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Kinetic Modeling Reveals the Roles of Reactive Oxygen Species Scavenging and DNA Repair Processes in Shaping the Dose-Response Curve of KBrO₃-Induced DNA Damage.

Spassova MA, Miller DJ, Nikolov AS - Oxid Med Cell Longev (2015)

Bottom Line: We used as an example chemical KBrO3 which is activated by glutathione and forms reactive intermediates that directly interact with DNA to form 8-hydroxy-2-deoxyguanosine DNA adducts (8-OH-dG).Our modeling revealed that sustained exposure to KBrO3 can lead to fast scavenger exhaustion, in which case the dose-response shapes for both endpoints are not substantially affected.The results are important to consider when forming conclusions on a chemical's toxicity dose dependence based on the dose-response of early genotoxic events.

View Article: PubMed Central - PubMed

Affiliation: National Center for Environmental Assessment, Office of Research and Development, U.S. Environmental Protection Agency, Washington, DC 20460, USA.

ABSTRACT
We have developed a kinetic model to investigate how DNA repair processes and scavengers of reactive oxygen species (ROS) can affect the dose-response shape of prooxidant induced DNA damage. We used as an example chemical KBrO3 which is activated by glutathione and forms reactive intermediates that directly interact with DNA to form 8-hydroxy-2-deoxyguanosine DNA adducts (8-OH-dG). The single strand breaks (SSB) that can result from failed base excision repair of these adducts were considered as an effect downstream from 8-OH-dG. We previously demonstrated that, in the presence of effective base excision repair, 8-OH-dG can exhibit threshold-like dose-response dependence, while the downstream SSB can still exhibit a linear dose-response. Here we demonstrate that this result holds for a variety of conditions, including low levels of GSH, the presence of additional SSB repair mechanisms, or a scavenger. It has been shown that melatonin, a terminal scavenger, inhibits KBrO3-caused oxidative damage. Our modeling revealed that sustained exposure to KBrO3 can lead to fast scavenger exhaustion, in which case the dose-response shapes for both endpoints are not substantially affected. The results are important to consider when forming conclusions on a chemical's toxicity dose dependence based on the dose-response of early genotoxic events.

No MeSH data available.


Related in: MedlinePlus

Base model. Our model consists of a series of reactions based on a mechanism proposed by Kawanishi and Murata [17]. Bromate (BrO3−) reacts with GSH to form a reactive intermediate complex. (BrOI1). This reactive intermediate receives an electron from Guanine, leading to base oxidation and formation of an 8-OH-dG adduct. This damage cycle can repeat twice more with BrO2− and BrO−, giving each bromate molecule three opportunities to form an adduct. Adduct repair is handled by a base excision repair (BER) mechanism that has a small chance of repair failure resulting in single strand breaks (SSB). A scavenger of the reactive intermediates (⦰) and additional SSB repair mechanisms (break repair) were added in specific cases. Round-end arrows indicate enzymatic participation in a reaction.
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fig1: Base model. Our model consists of a series of reactions based on a mechanism proposed by Kawanishi and Murata [17]. Bromate (BrO3−) reacts with GSH to form a reactive intermediate complex. (BrOI1). This reactive intermediate receives an electron from Guanine, leading to base oxidation and formation of an 8-OH-dG adduct. This damage cycle can repeat twice more with BrO2− and BrO−, giving each bromate molecule three opportunities to form an adduct. Adduct repair is handled by a base excision repair (BER) mechanism that has a small chance of repair failure resulting in single strand breaks (SSB). A scavenger of the reactive intermediates (⦰) and additional SSB repair mechanisms (break repair) were added in specific cases. Round-end arrows indicate enzymatic participation in a reaction.

Mentions: Here we developed further our kinetic model of KBrO3-induced DNA damage [14]. Our aim was to have a realistic model that reflects the experimental evidence in the literature in order to investigate the role of scavengers and DNA repair processes in shaping the dose-response. However, we needed a highly simplified model that can demonstrate the basic features of the dose-response for different types of DNA damage. Therefore, we did not consider special distribution/localization of different compounds or compartmentalization. This model is not intended to be predictive of rates of DNA damage in vivo or in vitro, as the current understanding of the processes involved is insufficient to support a predictive model. Rather, the model is intended to support further understanding of how different processes involved in this system can interact and how this may influence shapes of dose-response relationships. A number of studies have revealed that KBrO3 can cause DNA damage through oxidative stress [8, 17, 23, 24]. It is well documented that at least one of the oxidative DNA damage pathways involves generation of 8-OH-dG adducts. Evidence suggests that KBrO3 forms reactive metabolites by interaction with glutathione. Glutathione is considered to undergo redox cycle fast and the redox reactions are not included in our model for simplicity. The reactive metabolites can directly oxidize DNA Guanine residues. In the biochemical model that we developed here after Kawanishi and Murata [17] and used for our simulations, several consecutive oxidation steps are considered (Figure 1). In our model, via interaction with glutathione (GSH), an intermediate product is formed, which can itself form the oxidative lesion 8-OH-dG on DNA. One molecule of bromate can oxidize several Guanine residues in several consecutive steps, where the BrO3− ion is reduced to BrO2−, to BrO−, and finally to Br• (Figure 1). The 8-OH-dG lesions can subsequently be repaired by an appropriate base repair mechanism (BER). Alternatively, this repair process can also result in error leading to the production of single strand breaks (SSB), each due to a failed repair attempt, although at a much lower rate than successful repairs (Figure 1) [25]. The 8-OH-dG repair mechanism has been studied using mice knockout model [26, 27]. It has been demonstrated that the knockout Ogg1−/− mice have elevated mutation rates in proliferating liver cells due to a higher presence of 8-OH-dG after exposure to KBrO3 [26], suggesting involvement of OGG1 in the repair process. Furthermore, there is some evidence that KBrO3 can directly cause SSB [28], but, for simplicity, we did not include that pathway in our model. The schematics in Figure 1 reflect the biochemical reactions that are included in our computational model in the form of differential equations. The reactive intermediates labeled here BrOI1, BrOI2, and BrOI3 can oxidize Guanine residues to form 8-OH-dG. In some versions of the models we have also included an additional SSB repair mechanism in order to investigate how this step can affect the dose-response dependence of 8-OH-dG and SSB levels. The model elements Guanine, GSH, and BER were given appropriate initial values and KBrO3 was dosed at time zero. The reactions were modeled using simple mass action kinetics, with all simulations carried out using MATLAB SimBiology software. The list of the chemical reactions included in the model and modeled as a system of differential equations is as follows: (1)KBrO3⟶BrO3−GSH+BrO3−⟷GSH·BrO3−GSH·BrO3−⟶GSH+BrOI1BrOI1+S⟶BrOI1Sscavenger  variant  onlyBrOI1+Guanine⟷BrOI1·GuanineBrOI1·Guanine⟶BrO2−+8-OH-dG8-OH-dG+BER⟷8-OH-dG·BER8-OH-dG·BER⟶BER+Guanine8-OH-dG·BER⟶BER+SSBSSB+BR⟷SSB·BRBreak  Repair  variant  onlySSB·BR⟶BR+GuanineBreak  Repair  variant  onlyGSH+BrO2−⟷GSH·BrO2−GSH·BrO2−⟶GSH+BrOI2BrOI2+S⟶BrOI2Sscavenger  variant  onlyBrOI2+Guanine⟷BrOI2·GuanineBrOI2·Guanine⟶BrO−+8-OH-dGGSH+BrO−⟷GSH·BrO−GSH·BrO−⟶GSH+BrOI3BrOI3+S⟶BrOI3Sscavenger  variant  onlyBrOI3+Guanine⟷BrOI3·GuanineBrOI3·Guanine⟶Br+8-OH-dGwhere the dot notation signifies a bound complex of two participants, S is scavenger, BROI1S, BROI2S, and BROI3S are inactive compounds that cannot oxidize DNA and are removed from the system, labeled as (⦰) in Figure 1, BER is base excision repair mechanism, and BR is break repair mechanism. The MATLAB script is available upon request.


Kinetic Modeling Reveals the Roles of Reactive Oxygen Species Scavenging and DNA Repair Processes in Shaping the Dose-Response Curve of KBrO₃-Induced DNA Damage.

Spassova MA, Miller DJ, Nikolov AS - Oxid Med Cell Longev (2015)

Base model. Our model consists of a series of reactions based on a mechanism proposed by Kawanishi and Murata [17]. Bromate (BrO3−) reacts with GSH to form a reactive intermediate complex. (BrOI1). This reactive intermediate receives an electron from Guanine, leading to base oxidation and formation of an 8-OH-dG adduct. This damage cycle can repeat twice more with BrO2− and BrO−, giving each bromate molecule three opportunities to form an adduct. Adduct repair is handled by a base excision repair (BER) mechanism that has a small chance of repair failure resulting in single strand breaks (SSB). A scavenger of the reactive intermediates (⦰) and additional SSB repair mechanisms (break repair) were added in specific cases. Round-end arrows indicate enzymatic participation in a reaction.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig1: Base model. Our model consists of a series of reactions based on a mechanism proposed by Kawanishi and Murata [17]. Bromate (BrO3−) reacts with GSH to form a reactive intermediate complex. (BrOI1). This reactive intermediate receives an electron from Guanine, leading to base oxidation and formation of an 8-OH-dG adduct. This damage cycle can repeat twice more with BrO2− and BrO−, giving each bromate molecule three opportunities to form an adduct. Adduct repair is handled by a base excision repair (BER) mechanism that has a small chance of repair failure resulting in single strand breaks (SSB). A scavenger of the reactive intermediates (⦰) and additional SSB repair mechanisms (break repair) were added in specific cases. Round-end arrows indicate enzymatic participation in a reaction.
Mentions: Here we developed further our kinetic model of KBrO3-induced DNA damage [14]. Our aim was to have a realistic model that reflects the experimental evidence in the literature in order to investigate the role of scavengers and DNA repair processes in shaping the dose-response. However, we needed a highly simplified model that can demonstrate the basic features of the dose-response for different types of DNA damage. Therefore, we did not consider special distribution/localization of different compounds or compartmentalization. This model is not intended to be predictive of rates of DNA damage in vivo or in vitro, as the current understanding of the processes involved is insufficient to support a predictive model. Rather, the model is intended to support further understanding of how different processes involved in this system can interact and how this may influence shapes of dose-response relationships. A number of studies have revealed that KBrO3 can cause DNA damage through oxidative stress [8, 17, 23, 24]. It is well documented that at least one of the oxidative DNA damage pathways involves generation of 8-OH-dG adducts. Evidence suggests that KBrO3 forms reactive metabolites by interaction with glutathione. Glutathione is considered to undergo redox cycle fast and the redox reactions are not included in our model for simplicity. The reactive metabolites can directly oxidize DNA Guanine residues. In the biochemical model that we developed here after Kawanishi and Murata [17] and used for our simulations, several consecutive oxidation steps are considered (Figure 1). In our model, via interaction with glutathione (GSH), an intermediate product is formed, which can itself form the oxidative lesion 8-OH-dG on DNA. One molecule of bromate can oxidize several Guanine residues in several consecutive steps, where the BrO3− ion is reduced to BrO2−, to BrO−, and finally to Br• (Figure 1). The 8-OH-dG lesions can subsequently be repaired by an appropriate base repair mechanism (BER). Alternatively, this repair process can also result in error leading to the production of single strand breaks (SSB), each due to a failed repair attempt, although at a much lower rate than successful repairs (Figure 1) [25]. The 8-OH-dG repair mechanism has been studied using mice knockout model [26, 27]. It has been demonstrated that the knockout Ogg1−/− mice have elevated mutation rates in proliferating liver cells due to a higher presence of 8-OH-dG after exposure to KBrO3 [26], suggesting involvement of OGG1 in the repair process. Furthermore, there is some evidence that KBrO3 can directly cause SSB [28], but, for simplicity, we did not include that pathway in our model. The schematics in Figure 1 reflect the biochemical reactions that are included in our computational model in the form of differential equations. The reactive intermediates labeled here BrOI1, BrOI2, and BrOI3 can oxidize Guanine residues to form 8-OH-dG. In some versions of the models we have also included an additional SSB repair mechanism in order to investigate how this step can affect the dose-response dependence of 8-OH-dG and SSB levels. The model elements Guanine, GSH, and BER were given appropriate initial values and KBrO3 was dosed at time zero. The reactions were modeled using simple mass action kinetics, with all simulations carried out using MATLAB SimBiology software. The list of the chemical reactions included in the model and modeled as a system of differential equations is as follows: (1)KBrO3⟶BrO3−GSH+BrO3−⟷GSH·BrO3−GSH·BrO3−⟶GSH+BrOI1BrOI1+S⟶BrOI1Sscavenger  variant  onlyBrOI1+Guanine⟷BrOI1·GuanineBrOI1·Guanine⟶BrO2−+8-OH-dG8-OH-dG+BER⟷8-OH-dG·BER8-OH-dG·BER⟶BER+Guanine8-OH-dG·BER⟶BER+SSBSSB+BR⟷SSB·BRBreak  Repair  variant  onlySSB·BR⟶BR+GuanineBreak  Repair  variant  onlyGSH+BrO2−⟷GSH·BrO2−GSH·BrO2−⟶GSH+BrOI2BrOI2+S⟶BrOI2Sscavenger  variant  onlyBrOI2+Guanine⟷BrOI2·GuanineBrOI2·Guanine⟶BrO−+8-OH-dGGSH+BrO−⟷GSH·BrO−GSH·BrO−⟶GSH+BrOI3BrOI3+S⟶BrOI3Sscavenger  variant  onlyBrOI3+Guanine⟷BrOI3·GuanineBrOI3·Guanine⟶Br+8-OH-dGwhere the dot notation signifies a bound complex of two participants, S is scavenger, BROI1S, BROI2S, and BROI3S are inactive compounds that cannot oxidize DNA and are removed from the system, labeled as (⦰) in Figure 1, BER is base excision repair mechanism, and BR is break repair mechanism. The MATLAB script is available upon request.

Bottom Line: We used as an example chemical KBrO3 which is activated by glutathione and forms reactive intermediates that directly interact with DNA to form 8-hydroxy-2-deoxyguanosine DNA adducts (8-OH-dG).Our modeling revealed that sustained exposure to KBrO3 can lead to fast scavenger exhaustion, in which case the dose-response shapes for both endpoints are not substantially affected.The results are important to consider when forming conclusions on a chemical's toxicity dose dependence based on the dose-response of early genotoxic events.

View Article: PubMed Central - PubMed

Affiliation: National Center for Environmental Assessment, Office of Research and Development, U.S. Environmental Protection Agency, Washington, DC 20460, USA.

ABSTRACT
We have developed a kinetic model to investigate how DNA repair processes and scavengers of reactive oxygen species (ROS) can affect the dose-response shape of prooxidant induced DNA damage. We used as an example chemical KBrO3 which is activated by glutathione and forms reactive intermediates that directly interact with DNA to form 8-hydroxy-2-deoxyguanosine DNA adducts (8-OH-dG). The single strand breaks (SSB) that can result from failed base excision repair of these adducts were considered as an effect downstream from 8-OH-dG. We previously demonstrated that, in the presence of effective base excision repair, 8-OH-dG can exhibit threshold-like dose-response dependence, while the downstream SSB can still exhibit a linear dose-response. Here we demonstrate that this result holds for a variety of conditions, including low levels of GSH, the presence of additional SSB repair mechanisms, or a scavenger. It has been shown that melatonin, a terminal scavenger, inhibits KBrO3-caused oxidative damage. Our modeling revealed that sustained exposure to KBrO3 can lead to fast scavenger exhaustion, in which case the dose-response shapes for both endpoints are not substantially affected. The results are important to consider when forming conclusions on a chemical's toxicity dose dependence based on the dose-response of early genotoxic events.

No MeSH data available.


Related in: MedlinePlus