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Although organisms respiring air oxygen use their energy sources in an optimal way they are threatened by the compulsory formation of reactive oxygen species (ROS). A great variety of ROS sources have to be considered. Mitochondria, which are assumed to be mainly involved, produce ROS under pathophysiological conditions. Other cell organelles such as lysosomes and endoplasmatic reticulum may contribute to cellular ROS formation. Numerous oxidases and non-enzymatic compounds generate ROS. Nitric monoxide is also an important biomolecule with radical character. It is involved in a great variety of physiological and pathophysiological events.

Environmental factors are known sources for oxidative stress. In consequence of the numerous influences that define our environment, environmental oxidative stress can derive from several sources. Such sources can be categorised with respect to their mechanisms of action: Where are the reactive oxygen species generated? Where do they take effect? Are they generated chemically by a noxa, via a target biomolecule or are they physiologically generated by cells? Pollution, non-ionising (ultraviolet and infrared) and ionising radiations are known sources of environmental oxidative stress.

There is strong evidence in support of the oxidation of biomolecules during normal physiology and under conditions of environmental or pathological stress. Normally, the regulation of reduction/oxidation is vital for maintenance of cellular homeostasis. However, in conditions of excess free radicals, most intracellular substrates can be damaged, frequently leading to dysfunction and possibly disease. Whilst oxidants such as the hypohalous acids and reactive nitrogen species can cause a specific fingerprint of biomolecular damage, other commonly derived oxidants such as singlet oxygen, hydrogen peroxide and peroxy radicals cause generic types of damage. Herein, the chemistry of oxidation of DNA, lipids and proteins is considered in the context of our current knowledge on radical-specific oxidation, Furthermore, the formation of secondary oxidation products, arising from radical transfer between molecules and interaction between newly generated carbonyl groups and constituent amines, is examined.

Proliferating mammalian cells exhibit a broad spectrum of responses to oxidative stress, depending on the stress level encountered. Very low levels of hydrogen peroxide, for example 3–15 μM, or 0.1–0.5 mmol 10 cells, cause a significant mitogenic response with 25–45% growth stimulation. Higher HO concentrations of 120–150 μM, or 2–5 μmol 10 cells, cause a temporary growth-arrest that appears to protect cells from excess energy usage and DNA damage. After 2–4 h of temporary growth-arrest many cells begin to exhibit up to a 40-fold transient adaptive response in which genes for oxidant protection and damage repair are preferentially expressed: This transient adaptation is maximal at approximately 18 h after peroxide addition. The HO originally added is metabolized within 30–40 min and if no more is added the cells will gradually de-adapt, so that at 36 h after the initial HO stimulus they have returned to their original level of HO sensitivity. At levels of HO of 250–400 μM, or 9–14 μmol 10 cells, mammalian fibroblasts are not able to adapt, but instead enter a permanently growth-arrested state in which they appear to perform most normal cell functions, but never divide again. This state of permanent growth-arrest has often been confused with “cell death” in toxicity studies that have relied solely on cell proliferation assays as measures of viability. If the oxidative stress level is further increased to 0.5–1.0 mM HO, or 15–30 μmol 10 cells, apoptosis results. This oxidative stress-induced apoptosis involves nuclear condensation, loss of mitochondrial transmembrane potential, degradation/down regulation of mitochondrial mRNAs and rRNAs, and degradation/laddering of both nuclear and mitochondrial DNA. At very high HO concentrations of 5.0–10.0 mM, or 150–300 μmol 10 cells and above, cell membranes disintegrate, proteins and nucleic acids denature, and necrosis swiftly follows. Cultured cells grown in 20% oxygen are essentially pre-adapted or pre-selected to survive under conditions of oxidative stress. If cells are instead grown in 3% oxygen, much closer to physiological cellular levels, they are more sensitive to an oxidative challenge but exhibit far less accumulated oxidant damage.

Low molecular weight antioxidants are an important part of the antioxidative defense mechanisms of cells and organisms. This chapter gives a short overview of the actions of the main antioxidants, including uric acid, ubichinones, lipoic acid, vitamins C and E, carotenoids, and phenolic compounds. The antioxidative properties of these endogeous and nutritional compounds are discussed in this chapter. However, it is critically discussed whether the antioxidative properties of some of these compounds are really important in vivo.

Glutathione is the most abundant non-protein thiol in cells. It is a tripeptide with two important structural features: the thiol group and the gamma-glutamyl peptide bond between glutamate and cysteine. It is a major antioxidant, able to reduce peroxides (due to its action as substrate of glutathione peroxidases). As a result it is oxidised to the disulphide form (GSSG), which in turn is reduced back to GSH by glutathione reductase. Thus glutathione is a major player in maintaining physiological redox status in cells.

Some of the functions of glutathione depend on the presence of the gamma-glutamyl bond, for instance its role in the regulation of amino acid transport. But the majority of the functions of glutathione are related to its role in redox regulation in cells and in detoxification of xenobiotics.

Some areas of research of special interest on this molecule are glutathionylation of proteins, the cellular compartmentation and the role of this interesting molecule in disease.

Superoxide dismutases (SODs) and catalase represent the primary enzymatic defense against reactive oxygen species. Both enzymes are present in virtually all types of aerobic cells. Both are metalloproteins, employing efficient dismutation reactions to dispose of the two most common reactive oxygen species formed, the superoxide radical anion and hydrogen peroxide. These very fast reactions do not require reducing equivalents and thus energy input. Induction of expression of these enzymes in cell culture and in whole organisms provides protection against deleterious effects of oxidative stress in various situations. Transgenic organisms provided many clues concerning the physiological significance of superoxide dismutases. In the human, mutations of CuZn-superoxide dismutase underly cases of familial amyotrophic sclerosis; pathological aspects of polymorphism of superoxide dismutases have also been extensively studied. Application of the enzymes in therapy and for analysis of the level of reactive oxygen species has been suggested.

The early findings that significant amounts of modifications are induced to the cellular DNA both spontaneously and as a consequence of metabolism and environmental exposures have led to the discovery of the existence of multiple highly efficient repair mechanisms to maintain the integrity of DNA.

There is increasing evidence of considerable variation between individuals with regard to activity of DNA repair mechanisms and there are examples of diseases that have deficiencies in DNA repair as a risk factor. It is anticipated that detailed understanding of the DNA repair mechanisms will lead to insights for prevention of diseases, for diagnosis, into resistance to chemotherapeutic agents, and for development of therapeutic tools. DNA repair is also considered to be an important factor in aging.

This chapter briefly summarizes the known DNA repair pathways and reviews the approaches for clarifying and measuring DNA repair in vivo and in vitro.

Protein oxidation is one of the important processes taking place during oxidative stress. Numerous amino acids can be modified within proteins resulting in a large variety of oxidatively modified proteins. These oxidation reactions are accompanied by secondary unfolding of the proteins and modification by non-protein oxidation products. For maintaining the protein pool two processes must be taken into account: protein repair and protein degradation. The known protein repair mechanisms are limited to a few amino acid modifications. However, special removal systems exist in all cellular compartments for degrading oxidized proteins. The major removal system for oxidized proteins in the nucleus and the cytosol is the proteasomal system. The structure, composition, and function of this proteolytic system will be reviewed here. Severe oxidation, however, leads to the formation of insoluble, non-degradable aggregates. These aggregates accumulate within cells, especially during neurodegenerative diseases and the aging process.

Exposure of cells to reactive oxygen species (ROS) may result not only in cell death by excessive oxidation of biomolecules but also cause the activation of cellular stress signaling pathways. The modes of activation of these signaling cascades include the direct interaction of ROS with signaling proteins, i.e., their oxidation, or the indirect action of ROS via biomolecule oxidation products that, in turn, modulate the activities of signaling proteins. The oxidation of ROS target molecules such as glutathione, tyrosine phosphatases, and thioredoxin may result in modulation of signaling cascades, including the mitogen-activated protein kinase (MAPK) pathways. The biological consequences of ROS-induced signaling depend upon which subtypes of MAPK (ERK, JNK, or p38) are predominantly activated.