Reactive oxygen species (ROS)

Reactive oxygen species (ROS), such as hydrogen peroxide (\(\ce{H_{2}O_{2}}\)), superoxide (\(\ce{O_{2}^{.-}}\) and \(\ce{HO_{2}^{.}}\) ), hydroxyl radical (\(\ce{^{.}OH}\)), and hypochlorite anion (OCl-), are formed in the normal metabolism of oxygen and have important roles in intra- and intercellular signalling. Superoxide reacts with nitrogen oxide (NO), limiting the bioavailability of NO, facilitating vasoconstriction and reducing blood flow. Diffusion of hydrogen peroxide across lipid membranes is limited, but it can be transported by aquaporin-3 and aquaporin-8. Under normal conditions, ∼0.1-0.2% of O2 consumption in mitochondria results into production of ROS. ROS levels in mitochondria are 5-10 times higher than in elsewhere in cytosol or nucleus. Additionally, auto-oxidation of some compounds, including monoamines dopamine and noradrenaline, leads to ROS formation. Free iron, copper, and many other metal ions in the body react uncontrollably with oxygen producing ROS, and are therefore bound to carrier proteins. Increased ROS levels can cause substantial damage to cell structures (“oxidative stress”) and initiate apoptosis and tissue injury, possibly leading to fibrosis, or complications such as atherosclerosis. Radiation therapy is based on ROS directly generated by ionizing radiation. Activated granulocytes use NADPH oxidases to produce superoxide, which is transformed to hydrogen peroxide, and with myeloperoxidase (MPO) to hypochlorite; these ROS can kill pathogens or cancer cells at the site of inflammation. Activated microglia in neuroinflammation also use NADPH oxidases, especially NOX2, to produce ROS.

ROS concentrations are kept under control with enzymes such as catalase, superoxide dismutases (SOD1-3), and NADH peroxidase. Cells can regulate the number of proton leaking ion channels on the inner mitochondrial membrane. These uncoupling proteins (UCPs) reduce the proton gradient and production of ATP, and also the production of ROS. UCP1 is present in brown adipose tissue, required for its thermogenic function. UCP2 is present in tissues such as kidneys that have high ATP production. Increased ROS production activates UCP2 and UCP3. However, chronic overexpression of UCPs may lead to increased ROS production and hypoxia, as mitochondrial respiration is increased to sustain ATP levels.

Superoxide generation can be detected in vivo in peripheral organs with PET tracer [18F]DHMT (Chu et al., 2014; Zhang et al., 2016; Boutagny et al., 2018). This dihydroethidium (DHE) analogue tracer is selectively oxidized by superoxide, not by other ROS, and its oxidation products can bind to DNA, leading to trapped tissue uptake (Chu et al., 2014). This tracer, however, does not pass the blood-brain barrier, and therefore another tracer, [18F]ROStrace, has been developed for imaging superoxide production in the CNS (Hou et al., 2018).

Hydromethidine (HM) is oxidized both specifically by superoxide anion and non-specifically by any ROS, leading to products that are trapped by binding to DNA or by their charge in the CNS. [11C]HM penetrates BBB rapidly, and is trapped in the brain in LPS animal model of oxidative stress (Wilson et al., 2017).

PC-[18F]FLT-1 is a prodrug of FLT: upon exposure to hydrogen peroxide it releases [18F]FLT, which is transported and trapped in cells with active DNA synthesis (Carroll et al., 2014); since both cell proliferation and ROS production are typically increased in tumours, the tracer may be useful in cancer imaging.

Dihydroquinoline [11C]DHQ1 may be useful for assessing the redox status, even in the brain (Okamura et al., 2015).

A 18F-labelled hydrocyanine dye derivative can be used as NIR probe and PET tracer, but has very slow clearance from blood (Al-Karmi et al., 2017).

Glutathione (GSH) is the predominant intracellular antioxidant, which via antiporter system xC- also affects extracellular cysteine-cystine redox cycle and glutamate concentration in the brain. L-aminosuberic acid is an efficient system xC- substrate, and 5-[18F]fluoro-aminosuberic acid may be useful in imaging cellular response to oxidative stress (Webster et al., 2014; Yang et al., 2017). Gamma glutamyl transferase (GGT) catalyzes the breakdown of GSH. GGT is upregulated in malignant cells and is in part responsible for increased ROS production in tumours. Glutathione monoethyl ester (GSHMe) is a substrate of GGT. DTPA-conjugated GSHMe, DT(GSHMe)2, is accumulated in tumours, and [68Ga]DT(GSHMe)2 tumour uptake has been shown to be specific to GGT activity (Khurana et al., 2015).

Ascorbic acid (Vitamin C) is an important antioxidant, which also has roles in synthesis of collagen and catecholamines. Ascorbic acid is transported into cells via sodium dependent vitamin C transporters (SVCT1 and SVCT2). ROS oxidate ascorbic acid into dehydroascorbic acid (DHA), which is a substrate for glucose transporters (GLUT1, GLUT3, and GLUT4). Ascorbic acid and DHA have been labelled with 11C (Caroll et al., 2016). 6-deoxy-6-[18F]fluoro-L-ascorbic acid ([18F]DFA) distributes in tissues like ascorbic acid, and could be used for studying ascorbate recycling (Yamamoto et al., 1992; Yamamoto et al., 1996; Yamamoto et al., 2005).

Many of the PET tracers aimed for imaging tissue hypoxia are actually assessing redox state, and the uptake is related to the oxidative stress state.

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Cheng G, Zielonka M, Dranka B, Kumar SN, Myers CR, Bennett B, Garces AM, Dias Duarte Machado LG, Thiebaut D, Ouari O, Hardy M, Zielonka J, Kalyanaraman B. Detection of mitochondria-generated reactive oxygen species in cells using multiple probes and methods: potentials, pitfalls, and the future. J Biol Chem. 2018; 293(26): 10363-10380. doi: 10.1074/jbc.RA118.003044.

Hou C, Hsieh C-J, Li S, Lee H, Graham TJ, Xy K, Weng C-C, Doot RK, Chu W, Chakraborty SK, Dugan LL, Mintun MA, Mach RH. Development of a positron emission tomography radiotracer for imaging elevated levels of superoxide in neuroinflammation. ACS Chem Neurosci. 2018; 9: 578-586. doi: 10.1021/acschemneuro.7b00385.

Wilson AA, Sadovski O, Nobrega JN, Raymond RJ, Bambico FR, Nashed MG, Garcia A, Bloomfield PM, Houle S, Mizrahi R, Tong J. Evaluation of a novel radiotracer for positron emission tomography imaging of reactive oxygen species in the central nervous system. Nucl Med Biol. 2017; 53: 14-20. doi: 10.1016/j.nucmedbio.2017.05.011.

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Updated at: 2018-11-04
Created at: 2018-08-09
Written by: Vesa Oikonen