Indian Journal of Experimental Biology Vol. 40, November 2002, pp. 1233- I 239

Review Article

Oxidant-antioxidant system: Role and significance in human body

M Irshad & P S Chaudhuri

Clin ica l Biochemistry Di vision, Department of Laboratory Medicine, All Ind ia Inst itute of Med ical Sciences, New Delhi 110029, India

Fax: 9 I - I 1-6862663

Present arti cle gives a holi sti c view of the causes, role and conrol of ox idati ve st ress in the development and progre -sion of vari ous human diseases. Several types of reacti ve species arc generated in the body as a result of metabolic reactions in the form of free radicals ol"non-radicals. These spec ies may be ei ther oxygen derived or nitrogen derived and called pro­oxidants. They allack macromolecules including protein, DNA and lipid etc. causing cellular I ti ssue damage. To counter thei r effect, the body is endowed with anot her category of compounds called antiox idant s. These anti ox idants are produced either endogenous ly or received from exogenous sources and include enzy mes like superox ide dismutase, catalase, glu­tathione peroxidase and gl utathi one reductase, minerals like Se, Mn, Cu and Zn, and vitamins like vitamin A, C and E. Other compounds wi th antioxidant ac ti vity include glutathione, flavonoids, biliru bin and uric acid etc .. In a hea lthy body, pro­ox idants and an ti oxidants maintain a ratio and a shift in thi s ratio towards prooxidants gives ri se to ox idative stress . This ox idat ive st ress may be either mild or severe depending on the ex tent of shift and remains the cause of several diseases such as cardiovascular diseases, neurologica l diseases, malignancies, renal di seases, diabetes, infl ammatory problems, sk in dis­eases, aging, respiratory diseases, li ver diseases and different types of viral infections. As more and more reports arc pou r­ing-in , a lot of information is being unfolded about ox idat ive stress in relation to several other diseases.

Chemical compounds and reactions capable of gener­ating potenti al tox ic oxygen spec ies/free radicals are referred to as pro-ox idants. On the other hand, com­pounds and reacti ons di sposing off these species, scavenging them, suppressing their formation or op­posing their actions are called anti -ox idants. In a nor­mal cell , there is an appropriate prooxidant : antioxi­dant balance. However, thi s balance can be shifted towards the proox idant when producti on of oxygen species is increased or when levels of anti-ox idants are diminished. This state is ca lled ox idat ive stress and can result in se rious cell damage if the stress is mass ive or prolonged. Oxidative stress is implicated in the etiopathogenesis of a variety of human di s­eases' -4

During the last two decades, there has been a grow­ing interest in studies that concern with the prevention of uncontrolled ox idati ve process lead ing to various diseases in living system. Several studi es have shown the role of ox idati ve stress in the causation and pro­gression of different diseases including atherosc lero­sis, carcinogenesis, neurodegenerati ve di seases, chronic inflammatory di seases, radiation damage, ag­ing and various other pathobiological effects 1- 6. This definitely developed a responsibility to scienti sts, med ical practitioners and clinical epidemiologists to findout the exact rol e and the control of oxidant­antiox idant system in human di seases.

In view of an enormous data from several studi es showing a clear implication of ox idative stress in cau­sation and progress ion of various di seases globally , it appeared worth to have an updated knowledge in thi s area. Present rev iew presents a holistic picture of various findings on the role and significance of oxi­dant-antioxidant system in various important di seases

The oxidants/free radicals are spec ies with very short half life, hi gh reacti vity and damaging act ivity towards macromolecules like protein s, D A and li p­ids. These species may be either oxygen derived (ROS, reacti ve oxygen species) or nitrogen derived (RNS , reactive nitrogen species). The oxygen derived spec ies include O2- (superoxide), HO (Hydroxyl), HO~

(Hydroperox yl), ROO (Peroxy l),RO (alkoxy l) as free radica ls and H20 2 (Hydrogen peroxide), HOCL (Hy­pochlorous acid), 0 3 (Ozone), and '0 2 (Singlet oxy­gen) as non-radicals. Similarly, nitrogen derived oxi­dant species are mainly NO ( itric oxide), ONOO­(Peroxynitrite), O2 ( itrogen diox ide) and 20 3

(Dinitrogen tri ox ide{

Reactive Oxygen Species The exogenous sources of ROS include electro­

magnetic radi ation , cosmic radi ati on, cigarette smoke_ car exhaust, UV li ght, ozone (0 3), and low wave length electromagnetic radiati ons. Similarly, the endogenous sources of ROS are mitochondrial electron

1234 INDIAN J EXP BIOL. NOVEMBER 2002

transport chain, respiratory burst by phagocy tes, beta ox idation of fat in peroxi ome, auto-oxidation of amino acids, ca techolamines, haemoglobin and is­chaemia reperfusion injury. Superoxide anion radical (02 - ) regulates metabol ites capable of signal I ing and communica ting important informations to the cellular genetic machiner/' Over production of 0 ]- takes place in vari ous chronic inflammatory cases, induced by drug, tox in, stress, ti ssue injury and heavy exer­cises2

.9

. Hydroxy l radica l is another damaging radical with a half li fe of 10-5 sec. and produced from H20 2

and O2- by Haber-Weiss Reacti on 10. Some HO may be produced from hypochlorous acid in phagocyt ic cells . Similarly, H20 2 is a relatively stable, poorly reactive, non-radical oxygen spec ies which eas il y crosses cell membrane and attacks different sites by converting into HO. This is produced by dismutation of O2- by superox ide di smutase (SOD) and final ly meets many fates including i ts reduct ion to water. H20 2 is also involved in the generation of free radica ls in presence of transitional metal ions.

Of the reacti ve nitrogen species (R S), nitr ic oxide is the most important nitrogen derived physioloi ca l free rad ical and one of the 10 smallest molecules in nature wi th molecular weight of 30 Da. It was once considered as a mere tox ic pollutant.However, later studics reported it to relax smooth musclell and in­volved either in the causation or recovery of several diseases l 2

. NO is synthes is d from L-argin ine by a fam ily of enzymes, OS (Nitric ox ide synthase), in a two step process l3

.

Reactive Nitrogen Species o rapidly undergoes addit ion, substitut ion, redox

and chain terminating reactions. These reacti ons serve as the molecular basis for its biological effects in hu­man body. The target molecules of NO are intraccllu­lar thi ol and metal containing protei ns and low mo­lecular weight thiols like glutathi one and cysteine etc. 1-1 0 acts as a " Double edged Sword" in health and disease. The main ph ysiological ro le of NO is contro lled by Type [ and II[ NOS expression via in­tracellul ar Ca-calmodulin complex dependant mecha­nism. 80th deficiency and excess of NO are believed to be involved in different pathophysiologica l states like stroke of brain , ischaemia, gastro intestinal dys­functions, achalas ia, Hirschprung's disease, congeni-

I h I . I ' . 15 ta ypertrop llC py onc stenosIs etc . Increased type II NOS (iNOS) ex press ion and NO

production occur in bacterial sepsis (septi c shock), eclampsia and hyperdynamic states like laenne ' s cir-

rhosis. Thi s also plays a role in inflammatory diseases like bronchiai asthma, arthriti s and ulcerati ve coliti s via different cytoki nes and Th2 cell stimulation I2

-13

. [n diabetes mellitus, inducible OS ( iNOS)over expres­

sion plays a ro le in ~ cell destructi on by the inhibi tion of mitochondrial enzymes and via di fferent cytokines release from acti vated lymphocytes 15

Perox ynitrite (0 00-) is another powerful ox idan t that interacts w ith a wide range of targets to cause tyrosine nitration , thiol oxidation, l ipid peroxidation, DNA strand break, guanos ine nitration/ox idati on and ultimately cell death .The reaction of 0 100- with excess NO generates O2, which can combine with more NO to form N20 3 to cause nitrosati ve stres . 16.

When overa ll generat ion of ROS and RNS exceeds the total antiox idant act ivity in the body, the resulting condition is ca lled oxidative stress. Th is stress may be

'1 I 8 d . 17-1 5 h ml c or severe. ase on vanous reports - , t e causes and after effects of mi Id and severe ox idat i ve stresses are shown in the fl ow chart .

Preventive Antioxidants, First Line Defense To counter the harmful effects of ROS and R S,

antioxidant defense mechanism operates to detoxify or scavenge these reacti ve oxygen species . The anti­oxidant system compri ses di fferent types of functional components classi fi ed as first line, second line, third line and fourth line defenses. The f irst line defense comprises preventive antioxidants that act by quench­ing of O2 - , decomposition of H20 2 and sequestrat ion of metal ions_ The antiox idants belong ing to thi s cate­gory are enzymes, like superoxide dismutase (SOD), catalase, glutathione peroxidase, and glutathi one re­ductase and non-enzy matic molecules like minerals and some proteins. Superoxide dismutase muinly act26

-30 by quench ing of superox ide (0 2- ) , an acti ve

oxygen radical, produced in different aerobic mewbo­li sm. Catalase is a tetrameric enzyme. present in most of the cells and acts by catalys ing the decompos ition of H20 ] to water and oxygen. Glutathione perox idase (GPx) is a selenium containing enzyme which cata ly­ses the reducti on of H20 2 and li pid hydroperox ide (L02H), generated during lipid perox idati on, to water using reduced glutathione as substrate.

The antioxidant minerals include Se, Mn, Cu, and Zn and function primarily in the metalloenzymes. Se is required in some of the immune mechanisms and biosynthes is of ubiquinone and ATP in mitochondri a. The renal cortex, pancreas, pituitary and li ver contain high amounts of selenium. A defic iency of selenium produces hepatic necrosis, muscular dystrophy, necrosis

IRSHAD & CHAU DHURI : OXIDANT-ANTIOXIDANT SYSTEM 1235

of cardi ac muscle and other di sorders in varIOUS ex­perimental animals. Selenium and vitamin E both ap­pear to be necessary for efficient scavenging of perox­ides from cytosol and cell membrane, respectivel/ I

.

Mn exerts its antiox idant action through mitochondri al SOD (MnSOO) which catalyses the di smutation of oxygen radi al (0 2- ) produced during aerobic metabo­lism in mitochondri a. Cu is present in a number of important metall oenzy mes including cytosoli c SOD, cytochrome ox idase, dopamine ~-hydroxy l ase, lyoy l oxidase and ascorbic ac id ox idase etc. This exerts its anti ox idant acti vity th rough the cytoso lic superoxide di smutase. Zinc is an element essenti al for normal growth, reproduction and other different functi ons of the bod/ 2

.33

. It is a component of several enzy mes li ke cytosolic superox ide di smutase, alcohol dehydro-

genase, alkaline phosphatase, carbonic anhydrase etc. Radical Scavenging Antioxidants, Second Line Defense

The antiox idants belonging to second line defense include glutathi one (GSH ), vitamin C, uri c ac id, al­bumin, bilirubin , vitamin E (mainly a -tocopherol), carotenoids, fl avonoid and ubiquinol. Glutathione (GSH~ gamma glutamyl cystein ylglycine) is the most abundant non-protein thiol, synthesised in the li ver and acts as a substrate for glutathione peroxidase en­zy me. This also serves as a scavenger of different free radicals34 Similarly, ~-carotene (Pro-vitamin A), vi­tamin C and vitamin E are some important scavenging antiox idant vitamins which can not be synthes ized by most mammals including human beings and therefore. are required fro m diet.

Responses and signals during oxidative stress

Mild oxidative stress

(positive effect)

+ Phys iolog ica l signal fo r beni ficial ce llular res ponse

+ Acti va ti on of signal transdu cti on mechani sm

+ Cell di ffe renti ati on Apoptos is D A repair

Cell cycle arrest Inducti on of antiox idants

+ Tissue protec ti on

+ Host defense

Severe oxidative stress

(negative effect)

+ Excess ROS/RNS & Low anti oxidant defense

+ Damage to biomolecules (Lipid , DN A, Protein )

Lipid peroxidati on (Damage to membrane ion channel, ion transporters.

DN A damage (Strand breakage Base Modi ficati on)

Ra i sedin~l

Protein damage (Damage to receptor,

enzy me, ion channel. Raised intrace llul ar Ca~+

Cellular damage Wilrelease of more radica ls

Ce ll death and ti ssue damage

+ Carcinogenes is , atherosc leros is, ag in g etc.

1236 INDIAN J EXP BIOL, NOVEMBER 2002

The retinoids like Beta-carotene, provitamin A and Vitamin A, are essential for vision, reproduction , growth and maintenance of epithelial ti ssues3:'-37. ~­carotene is an establi shed and excellent scavenger of singlet oxygen, produced during photosensitivity 38.39. Vitamin C and dehydroascorbic ac id interact directly with radicals like O2- and HO in plasma, thus prevent­ing damage to red cell membrane40. It probably assists a-tocopherol in inhibition of lipid peroxidation by recycling the tocopherol radical. Ascorbate (radica l) is reduced by intracellular GSH. It is a good scavenger of many free radicals like O2-, HO and various lipid hydroperox ides and may help to detoxify many in­haled ox idi zing ai r pollutants like ozone and N02 and free radicals in cigarette smoke in the respiratory tract 41.42.

Vitamin E compri ses a group of eight naturall y oc­curring related tocopherols, most important being a ­tocopherol.This is one of the most important and least tox ic of all lipid soluble anti-ox idant vitamins. It scavenges peroxy l radical intermediates in lipid per­ox idati on and is responsible fo r protect ing PUFA (poly unsaturated fatty ac id) present in cell membrane and low density lipoprotein (LDL), against lipid per­ox idation.J3-45.

Flavonoids are phenolic compounds, present in several plants, which inhibit lipid perox idation and lipoxygenases in vitro and in presence of free meta l ion (Fe3+). They act as pro-oxidant. Bilirubin and uric ac id also act as antiox idants46,47.

Repair and De-Novo Enzymes, Third Line Defense Third line antioxidants are a complex group of en­

zy mes for repair of damaged D A 4R,damaged protein, ox idi zed lipids and perox ides and also to stop chain propagation of peroxyl lipid rad ica l (Tab le I). These enzy mes repair the damage to biomolecules and re­constitute the damaged cell membrane e.g . Lipase, proteases, DNA repair enzy mes, transferase, methion-

ine sulphoxide reductase etc_

Cardiovascular Diseases Epidemiological studies have indicated that vita­

mins C and E exert protecti ve effect against cardio­vascular di seases. Low plasma level or low dietary intake of vitamin C are associated with hi gh blood pressure and unstable coronary syndrome. Taddei et 01.49 concluded that in essenti al hypertension, vita­min C improves the endothelium dependant vasodi la­tion by increasing type III NOS (eNOS) ex pression and ensu ing 0 production. Similarly, Reimersma et 01.50 showed that plasma levels of viramin E was associated with ri sk of angin a. Vitamin E supplemen­tati on red uced non-fatal heart attack. ri sk of coronary di seases and atherosc lerosis by inhibiting ox idat ion of LDL by free radicals51-57 .

Central Nervous System Diseases Due to hi gh oxygen consumption, low glutathione

(anti ox idant Status) content, hi gh levels of free iron and oxidizable substances like PUFA in the central nervous system, neuronal cells appear to be particu­larly vulnerable to oxidative stress. This oxidat ivc stress with decreased anti ox idan t defense does have a role in the genes is and progression of Alzheimer's di sease5H.59, Parkinson's disease6o

, Brai n neoplasm61

Down sy nd rome62 and also some other neurodcgen­erative di sorders63-65.

Carcinogenesis A major development of carcinogenes is research

during the last two decades has been the discovery of significant levels of DNA damag by oxyradica ls of endogenous cellular source. These oxyrad ica ls attack DNA causing change in genomic sequences leading to mutation, deletion, gene-amp I i fication or rearrange­ment. In thi s manner, ox idati ve DNA damage was found important in the etiology of many human cancers66-71 .

Site of damage

Table I-- Repa ir of ox idati ve damage

Repair system

DNA: All components can be attacked by HO, ONOO-, H O2,

HOCr that cause base damage.

Protein : ROS attack SoH group and HO modifies amino acid residues.

Lipid: Some ROS/RNS can ini tiate and propagate lipid peroxidatioll _

A wide range of enzymes remove abnormali tics in DNA by exc ision, resy nthesis or strand rejoi ning_

Methioninc sulphox ide reductase repairs ox idizcd 'methionine rcsidues. Damagcd proteins may be recogni ~eJ and preferen ti ally degradcd by protcosomc enzyme complex.

Chain breaking antiox idan ts remove chain propagat ing pcroxy l rad icals. Phospholi pid hydroperoxidc GPx and somc phospholipases can remove perox ides from membranc_ Damaged lipid may be releascd from cellmembranc_

IRSHAD & CHAUDHURI : OXIDANT-ANTIOXIDANT SYSTEM 1237

Inflammatory Diseases In chronic inflammatory conditi ons phagocyte de­

rived ROS have been implicated in infl ammation re­lated injury . Intracellular protection of cytoplasmic component against phagocyte derived ox idative injury is medi ated predominantly by anti ox idant enzy mes like SOD, catalase and glutathione perox idase. The development of mutations in pS3 tumour suppressor gene and other key regulatory genes promotes in­flammation into chronic disease in rheumato id arthri-. d I . fI d' d 7?·74 ti S an ot ler 111 ammatory Isor ers - .

Renal Diseases Free radical injury and ox idat ive stress have been

implicated in many renal diseases like acute renal fa ilure (A RF), IgA nephropathy, anaemia of chroni c renal fa ilure (CRF) and ischaemic kidney. Pl asma glutathione perox idase whi ch has been suggested as an index of rena l anti ox idant defense, is reported to be markedly depressed in CRF and patients undergoing hemodialysis75

.76

.

Diabetes Mellitus Diffe rent studies have given a lot of ev idence of

increased ox idative stress with dep leted anti ox idant enzymes and vitamins, in both type I and type 2 dia­betes mellitus. Hyperglycem ia can increase oxidative stress and change the redox potential of glutathione.

ow ox idati ve stress is acknowledged as a pathoge­netic mechanism in diabetic compl ications like dia­betic retinopathy, nephropathy, microangiopathy 77.

Respiratory Diseases Oxidative stress is assumed to play an important

ro le in the pathogenesis of a number of lung diseases, like-chronic obstructi ve pulmonary diseases (COPD), bronchial asthma & acute respiratory distress syn­drome, not only th rough direct injurious effects but also by in volvement in the molecular mechanisms that control lung inflammation 7R .I mportant consequences of ox idative stress in the pathogenesis of COPD in­clude ox idati ve i nacti vati on of anti protei nascs, ai r­space epithelial injury, increased sequestrat ion of neu­trophils in the pulmonary mi crovascul ature and gene expression of proinflammatory mediators.

Infectious Diseases A hallmark of HIV infection is immunodeficiency

with progressive CD4+ lymphocyti c cell depletion. HIV and opportuni stic infections directly or indirectl y lead to an oxidative stress, caused by the excess gen­eration of ROS/RNS by activated cells of immune

system. Diffe rent studi es have shown th at even in early stages of HIV infecti on, pl asma I lymphocyte SOD level is decreased parallel to the decrease of CD4+ cell s number 5. Another study has shown that HIV Tat protein downregulates MnSOD (mitochon­dria l SOD) to cause decreased SOD level.

Aging Aging in humans is assoc iated with changes in

physical characteristics and the decline of many physiological functions. Increased accumulat ion of free rdica ls heightens the vulnerability of older indi­viduals to a vari ety of oxidati ve insults. These radicals are capable of causing apoptosis , necros is and cell death79

.xo .

Liver Diseases The in volvement of free rad icals in the pathogene­

sis of li ver inj ury has been in vest igated for many years in a few well defined experimen tal systemx,. Several studies were conducted to fi nd out the role of ox idant stress and AOS in alcohol induced cirrhosis 82.XJ which is considered as the terminal irreversible stage of li ver diseases. These studies have shown in­creased lipid peroxidation by HO radical and hydrop­erox ides in experimental acute and chroni c alcoholic li ver diseases . Also it has been suggested that ROS and lipid peroxidation may playa role in pathogenesis of hepatic fibros is with loss of normal li ver architec­ture. In another stud/4 it is found that plama levels of AO vitam ins are low in patients with chron ic choles­tatic li ver diseases.

Hepatiti s viruses are taxonom icall y quite diverse and include Picorna, Flavi, Hepadna and ca lciviruses as well as delta agent assoc iated with hepatiti s B vi ru s. Viral hepatiti s is caused by both RN A (hepatit is A,C,D,E) viruses and DN A virus (HBV ). In these in­fections , there is an assoc iated enhanced production of ROS/RNS 2 via long term oxidant stress .In another

d 85 86 . d . b d . . h 1 stu y . OX I at lve st(ess was 0 serve In pel'lp era blood mononuclear cell s from chroni c hepatiti s C pati ents.

The preliminary studies conducted by us at our set­up concluded that there is a mild impact of hepatitis viruses in disturbing the anti ox idant level in the body. In spite of act ive diseases caused by hepatiti s viruses, the superox ide di smutase and total antioxidant levels in the body do not show signifi cant changes. The be­haviour of all the viruses remains si milar and does not exacerbate with increas ing severity of disease. Th is means that during viral hepatitis, protective calibre of body against oxidative stress remains intac tX7

.

1238 INDIAN J EXP BIOL, NOVEMBER 2002

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