RESULTS

Table No. 1 display serum total lipid peroxide (MDA) levels in healthy controls and smokers’    

Significantly higher levels of serum total lipid peroxide (MDA) (P<0.001) were observed in smokers’ as compared to healthy controls .The elevated MDA levels observed in the present study suggest increased ROS production and thereby lipid peroxidation in smokers’.

Reactive oxygen species (ROS), such as the superoxide anion liberated by phagocytes recruited to sites of inflammation, are proposed to be a major cause of the cell and tissue damage, including apoptosis, associated with many chronic inflammatory diseases Lung cells, in particular alveolar epithelial type II cells, are susceptible to the injurious effects of oxidants. Lungs are continuously exposed to oxidants, either generated endogenously by metabolic reactions or exogenously, such as air pollutants or cigarette smoke.

Cigarette smoking a environmental hazard, also delivers oxidants and free radicals to the lungs. Cigarette smoke contains many oxidants and free radicals, both in the gas and the tar phase (13) and causes sequestration of neutrophils into the pulmonary microcirculation and accumulation of macrophages in respiratory bronchioles (14).  Once recruited, these cells become activated and generate ROS in response to a sufficient level of stimulus (threshold concentration). The mechanism for this may involve neutrophil adhesion to endothelium and upregulation of CD18 integrins (15,16), which is known to upregulate the reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, hydrogen peroxide-generating system.

Activation of macrophages, neutrophils and eosinophils generates superoxide anion, which is rapidly converted to H2O2 by superoxide dismutase (SOD), and hydroxyl radicals, formed nonenzymatically in the presence of Fe2+ as a secondary reaction. In neutrophils, myeloperoxidase also catalyses the formation of the potent oxidant hypochlorous acid from H2O2 in the presence of chloride ions. ROS, which may also be released by lung epithelial cells, may also stimulate inflammatory cells directly, thereby amplifying lung Inflammatory and oxidant events. Changes induced by reactive oxygen species may result in inactivation of antiproteases, epithelial cell injury, apoptotic and necrotic cell death, mitochondrial dysfunction, disturbance of extracellular matrix repair and maintenance of airway inflammation ⁽¹⁷⁾.

Thus, smokers’ are clearly subjected to oxidative stress as indicated by increased lipid peroxidation.

To prevent free radical formation or limit their damaging effects, cells have developed a comprehensive array of defenses i.e. antioxidants. Table No. 1 show superoxide dismutase activity in healthy controls and smokers’ significantly diminished SOD activity (P<0.001) was observed in smokers’.         

In smokers’ the excessive production of superoxide anions, by polymorphonuclear leukocytes, macrophages and monocytes disturb the oxidant and antioxidant balance. To scavenge these superoxide anions more amount of SOD gets utilized.

Cu-Zn-SOD has two identical subunits of about 32 kDa, each containing a active site constituted by a copper and a zinc atom bridged by a common ligand histidine ^{( 18).}

It has been reported that Cu-Zn SOD is irreversibly inactivated by its product H₂O₂, and it is well known that mature erythrocytes are unable to synthesize de novo SOD (or other proteins). Under normal circumstances a minor loss of SOD due to oxidative inactivation may not be of physiological significance. During exposure to oxidative stress, however, substantial SOD inactivation may occur, thus seriously compromising the antioxidant defenses of the RBC ⁽¹⁹⁾.

SOD inactivation by H₂O₂ has been shown to involve oxidative modification of a histidine residue which is important in the binding of the copper moiety to the active site of the enzyme. H₂O₂-dependent structural changes in the active site region of SOD lead to a conformational change which is susceptible to proteolysis. Such modified SOD undergoes

further reactions to form protein fragments, which are rapidly degraded by intracellular peptidase. The low SOD activity could contribute to a failure in plasma antioxidant defense.

Table No.1depicts the levels -SH proteins in healthy controls and smokers’ significantly lower levels of -SH proteins (P<0.001) were observed in smokers’ as compared to healthy controls.

Protein sulphydryl groups contribute physiologically to overall redox balance and modulate oxidative stress by generating reversible semi-oxidized species (mixed disulphides with non-protein low molecular weight thiols). Protein sulphydryl groups are also responsible for antioxidant action in plasma because they can react with several oxidant radicals ^(20).

In smokers’, oxidative stress oxidize thiols of various proteins and glutathione. Two series of oxidized species arise depending on the starting target, oxidation / nitration leads to the formation of protein thiolates (cys–S^-), can be readily oxidized to a sulphenic acid (cys-SOH), which is a relatively reactive form that can quickly form a disulphide with a nearby -SH. Strong oxidants will oxidize either cys–S^- or cys–SH to sulphinic (cys–SO₂H) and /or sulphonic (cyss–SO₃ H) acid derivatives. This difference in the generation of a particular cysteine -SH species provides a basis for distinguishing redox signaling from oxidative stress. While oxidative stress generally involves nonspecific oxidation of those cysteines which are located in an environment promoting dissociation of -SH. The higher oxidation states in the form of sulphinic and sulphonic derivatives have essentially been considered as irreversible modification under biologically relevant condition and associated with oxidative injury thus lowering the -SH levels in smokers’^(21).

It has been hypothesized that cigarette smokers were found to have 20 % more oxidized proteins in their plasma compared with non smokers’. Therefore -SH proteins measurement may be used as a prognostic marker test for cigarette smoke dependent lung diseases.

Total antioxidant capacity and smokers- The total antioxidative potential of the plasma reflects the ability of an individual to resist the oxidative stress. Ferric reducing ability of plasma (FRAP) evaluates plasma total antioxidant capacity due to known and unknown antioxidants in the plasma ⁽²²⁾.

Table No. 1 describe total antioxidant capacity in healthy controls and smokers. As compared to healthy controls total antioxidant capacity of smokers was significantly decreased (p<0.001).

 

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