Neuronal membrane lipid peroxidation. Lipid peroxidation (LPO)

MINISTRY OF EDUCATION OF THE REPUBLIC OF BELARUS

GOMEL STATE MEDICAL UNIVERSITY

Faculty of Medicine and Diagnostics

Department of Clinical Laboratory Diagnostics

Laboratory diagnostics of the intensity of lipid peroxidation

Course work

Performer: Podstrekha Elena Stanislavovna

student of group D-503

Scientific supervisor: head of the department, Doctor of Medical Sciences, Associate Professor Novikova Irina Aleksandrovna

GOMEL 2016

Essay

The work analyzes the literature on the study of enzymatic and non-enzymatic pathways for the formation of reactive oxygen species, the mechanisms of their damaging effects on living cells, in particular the initiation of free radical lipid peroxidation. Anti- and pro-oxidant systems of the body's defense are considered, balancing the formation, metabolism and utilization of reactive oxygen species.

Coursework 35 pages, 1 table, 17 sources.

List of keywords: lipid peroxidation, free radical oxidation, reactive oxygen species, antioxidant protection, malondialdehyde.

List of abbreviations

Introduction

The role of free radical oxidation and the antioxidant system in the human body

1 Forms of free radicals in the body

2 General characteristics of the main ROS, their biological role

3 Physiological role of free radical oxidation

4 Regulation of free radical oxidation

5 The body's antioxidant defense system

6 Pathogenetic mechanisms of disorders that develop when the balance of antioxidant and prooxidant systems is disturbed

Free radical (lipid peroxidation)

1 Participation of ROS and lipid peroxidation products in the pathogenesis of human diseases

2 Diagnostics of peroxidation processes

Conclusion

List of symbols

AOA - antioxidant activity

AO - antioxidant

ROS - reactive oxygen species

GR - glutathione reductase

GP - glutathione peroxidase

MP - myeloperoxidase

LPO - lipid peroxidation

SOD - superoxide dismutase

FRO - free radical oxidation

CL - chemiluminescence

EPR - electron paramagnetic resonance

Introduction

lipid peroxidation active oxygen

In the body, as a result of redox reactions, the generation of reactive oxygen species (ROS) constantly occurs, which are highly reactive, causing, in particular, the oxidative modification of biopolymers: proteins, lipids, nucleic acids, carbohydrates. Oxygen radicals, despite their reactivity and potential toxicity, in low concentrations are normal metabolites of many biochemical reactions in the cell. Under physiological conditions, free radical reactions occur at a low level. The processes occurring with the participation of oxygen radicals indicate the important role of these compounds in maintaining homeostasis, forming the body's resistance to infections, and ensuring the regeneration of tissues and organs. If the process of ROS generation intensifies, this can be and is a trigger factor for the development of a whole list of various pathological processes.

The relevance of in-depth development of the problem of the pathogenetic significance of free radical peroxidation is determined by increasing environmental distress. The study of this important link in homeostasis has direct practical significance, since it allows us to develop and apply adequate preventive approaches that prevent the start of a chain reaction of free radical oxidation or neutralize the toxicity of lipid peroxidation products.

1. The role of free radical oxidation and the antioxidant system in the human body

Systems involved in the formation of ROS and processes associated with the oxidative alteration of biological compounds are conventionally united by the concept of a pro-oxidant system.

Pro-oxidants in a living cell include high concentrations of oxygen (for example, during prolonged hyperbaric oxygenation of a patient), enzyme systems that generate superoxide radicals (for example, xanthine oxidase, enzymes of the plasma membrane of phagocytes, etc.), ferrous ions.

Oxidative reactions are the basis for energy production and vital activity of all cells of the human body. They can occur without the addition of oxygen (oxidase reactions) and with the addition of molecular or atomic oxygen - oxygenase reactions. The intermediate products of the latter are peroxides and epoxides, therefore such reactions are called peroxidation. It is induced by highly reactive free radicals.

Under physiological conditions, the intensity of peroxide processes is insignificant and is maintained at a stationary level thanks to a multicomponent system for neutralizing constantly formed free radicals - the antioxidant system.

The formation of pro-oxidants in tissues is balanced by the activity of intra- and extracellular antioxidants, forming a certain optimal level of pro-oxidant-antioxidant balance.

1.1 Forms of free radicals in the body

There are many known redox reactions that produce various types of free radicals. Free radicals were discovered at the end of the last century. To date, more than 8000 of them have been described. Free radicals are highly active compounds that are formed under physiological conditions as secondary products during metabolism, as well as in other ways, including redox reactions carried out by single-electron transfer; homolysis of initiator molecules with weak covalent bonds, radiolysis; photolysis, thermolysis.

Many xenobiotics, including various drugs, alcohol, etc., are metabolized in the body, generating free radicals. Despite the great diversity of their origin and structure, free radicals are divided into 3 large groups: 1) reactive oxygen radicals (ROS); 2) reactive nitrogen radicals (RNS); 3) reactive chlorine radicals (RCS).

A free radical is a molecule, atom, or group of atoms that has an unpaired electron in its outer atomic orbital. ROS are active participants in a large number of chemical reactions in cells, exerting a variety of physiological effects.

Distinctive features of free radicals:

the presence of an unpaired electron at the outer energy level;

own magnetic moment;

high chemical activity and short lifetime;

the ability to initiate oxidation chain reactions;

The most likely occurrence of free radicals in the body is during the sequential addition of electrons to oxygen and during free radical lipid peroxidation.

The main processes leading to the formation of free radicals in the body:

sequential addition of electrons to oxygen in the presence of metals of variable valency;

microsomal and mitochondrial oxidation, phagocytosis;

enzymatic reactions involving hydrolases, oxidases, dehydrogenases;

reactions of autoxidation and biosynthesis (thiols, catecholamines, etc.);

oxidation of foreign compounds - xenibiotics, some medications;

the effect of negative environmental factors (physical and chemical oxidation initiators);

photochemical processes;

lipid peroxidation.

modification of physicochemical properties of biological membranes;

protective functions, oxidation of foreign compounds, microbicidal effect;

metabolism, accumulation and biotransformation of energy;

influence on immunity, information transfer.

The most common forms of free radicals in the body are:

Reactive oxygen species:

ABOUT ˙ 2 - superoxide anion radical;

¹ O2 - singlet form of oxygen;

OH ˙ - hydroxyl radical;

H2O2 - hydrogen peroxide;

Oxidized halogens: CLO ˙ - hypochloride, chloramines;

Nitrogen oxides: NO˙ - Nitric oxide;

Free radicals formed during lipid peroxidation: RO ˙ ,RO2 ˙ - mono-, dimeric, polymeric, cyclic, alkoxy and peroxide radicals of fatty acids.

2 General characteristics of the main ROS, their biological role

Oxygen radicals are formed during processes associated with the transport of electrons along the respiratory chain. Under normal conditions, from 1 to 5% of consumed oxygen is consumed for the generation of ROS. However, this value can increase significantly when the body's oxygen budget changes - during hyperoxia or hypoxia. As a result of the sequential reduction of molecular oxygen, the formation of superoxide anion, hydrogen peroxide, and hydroxyl radical occurs.

Superoxide anion radical (O ˙ 2) is formed when one electron is added to an oxygen molecule in the ground state. The radical is a relatively weak oxidizing agent and in many biological systems acts as an electron donor, reducing a number of compounds. When interacting with a proton, O2 transforms into a hydroperoxide radical. In addition, it is a potential source of hydroxyl radical and hydrogen peroxide.

Superoxide anion radical is a more reactive compound than oxygen. In the body, superoxide anion radical is an intermediate product of many biochemical reactions, such as the oxidation of thiols, flavins, quinones, catecholamines, pterins, as well as the metabolism of xenobiotics. Of the sources of superoxide anion radical, the most interesting are hemoglobin, myoglobin, reduced cytochrome C, NADPH oxidases of phagocytic cells, etc. The main source of the radical in the blood is neutrophils, which generate it during a number of reactions of specific and nonspecific immunity. Another enzyme specialized in the formation of superoxide anion radical is xanthine oxidase, which under normal conditions is predominantly in the dehydrogenase form and can reversibly and irreversibly transform into the oxidase form, which occurs during ischemia. It is believed that the generation of the radical by xanthine oxidase is necessary for iron metabolism, regulation of vascular tone and cell proliferation, and provision of the microbicidal potential of neutrophils.

Superoxidation radical is the trigger of a cascade of free radical reactions leading to the emergence of most ROS and lipid peroxidation products. It is involved in the synthesis of chemotactic peptides, enhances mitogen-stimulated proliferation of lymphocytes, inhibits the action of endothelial relaxing factor, can damage erythrocyte membranes, inhibit Ca2+-ATPase, RNA and protein synthesis of endothelial cells, oxidize serum proteins, while at the same time its direct cytotoxicity is low.

To regulate O levels ˙ 2 in cells there is a highly specific antioxidant enzyme - superoxide dismutase, which has the ability to significantly accelerate the reaction of dismutation of the radical into hydrogen peroxide.

Hydrogen peroxide (H2O2). The addition of two electrons to an oxygen molecule or one electron to an O2 anion is accompanied by the formation of a doubly charged O22ˉ anion, which transforms into HO2 ˙ or hydrogen peroxide. Hydrogen peroxide is classified as a medium strength oxidizing agent; in the absence of enzymatic antioxidants and metal ions of variable valence, it is relatively stable and can migrate into cells and tissues. Hydrogen peroxide has a limited damaging effect, causing, in particular, disruption of calcium homeostasis in the cell. In the body, its sources are reactions involving oxidases (xanthine oxidase, L-amino acid oxidase and a number of others), transferring two electrons to an oxygen molecule, as well as the dismutation reaction of the superoxide anion radical, catalyzed by superoxide dismutase. This reaction is the source of almost 80% of the hydrogen peroxide at the site of inflammation.

Hydrogen peroxide is an intermediate product in the formation of most ROS. H2O2 is one of the sources of the most toxic ROS - hydroxyl radical. In the presence of myeloperoxidase, highly reactive hypohalides are formed from it - HOC1, HOBr, HOJ, HOSCN.

Under normal conditions, mammalian cells are quite resistant to the effects of hydrogen peroxide due to the presence of enzymes - glutathione peroxidase and catalase. H2O2 molecules perform a number of regulatory functions in the cell. Peroxide can serve as a metabolic signal to induce the expression of genes that synthesize structural and functional proteins in the cell.

Hydroxyl radical (HO ˙) is the most reactive and, accordingly, toxic of all ROS formed in biological systems. The radical can break any hydrocarbon bond, and the rate of its interaction with organic substrates reaches values ​​equal to the diffusion rate (i.e. 107-1010 mol/s, which is 106 times higher than for the superoxide anion radical and hydrogen peroxide). Compared to other AFK, BUT ˙ has the highest redox potential, equal to + 2.7, which allows it to attack and destroy any macromolecules almost on the spot at the moment of their appearance.

The main source of hydroxyl radical is the Fenton reaction with the participation of metals of variable valence, mainly with Fe2+, according to the scheme H2O2 + Fe+2 → Fe+3 + OH + OH ˙. The formation of the radical also occurs during the oxidation of arachidonic acid, in the Haber-Weiss, Osipov reaction, during microsomal oxidation, in reactions with flavin enzymes and CoQ. Reverse reduction of Fe3+ is possible in reaction with O2, as well as in interaction with ascorbic acid, glutathione, cysteine ​​and other oxidizing compounds. It has been shown that the cytotoxic and carcinogenic effects of ionizing radiation are directly related to the formation of hydroxyl radicals during the radiolysis of water. BUT ˙ also participates in the microbicidal and cytotoxic effects of granulocytes, monocytes and T-lymphocytes. Hydroxyl radicals cause damage to nucleic acids, proteins, as well as other cellular structures, and inhibit a number of complement fractions. They induce the formation of organic radicals and thus trigger lipid peroxidation processes. Due to the high nonspecificity of the reactions of the radical with various organic molecules, its interaction is of a chain nature. It is important to note that the body does not have specialized enzyme systems that have the ability to inactivate hydroxyl radicals. Low molecular weight compounds, such as uracil, uric acid, salicylates, glucose, dimethyl sulfoxide, have the ability to inhibit OH ˙ radical only at sufficiently high concentrations. Thus, in a number of pathological conditions accompanied by excessive formation of ROS and, accordingly, hydroxyl radicals, the body becomes practically defenseless against the damaging effects of this compound. Prevention of damage to cellular structures is carried out only by reducing the concentration of radicals - OH precursors, in particular, superoxide anion radical and hydrogen peroxide. SOD and catalase have the ability to destroy these precursor radicals.

Singlet oxygen( ¹O 2). It is formed when the spin of one of the electrons of the p-orbital in the oxygen molecule changes. Emergence ¹O 2 as a by-product was noted in many enzymatic reactions involving SOD, catalase and peroxidases, as well as in reactions involving most ROS. Thus, in the reaction of hydrogen peroxide decomposition by catalase to 1%, the resulting oxygen appears in the singlet state. Singlet oxygen is highly reactive and easily enters into oxidative reactions with organic compounds. It often acts as an inducer of LPO reactions. Although ¹ O2 has a cytotoxic effect and takes part in the microbiocidal action of granulocytes; its contribution to these processes is not decisive. One of the most effective quenchers of singlet oxygen in the cell is beta-carotene, one molecule of which can quench about 1000 of its molecules before undergoing oxidative destruction.

ROS act as secondary messengers in cell life processes. By being involved in signal transduction, ROS affects key components of metabolic processes: phosphorylation, Ca2+ metabolism, modulation of transcription factors, hydrolysis of phospholipids. During any stress reactions of the body, accompanied by a state of oxidative stress, ROS are involved in signal transmission from primary messengers to trigger a cascade of reactions necessary for adaptation and survival in extreme conditions.

Each tissue has a certain AOP buffer capacity. It depends on the state of AOP of the intercellular fluid and the cell itself, and its individual components. Some tissues, due to the characteristics of their functional and metabolic activity, are highly sensitive to the state of oxidative stress; this is due to the high potential power of the pro-oxidant system and the low buffer capacity of AOP. Such tissues include the brain, retina, and lungs. This is due to the important regulatory function that generated ROS and radical metabolites perform in these tissues. In brain tissue, this is associated with the transmission of excitation signals, the occurrence of action potentials and the activation of synapses.

AFK - secondary messengers.

The metabolic background of any cell depends on the nature of the information coming from the environment. The carriers of this information are primary messengers: hormones, cytokines, neurotransmitters. This process is carried out through cellular signaling or signal transduction. And secondary messengers are included in signal transmission through the cell membrane. ROS actively participate as secondary intermediaries. They play a regulatory role in cell growth, apoptosis, cell adhesion, blood coagulation, etc. Low (micromolar) concentrations of ROS increase growth or enhance the response to growth stimulation in many cell types, and antioxidants suppress normal cell proliferation. Low concentrations of H2O2 stimulate fibroblast growth. Inhibition of SOD or glutathione peroxidase increases cell proliferation. HE ˙ , is a factor that enhances cell proliferation and the activity of mitogen-activated protein kinase (MAP kinase).

At physiological concentrations, ROS in the role of second messengers is formed indirectly through ligand-receptor interaction. Hormones (insulin, angiotensin, parathyroid hormone, vitamin D), cytokines, and growth factors can act as such ligands. The formation of ligand-receptor complexes is accompanied by the formation of ROS, which are actively involved in signal transduction, affecting key components of metabolic processes in the cell.

Primary messengers regulate the level of ROS in the cell by activating the processes of their generation on the one hand and reducing the activity of individual AOP units on the other. Cytokines take an active part in this process. Cytokines stimulate the release of ROS from many cell types, including human fibroblasts, epithelial and endothelial cells. ROS is associated with signal transmission from platelet-derived growth factor, epidermal growth factor, transforming growth factor β -1, tumor necrosis factor. The participation of interleukin-1 and interferon in signal transduction is associated with the formation of O2ˉ, and TNF - with H2O2.

ROS also act as secondary messengers in bone cells. TNF, interleukin-1, parathyroid hormone and vitamin D stimulate the formation of ROS due to the presence in

osteoclasts NADPH oxidase. Vasoactive peptide (angiotensin II) exerts its effect on the processes of muscle contraction and cellular growth of vascular smooth muscles through the generation of intracellular oxygen ˙ 2ˉ. Source O ˙ 2ˉ were NADH and NADPH oxidases, since both enzymes are activated by angiotensin.

ROS, as secondary messengers, are involved in the regulation of Ca2+ metabolism, stimulation of protein phosphorylation and activation of transcription factors.

In the presence of oxidants, Ca2+ transport through calcium channels increases and the ATP-dependent Ca2+ pump is inhibited.

Oxidants, increasing the activity of various

protein kinases are involved in the regulation of numerous cellular processes, such as mitogenesis, cell adhesion, apoptosis, etc.

Also involved in the action of oxidants as secondary messengers is phospholipase A2. Its activation by oxidants involves the involvement of many signal transduction pathways. Arachidonic acid, as a product of phospholipase A2, is an important mediator of processes such as inflammation, immune processes, NADPH oxidase activity, and blood coagulation.

Thus, there are 3 possible ways of action of oxidants as secondary messengers on processes in the cell associated with the formation of signaling molecules:

Influence on the structure of cell membranes.

Influence on the state of Ca2+ depot, which is accompanied by its mobilization from the depot and entry into the cytosol.

Activation of phospholipase A2.

1.3 Physiological role of free radical oxidation

Any radical is an inducer of free radical reactions. The processes of free radical oxidation (FRO) with the participation of ROS at a fairly low intensity are normal metabolic processes. Oxygen radicals induce lipid peroxidation processes necessary for the processes of phospholipid renewal and regulation of cell membrane permeability. An important physiological function of ROS is the activation of a number of membrane proteins and immunoglobulins, as well as enzymes that regulate the switching of metabolic pathways and the synthesis of high-energy compounds in the cell. Oxidative phosphorylation and the rate of cell division are directly related to FRO processes. Hydrogen peroxide can act as a metabolic signal for intracellular processes leading to the oxidation of specific SH groups of protein kinases. Once activated, these proteins translocate into the nucleus and induce the expression of groups of genes, the expression products of which are responsible for various forms of cell defense reactions. In addition, peroxide has an insulin-like effect.

The products of free radical reactions and LPO are involved in the biosynthesis of progesterone, steroid and thyroid hormones, leukotrienes, thromboxane A2, prothrombin. An important property of active oxygen metabolites and, in particular, superoxide anion radical, is the regulation of connective tissue metabolism. ROS stimulate the proliferation of fibroblasts, the synthesis and breakdown of collagen and tryptophan, and are involved in iron metabolism. Some of the oxygenases, namely a group of enzymes called cytochrome P-450 (currently there are more than a hundred isoforms), in addition to hydroxylation of endogenous compounds, use oxygen and a number of ROS for the detoxification of lipophilic xenobiotics. Cytochrome P-450 dependent monooxygenases and the electron transport system associated with them introduce ROS directly into the substrate molecule, which leads to the formation of an oxidized, more hydrophilic product. In the body, cytochrome P-450-dependent monooxygenases perform a number of important functions. Firstly, this is the oxidative biotransformation (biosynthesis or degradation) of endogenous lipophilic endobiotic molecules (steroids, retinoids, arachidonic acid metabolites), secondly, the biotransformation of xenobiotic chemical compounds coming from outside, which are not participants in normal biochemical processes in the cell and are subject to removal. The main monooxygenase reaction is always accompanied by the formation of superoxide anion radical, hydrogen peroxide, and sometimes active metabolites of the oxidized substrate. These enzymes are predominantly present in the endoplasmic reticulum of cells; the enzyme system is maximally expressed in hepatocytes, adrenal glands and gonads.

ROS are involved in the regulation of vascular tone by inhibiting endogenous nitric oxide. One of the radical forms of oxygen - peroxynitrile, which is easily converted into the starting products - nitrogen oxide and superoxide anion radical, is a transport form for NO ˙, as a result, the range of action of this signaling molecule can increase significantly. The free radical nature of peroxynitrile is the reason for the weakening and leveling of numerous physiological effects of nitric oxide. The interaction of nitrous oxide and superoxide anion radical is accompanied by a sharp increase in the oxidative potential of the latter.

Oxygen metabolites participate in the reactions of cellular and humoral immunity as regulators and effectors of these processes. In particular, oxygen radicals generated by oxidases of neutrophil granulocytes and mononuclear phagocytes play a major role in the microbicidal, cytotoxic and immunoregulatory effects of these cells. In this case, the main effectors of microbicidal action are hydroxyl radical, hydrogen peroxide and hypohalides.

ROS stimulate the proliferation of immunocompetent cells. The direct participation of the superoxide anion radical in the formation of chemotactic factors that cause activation and migration of leukocytes to the site of inflammation has been proven. The hydroxyl radical also has the ability to induce the synthesis of chemotactic peptides, which at the same time enhances mitogen-stimulated proliferation and subsequent differentiation of lymphocytes.

Many important processes, such as the generation of end products of purine metabolism and the breakdown of dopamine, are accompanied by the production of ROS.

Adrenergic stimulation physiologically leads to an increase, and cholinergic stimulation - to a weakening of the production of endogenous ROS, oppositely changes the redox potential of the cell and creates conditions for a permissive effect, when the same signal causes a different response of cells, depending on their redox state. For example, TNF causes either cell death or proliferation, because the transcription factor dependent on it is triggered only when the oxidative potential of target cells shifts.

Carrying out protective reactions, cells (macrophages and histiocytes) can repeatedly increase the production of ROS. During phagocytosis, a “metabolic explosion” occurs in phagocytes, i.e. multiple increase in energy consumption by the phagocytic cell. A significant part of this energy is spent by NADPH-dependent oxidases on the formation of superoxide radical.

ROS exert a bactericidal effect in phagolysosomes, because Unlike lysosomal hydrolases, ROS are capable of destroying intact bacterial cell walls and intact cell membranes; the oxygen-dependent mechanism of the final stage of phagocytosis is more important than the hydrolytic one.

ROS are also secreted externally during the process of exacytosis, relying on their ability to destroy the causative agent of primary alteration, by peroxidation of the membranes of neighboring cells, carry out secondary self-damage and promote the production of eicosanoid inflammatory mediators.

1.4 Regulation of free radical oxidation

The FRO rate and the content of free radicals in the body are normally maintained at a certain level by a complex, multi-stage regulation system. It can conditionally distinguish specific and nonspecific factors, the significance and contribution of which changes at different stages of oxidation.

Nonspecific factors:

mechanisms regulating the quantity and quality of the oxidation substrate and its availability;

factors affecting oxidation initiators, in particular, the state of metals of variable valence;

physicochemical properties of biological membranes;

mechanisms that maintain low O2 content in tissues.

Specific mechanisms:

enzymes responsible for the formation and metabolism of ROS (SOD, catalase, etc.);

systems that utilize peroxide products (glutathione peroxidase, glutathione reductase, etc.);

ROS interceptors (methionine, histamine, etc.);

bioantioxidants (tocopherol, ubiquinone, ceruloplasmin).

1.5 The body's antioxidant defense system

Based on the properties of radical derivatives of molecular oxygen and organic peroxides, protecting the body from their harmful effects is one of the most important tasks in maintaining homeostasis. The system for protecting tissues and cells from toxic oxygen metabolites and lipid peroxidation products can be divided into physiological (mechanisms that regulate the delivery and supply of oxygen to cells) and biochemical (the body’s own antioxidant system, i.e., a wide class of chemical compounds that reduce the activity of radical oxidative agents). processes). The physiological component of the body's AO defense system ensures a balance between the intensity of oxygen transport to cells and metabolic processes for its beneficial and safe utilization. These mechanisms for limiting free radical reactions are provided by:

the presence of a cascade of oxygen partial pressure levels, decreasing from the alveoli to the cells from 100-105 to 8-10 mmHg. Art., that is, 10-13 times.

reducing the oxygen tension in some subcellular structures by 100-1000 times compared to the partial pressure of oxygen in the capillaries. The process is mediated by a relatively large intercapillary distance and high affinity of cytochrome oxidase for oxygen;

reduction of microcirculation in tissues with an increase in the partial pressure of oxygen in arterial blood. The so-called “hyperoxic vasospasm” that occurs has several causes. Significant importance is attached to removing the vasodilating effect of CO, as well as reducing the activity of NO synthases and, accordingly, reducing the production of the main factor for vasodilation - nitric oxide. The possibility of a direct vasoconstrictive effect of oxygen is also recognized.

The biochemical AO system of the body can be divided into specific and nonspecific. A specific AO system is aimed at destroying ROS and the products of their further transformations. The action of the nonspecific AO system is associated with the prevention of conditions and possibilities for electron leakage and ROS generation during redox reactions (as part of oxidative phosphorylation) or in the process of autoxidation of substrates (microsomal oxidation).

The pathogenic effects of LPO are resisted by specialized enzyme systems and a number of non-enzymatic compounds. Specific AO enzymes include superoxide dismutase, catalase, glutathione-dependent peroxidases and transferases. This group of enzymes, localized primarily intracellularly, has the ability to destroy free radicals and also participate in the decomposition of hydroperoxides in a non-radical way. Antiradical defense enzymes are characterized by high selectivity of action directed against certain radicals; specificity of cellular and organ localization, as well as the use of metals as stabilizers, which include copper, zinc, manganese, iron and a number of others.

OxidantsAntioxidantsReactive oxygen species: O ˙ 2 - superoxide anion radical; ¹ O2 - singlet form of oxygen; OH ˙ - hydroxyl radical; H2O2 - hydrogen peroxide; Oxidized halogens: CLO ˙ - hypochloride, chloramines; Nitrogen oxides: NO˙ - Nitric oxide; Free radicals formed during lipid peroxidation: RO ˙ ,RO2 ˙ - mono-, dimeric, polymeric, cyclic, alkoxyl and peroxide radicals of fatty acids [.Enzymatic Superoxide dismutases Catalase Glutathione (GSH-)-peroxidases GSSG reductases Glutathione-S-transferases UDP-glucuronyltransferases NADP-N-quinone oxidoreductase NADP-N-quinone oxidoreductase Non-enzymatic Glutathione a-Tocopherol (vitamin E) β- Carotene Urates Bilirubin Flavonoids Albumin Ceruloplasmin Transferrin Table 1. Summary table of oxidants and antioxidants.

The content of AO enzymes in different tissues of the body varies significantly. Thus, their level and activity in connective tissue cells is several times lower than in parenchymal organs. The level of enzymatic AO in cells is under genetic control. Thus, an increase in the content of superoxide anion radical or hydrogen peroxide in the cytosol of cells is accompanied by activation of transcription of genes that trigger the synthesis of about 40 proteins, including catalase, superoxide dismutase, hydroperoxidase, and DNA repair endonuclease. Almost all influences leading to increased formation of ROS in cells induce the synthesis of enzymatic AO.

Under conditions of hypoxia and hyperoxia, i.e., conditions that enhance the formation of ROS, the level of enzymatic AOs inside cells increases. Under normal conditions, the content of enzymatic AO is relatively constant and depends little on gender; there is a slight decrease in their level with age. At the same time, the normal functioning of the body's AO system and the content of enzymatic AO are sharply disrupted in critical conditions (wounds, injuries, as well as extensive skin burns. Severe injuries are accompanied by a violation of the dynamic balance of pro-oxidant and antioxidant systems, while the content and activity of endogenous antioxidants, in including AO-enzymes, decreases sharply inside and outside cells.

As already noted, the key enzyme of AO defense is superoxide dismutase (SOD), discovered by McCord J. and Fridovich I. in 1969. SOD belongs to the class of redox enzymes and is present in significant quantities in almost all pro- and eukaryotic cells. In the human body, SOD is found in all organs and tissues, with particularly high intracellular concentrations.

In the blood, the enzyme is found in trace concentrations; when administered exogenously, it is excreted unchanged by the kidneys within 5-10 minutes. High concentrations of SOD, comparable to intracellular levels, are observed in bronchoalveolar fluid. In the extracellular space, the enzyme is present in very small quantities. As a consequence, in the case of the formation of a large number of superoxide anion radicals in the extracellular space, for example, in the focus of inflammation, the body is practically defenseless against the damaging effects of radicals.

The main function of SOD is to accelerate the dismutation reaction of superoxide radicals formed during biological oxidation. The rate of spontaneous reaction at neutral pH values ​​does not exceed 7 x 105 Mˉ1 sˉ1; in the presence of SOD, it increases to 2 x 109 Mˉ1 sˉ1 and higher, i.e. the enzyme accelerates the reaction by three to four orders of magnitude.

SOD has several isoforms that differ in the structure of the active center. The iron-containing isoenzyme is characteristic only of microorganisms; two other varieties are characteristic of human cells. Manganese-containing enzyme (Mn-SOD) is localized in mitochondria and is cyanide-resistant. In the total volume of superoxide dismutase activity, the share of Mn-SOD is small and amounts to about 15%. Up to 40% of SOD concentrated in cell nuclei comes from the manganese form of the enzyme, which dismutates up to 20% of superoxide radicals produced in the cell nucleus. The copper-zinc form of the enzyme (Cu, Zn-SOD), sensitive to cyanide, is found in the cytosol and intermembrane space of mitochondria. It is believed that the copper atom provides catalytic activity, and the zinc atom stabilizes the structure of the cytosolic isoform of the enzyme. Low molecular weight thiols, by reducing the Cu2+ ion present in the active center of SOD, activate the enzyme and accelerate the dismutation of oxygen.

In most cases, the enzyme consists of two identical subunits (molecular weight ranges from 30-40 kDa), each of which contains a catalytically active copper ion and a zinc ion, bound by a common ligand - histidine imidazole 61.

The most important role of SOD for cellular metabolism is predetermined not only by the wide distribution of this enzyme, but also by a number of its unique properties: high thermal stability, resistance to the action of proteases, denaturing agents, and a wide pH optimum of catalytic activity. The enzyme is quite stable in solutions - it can withstand heating up to 100° C for a minute, and does not lose activity in the pH range from 2 to 12.

Catalase is a heme-containing enzyme with a molecular weight of 250 kDa. Just like SOD, catalase is present in all cells of the body, but its level in different tissues varies. High levels of catalase are found in red blood cells, kidneys and liver. In the latter peroxisomes, the enzyme makes up up to 40% of the total protein. The reaction catalyzed by catalase is generally as follows.

The speed of catalysis is extremely high: one molecule of catalase per second decomposes up to 44,000 molecules of hydrogen peroxide. The activity of catalase depends on the ratio of the number of disulfide bonds to the number of sulfhydryl groups in the enzyme molecule, which participate in the formation of its active center. Because of this, catalase is sensitive to the action of thiol poisons. Due to the large molecular weight of the molecule, the enzyme practically does not penetrate cell membranes. This is some obstacle to the use of catalase drugs in practice.

The AO enzymes SOD and catalase, functioning together, promptly inactivate ROS, superoxide anion radical and hydrogen peroxide formed during normal cell metabolism, as well as during significant intensification of lipid peroxidation processes. However, these enzymes have weak activity towards lipid peroxides formed during chain reactions of lipid peroxidation. The destruction of these products is carried out with the participation of the glutathione enzyme system.

Glutathione enzyme system. Selenium-containing glutathione peroxidase can effectively decompose lipid hydroperoxides and hydrogen peroxide. Its affinity for hydrogen peroxide is higher than that of catalase, so the former works more effectively at low concentrations of peroxide, while at the same time, catalase plays a key role in protecting cells from oxidative stress caused by high concentrations of hydrogen peroxide. Glutathione peroxidase catalyzes the oxidation reaction of glutathione with the formation of its conjugated form; during the reaction, hydrogen peroxide decomposes to water.

2G-SH + H2O2 → G-S-S-G + 2H2O

The enzyme also catalyzes the reaction of reduced glutathione with lipid hydroperoxides, with the latter being converted into fatty hydroxy acids:

2G-SH + ROOH → G-S-S-G + R-OH+ H2O

Along with this, glutathione peroxidase is capable of reducing peroxides of protein and nucleic acid origin. Glutathione peroxidase is localized in the cytosol (about 70%) and mitochondria (20-30%) of all mammalian cells. The active site of the enzyme contains four selenium atoms covalently linked to form selenocysteine. A lack of selenium in the diet is accompanied by a decrease in the activity of glutathione peroxidase in cells and, accordingly, a decreased resistance of the body to oxidative damage. In severe cases, this can lead to the development of a free radical pathology similar to vitamin E deficiency, which is characterized by obesity, necrobiotic changes in the liver and hemolysis of red blood cells (Keshan disease).

Another enzyme that uses glutathione to carry out AO protection is glutathione transferase. Glutathione transferase is found predominantly in the cytosol of cells; in the human liver it makes up 2-4% of the total amount of cytosolic protein. The enzyme, or more precisely, a group of enzymes, carries out the conjugation of reduced glutathione with hydrophobic compounds and the reduction of organic peroxides. Elimination of derivatives of lipophilic xenobiotics, as well as LPO products, including fatty acid hydroperoxides, is carried out through their reduction, nucleophilic substitution or addition to the glutathione molecule:

ROOH + 2G - SH → ROH +G - S - S - G + H 2O+G - SH →R- S - G + XH+ G - SH → HR - SG

Unlike glutathione peroxidase, for which the best substrates are hydrophilic hydroperoxides with a small molecule size, glutathione transferase does not interact with hydrogen peroxide, but reduces hydrophobic hydroperoxides with a large molecule volume: hydroperoxides of polyunsaturated fatty acids - linolenic and arachidonic acids, as well as phospholipids. In addition, the enzyme restores hydroperoxides of mononucleotides and DNA, thereby participating in their repair. Glutathione transferase is an important component of AO defense, ensuring the removal of many oxidative stress metabolites. Reduced glutathione is necessary for the normal functioning of glutathione-dependent enzymes. Maintaining its sufficient level is carried out through synthesis, due to its desorption from bonds with proteins, or through recovery from oxidation of the form. This reaction is carried out by the enzyme glutathione reductase, a flavoprotein.

G-S-S-G + NADPH + H+ → 2G - SH + NADP +

The glutathione reductase system allows you to quickly replenish the pool of reduced glutathione in tissues. The main source of NADPH for this reaction is the pentose cycle of glucose oxidation, while NADPH-dependent dehydrogenases of the pentose cycle are activated by oxidized glutathione.

A decrease in the content of reduced glutathione in tissues creates favorable conditions for blocking SH groups that are part of enzymes and structural proteins of the cell. Glutathione is believed to be the main agent protecting thiol enzymes from oxidation. Thiol enzymes include many oxidoreductases, transferases, hydrolases, lyases and ligases. Oxidation of SH groups disrupts the functioning of glucose-6-phosphate dehydrogenase, lactate dehydrogenase, succinate dehydrogenase, ATPase, monoamine oxidase, xanthine oxidase, glutathione reductase and catalase. The above facts confirm the opinion that thiol compounds take the most direct and broad part in the mechanism of functioning of the enzymatic link of the AO system, performing not only an antiradical, but also an antiperoxide effect.

Water-soluble low molecular weight thiol compounds: glutathione and ergothioneine. The first of them is a component of the non-protein thiol disulfide redox system and is a tripeptide formed by the amino acids cysteine, glutamic acid and glycine. Glutathione exists in two forms - reduced (up to 97% of the total) and oxidized. Along with other sulfur-containing compounds, glutathione is an inhibitor of ROS and stabilizes cell membranes. It plays a leading role in neutralizing the hydroxyl radical formed in the Fenton reaction or as a result of rhydiolysis of water under the influence of ionizing radiation.

1.6 Pathogenetic mechanisms of disorders that develop when the balance of antioxidant and prooxidant systems is disturbed

An imbalance between the antioxidant (AOS) and prooxidant systems (POS) causes the development of oxidative stress (OS). The toxic effect of ROS manifests itself in OS conditions, which is accompanied by a sharp intensification of free radical processes in tissues. This is the most important pathogenetic link in the development of many inflammatory processes, radiation injuries, cardiovascular diseases, cancer, chemical and other intoxications. The mechanism of ROS generation in many pathological conditions is general. Some distinctive features can be identified only in the initial stages. Thus, during inflammatory processes, the triggering factor for the intensification of free radical processes is a respiratory explosion; during hypoxia, a disruption primarily of the tissue respiration system; and during chemical lesions, activation of the microsomal oxidation system. Thus, the reasons causing the intensification of free radical processes may be different, but changes at the molecular level are of the same type and the processes of ROS generation are interrelated. Some antioxidants under OS conditions can act as pro-oxidants. In OS conditions, the recovery potential of the cell increases due to substrates and coenzymes in a reduced state, which leads to a decrease in pH in areas of ischemia of heart and brain tissue. This creates conditions for increasing the pool of “active forms” of metals of variable valence. Under conditions of increased ROS generation, they can participate in reactions associated with the generation of radical products. Thus, in the presence of Fe/Cu and O2, thiols (RSH) are sources of radicals, RS , O2 -, H2O2 and OH , NADPH radicals NAD(P) , ascorbic acid is a semidehydrascorbate radical. An increase in the level of ROS is associated with an intensification of the processes of oxidative destruction of lipids, proteins, nucleic acids, and carbohydrates. It is the intensification of these processes that is the main cause of cytotoxic tissue damage.

As already indicated, the reason for the accumulation of reactive oxygen species is disturbances in the normal oxidative metabolism of cells during pathology (shunting by oxygen of electrical transport pathways in mitochondria and in the macrosomal oxidation system due to “leakage” of electrons, transformation of dehydrogenase pathways into oxidase pathways, autoxidation reactions, for example, catecholamines, etc. ).

In concentrations exceeding physiological ones, all these compounds are highly toxic for biological systems at all levels, from molecular-cellular to organismal. The ability to carry out oxidative damage and destruction of components of living systems, caused by active oxygen derivatives, is designated as “oxygen toxicity”.

Free oxygen radicals are unstable chemical compounds and easily react with biomolecules, causing their modification or destruction. The objects of nonspecific unauthorized oxidative reactions involving oxygen and its chemically active derivatives can be molecules of various chemical natures. ROS are capable of breaking any hydrocarbon bond and easily destroy high-molecular compounds: hyaluronic acid, proteoglycans, collagen, immunoglobulins. In the presence of ferrous ions, ROS convert oxyhemoglobin into methemoglobin. Oxidation of functional groups of biologically active substances causes degradation of structural proteins and lipids of cell membranes and nucleic acids, inhibition of enzymes, changes in the structure and properties of hormones and their receptors.

The result of the interaction of active oxygen derivatives with the DNA molecule is the structural modification of nitrogenous bases, the decomposition of the five-membered deoxyribose ring, as well as the cleavage of the sugar-phosphate backbone, which ultimately leads to the fragmentation of this polymer. It has been proven that excessive production of ROS, in particular, superoxide anion radical, singlet oxygen and hydroxyl radical, can cause a number of chromosomal aberrations and mutations of a number of genes in human lymphocytes. The interaction of ROS with proteins leads to structural changes in this type of biomolecule, consisting of modification of side groups of amino acid residues, fragmentation of polypeptide chains and the formation of covalent bonds within and between molecules. The tolerance of proteins to ROS depends on their amino acid composition; cyclic and sulfur-containing amino acids are more sensitive to free radical damage. Of the 20 essential amino acids, the most vulnerable to ROS, in particular to singlet oxygen, are histidine, tryptophan, methionine, tyrosine and cysteine. A distinctive feature of oxidative damage to lipids in cell membranes is that molecular oxygen can accumulate in the hydrophobic region of their fatty acid residues. As a result of this, as well as the close and parallel arrangement of neighboring fatty acid residues in relation to each other, conditions are created for the development of oxidative damage like chain reactions. Because of the key role of organic hydroperoxides in this expansion, this process is called lipid peroxidation.

2. Free radical (lipid peroxidation)

The main substrates of LPO are polyunsaturated higher fatty acids (HFA), located in the structure of membrane phospholipids. At different stages of VFA peroxidation, diene and triene conjugates, BFA peroxides (R-OO) are formed ˙), VFA hydroperoxides (R-OO), endoperoxides, malondialdehyde and new free radicals. The strongest catalyst for the process are metal ions (Fe2+). The process may be interrupted by the formation of products that do not contain free radicals.

LPO processes can be conditionally divided into three successive stages, or phases of development: processes of chain initiation, processes of development of chain reactions, and chain termination. At the stage of chain nucleation, under the influence of oxygen free radicals, ionizing radiation, ultraviolet irradiation and a number of chemical substances related to pro-oxidants, the formation of organic radicals (R) occurs.

O2 + R-H→R ˙ + HO2

O2 + R-H→R ˙ + HO2

HE ˙ + R-H → R ˙ + H2O

In the next stage, the radical quickly interacts with oxygen, which acts as an electron acceptor. As a result, the formation of peroxy radical (RO2) occurs, which attacks unsaturated lipids. The emergence of organic peroxides and a new radical (R) as a result of this reaction contributes to the continuation of oxidative reactions that acquire a chain character:

R + O2 → RO2;+ RH → ROOH + R

Organic peroxides (ROOH) are included in the process of generating radicals; in the presence of metals of variable valency (copper, cobalt, manganese, iron), a reaction alkoxy radical is formed:

ROOH + Me n+→RO + OH ˉ + Me (n+1)-

Some of the resulting organic radicals interact with each other, resulting in the formation of inactive molecules, which interrupts the course of free radical oxidation reactions:

RO2 + R → ROOR

Lipid hydroperoxides are capable of undergoing non-radical oxidative transformations, which leads to the formation of primary (diene conjugates, dialdehydes), intermediate (Schiff bases) and final lipid peroxidation products, as well as alcohols, ketones and aldehydes. Breaking chain reactions of peroxidation is possible through the interaction of radicals with specialized enzyme systems, as well as with a number of low-molecular substances that collectively form the biochemical component of the body's antioxidant system

Excessive activation of LPO can occur as a result of sudden changes in the oxygen regime of the cell. In this case, hyperoxia causes a temporary increase in lipid peroxidation processes, and persistent hypoxia leads to an avalanche-like accumulation of toxic peroxidation products. Excessive intensity of lipid peroxidation reactions develops under the influence of external influences, which include ionizing and ultraviolet radiation, as well as a number of chemical substances related to pro-oxidants. The production of ROS and, accordingly, the intensity of lipid peroxidation processes in cells sharply increase as a nonspecific reaction when the body is exposed to various stress factors of a chemical, physical and biological nature. Ultimately, all these effects can lead to tension and subsequent decompensation of the body's antioxidant defense mechanisms and the development of oxidative stress, manifested at the cellular, tissue and organismal levels. Typical pathological processes, such as hypoxia and inflammation, characteristic and developing in most somatic and infectious diseases, severe injuries and wounds, are always accompanied by excessive formation of ROS and lipid peroxidation products

Long-term activation of lipid peroxidation processes is accompanied by the development of lipid peroxidation syndrome. With excessive activation of lipid peroxidation, when a significant part of membrane phospholipids undergoes oxidative degradation, the lipid phase of membranes becomes more rigid. This limits the conformational mobility of the polypeptide chain, as a result of which the functional activity of enzymes, receptors and channel-forming proteins built into the membranes is reduced, which in turn prevents the removal of Ca+2 from the sarcoplasm and ensures the damaging effect of calcium on cellular organelles. To date, the ability of LPO metabolites to inhibit the K+/Na+-ATPase of the plasma membrane and cause cytolysis of hepatocytes with the development of fermentemia has been proven. LPO products disrupt the normal functioning of L-type calcium channels, which is accompanied by electrolyte disturbances in myocardial cells. Their excess leads to a shift in the metabolism of arachidonic acid, as a result of which one of the resulting compounds (15-hydroarachidonic acid) reduces β- adrenergic stimulation due to inhibition of this type of adrenergic receptors. The products of these reactions damage the membranes of erythrocytes, oxidize serum albumin, and disrupt the synthesis of nucleic acids in cells.

Activated oxygen species and lipid peroxidation products damage hyaluronic acid, proteoglycans, collagen, and immunoglobulins. The generation of activated oxygen species that is not compensated by the antioxidant system can cause a damaging effect on the endothelium and vascular basement membrane directly or indirectly through the inactivation of antioxidant systems. Morphologically detectable lipofuscin granules, consisting of deposits of insoluble lipoperoxides and proteins, are products of lipid peroxidation.

ROS and lipid peroxidation products contribute to the excessive production and release of a number of pro-inflammatory cytokines (tumor necrosis factor, interleukin-1, interleukin-6) and inflammatory mediators (histamine, bradykinin, serotonin), arachidonic acid derivatives - leukotrienes, prostaglandins and thromboxanes. Excessive production of ROS and lipid peroxidation products causes a sharp activation of the complement system along the alternative pathway with the release of anaphylatoxin.

2.1 Participation of ROS and LPO products in the pathogenesis of human diseases

Most or almost all pathological conditions, the course of which is accompanied by the participation of ROS, are characterized by the so-called state of oxidative stress, characterized by intensified production of these substances. It should be noted that the state of oxidative stress is characteristic not only of diseases, but also occurs in physiological states close to extreme - intense physical and psycho-emotional stress, overwork, as well as during the aging of the body. There is a “free radical theory of aging”, its creator D. Harman, who first outlined the main provisions of the theory in 1955. Harman’s theory is based on a system of arguments associated with the ability of free radicals to nonspecifically damage macromolecules such as DNA, lipids and proteins, and the aging process is associated with the accumulation of such damage in the body. Within the framework of this theory, not only the aging of the body is explained, but also a wide range of pathological processes associated with aging: cardiovascular diseases, age-related brain dysfunctions, immunosuppression, carcinogenesis and other types of pathology.

In all diseases where an inflammatory reaction is present as an obligatory component, the leading role in damage to cells and tissues of the body is played by ROS produced by leukocytes, mainly nertrophils. Although the most destructive forms of ROS generated by myeloperoxidase act in the phagolysosomes of the leukocyte, at the site of inflammation, neutrophils are able to secrete this enzyme into the extracellular space. The destruction of an infectious agent during the development of the inflammatory process can also be achieved at the cost of collateral damage to one’s own tissues at the site of inflammation, while oxygen-dependent tissue damage is both direct and indirect. The latter is realized when ROS influences regulatory and effector molecules, for example, proteinases and proteinase inhibitors. ROS can directly activate latent forms of collagenase secreted by neutrophils, as well as inactivate the universal proteinase inhibitor - α 2-macroglobulin and specific inhibitor of serine proteinases - α ı- antitrypsin. ROS, generated intracellularly and also penetrating through the cell membrane, are triggers for the induction of apoptosis.

The free radical mechanism of damage to plasma, mitochondrial and nuclear membranes, the nuclear and mitochondrial genome, blood lipoproteins leads to damage to blood vessels and histohematic barriers, which plays an important, often decisive role in the pathogenesis of the most common diseases of inflammatory, toxic and autoimmune nature.

The pathogenetic role of ROS has now been identified for approximately hundreds of human diseases. This occurs in cardiovascular pathology - coronary heart disease, myocardial infarction, acute arterial obstruction of a limb segment, ischemic and reperfusion damage to the kidneys, brain and other tissues, in the development of cataracts and atherosclerosis. It has been shown that the formation of atherosclerotic plaques in the vascular intima occurs with the active participation of free radicals interacting with polyunsaturated fatty acids. The pathogenesis of systemic lupus erythematosus is based on the increased sensitivity of nucleic acids (in particular, nuclear DNA) to the damaging effects of free radicals. A predisposing factor for the disease is a diet with excess amounts of polyunsaturated fatty acids.

ROS play an important role in bronchopulmonary pathology, both in acute and chronic diseases (for example, emphysema, asthma and chronic bronchitis). Free radicals are extremely important in the pathogenesis of respiratory distress syndrome. It is ROS produced by blood neutrophils that play the leading role in damage to the endothelial-alveolar barrier, the development of interstitial edema and the full-blown clinical picture of this syndrome. Excessive and prolonged stimulation of enzyme systems responsible for the generation of active radicals of phagocytes underlies the mechanisms of formation of occupational dust disease and its complications such as systemic collagenosis and malignant degeneration of cellular elements of the lungs and pleura. A long-term retrospective study conducted in 23 countries under the auspices of WHO revealed a direct dependence of the incidence of certain types of malignant neoplasms of the lungs, breast, ovaries and rectum on the amount of animal fat consumed and the intensity of lipid peroxidation processes in the body.

Numerous studies indicate that free radical oxidation processes underlie the pathogenesis of many chronic liver diseases, and excessive formation of ROS and lipid peroxidation products manifests itself at the earliest stages of the process. The hepatotoxic effect of oxygen radicals manifests itself on cell membranes, which are the main site of superoxide anion radical synthesis. Increased ROS production as a result of induction of xanthine oxidase by interferon occurs even during viral infection. Oxygen radicals damage the membrane apparatus of the hepatocyte, destroy the lipid layer of its membranes, and also damage proteins - enzymes of the monooxygenase system. The severity and prognosis of chronic liver disease are closely related to the functional state of the antioxidant system of hepatocytes.

Excessive activation of LPO is the cause of the onset and progression of hypertension. It has been established that under the influence of ROS and lipid peroxidation products, the permeability of cell membranes and the activity of Na+/K+-ATPase are disrupted, which leads to the accumulation of ionized calcium in cells. In addition, secondary LPO products can directly affect the tone of arterial vessels. In addition to the vasoconstrictor effect, oxygen radicals activate the work of connective tissue fibroblasts, resulting in the development of arteriocapillary fibrosis, i.e., irreversible changes in blood vessels. Oxidative stress almost always develops in infectious diseases. Excessive formation of oxygen free radicals is one of the leading mechanisms in the pathogenesis of influenza. The generation of ROS determines the mutagenesis and proteolytic activity of the influenza virus, the cytopathic effect of viral infection, the destruction of the capillary network and capillary walls, the development of confluent pneumonia and pulmonary edema.

Toxic damage to neurons by ROS, lipid peroxidation products and oxidative stress are considered as the leading mechanism in neurodegenerative diseases. One of the reasons for the development of multiple sclerosis may be mitochondrial disorders accompanied by excessive production of ROS. Active oxygen in multiple sclerosis has a direct damaging effect on the central nervous system and is a causative factor in the development of the disease. Oxidative damage leads to the death of nerve and glial cells, which is manifested by demyelination and the formation of foci of gliosis in the central nervous system. In 37% of people with Parkinson's disease, there is a defect in the first complex of the respiratory chain in the cells of the substantia nigra. The gradual accumulation of oxygen metabolites, lipid peroxidation products, metalloproteins and the development of oxidative stress in various parts of the central nervous system are of particular importance in Alzheimer's disease and Huntington's chorea. The accumulation of lipofuscin in CNS cells under these conditions correlates with the level of antioxidant enzymes in brain tissue.

ROS and lipid peroxidation products have a direct destructive effect on internal organs and lead to the development of multiple organ failure after severe injuries and extensive burns. Radiolysis of water, the formation of free oxygen radicals and various lipid peroxidation metabolites underlie the pathogenesis of acute and chronic radiation injuries in humans.

2.2 Diagnostics of peroxidation processes

There are no clear clinical signs of excessive activation of free radical oxidation in the human body. The development of oxidative stress in a patient is accompanied by:

signs of respiratory distress and failure

blood circulation (a hyperdynamic type of blood circulation often occurs);

psychomotor agitation;

an increase in temperature of more than 37.5° C;

pronounced pain reaction.

The authors consider laboratory signs of oxidative stress to be:

increase in the number of leukocytes in the blood (> 10 thousand) without band shift in the near future after injury (10-12 hours);

increased levels of alanine and aspartic transaminases in the blood;

increased plasma glucose concentration > 7.0 mmol/l;

uncompensated metabolic acidosis (pH< 7,2);

For laboratory diagnosis of the severity of oxidative stress and determining indications for the use of drugs with antioxidant properties, it is advisable to evaluate the state of the antioxidant system, as well as the intensity of free radical reactions (LPO processes) in the patient’s body. Assessment of the state of the antioxidant system usually includes determination of the activity of the main enzymatic antioxidants (activity of SOD, catalase and glutathione peroxidase of erythrocytes), as well as the content of ascorbic acid, tocopherol and selenium in the blood. Instrumental assessment of the severity of free radical reactions involves the determination of peroxidation reaction products in blood or tissue samples, which include diene conjugates, malondialdehyde, Schiff bases and reactive oxygen species. The combination of these indicators makes it possible to assess both the state of the main links of the AO system of the body, i.e. its ability to prevent excessive reactions of free radical nature, and the actual intensity of LPO reactions and ROS synthesis. Based on this, these tests can be considered as laboratory criteria for diagnosing oxidative stress.

Laboratory diagnostics of the intensity of lipid peroxidation is necessary to judge the AO activity of a particular drug, since based on clinical signs it is impossible to adequately assess the effect of the AO drug. Note that using only one of the above tests does not provide complete information. The dynamics of indicators of the intensity of free radical reactions and the content of AO in the body sometimes have the opposite direction. In some cases, fluctuations in the content of free radical reaction products are not accompanied by changes in the levels of reactive oxygen species, and vice versa. A sharp intensification of free radical reactions and accumulation of the products of these reactions in the blood may not lead to a change in the content and activity of the main components of the AO system. For reliable and adequate clinical interpretation of the results obtained, not only the absolute value of the indicators is of fundamental importance, but their dynamics against the background of treatment measures.

Laboratory criteria for diagnosing oxidative stress

Assessing the state of the patient's antioxidant system Determining the intensity of free radical reactions Content and activity of antioxidant enzymes in erythrocytes or other cells available for study: Superoxide dismutase Catalases Glutathione peroxidaseContent in blood plasma (serum) of primary, secondary and intermediate products of lipid peroxidation: Diene conjugate Malondialdehyde Schiff basesContent of low molecular weight antioxidants in the blood: Ascorbic acid Tocopherol SeleniumContent of reactive (radical) forms of oxygen in the blood: Superoxide anion radical Singlet oxygen Total level of radicals Each of the proposed indicators has its own advantages and disadvantages. The main requirements for laboratory methods are accuracy, ease of implementation and good reproducibility. The ease of preparing samples for testing and the possibility of storing them are desirable, which makes it possible not to carry out all tests ex tempore. The most widely used method in experimental research and clinical practice for determining malondialdehyde in the reaction with thiobarbituric acid is not an accurate indicator reflecting the course of lipid peroxidation processes, since during the reaction thiobarbituric acid interacts with non-lipid substances containing a keto group, for example , with glucose. The interaction of thiobarbituric acid with malondialdehyde, initially contained in lipid systems, plays an insignificant role in quantitative terms. Nevertheless, for almost thirty years, the method has been the most used by most researchers when studying lipid peroxidation processes in various biological systems. Its advantages include simplicity and relative speed of execution, as well as good reproducibility of results. Other common methods for assessing the intensity of LPO reactions are to determine the level of primary and intermediate products of these reactions, i.e., diene conjugates and Schiff bases. The methods are accurate and relatively easy to reproduce. Their disadvantages include the impossibility of storing selected samples, so studies must be carried out immediately or within half an hour after their collection. A more accurate and adequate indicator of the intensity and severity of free radical reactions and, accordingly, the state of the body's antioxidant system is the determination of ROS - the main precursors of LPO reactions; fast and accurate measurement of the level of which in whole blood is carried out by the luminescent method. Depending on the type of phosphor used (luminol, lucigenin, etc.), it is possible to determine both individual types of ROS and their total quantity. The method is fast and can be recommended for express diagnostics. The technique allows you to store heparinized blood samples for research for up to 12 hours at a temperature of +4˚C, i.e. in a regular household refrigerator.

Main indicators characterizing the oxidant-antioxidant system.

The intensity of LPO is assessed by the concentration in the blood and other biological fluids of intermediate and final reaction products.

Malondialdehyde (MDA) is the end product of LPO.

Normal concentration in the blood is 2.5-6.0 µM/l. Depends on the determination method, so each laboratory has its own reference interval.

An increase in concentration is evidence of increased LPO and disruption of antioxidant protection.

Determination of malondialdehyde in blood by fluorimetric method (Fedorova T. N., Koryatsiva T. S., Larsky E. G.)

The principle of the method: thiobarbituric acid (TBA) in an acidic environment reacts with low molecular weight dialdehydes (mainly malonic acid) to form a pink-colored complex.

Standard values: 3.7±0.12 units.

Modification of the determination of lipid peroxidation products in the reaction with thiobarbituric acid (Korobeinikov E. N.)

The principle of the method: when heated in an acidic environment, part of the LPO products belonging to the class of hydroperoxides decomposes with the formation of malondialdehyde, the interaction of a molecule of which with two molecules of thiobarbituric acid leads to the formation of a colored complex.

Standard values: 3.69±0.14 nmol/l.

Determination of lipid peroxides in the reaction with thiobarbituric acid (Gavrilov V. B., Gavrilova A. R., Mazhul L. M.)

Principle of the method: thiobarbituric acid reacts with malondialdehyde, which is formed during the peroxidation of unsaturated fatty acids having 2-3 diene bonds, to form a pink product with a maximum absorption at 535 nm.

Standard values: 100-120 nmol/l.

Determination of diene conjugates in blood plasma by UV absorption

heptane and isopropanol extracts (Gavrilov V.B.,

Gavrilova A. R., Khmara N. F.)

Principle of the method: the method is based on measuring the absorption intensity in the region of 223-234 nm, caused by conjugated diene structures (previously extracted from plasma) that arise during the formation of hydroperoxides of polyunsaturated fatty acids.

Spectrophotometric determination of the content of acyl hydroperoxides (diene conjugates) in blood plasma (serum).

The principle of the method is based on determining the content of primary lipid peroxidation products in the blood by the absorption of a monochromatic light flux by a lipid extract in the ultraviolet region of the spectrum (233 nm).

Vitamins with antioxidant properties.

Vitamin A (retinol) is a fat-soluble vitamin found in animal products in the form of A and A2, in plants in the form of provitamin - carotene, the most active of which is beta-carotene.

Normal serum concentration is 1.05-2.27 µM/L or 300-650 µg/L.

Vitamin A deficiency, which can occur not only with a deficiency of retinol or carotene in food, but also due to impaired absorption due to intestinal diseases, as well as with a small amount of fat and protein in food, contributes to increased LPO. The epithelium of various organs and the eye suffers to a greater extent (slow recovery of visual purple - rhodopsin). This is manifested by increased susceptibility to bronchopulmonary diseases, urinary tract infections, mucous membranes of the eyes, skin, and decreased visual acuity in the dark (“night blindness”).

Vitamin E (tocopherols) is a group of substances, among which alpha-tocopherol has the greatest activity. A fat-soluble vitamin that acts in conjunction with vitamin A. In blood serum and cells, the main part is associated with lipoproteins.

Determination of the level of alpha-tocopherol in blood plasma.

Principle of the method: Plasma alpha-tocopherol is extracted with an ethanol-hexane mixture. The selected hexane phase is evaporated. Ferric chloride is added to the resulting residue, which is capable of being reduced from the trivalent state to the divalent state under the influence of antioxidants (and primarily alpha-tocopherol). The amount of alpha-tocopherol is judged by the content of reduced iron, which is determined by the reaction with alpha-2-, alpha-2-dipyridyl, accompanied by the formation of a red complex.

Normal concentration in blood serum is 5-20 mg/l.

The degree of supply of the body with vitamin E can be characterized by directly determining its concentration in the blood, and indirectly by determining the peroxide resistance of erythrocytes.

Peroxide resistance of erythrocytes (PRE) is an indicator of the provision of membranes with antioxidants, primarily vitamin E, and their resistance to the damaging effects of peroxides.

The normal value is up to 10%.

The increase indicates insufficient antioxidant protection of cell membranes, high susceptibility to lipid peroxidation and lability. There are seasonal fluctuations in ERP with an increase in the percentage of peroxide hemolysis in the spring. .

Vitamin C (ascorbic acid) is a water-soluble vitamin that, unlike animals, is not synthesized in the human body and comes only with food. Prevents the oxidation of vitamins A and E, restores oxidized forms of enzymes and various substrates, and participates in a large number of metabolic reactions. Regulates the permeability of the vascular wall, significantly increases the body's resistance to infections, and is necessary for the formation of collagen, healing of wounds and burns.

Normal concentration:

in blood -6-20 mg/l,

in urine -20-30 mg/day.

Daily requirement - 45-60 mg; increases significantly with physical and neuropsychic stress, low and high temperatures, radiation, smoking (1 cigarette destroys 25 mg of vitamin C), pregnancy and lactation, chronic diseases of the gastrointestinal tract, taking antibiotics and sulfonamides. Ascorbic acid is not recommended for cancer patients during radiation and chemotherapy.

Lack of vitamin C in food contributes to increased lipid peroxidation, metabolic disorders of many substances, functions of the nervous and endocrine systems, decreased resistance to infections and is manifested by weakness, increased fatigue, irritability, pain in the limbs and other nonspecific symptoms. Long-term vitamin deficiency causes the development of scurvy.

Antioxidant enzymes.

Superoxide dismutase (SOD) is an enzyme that catalyzes the reaction of neutralizing the superoxide radical O2. It is the main enzyme of intracellular antiradical defense. Normal activity in erythrocytes (NBT recovery method) is 1.04 ± 0.05 arb. units/mg hemoglobin. Absolute values ​​vary depending on the reagents used. The most accessible for analysis is erythrocyte SOD containing CuZn.

Determination of erythrocyte SOD activity by the R. Fried method. Principle of the method: the method is based on determining the degree of inhibition of the reduction reaction of nitroblue tetrazolium by superoxide radicals under the influence of erythrocyte SOD.

The study of SOD activity is performed at room temperature. Blood should be drawn with heparin. The enzyme can remain stable for up to 1 week when stored at +4 C.

Enzyme activity is expressed in arbitrary units (arbitrary units). 50% inhibition of the reduction process of nitroblue tetrazolium compared to the control sample is considered to be 1 arb. units

Standard values: 1.04±0.3/mg hemoglobin.

During the initial period of increased formation of free radicals, SOD activity increases to neutralize them. If the process of radical formation continues to intensify, then at a certain stage, compensatory reserves are depleted and SOD activity decreases below normal, which indicates decompensation of this protective mechanism.

An increase in SOD activity is observed in poisoning with hashish, carbon tetrachloride, amitriptyline, organophosphorus compounds, acetic acid, septicemia, focal tuberculosis, leukemia, Duchenne muscular dystrophy, cystic fibrosis, thalassemia, myocardial infarction, and mental illness.

An increase in SOD activity without corresponding activation or absence of other antioxidant enzymes - catalase (hypo-, acatalasemia), peroxidases is an unfavorable change for the body due to the accumulation of hydrogen peroxide - a product of the superoxide dismutase reaction.

A decrease in SOD activity is observed in case of poisoning with carbon monoxide, lead compounds, cadmium, with septicopyemia, with ischemic heart disease, cerebral atherosclerosis, with epilepsy, in newborns with respiratory distress syndrome, with retinopathy, in pregnant women with late toxicosis (diagnostic test for determining intrauterine hypoxia fetus). Low levels of enzyme activity should be considered as an unfavorable prognostic sign, indicating a decrease in the nonspecific resistance of the body.

It is advisable to determine SOD activity with simultaneous study of other AOD components and LPO indicators.

Catalase is an enzyme that decomposes hydrogen peroxide into oxygen and water. The maximum amount is found in red blood cells. It has a specific antioxidant protective function against endothelial cells.

Normal activity is 18.4 - 25.0 µU/erythrocyte.

In the initial compensatory phase of radical formation there is an increase in enzyme activity, and in the decompensation phase there is a decrease. With age, enzyme activity decreases.

Determination of catalase activity using the E. Beutler method.

Principle of the method: the method is based on determining the rate of decomposition of hydrogen peroxide in mM/min spectrophotometrically at a wavelength of 230. Ethanol is added to stabilize the hemolysate and decompose the catalase-hydrogen peroxide complex.

Determination of blood catalase activity using the Bach and Zubkova method.

Principle of the method: the method is based on determining the amount of hydrogen peroxide that is destroyed by the enzyme in 30 minutes. In the experiment, the amount of hydrogen peroxide not destroyed during the catalase reaction is determined; in the control, the amount of peroxide taken in a sample with the enzyme inactivated by boiling is determined. Hydrogen peroxide is titrated with potassium permagane in an acidic medium. The reaction follows the equation:

2KMnO + 5H2O2 + 4 H2 SO + 2 KMnSO + 8H2O +5O2

The difference between the titration numbers of the control and experiment corresponds to the amount of hydrogen peroxide destroyed under the action of catalase.

Standard values:

Titrometry method: catalase number 12-20. In one µl. Men's blood contains 4·10-5·10 erythrocytes, women's blood contains 3.9·10 - 4.7·10 erythrocytes.

Photometric method: according to E. Beutler 15.31·10 ± 2.39·10 IU/g hemeglobin.

An increase in catalase activity is observed in hemolytic conditions, during surgical interventions performed under local anesthesia, in children with bronchopulmonary pathology, rheumatoid arthritis, and thyrotoxic goiter.

A decrease in catalase activity is observed in infectious diseases, iron deficiency anemia, malabsorption syndrome, carcinoma, in newborns with respiratory distress syndrome, chronic poisoning with phosphorus, arsenic, lead, mercury, general anesthesia, and the prescription of antibiotics. It is advisable to determine the activity of catalase with a simultaneous study of the activity of other AO enzymes, the content of methemoglobin and lipid peroxidation products.

The most common method for determining GR activity is the spectrophotometric method. Among other methods, noteworthy is the colorimetric determination of GR activity by the rate of formation of reduced glutathione during iodometric titration or using a color reaction with nitropruside.

Determination of GR activity by the Beutler method.

Principle of the method: the method is based on the spectrophotometric determination of the amount of coenzyme NADPH consumed during the enzymatic reaction, taken into account by the change in absorption at 340 nm. The wavelength of 340 nm corresponds to the maximum absorption of the reduced coenzyme. During the reaction, as NADPH is oxidized, the optical density of the incubation sample decreases.

GR activity is significantly increased in the serum of patients with sarcoma, in patients with breast carcinoma and in other neoplastic diseases. An increase in GH is observed in hepatitis, obstructive jaundice and, less commonly, in cirrhosis. High activity values ​​can be determined during acute myocardial infarction. As a rule, megaloblastic anemia is accompanied by an increase in the level of GH activity.

Glutathione peroxidase (GP). Catalyzes the oxidation reaction of glutathione G-SH with hydrogen peroxide (1st reaction) or ROOH hydroperoxides (2nd reaction), formed as a result of LPO.

The contribution of HP to the neutralization of hydrogen peroxide is more significant compared to catalase. It has a greater affinity for hydrogen peroxide and decomposes it even at low concentrations. The active site of the enzyme contains selenium. There are 4 grams of selenium atom per 1 mole of enzyme.

Determination of GP activity using the E. Beutler method.

Principle of the method: the method is based on determining the rate of formation of oxidized glutathione, the content of which is determined in the coupled glutathione reductase reaction by the degree of oxidation of NADPH at 340 nm.

G - S - S - G + NADPH→2G - SH + NADP

T-butyl hydroperoxide is used as hydroperoxide.

Standard values: According to Beutler 30.8±4.73 IU/g hemoglobin (when taking blood with EDTA) and 34.2±3.84 IU/g hemoglobin (in heparinized blood). According to Titz: 19.9±0.31 IU/mol hemoglobin and 0.89±0.14 WEEK/erythrocyte.

Increased activity is observed with glucose-6-phosphate dehydrogenase deficiency , α- thalassemia, acute lymphocytic leukemia.

A decrease in GP activity is observed in iron deficiency anemia, lead poisoning, sickle cell anemia, and selenium deficiency.

Ceruloplasmin (copper oxidase). Copper-containing glycoprotein α2- globulin fraction. The functions of ciruloplasmin are diverse: it is the main oxidase of blood plasma, an acute phase reactant, and transports Cu for the synthesis of SOD and cytochrome oxidase. Ciruloplasmin exhibits its AO properties extracellularly without the formation of any radicals.

Pure ceruloplasmin protein has an intense blue color.

The simple colorimetric method proposed by Ravin is widely used in modification (S.V. Bestuzhevoy and V.G. Kolba).

There are other research methods: nephelometric, manometric and immunological. With the manometric method, the reaction takes place in a Warburg apparatus; the rate of oxygen consumption is measured (μmol of oxygen consumed in 1 minute per 1 liter of serum under certain conditions). An excellent correlation was noted between the values ​​obtained by the Revin colorimetric method and the monometric method. The colorimetric method has been modified for use in an autoanalyzer.

Determination of ciruloplasmin activity using the modified Revin method.

Principle of the method: the method is based on the non-enzymatic oxidation of n-phenylenediamine with ceruloplasmin. The reaction is stopped by adding sodium fluoride. The optical density of the resulting colored products is used to judge the concentration of ceruloplasmin.

The level of ceruloplasmin in serum increases in various infectious diseases, in acute and chronic inflammatory processes accompanied by destructive and necrotic changes in tissues, in malignant growth and schizophrenia.

Antioxidant activity (AOA) is a complex of enzymatic and non-enzymatic reactions of binding and decomposition of intermediate peroxidation products that inhibit free radical oxidation of lipids. It is most often determined by chemiluminescence in model systems.

The normal value for blood serum is 60-75%.

A high level of AOA ensures resistance to peroxide damage to cell membranes and a low level of lipid peroxidation.

A low level of AOA promotes increased lipid peroxidation, inhibition of proliferation and regeneration processes.

The clinical significance of determining AOA is that some pathological processes develop against the background of increased AOA, while others develop against the background of decreased AOA and therefore require multidirectional correction. .

Conclusion

Thus, the basis for maintaining free radical homeostasis is the balance between pro-oxidant and antioxidant processes that maintain peroxidation within limits that are not only compatible with life, but also beneficial for it. Violation of this balance is the starting point in the initiation of “free radical pathology.” The breakdown of antioxidant protection is characterized by the development of free radical damage to various components of cells and tissues. All cellular components are susceptible to peroxidation to one degree or another, but this process is most pronounced in lipid (phospholipid) structures, primarily in the lipid bilayer of membranes. Violation of pro- and antioxidant balance, accompanied by an increase in the concentration of lipid peroxidation products in the tissues and fluids of the body, has been noted in a wide variety of diseases: inflammatory, cardiovascular, oncological, infectious, burn and radiation diseases, with various toxic effects, as well as with aging of the body. All this allows researchers to consider LPO activation as a universal component of the body’s nonspecific response to extreme influences, i.e. as a part of the stress response.

From the above it follows that the mechanism of maintaining pro-oxidant-antioxidant balance is quite complex. On the one hand, oxidizing and pro-oxidizing factors and substrates act: molecular oxygen, OH ˙, hydroperoxides, organic peroxides, epoxides, easily oxidized substrates (lipids), oxidative enzymes and free metal ions with variable valency, neurotransmitters (catecholamines). On the other hand, antioxidant components; enzymes (SOD, GPO, glutathione transferase, catalase), hormones (steroid and thyroid), bioamines (serotonin, histamine), fat-soluble antioxidants - membrane components (tocopherols, ubiquinones, retinoids, carotenoids, phenolic compounds); water-soluble antioxidants (thiol compounds, ascorbate, water-soluble phenols); Selenium ions - free and as part of antioxidant enzymes.

For laboratory diagnosis of the severity of oxidative stress and determining indications for the use of drugs with antioxidant properties, it is advisable to evaluate the state of the antioxidant system, as well as the intensity of free radical reactions (LPO processes) in the patient’s body. As a result of the analysis of the literature, we can conclude that in order to assess the state of the intensity of lipid peroxidation in the human body, it is necessary to use integrative approaches; the use of only one of the above tests does not provide complete information. Also, laboratory diagnostics of the intensity of lipid peroxidation is necessary to judge the AO activity of a particular drug, since based on clinical signs it is impossible to adequately assess the effect of the AO drug.

List of sources used

Dubinina, E. E. Oxidative stress as a reaction of the body’s adaptation to extreme conditions / E. E. Dubinina // Issues of medical chemistry. - 2001. - No. 6., volume 47 - P. 561-581.

Balabolkin, M. I. The role of oxidative stress in the pathogenesis of vascular complications of diabetes / M. I. Balabolkin, E. M. Klebanova // Problems of endocrinology. - 2000. - No. 6. - P. 29-34.

Vladimirov, Yu.A. Free radicals in major systems / Yu.A. Vladimirov, O.A. Azizova; ed. A.I. Deev. - M.: VINITI, biophysics series, 1991. - 252 p.

Free radicals. Definition, nomenclature, classification [Electronic resource] / Academician of the Russian Academy of Medical Sciences, Professor Vladimirov Yu.A. - 2006. - Access mode: #"justify">Vladimirov, Yu.A. Lipid peroxidation in biological membranes / Yu.A. Vladimirov, A.I. Archakov. - M.: Medicine, 1972. - 252 p.

Shanin, Yu.N. Antioxidant therapy in clinical practice / Yu.N. Shanin, V. Yu Shanin, E. V. Zinoviev. - St. Petersburg, ELBI - SPb., 2003 - 128 p.

Zaichik, A.Sh. General pathophysiology (with basics of immunology) / A.Sh. Zaichik, L.P. Churilo. - St. Petersburg, ELBI - St. Petersburg, 2005. - 656 p.

Kashulina, A.P. The role of free radical peroxide oxidation in pathology and methods of its study / A.P. Kashulina, E.N. Sotnikova // Med. consulting. - 1996. - No. 2. - P. 20-24.

Bueverov, A. O. Oxidative stress and its role in liver damage / A. O. Bueverov. // Russian Journal of Gastroentorology. - 2002. - No. 4. - P. 21-25.

Khutsishvili, M. B. Free radical processes and their role in the pathogenesis of certain diseases / M. B. Khutsishvili, S. I. Rapoport. // Clinical medicine. - 2002. - No. 10. - P. 10-16.

Troy, C.M. Down-regulation of copper zinc superoxide dismutase causes apoptotic death in PC 12 neuronal cells / C.M. Troy // Proc. Natl. Acad. Sci. USA. - 1994. - No. 14. - P. 6384 - 6387.

Boldyrev, A. A. Introduction to biomembranology: textbook. allowance / Ed. A. A. Boldyreva. - M.: Moscow State University Publishing House, 1997. - 208 p.

Damage to components of biological membranes during pathological processes [Electronic resource] / M.: - 2006. - Access mode: #"justify">Radi, R. Peroxynitrite-induced membrane lipid peroxidation: the cytotoxic potential of superoxide and nitric oxide / R. Radi //Arch. Biochem. Biophys. - 1991. - No. 2. - P. 481-487.

Danilova, L. A. Collection of laboratory research methods / Ed. L. A. Danilova. - St. Petersburg: Peter, 2003. - 736 p.

Lifshits, V.M. V. I. Sidelnikova. ″Medical laboratory tests″. Directory. Second edition, corrected and expanded / V.M. Lifshits, V.I. Sidelnikova. - M.: ″Triad - X″, 2003. - 312 p.

Nikolaev, A. Ya. Biological chemistry / Nikolaev A. Ya. - M.: Medical Information Agency, 2001. - 496 p.

Similar works to - Laboratory diagnostics of the intensity of lipid peroxidation

Peroxide oxidation

a complex multi-stage chain process of oxygen oxidation of lipid substrates, mainly polyunsaturated fatty acids, including the stages of interaction of lipids with free radical compounds and the formation of free radicals of lipid nature. O. p. phospholipids of biological membranes play an important role in the life of living organisms. Strengthening the processes of O. p. is essential in the etiology and pathogenesis of many diseases and the development of the consequences of various extreme influences.

Peroxide oxidation is a special case of liquid-phase oxidation of hydrocarbons. It is a typical chain process with pronounced branching. O. p. can include stages of non-enzymatic autoxidation and enzymatic reactions. The enzymatic and non-enzymatic pathways of oxygen production lead to the formation of free radicals of lipids in several main stages: initiation (nucleation of the chain), continuation of the chain; chain branch ; chain termination molecular products, molecular products, molecular products, where RH is the oxidation substrate (polyunsaturated fatty acid). The so-called reactive oxygen species, primarily oxygen radicals containing unpaired electrons, play a decisive role in the initiation of opacification. As a result of the one-electron reduction of molecular oxygen O 2 in cells, a superoxide anion radical is formed, which occurs in the electron transfer chain of mitochondria, chloroplasts, in reactions catalyzed by certain oxidative enzymes, during the autoxidation of monoamines and other compounds. During the dismutation reaction of two superoxide radicals, a molecule of hydrogen peroxide H 2 O 2 is formed; Other sources of hydrogen peroxide include reactions catalyzed by certain oxidases. Cells have special systems for neutralizing toxic oxygen radicals, in particular enzymatic systems: superoxide dismutase, which catalyzes the conversion of superoxide into hydrogen peroxide, and peroxidases, which catalyze reactions in which hydrogen peroxide is reduced to water. The most reactive and therefore the most dangerous oxygen radicals include the hydroxyl radical OH - one of the main damaging factors when exposed to ionizing radiation (ionizing radiation). A significant part of OH radicals in living organisms is generated as a result of reactions of hydrogen peroxide and superoxide radicals with catalytic amounts of metals of variable valence, primarily with ions and copper. Relatively low-active and long-lived H 2 O 2 can serve as a source of the OH radical, which interacts with almost all classes of biomolecules in the presence of trace amounts of free iron or copper. Along with the OH radical, other free radicals, for example, protonated superoxide anion, as well as singlet and a number of other reactive oxygen species, can be direct initiators of oxygen reactive reactions.

O. p. products, in particular lipid peroxides, are used in the body for the synthesis of biologically active substances - prostaglandins (Prostaglandins), thromboxanes, steroid hormones (Hormones), etc. The intensity of O. is directly related to the processes of renewal of the composition of phospholipids in biological membranes, changes in the relative content of lipids and proteins, and, as a consequence, changes in the structure of biological membranes and their functioning. In living organisms, there is a complex regulation of the intensity of the oxygen production process. Normally, the processes of formation and consumption of oxygen products are well balanced, which determines their relatively low content in cells. The rate of oxygen production at the levels of initiation, continuation, and chain termination is largely determined by the structural organization of lipids in the biological membrane. which affects the residues of unsaturated fatty acids (fatty acids) for oxygen. Factors that disrupt the “packing” of lipids in a biological membrane accelerate, and factors that support the structuring of lipids (for example), inhibit O. p. Another regulatory component of the O. p. system are those involved in the formation (for example, some) or death (superoxide dismutase ) reactive oxygen species and free radicals, as well as in the decomposition of peroxides without the formation of free radicals (catalase, peroxidases). These enzymes may also depend on the structuring of the lipid bilayer of the biological membrane. At almost all stages of opiation, a significant modulatory role is played by factors that regulate the phospholipids of biological membranes and influence the rate of oxidation by changing the lipid composition of the membranes. Numerous low-molecular compounds that perform the functions of initiators, catalysts, inhibitors, quenchers, and synergists of this process are extremely important in the regulation of opacities. Among the most important stabilizers of biological membranes is the natural antioxidant (O.p.) E; other natural antioxidants are thyroxine and vitamin K. Metal ions of variable valency, C, D, etc., have the properties of prooxidants (substances that enhance oxygen absorption).

With the development of a pathological process, the balance of formation and consumption of peroxides and other oxygen products may be disrupted; oxygen products accumulate in tissues and biological fluids, which leads to serious disturbances, primarily in biological membranes. The consequence of activation of O. p. may be a change in the physicochemical properties of membrane proteins and lipids, a change in the activity of membrane-bound enzymes, a violation of membrane permeability (including for protons and calcium ions), ion transport (for example, inhibition of the sodium pump) , reducing the electrical stability of the lipid bilayer of membranes. Activation of O. p. leads to changes in the structure of serum lipoproteins and hypercholesterolemia, disrupts various processes of cellular metabolism at almost all levels.

Not only the peroxides formed as a result of opiation are toxic to the body, but also the products of deeper oxidation of lipids, aldehydes and acids. Carbonyl products of O. p. inhibit a number of enzymes, suppress DNA, enlarge capillaries, modify platelet aggregation, and exhibit a number of other undesirable effects. The reactive free radicals that initiate oxygen and arise during the oxidation process cause the structures of nucleic acids (Nucleic acids), primarily DNA, the destruction of nucleotide coenzymes (Coenzymes), disruption of the functioning of enzymes (primarily SH enzymes), and covalent modification of various biomolecules. The consequence of excessive generation of free radicals can be pathological changes in the properties of blood vessels.

For the prevention and treatment of conditions associated with excessive activation of O. p., substances that specifically react with certain free radicals (traps or interceptors), specific substances that form complex compounds with metals of variable valence, as well as various ways of activating endogenous systems of antiradical defense of the body (for example, gradual to hypoxia or other factors).

In connection with the important role of O. in the pathogenesis of various diseases, the determination of the products of this process (mainly conjugated dienes, malondialdehyde), spontaneous and induced chemiluminescence in biological material (serum and blood plasma, red blood cells, urine, exhaled air condensate, etc.) d.) has an ever-increasing diagnostic and prognostic significance.

1. Small medical encyclopedia. - M.: Medical encyclopedia. 1991-96 2. First aid. - M.: Great Russian Encyclopedia. 1994 3. Encyclopedic Dictionary of Medical Terms. - M.: Soviet Encyclopedia. - 1982-1984.

See what “peroxide oxidation” is in other dictionaries:

lipid peroxidation- The process of interaction of lipids (their unsaturated sections), which are part of cell membranes, with oxidizing agents (O2 anion, HO radical, etc.) formed under the influence of ionizing radiation and in the metabolic processes of certain substances; ... Technical Translator's Guide

Mechanism of LPO Lipid peroxidation (LPO) is the oxidative degradation of lipids, occurring mainly under the influence of free radicals. One of the main p... Wikipedia

Lipid peroxidation lipid peroxidation. The process of interaction of lipids (their unsaturated sections), which are part of cell membranes, with oxidizing agents (O2 anion, HO radical, etc.) formed under the influence of ionizing... ... Molecular biology and genetics. Dictionary.

The set of processes of digestion and absorption of neutral fats (triglycerides) and their breakdown products in the gastrointestinal tract, intermediate metabolism of fats and fatty acids and the removal of fats, as well as their metabolic products from the body.... ... Medical encyclopedia - a set of processes of absorption, distribution, assimilation and excretion of mineral substances found in the body mainly in the form of inorganic compounds. Minerals play a major role in maintaining acid-base... ... Medical encyclopedia

POL mechanism. Lipid peroxidation (LPO) is the oxidative degradation of lipids, occurring mainly under the influence of free radicals. One of the main consequences of radiation. One of the products of this process is malondialdehyde. Literature Yu ... Wikipedia

Young trees in a botanical garden ... Wikipedia

- (synonym: neutral fats, triglycerides) esters of triatomic alcohol glycerol and higher or medium fatty acids, the main component of animal fats and vegetable oils, are present in all animal and plant tissues, in nutrition... ... Medical encyclopedia

The chaotropic effect of excess fatty acids and lysophosphatides supports activation lipid peroxidation (LPO), initiated by the accumulation of reactive oxygen species (ROS) in a hypoxic cell. The generation of the latter is associated with Ca 2+ -dependent damage to mitochondria and the formation of an excess of electron donors - reduced cofactors.

Formation of reactive (toxic) oxygen species (in an unexcited state, oxygen is non-toxic) is associated with the peculiarities of its molecular structure: O 2 contains two unpaired electrons with parallel spins, which cannot form a thermodynamically stable pair and are located in different orbitals. Each of these orbitals can accept one more electron. Thus, complete reduction of an oxygen molecule occurs as a result of four one-electron transfers:

E - e - e - e - , H +

O 2 O 2 - H 2 O 2 `OH + H 2 O 2H 2 O

Superoxide formed during incomplete reduction of oxygen molecules (O 2 -), peroxide (H 2 O 2) and hydroxyl radical (`ON) – reactive oxygen species , are oxidizing agents, which pose a serious danger to many structural components of the cell (Avdeeva L.V., Pavlova N.A., Rubtsova G.V., 2005). The hydroxyl radical (OH) is especially active, interacting with most organic molecules. It takes away an electron from them and thus initiates oxidation chain reactions.

The main route of ROS formation in most cells - leakage of electrons from their transmission chain (respiratory chain) and the direct interaction of these electrons with oxygen (Gubareva L.E., 2005). As two more sources can perform reactions involving oxidases , using molecular oxygen as an electron acceptor and reducing it to H 2 O or H 2 O 2 and reactions involving oxygenases, including one (monooxygenase) or two (dioxygenase) oxygen atoms in the resulting reaction product. Under conditions of oxygen deficiency in tissues, i.e. in a situation where “demand” (reduced cofactors) exceeds “supply” (the number of oxygen molecules), the likelihood of increased ROS formation increases sharply. The free radical reactions they initiate lead to damage to cellular and subcellular structures, including mitochondria, DNA and protein molecules. And although the contribution of ROS to the development of hypoxic necrobiosis (as opposed to reperfusion syndrome) is not regarded as the dominant mechanism by all authors (Zaichik A.Sh., Churilov L.P., 1999), nevertheless, their participation in the activation of free radical processes in the cell, including LPO, is decisive.

It should be noted that LPO, being a self-developing chain reaction, constantly occurs in the cell, playing the role of a necessary link in its life activity and in adaptive reactions. Due to peroxidation, polar hydroperoxide groups (lipid hydroperoxides), which have a detergent effect, appear in the phospholipid molecule of cell membranes containing a fatty acid in the second position. The appearance of such groups increases the mobility of polypeptide chains, i.e. facilitates conformational changes in protein molecules, which is accompanied by an increase in the activity of membrane-bound enzymes, which essentially include all enzyme systems of the cell. And only excessive activation of LPO, affecting more than 3-5% of membrane phospholipids, turns it from a regulatory mechanism into a link in the pathogenesis of their damage during cell death (Yu.A. Vladimirov, 1987; 2000).

As a result of the activation of LPO initiated by ROS, and primarily by the hydroxyl radical (OH), the formation of new secondary radicals occurs: lipid (L), alkoxy (LO), peroxide (LOO). Rice. 28.

Rice. 28. Lipid peroxidation and formation of secondary radicals

(Yu.A. Vladimirov, 2001)

The chemical activity of these secondary organic radicals is lower than that of the hydroxyl radical (OH), but they are actively involved in the lipid peroxidation chain reaction, maintaining and aggravating damage to the lipid bilayer of cell membranes.

The modifying effects of LPO on phospholipids determine the chain of further events (Arkhipenko Yu.V. et al., 1983; Meerson F.Z., 1989; Vladimirov Yu.A., 2001). First of all, in phospholipid molecules containing a fatty acid in the second position, a polar hydroperoxide group appears (Fig. 29).

In this case, the accumulation of lipid hydroperoxides is accompanied by a decrease in the amount of unsaturated lipids. At moderate activation LPO, as noted above, the appearance in the microenvironment of integral proteins of polar products of LPO, which have a detergent effect, causes an increase in the mobility of the polypeptide chain, which is usually accompanied by an increase in the catalytic activity of enzymes. At excessive activation LEXURE, the main importance is to reduce the amount of unsaturated phospholipids.

Rice. 29. Formation of phospholipid hydroperoxide, the initial stage of the lipid peroxidation process

(F.Z. Meerson, 1984).

· A significant decrease in the content of unsaturated phospholipids in the membrane under the influence of LPO increases the rigidity (microviscosity) of its lipid bilayer, which is accompanied by a decrease in the conformational mobility of the polypeptide chains of proteins embedded in the membrane (the “freezing” effect). Since such mobility is necessary for the normal functioning of enzymes, receptors and channelformers, their functional response is inhibited(Fig. 30) .

Rice. thirty Changes in Ca-ATPase activity in sarcoplasmic membranes

reticulum as a result of modification of the lipid environment of this enzyme

LPO process(F.Z. Meerson, 1984)

A - initial state; B - moderate activation of Ca-ATPase; B - inhibition of Ca-ATPase.

· Phospholipids oxidized during LPO activation undergo lateral diffusion along the membrane and form associates (clusters) fixed by the interaction of phospholipids with each other and with water molecules. These areas of the membrane become hydrophilic. Located opposite each other in each of the monolayers of the lipid bilayer, such associates form channels in the membrane, increasing its permeability to water, calcium and other ions(Fig. 31).

Rice. 31. Scheme of formation of peroxide clusters and membrane fragmentation during the induction of lipid peroxidation (F.Z. Meerson, 1984)

The light triangle is the hydroperoxide group.

· The resulting decomposition products of phospholipid hydroperoxides (malonic, glutaric and other dialdehydes) interact with free amino groups of membrane proteins, forming intermolecular cross-links and inactivating these proteins(Fig. 32) . In vivo, this process leads to the formation of the so-called. Schiff bases of lipofuscin wear pigment.

Rice. 32. Formation of cross-links and inhibition of membrane enzyme proteins as a result of LPO activation(F.Z. Meerson, 1984)

The latter is a mixture of lipids and proteins interconnected by cross-sectional covalent bonds and denatured as a result of interaction with chemically active groups (dialdehydes) of lipid peroxidation products. This pigment is phagocytosed, but is not hydrolyzed by lysosome enzymes, and therefore accumulates in cells in the form of pigment spots, especially on the dorsal surface of the palms of older people.

Hydroperoxide (2), formed as a result of the reaction of phospholipids (1) with molecular oxygen, decomposes into a phospholipid with a shortened hydrocarbon chain in the second position, similar to lysophospholipids (3) and a short hydrocarbon fragment - dialdehyde (4). The interaction of a dialdehyde molecule, bifunctional in nature, with the amino groups of two protein molecules simultaneously leads to the formation of a cross-link (5).

· Under the influence of LPO, sulfhydryl (-SH) groups of membrane proteins: enzymes, ion channels and pumps are oxidized, which leads to a decrease in their activity.

· The formation of polar oxidation products contributes to an increase in the negative surface charge on the membrane, which causes the fixation of polyelectrolytes on it. Among the latter are some proteins and peptides that form protein pores - one of the factors reducing the electrical stability of membranes.

· An increase in the polarity of the inner membrane of the membrane causes the penetration of water into the lipid bilayer - the so-called. "water corrosion of the membrane."

· “Pushing out” some of the oxidized polyunsaturated fatty acids from the membrane leads to a decrease in the area of ​​its lipid bilayer.

Thus, at this stage of development of hypoxic cell damage, the key link in pathogenesis is the disorganization of the lipid bilayer of membranes, carried out with the participation of calcium ions and the lipid triad: activation of lipases and phospholipases; detergent action of excess fatty acids and lysophospholipids, and activation of lipid peroxidation.

A significant contribution to this disorganization is also made by: mechanical (osmotic) stretching of membranes and adsorption of polyelectrolytes on the lipid bilayer , contributing to an increase in their porosity. Taken together, these violations cause a decrease in the electrical strength of membranes and the occurrence of electrical breakdown of the lipid bilayer by its own membrane potential(Fig. 33). The latter is considered as a terminal mechanism for disrupting the barrier function of the membrane (Vladimirov Yu.A., 2001).

This stage of the pathogenetic chain of cell damage during hypoxia, characterized increasing loss of barrier and matrix functions of membranes, determines transition of reversible changes in the cell into irreversible ones.

Subsequent developments of events are associated with the formation damage to cellular structures, directly leading to cell death. It is significant that the mechanisms of these damaging effects are also closely related to the increased content of Ca 2+ ions in the cytosol.

The pathogenetic consequences of excess calcium ions in the final stage of hypoxic cell damage (necrobiosis stage) are not limited to activation of lipases and phospholipases. Ca 2+ ions are directly involved in the direct effects of damage to cellular structures and apoptotic cell death. These effects include:

· Destruction of the cytoskeleton, which is associated with Ca 2+ -dependent activation of calpains. Destruction of some cytoplasmic proteins (β-actin, fodrin) occurs, which causes cell deformation, limiting the possibility of their interaction with the microenvironment, as well as the ability to perceive regulatory signals. The weakness of the cytoskeleton contributes to the disintegration of some supramolecular complexes in the cell, in particular, the detachment of ribosomes from the membranes of the rough endoplasmic reticulum. As a result, the cytoplasm becomes saturated with protein molecules that undergo degradation.

· Mechanical damage to cellular structures, conditional Ca 2+ activation of the contractile function of myofibrils with a simultaneous loss of their ability to relax. Such contracture contractions accompanied by mechanical damage to the contractile structures of the cell.

· Saponification and endogenous detergent effect. The accumulation of fatty acids in the cell in the presence of excess Ca 2+ (and Na +) ions leads to soap formation – salts of higher fatty acids. For this reason, hydrolysis of ester bonds is called saponification . The formation of soaps in the cytosol sharply increases its detergent activity, which literally dissolves lipid membranes (Zaichik A.Sh., Churilov L.P., 1999). Soaps, destroying the membranes of organelles, attack the cell with hydrolases, active radicals and other metabolites, which until that moment were isolated in various compartments of the cell. This endogenous effect is critical in shaping the final stage of cell death.

· Along with participation in necrobiosis, calcium ions are involved in the implementation mechanisms of apoptotic cell death. Among the latest: increased activity of Ca 2+ -dependent endonucleases and calpains. Such activation poses a threat to the cell, initiating its apoptotic death or due to DNA fragmentation ( endonucleases ), or as a result of proteolysis of anti-apoptotic proteins (bcl-2) calpains . Apoptosis can be promoted by calpain-induced degradation of protein kinase C (PKC), realizing mainly anti-apoptotic effects and increasing cell resistance to toxic metabolic products.

· Moreover, excess Ca 2+ ions myself promotes the formation of toxic products, which can, in particular, be molecules nitric oxide in high concentrations, created by Ca 2+ -activation of inducible NO synthase. This effect manifests itself most clearly with the so-called. glutamate neuron death occurring during hypoxia (cerebral ischemia). The initiation of events in this case is associated with energy deficiency in neurons, the release of potassium ions, membrane depolarization and an increase in the intracellular Ca 2+ pool as a result of the prolonged opening of voltage-dependent calcium channels (Fig. 34).

Rice. 34. The mechanism of development of glutamate death of neurons during hypoxia

The consequence of an excess of calcium ions in the cytoplasm is an increased release of the neurotransmitter (glutamate) by glutamatergic neurons into the synaptic cleft. The perception of this signal by postsynaptic neurons is carried out using NMDA receptors (the most well studied subtype of glutamate receptors with high affinity for the synthetic amino acid N-methyl-D-aspartate), the sensitivity of which to the transmitter under hypoxic conditions increases significantly (Kryzhanovsky G.N., 1997 ). The result of “glutamate bombardment” (Akmaev I.G., 1996; Akmaev I.G., Grinevich V.V., 2001) of the postsynaptic neuron is the opening of ion channels in it, leading to an increase in the flow of calcium into the cell and activation of neuronal NO synthase (NOS). The nitric oxide produced under its influence, having a small size and lipophilic nature of the molecule, diffuses into the extracellular space and enters through membranes into nearby cells (neurons), exerting a toxic effect on them. The basis of this toxic effect is the energy deficiency of cells. The mechanism for the formation of such a deficiency is associated with the ability of NO to cause S- nitrosylation of cellular iron-containing proteins(aconitase TCA cycle, complexes I-III of the electron transport chain in MTX) and their inactivation. In addition, under the influence of NO, ribosylation And nitrosylationglyceraldehyde-3-phosphate dehydrogenase, causing inhibition of glycolysis. Finally, when NO interacts with another radical - O 2 - it forms peroxynitrite anion (ONOO -), causing irreversible inhibition of iron-containing proteins.

Due to the formation of ONOO, it is possible to turn on the apoptotic mechanism of cell death through the implementation of the following cascade:

A feature of glutamate neuron death is the absence of death of the NO-producing neurons themselves, which are protected from the toxic effects of NO. The mechanism of this protection is associated with the activation of superoxide dismutase (SOD) and (or) with the transition of NO to the oxidized form (NO +). In fact, there is a direct analogy with macrophages, which, while producing NO, themselves exhibit resistance to it.

Thus, cell death during hypoxia is a natural unfolding of a chain of events, including the formation of energy deficiency, inhibition of major metabolic pathways, activation of the lipid triad and subsequent irreversible damage to cellular structures. The central link in the pathogenesis of these events is an increase in the intracellular concentration of calcium ions, and the main target is cell membranes and, above all, mitochondria.

The sequence of changes considered during hypoxia (anoxia) is the same for a wide variety of tissues. This is evidenced by experiments with tissue sections, isolated cells and isolated organelles (Vladimirov Yu.A., 2001). Rice. 35.

The difference is only in the speed of these processes, which is 2-3 times higher at human body temperature. In addition, this speed is different for different tissues and these processes occur at the greatest speed in brain tissue, at a lower speed in the liver, and at an even lower speed in muscle tissue.

Rice. 35. Sequence of disorders in liver cells during anoxia

according to Yu.A. Vladimirov, 2001

XIV. HYPEROXIA

Hyperoxia – increased oxygen supply to the body . Unlike hypoxia, hyperoxia is always exogenous and practically never occurs in natural conditions. In this regard, adaptive mechanisms to this state are effective only under conditions of a relatively low oxygen load, determined by the magnitude of the partial pressure of oxygen and the duration of its action. An example of such a dependence is the curve of safe periods for breathing oxygen in humans (Fig. 36).

Rice. 36. The limit of oxygen action on humans(after Hartmann, 1966).

Quoted from A.G. Zhironkin (1979).

The x-axis is the duration of oxygen breathing, hours; along the ordinate - partial pressure of oxygen, atm.

As can be seen from the figure, zone so-called "physiological action of oxygen" lasts most at low values ​​of its partial pressure (about 0.5 atm.), when protective-adaptive reactions are able to ensure the preservation of normal oxygen tension in the tissues. These reactions are based on mechanisms aimed at limiting the supply and transport of oxygen. This is, in particular, aimed at primary reaction of the external respiratory system, in the form of a decrease in pulmonary ventilation and minute respiratory volume.

These changes are a consequence of the cessation of normal natural impulses from arterial chemoreceptors under conditions of increased oxygen supply. At the same time, limiting ventilation not only reduces the supply of oxygen to the body, but also leads to the development of hypercapnia. The latter determines the second phase of the respiratory system reaction, characterized by increased ventilation aimed at reducing PaCO 2 and eliminating gas acidosis. The most important shift in the circulatory system with hyperoxia there is a natural narrowing of small blood vessels, accompanied by an increase in peripheral resistance, a slowdown in general and local blood flow, and an increase in diastolic pressure. Another manifestation of the reaction from this system is bradycardia, recorded before signs of oxygen poisoning appear. Changes in the blood system in response to hyperoxia, they manifest themselves in the initial period as transient erythropenia and a decrease in hemoglobin levels, which is caused by the movement of tissue fluid into the blood and the deposition of red blood cells (Zhironkin A.G., 1979).

As the partial pressure of oxygen in the inhaled gas mixture increases, its toxic effect comes to the fore, since the protective effect of adaptive reactions is minimized. In this zone, oxygen already plays the role of a factor that does not provide, but inhibits oxidative processes in tissues. As for the mechanisms of the toxic effect itself, today the most accepted point of view is R. Gershman (1964), who connects this mechanism with the formation of reactive oxygen species and with the activation of free radical oxidation.

In conditions of tissue oversaturation with oxygen, i.e. in a situation where “supply” (excess oxygen) exceeds “demand” (the amount of reduced cofactors subject to oxidation), the likelihood of increased ROS formation increases. Accordingly, free radical oxidation increases, accompanied by damage to cellular and subcellular structures, and, above all, mitochondria.

It is obvious that disorganization and damage to mitochondria will be accompanied by disruption of the electron transport chain and oxidative phosphorylation. Those. disorders that define the essence of the concept of “hypoxia”. Respectively, this state is called hyperoxic hypoxia.

Damage to cellular and subcellular structures during activation of free radical processes leads to the development of numerous disorders of the specific functions of various organs and systems. Thus, inhibition of enzymes in the brain reduces the production of γ-aminobutyric acid, the most important inhibitory mediator, which serves as one of the development mechanisms in hyperoxia convulsive syndrome of cortical origin. Violation of surfactant production by the pulmonary epithelium causes a sharp decrease in the compensatory reserves of the external respiration system, increasing surface tension of the alveoli, and contribute to the appearance of microatelectasis. In severe cases, disruption of surfactant production may be accompanied by pulmonary edema. In some children in the first year of life, breathing pure oxygen leads to the development respiratory distress – bronchopulmonary dysplasia(Malyarenko Yu.E., Pyatin V.F., 1998) . Activation of free radical oxidation during hyperoxygenation underlies the formation visual defects in young children due to impaired maturation of photoreceptors.

Along with ROS, the toxic effect of oxygen is also mediated by excessive tension in some protective and adaptive reactions. Such reactions, in particular, include prolonged vasospasm (reaction to hyperoxia). In premature babies, it promotes the development retrolental fibroplasia(formation of fibrous tissue behind the lens), leading to blindness. A similar spasm of blood vessels in the lungs causes pulmonary hypertension, microcirculation disorders and damage to the pulmonary epithelium - disorders that predispose to the development of inflammation.

These circumstances force us to limit the use of oxygen for therapeutic purposes, in which PO 2 should not exceed 380 mm Hg. Art. (Berezovsky V.A., 1975).

Fetal brain tissue is particularly sensitive to the toxic effects of excess oxygen., which is characterized by a significantly lower oxygen tension than the cerebral structures of a mature organism . “This fact is not the result of imperfections in the processes of oxygen supply to the body in the prenatal period, but, on the contrary, reflects the balance of these processes, ensuring, on the one hand, adequate oxygenation of the brain, and on the other, protecting it from excess flow of O 2 "(Raguzin A.V., 1990). It has been experimentally established that oxygen tension of fetal brain tissue is a relatively stable parameter of homeostasis in the intrauterine developing organism, which changes little even with significant shifts in the oxygen regime of pregnant animals . Such constancy of PO 2 of fetal brain tissue with shifts in PaO 2 (from 50 to 370 mm Hg) of the maternal body is determined by mechanisms localized primarily in the uteroplacental region, but not by systemic reactions of respiration and blood circulation. By birth the formation of mechanisms for stabilizing oxygen homeostasis in the brain is not complete, which causes a more significant (than in adults) increase in PO 2 of the cerebral structures of newborns during inhalation of pure oxygen. Such an increase in PO 2 is accompanied by activation of free radical oxidation in brain tissue and the development of negative qualitative changes in the parameters of conditioned defensive reflexes in adulthood (Raguzin A.V., 1990). In connection with this situation, an approach to correcting severe hypoxia in newborns is substantiated using gas mixtures with a reduced content rather than pure oxygen for inhalation.

Convulsive form of oxygen poisoning occurs in acute oxygen poisoning and has been known since the end of the 19th century as Baer's sign, first discovered and described by this author. Convulsions usually occur when breathing oxygen under pressure exceeding 3-4 atm. and are very similar in their course to epileptic seizures.

Clinically, three stages of this process are distinguished (Chereshnev V.A., Yushkov B.G., 2001):

Stage I – increased breathing and heart rate, increased blood pressure, dilated pupils, increased activity with occasional muscle twitching.

Stage II is the stage of seizures similar to epileptic seizures with clonic and tonic manifestations.

Stage III – terminal – weakening of convulsions with respiratory distress, which progresses to individual breaths. Death occurs from paralysis of the respiratory center.

Products of this process include malondialdehyde and 4-hydroxynonenal.

Biological oxidation reactions are accompanied by the formation of free radicals, particles with an unpaired electron in the outer orbit. This causes the high chemical activity of these radicals. For example, they react with unsaturated fatty acids in membranes, disrupting their structure. Antioxidants prevent free radical oxidation.

Through the stage of peroxide derivatives of unsaturated fatty acids, the biosynthesis of prostaglandins and leukotrienes is carried out, and thromboxanes, which have a powerful effect on the adhesive-aggregation properties of blood cells and microcirculation, are themselves hydroperoxides. The formation of cholesterol hydroperoxides is one of the links in the synthesis of some steroid hormones, in particular progesterone.

Literature

• Vladimirov Yu.A., Archakov A.I. Lipid peroxidation in biological membranes. - M.: Nauka, 1972. - 252 p.
• Baraboy V.A., Orel V.E., Karnaukh I.M. Peroxidation and radiation. - K.: Naukova Dumka, 1991.
• Kovshevny V.V.- free radical oxidation

Notes

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