Inhibition, ECT complexes of the respiratory chain leads to Increase, Oxidative Stress

The biological plausibility for this KER is moderate, as some specific stressors showed the dependent change in both events, however there was an inconsistency with one article where KE1 preceded the MIE.

ROS formation occurs mainly in the mitochondria of a cell, specifically by the complexes of the electron transport chain (ETC) (Zhao et al., 2019; Yu et al., 2021; Shaki et al., 2012; Huerta-García et al., 2014). ROS formation is the result of the leaking of electrons from the ETC, which can then interact with oxygen molecules to form hydrogen peroxide and superoxide (Zhao et al., 2019). In particular the superoxide anion is created as a result of the reaction of oxygen with the iron-sulfur centers in complexes I and III (Kruidering et al., 1997). This is a normal function of the mitochondria when ROS formation is only produced at very low levels, as ROS molecules are involved in signalling pathways within the cell (Zhao et al., 2019). These molecules are then scavenged in the cell by antioxidants in order to maintain a balance of ROS levels in the cell (Kruidering et al., 1997, Zhao et al., 2019). However, complex inhibition in the ETC results in a disrupted electron flow and therefore leads to an increased incidence of electron leakage (Zhao et al., 2019). Complex I and III in particular are considered to be the most common sites of ROS formation within the mitochondria (Zhao et al., 2019, Kuridering et al., 1997, Shaki et al., 2012). Superoxide and hydrogen peroxide molecules then further the increase of oxidative stress by oxidizing lipid molecules and protein molecules while depleting antioxidant molecules (Shaki et al., 2012; Santos et al., 2007).

Dose concordance

Very few studies have been done to assess the dose concordance between mitochondrial electron transport chain inhibition and increased oxidative stress.

One article detailed the inhibition of complexes I through IV as well as the incidence of ROS formation all at varying concentrations of silver used to treat rat liver mitochondria (Miyayama et al., 2013). They found that inhibition of every complex occurred at a dosage of 0.01 μM while ROS formation did not occur until treatment with 0.1 μM. Similar results were reported in a study of lead treatment on rat livers, which found that mitochondrial electron transport complexes I to IV were all significantly inhibited by 10 μM treatment with lead, while GSH content was not significantly decreased before treatment with 40 μM (Ma et al., 2017). More research will be required to assess the dose concordance for the relationship between ETC inhibition and increased oxidative stress.

Temporal concordance

There are few studies regarding the time concordance for the relationship between ETC inhibition and increased oxidative stress. Results from a study of the effects of cisplatin on pig kidney mitochondria showed that mitochondrial electron transport chain inhibition of complexes III and IV occurred after 20 minutes of treatment with 100 μM of cisplatin, while ROS production did not begin until 40 min post-treatment with 100 μM (Kruidering et a., 1997). Another study found that complex I activity in rat liver mitochondria treated with chromium was significantly inhibited as early as 24 hours after treatment, while antioxidant activity was not inhibited until 48 hours after treatment for SOD, CAT, GR, and GST (García-Niño et al., 2013). Further research will be necessary to definitively confirm that ETC inhibition occurs before increased oxidative stress.

Incident concordance

There are some studies which are able to show incident concordance but lack information regarding different doses or time periods being examined. One study showed that inhibition of the mitochondrial ETC complexes after 12 weeks of arsenic treatment in rat brains caused a decrease of 42% in complex I activity, 32% in complex II activity, and 35% in complex IV, resulting in an overall mitochondrial ETC inhibition of 37% (Prakash, Soni, and Kumar, 2015). Meanwhile ROS generation after the same treatment length was only increased by 35% (Prakash, Soni, and Kumar, 2015).

Other evidence

There were also some studies which showed that significant perturbation of the MIE would also induce KE1. One such article showed dose-dependant inhibition of the mitochondrial ETC complexes I and III and then showed the induction of ROS occurring at a concentration above the IC50 value for complex III by lead (Wang et al., 2009). Another example of this was a study which showed the inhibition of complexes II, II-III and IV when rat kidneys were treated acutely with 10 nm gold nanoparticles, which then led to an increase in lipid peroxidation (Ferreira et al., 2015).

Essentiality

While the mitochondrial ETC is commonly accepted to be the main source of ROS production within a cell, metal-catalyzed oxidation can also result in ROS production (Zhao et al., 2019; Shaki et al., 2012). This process is a result of metals interacting with enzymes which produce ROS molecules, such as NADPH-oxidase, uncoupled endothelial nitric oxide synthase (NOS), and xanthine oxidase (Li et al., 2013). However, the ROS molecules that result from this would then go on to perturb the mitochondrial ETC and enter the positive feedback loop of ETC inhibition leading to oxidative stress which then causes further ETC inhibition (Li et al., 2013). Other articles also cite elevated inner membrane potential, elevated mitochondrial calcium concentration, and nitric oxide levels all playing roles as regulators of mitochondrial ROS production, with a possibility that lipid oxidation products are also exogenous stimulators of mitochondrial production (Huerta-García et al., 2014, Zmijewski et al., 2005).

1. One of the articles, the Shaki et al.’s 2012 article, did not show dose concordance for this KER when using uranium as a treatment, as oxidative stress was induced before mitochondrial electron transport chain inhibition occurred, at 50 and 100 μM respectively.

There are currently no articles detailing the response-response relationship between the inhibition of the mitochondrial ETC and an increase in oxidative stress. Further studies will need to be conducted in order to determine a response-response relationship.

There are currently not enough articles which investigate the time-scale over which inhibition of the mitochondrial ETC occurs and instigates oxidative stress and further research must therefore be conducted to identify the time-scale for this relationship.

There is a known feedback loop which influences this key event relationship. Inhibition of the mitochondrial electron transport chain results in increased oxidative stress, which in turn further inhibits the mitochondrial electron transport chain (Guo et al., 2013). The molecular basis behind this is that the ROS molecules are damaging to the macromolecules, such as DNA, proteins, and lipids that they interact with in the mitochondria (Guo et al., 2013). Unrepaired damage to mitochondrial DNA, which is known to be more sensitive than nuclear DNA to ROS molecules due to proximity to the ETC, leads to defective complex I and III function and results in increased reduction of oxygen to it’s reactive forms (Guo et al., 2013; Gonzalez-Hunt et al., 2018). Similarly, damage to the mitochondrial DNA coding for other critical proteins for electron transport can lead to further generation of ROS molecules, all leading to a cycle of ROS molecule generation and organelle dysfunction which ultimately results in the induction of apoptosis (Guo et al., 2013).