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Relationship: 2776

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, Endothelial Dysfunction

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, Endothelial Dysfunction

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 leads to vascular remodeling	non-adjacent	Moderate	Low	Cataia Ives (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
mouse	Mus musculus	Moderate	NCBI
rat	Rattus norvegicus	High	NCBI
pigs	Sus scrofa	Low	NCBI

Sex Applicability

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

Sex	Evidence
Male	High
Female	Low
Unspecific	Low

Life Stage Applicability

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

Term	Evidence
Adult	Moderate
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 describes the imbalances in reactive oxygen and reactive nitrogen species (RONS) radical formation as well as antioxidants and reactive oxygen species (ROS) scavengers (Beckhauser et al., 2016; Elahi et al., 2009; Ray et al., 2012). Oxidative stress can lead to endothelial dysfunction. Within the cardiovascular system, every vessel is lined with a single layer of endothelial cells (Augustin et al., 1994; Fishman, 1982). This endothelial layer plays a crucial role in the regulation of vascular homeostasis through controlling various factors such as vascular permeability, vasomotion, and immune response (Baran et al., 2021; Bonetti et al., 2003; Hughson et al., 2018; Slezak et al., 2017; Sylvester et al., 2018). Of the vascular wall components, the endothelium is also the most vulnerable to damage from ROS (Soloviev & Kizub, 2018). Endothelial cells normally exist in a quiescent state characterized by high nitric oxide (NO) bioavailability (Carmeliet & Jain, 2011); however, cells can become activated as part of a normal host-defence response following tissue injury or oxidative stress (Deanfield et al., 2007; Krüger-Genge et al., 2019). Sustained activation leads to the pathological state of endothelial dysfunction which is defined by decreased NO bioavailability, increased vessel permeability, altered vasomotion, and a pro-thrombotic and inflammatory environment (Baran et al., 2021; Bonetti et al., 2003; Deanfield et al., 2007; Schiffrin, 2008).

Shifting redox balance towards oxidation is known to indirectly lead to endothelial dysfunction through various mechanisms (Hughson et al., 2018; Ramadan et al., 2020; Soloviev & Kizub, 2018). There are several ways through which imbalanced ROS can affect endothelium function, including decreasing NO bioavailability through direct scavenging, which forms the RNS peroxynitrite (ONOO^-) (Hatoum et al., 2006; Li et al., 2002; Schiffrin, 2008; Soloviev & Kizub, 2018; Venkatesulu et al., 2018), as well as impeding NO production and diffusion (Hatoum et al., 2006; Li et al., 2002; Schiffrin, 2008; Soloviev & Kizub, 2018; Venkatesulu et al., 2018; Schiffrin, 2008; Soloviev & Kizub, 2018). Additionally, elevated ROS contribute to introducing a pro-inflammatory and pro-thrombotic milieu characteristic of dysfunction (Hughson et al., 2018; Schiffrin, 2008; Slezak et al., 2017; Tapio, 2016; Venkatesulu et al., 2018). It is also linked to decreased vasomotion (Schiffrin, 2008; Soloviev & Kizub, 2018; Venkatesulu et al., 2018) and finally the onset of endothelial cell apoptosis and premature senescence (Borghini et al., 2013; Hughson et al., 2018; Tapio, 2016; Wang et al., 2016).

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 on radiation stressors 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 Map 2.0

ID	Experimental Design	Species	Upstream Observation	Downstream Observation	Citation (first author, year)	Notes

Evidence Map

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

Title	First Author	Biological Plausibility	Dose Concordance	Temporal Concordance	Incidence Concordance

Biological Plausibility

Dose Concordance Evidence

Temporal Concordance Evidence

Incidence Concordance Evidence

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

Work by Ramadan et al. (2020) explored the use of TAT-Gap19 to block endothelial intracellular communication in order to modulate radiation response of intercellular connexin proteins. Overall, TAT-Gap19 was shown to reduce ROS production and subsequent senescence (SA β-gal activity) and apoptosis (Annexin V and Caspase 3/7) markers. However, treatment with TAT-Gap19 led to an increase in SA β-gal in non-irradiated control at the 9-day point. Additionally, the 0.1 Gy irradiated group showed persistent SA β-gal activity at all time points studied, while the 5 Gy group demonstrated an unexpected decrease before day 14.

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

Modulating factor	Details	Effects on the KER	References
Drug	MnTBAP (a superoxide dismutase mimetic)	Treatment with MnTBAP after irradiation was able to reduce superoxide and peroxide levels and restore vasodilation ability	(Hatoum et al., 2006)
Drug	Tempol (a superoxide dismutase mimetic)	Treatment with tempol after irradiation was able to restore vasodilation ability	(Hatoum et al., 2006)
Drug	TAT-Gap19 (inhibitor of connexin 43 which is associated with atherogenesis and endothelial stiffness)	Treatment with TAT-Gap19 led to a decrease in ROS and SA β-gal levels after irradiation	(Ramadan et al., 2020)
Drug	hBMSCs (protect against vascular damage through antioxidant properties)	Treatment with hBMSCs after irradiation caused increased catalase and HO-1, as well as decreased oxidative damage and apoptosis	(Shen et al., 2018)
Drug	Oxp (can inhibit XO, a source of ROS)	Treatment with Oxp showed decreased XO activity and ROS production along with increased vasodilation after irradiation	(Soucy et al., 2007; Soucy et al., 2010; Soucy et al., 2011)

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 are a few examples of quantitative understanding of the relationship. All data that is represented is statistically significant unless otherwise indicated.

Response-response Relationship

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

Dose/incidence concordance

Reference	Experiment Description	Result
Soucy et al. 2007	In vivo. Sprague-Dawley rats were whole-body irradiated with ¹³⁷Cs gamma radiation at 50, 160 and 500 cGy. XO is a primary source of cardiac ROS and was used as a measure of oxidative stress. Vasodilation response to ACh was used to evaluate endothelial function.	At 500 cGy, XO activity was found to be 2-fold elevated compared to control, and there was also an increase in XO quantity. Simultaneously, there was endothelial dysfunction as seen with a ~30 percentage point decrease in vasodilation response to ACh.
Soucy et al. 2010	In vivo. In Sprague-Dawley rats were whole-body irradiated with ¹³⁷Cs gamma radiation at 5 Gy. ROS were measured using dihydroethidium fluorescence. Aortic relaxation response to ACh was also measured.	After 5 Gy, ROS increased 1.7-fold and relaxation decreased 0.7-fold.
Soucy et al. 2011	In vivo. Wistar rats were exposed to 0.5 and 1 Gy doses of ⁵⁶Fe-ion radiation. ROS production rates were evaluated using dihydroethidium fluorescence. ACh-induced vasodilation responses were measured.	⁵⁶Fe irradiation at 1 Gy produced a 1.8-fold increase in ROS levels. At 10^-5 M ACh, aorta without irradiation relaxed by 87%, while aorta with 1 Gy irradiation had significantly lower relaxation of 76%.
Li et al. 2002	In vivo. Surgically exposed coronary arteries of yucatan pigs were irradiated with 20 Gy ³²P β-irradiation. Oxidative stress was evaluated through superoxide production. Endothelial function was evaluated through endothelial-dependent vasomotor response and morphological changes.	Superoxide production increased 3.5-fold between the control and 20 Gy irradiated groups. Contractile response to KCl dropped over 50% in the irradiated group. Morphological changes were also observed, with irradiated arteries seeing enlarged endothelial cells, formation of fibrin networks, activated platelets, leukocytes exhibiting membrane protrusions and pseudopodial extensions all indicative of an inflammatory and pro-thrombotic state of endothelial dysfunction.
Shen et al. 2018	In vivo. Male mice were irradiated with 18 Gy X-rays. Oxidative stress was measured with 4-HNE and 3-NE oxidative damage markers and antioxidant enzymes catalase and HO-1, measured by immunohistological staining. Endothelial dysfunction was determined through apoptosis.	4-HNE showed a maximum increase of ~1.8-fold, and 3-NT showed a maximum increase of ~2.2-fold. Apoptosis levels peaked at a ~5-fold increase above control levels.
Ramadan et al. 2020	In vitro. Telomerase-immortalized human Coronary Artery and Microvascular Endothelial cells (TICAE) and Telomerase Immortalized human Microvascular Endothelial cells (TIME) were exposed to X-rays (0.1 and 5 Gy). ROS production was measured using CM-H2DCFDA combined with Incucyte live cell imaging. Endothelial dysfunction was evaluated through: endothelial apoptosis Annexin V and Caspase 3/7 marker levels Dextran fluorescein dye uptake level by necrotic cells cell senescence SA β-gal activity IGFBP-7 and GDF-15 senescence marker levels	ROS production was increased in TIME cells after 0.1 and 5 Gy dose. In both coronary and endothelial cells, apoptosis mainly occurred after 5 Gy radiation. With coronary cells demonstrating an increase in Annexin V and Caspase 3/7 markers and endothelial cells showing elevated Annexin V and membrane leakage. SA β-gal activity significantly increased for both 0.1 and 5 Gy doses. IGFBP-7 and GDF-15 levels were also elevated in both cell types; GDF-15 increasing at both 0.1 and 5 Gy doses, while IGFBP-7 only showed significant elevation at the 5 Gy dose. With the 0.1 Gy dose, there was a significant increase in SA β-gal activity of ~3-fold, while at 5 Gy the activity increased ~5-fold. Endothelin-1 was found to be significantly elevated following 5 Gy irradiation in both cell types.
Ungvari et al. 2013	In vitro. Cerebral microvascular endothelial cells (CMVECs) from F344×BN rats were harvested and cultured. Following culture, cells were irradiated with ¹³⁷Cs gamma radiation in doses between 2-8 Gy. Oxidative stress was evaluated through cellular peroxide and superoxide production. Endothelial dysfunction was evaluated through cell senescence via SA-β-gal presence, and apoptosis via caspase 3/7 maker and ratio of apoptotic:viable cells.	Oxidative stress increased in a dose-dependent manner following irradiation. Change of ROS became significant after 4 Gy at a ~1.5-fold increase and reached ~3-fold increase at the highest studied dose of 8 Gy. Mitochondrial oxidative stress also became significant after 4 Gy and increased linearly for a peak of a ~1.5-fold increase at 8 Gy. Endothelial cell senescence and apoptosis were similarly found to increase in a dose dependent manner. With ~30% of cells being SA-β-gal positive after 8 Gy irradiation, signalling premature senescence. Ratio of dead cells peaked at 10% and Caspase 3/7 peaking at a ~5.5-fold change following 18h post irradiation.
Hatoum et al. 2006	In vivo. Effect of cumulative radiation doses on rat gut microvessels was studied. Rats were exposed to 1 to 9 fractions of 250 cGy for a total dose of up to 2250 cGy. Following exposure, the animals were euthanized, and submucosal vessels isolated. Oxidative stress was measured through superoxide and peroxide levels. Endothelial function was assessed through ACh vasodilation response.	After the final cumulative dose of 2250 cGy, superoxide was ~1.6-fold elevated and peroxides were ~1.7-fold elevated compared to non-irradiated controls. ROS levels increased sharply after the second dose, immediately preceding drop in ACh vasodilation response. Max dilation dropped from 87% to 3% between pre-irradiation and post-final radiation dose. ACh response remained within control levels following fractions 1 and 2, however following fraction 3, response dropped below 30% for all remaining doses.
Delp et al. 2016	In vivo. The effects of HU and 1 Gy dose of ⁵⁶Fe radiation of the gastrocnemius muscle feed arteries and coronary arteries of C57BL/6 mice was studied. Xanthine oxidase (XO) levels were used as a measure of ROS production and therefore oxidative stress. Endothelial function was evaluated through vasodilation response to ACh.	Following 2-week HU, there were no significant changes to XO levels or vasomotor response. Following total body irradiation with 1 Gy, there was a ~2-fold increase in XO activity in both gastrocnemius muscle feed and coronary arteries. Vasodilation response subsequently decreased ~10 percentage points. Combined HU and total body irradiation led to a ~2.2-fold increase in XO activity in both artery types and vasodilation response decrease of ~10 percentage points.

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

Time concordance

Reference	Experiment Description	Result
Soucy et al. 2007	In vivo. Sprague-Dawley rats were whole-body irradiated with ¹³⁷Cs gamma radiation at various doses. 2 weeks after irradiation, the animals were euthanized, and aortas were harvested. XO was used as a measure of oxidative stress. Dose-dependent vasodilation response to ACh was used to evaluate endothelial function.	XO activity was elevated 2-fold compared to control, and there was also an increase in XO quantity. Simultaneously, endothelial dysfunction was seen with a ~30% decrease in vasodilation response to ACh.
Soucy et al. 2010	In vitro. Sprague-Dawley rats were whole-body irradiated with ¹³⁷Cs gamma radiation. 2 weeks after receiving radiation dose, the animals were euthanized and aortas were harvested. ROS were measured using dihydroethidium fluorescence. Aortic relaxation response to ACh was also measured.	ROS increased 1.7-fold and relaxation simultaneously decreased 0.7-fold.
Soucy et al. 2011	In vivo. Wistar rats were exposed to ⁵⁶Fe-ion radiation. Rats were euthanized at 4 months post-irradiation and aorta was harvested. ROS production rates evaluated using dihydroethidium along the ACh-induced vasodilation response were measured.	ROS levels increased by 75% 4 months post-irradiation. ACh vasodilation response decreased by 13%.
Shen et al. 2018	In vivo. Male mice were irradiated with 18 Gy X-rays. Immunohistochemical staining assessed oxidative stress using 4-HNE and 3-NE as markers for oxidative damage. Endothelial dysfunction was determined through apoptosis.	Exposure to 18 Gy caused increased 4-HNE and 3-NT levels. 4-HNE showed a maximum ~1.8-fold increase at 14-days post radiation, and 3-NT showed a maximum ~2.3-fold increase 7 days post radiation. Apoptosis levels peaked at 7 days post-irradiation with a ~5-fold increase above control levels.
Ramadan et al. 2020	In vitro. TICAE and TIME cells were exposed to X-rays (0.1 and 5 Gy). Oxidative stress was evaluated through intracellular ROS production. Endothelial dysfunction was evaluated through: endothelial apoptosis Annexin V and Caspase 3/7 marker levels Dextran fluorescein dye uptake level by necrotic cells cell senescence SA β-gal activity IGFBP-7 and GDF-15 senescence marker levels Endothelin-1 levels	Highest response was observed for both doses at 45 minutes after irradiation followed by a decline at the 2- and 3-hour time points but remaining elevated above non-irradiated control levels. Endothelial cells studied produced more ROS than the coronary cells. Caspase 3/7 and annexin V increased linearly until 100h. SA β-gal activity significantly increased at 7 and 9 days. GDF-15 and IGFBP-7 were increased after 7 days. Endothelin-1 was found to be significantly elevated after 7 days.
Ungvari et al. 2013	In vitro. Cerebral microvascular endothelial cells (CMVECs) from F344×BN rats were harvested and cultured. Following culture, cells were irradiated with ¹³⁷Cs gamma radiation. Oxidative stress was evaluated through cellular peroxide and superoxide production. Endothelial dysfunction was evaluated through cell senescence via SA-β-gal presence, and apoptosis via caspase 3/7 maker and ratio of apoptotic:viable cells.	Superoxide and peroxide increased 1 day but not 14 days post-irradiation ~30% of cells were SA-β-gal positive after 8 Gy irradiation, measured 7 days post-irradiation. 24 h after irradiation, 10% of cells were dead. Caspase 3/7 increased from 2 to 18 h, peaking at a ~5.5-fold change following 18 h post-irradiation but decreased at 24 h.
Hatoum et al. 2006	In vivo. Effect of cumulative radiation doses on rat gut microvessels was studied. Rats were exposed to 1 to 9 cGy in 3 fractions per week on alternate days for 3 successive weeks for a total dose of up to 2250 cGy over a total time of 19 days. Oxidative stress was measured through superoxide and peroxide levels from various fluorescent markers. Endothelial function was assessed through ACh vasodilation response.	After 19 days, superoxide was ~1.6-fold elevated and peroxides were ~1.7-fold elevated compared to non-irradiated controls. ROS levels increased at day 5, at the same time as a drop in ACh vasodilation response. Max dilation dropped from 87% to 3% between day 1 and day 19.

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

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

The evidence is derived from rat in vivo and in vitro models. Mice cell-derived studies were also available but less in-vivo evidence was available from this species. There was a low number of studies containing human or pig models to support this KER. Males have been studied more often than females. There are a few studies with unspecified lifestage of models, while the studies with a defined age typically used adult models.