description. More help

Ahmadi, M. et al. (2021), “Early Responses to Low-Dose Ionizing Radiation in Cellular Lens Epithelial Models”, Radiation Research, Vol.197, Radiation Research Society, Indianapolis, https://doi.org/10.1667/RADE-20-00284.1. 

Aitken, R.J. and C. Krausz. (2001), “Oxidative stress, DNA damage and the Y chromosome”, Reproduction, Vol.122/2001, Bioscientifica, Bristol, https://doi.org/10.1530/rep.0.1220497. 

Annesley, S.J. and P.R. Fisher. (2019), “Mitochondria in Health and Disease”, Cells, Vol.8/7, MDPI, Basel, https://doi.org/10.3390/cells8070680. 

Brennan, L., R. McGreal and M. Kantorow. (2012), “Oxidative stress defense and repair systems of the ocular lens”, Frontiers in Bioscience – Elite, Vol.4/E(1), Frontiers in Bioscience, Singapore, https://doi.org/10.2741/365. 

Britton, S. et al. (2020), “ATM antagonizes NHEJ proteins assembly and DNA-ends synapsis at single-ended DNA double strand breaks”, Nucleic Acids Research, Vol.48/17, Oxford University Press, Oxford, https://doi.org/10.1093/nar/gkaa723. 

Cabrera, M. and R. Chihuailaf. (2011), “Antioxidants and the integrity of ocular tissues”, Veterinary Medicine International, Vol.2011, Hindawi Limited, London, https://doi.org/10.4061/2011/905153. 

Cannan, W. and D. Pederson. (2016), “Mechanisms and consequences of double-strand DNA break formation in chromatin”, Journal of Cell Physiology, Vol.231/1, Wiley, Hoboken, https://doi.org/10.1002/jcp.25048. 

Cencer, C. et al. (2018), “PARP-1/PAR Activity in Cultured Human Lens Epithelial Cells Exposed to Two Levels of UVB Light”, Photochemistry and Photobiology, Vol.94/1, Wiley-Blackwell, Hoboken, https://doi.org/10.1111/php.12814. 

Cervelli, T.et al. (2014), “Effects of single and fractionated low-dose irradiation on vascular endothelial cells”, Atherosclerosis, Vol.235/2, Elsevier, Amsterdam, https://doi.org/10.1016/j.atherosclerosis.2014.05.932.

Cervelli, T. et al. (2017), “A New Natural Antioxidant Mixture Protects against Oxidative and DNA Damage in Endothelial Cell Exposed to Low-Dose Irradiation”, Oxidative medicine and cellular longevity, Vol. 2017, Hindawi, London, https://doi.org/10.1155/2017/9085947.

Climent, M. et al. (2020), “MicroRNA and ROS Crosstalk in Cardiac and Pulmonary Diseases”, International Journal of Molecular Science, Vol.21/12, MDPI, Basel, https://doi.org/10.3390/ijms21124370. 

Dahm-Daphie, J., C. Sass, and W. Alberti. (2000), “Comparison of biological effects of DNA damage induced by ionizing radiation and hydrogen peroxide in CHO cells”, International Journal Radiation Biology, Vol.76/1, Informa, London, https://doi.org/10.1080/095530000139023. 

El-Missiry, M. A. et al. (2018), "Neuroprotective effect of epigallocatechin-3-gallate (EGCG) on radiation-induced damage and apoptosis in the rat hippocampus", International Journal of Radiation Biology, Vol. 94/9, https://doi.org/10.1080/09553002.2018.1492755.

Engwa, G.A., F.N. Nweke and B.N. Nkeh-Chungag. (2020), “Free Radicals, Oxidative Stress-Related Diseases and Antioxidant Supplementation”, Alternative therapies in health and medicine, Vol.28/1, InnoVision Health Media, Eagan, pp.114-128. 

Feng, C. et al. (2016), “Lycopene protects human SH-SY5Y neuroblastoma cells against hydrogen peroxide-induced death via inhibition of oxidative stress and mitochondria-associated apoptotic pathways”, Molecular Medicine Reports, Vol.13/5, Spandidos Publications, Athens, https://doi.org/10.3892/mmr.2016.5056. 

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