Changes in the activity of antioxidant enzymes and GSH level, in relation to age are commonly described in many articles and scientific publications. However, the specificity of the change in activity of these enzymes and GSH in relation to physical activity, diet or smoking is less known.
The literature describes the importance of oxidative stress and the reduced efficiency of repair processes in the process of aging. The most visible effects of these pathological conditions are revealed in DNA. DNA damage caused by the action of reactive oxygen species can lead to the formation of mutations, which in turn may be a cause of cancer development. Therefore, with increasing age, humans experience a higher incidence of various diseases, mainly including cancers but also neurodegenerative diseases and atherosclerosis, among other disease. Well-functioning repair systems remove damage and prevent harmful changes in cells. Unfortunately, with age, they are weakened, which contributes to an increase in the number of damaged cells .
Changes related to the aging of the body and increasingly becoming associated with the operation of ROS. In order to avoid the accumulation of ROS, the body has developed mechanisms of antioxidant defense, which include, among others, the action of enzymes such as CAT, SOD, GST, GPx, or non-enzymatic antioxidants such as GSH. Changes in their activity and concentration depend on the organ or subcellular location of the enzyme, as well as race and sex, and other variables. The reduction of their activity and concentration with age is caused by the modification of the enzyme molecule, which is caused directly or indirectly by ROS. Increasing their activity, on the other hand, should be treated as a compensatory response to overproduction of reactive oxygen species. With age, there may also be a decrease in GSH synthesis due to the much lower availability of methionine and cysteine, and the activity of γ-glutamyl cysteine synthetase and cystathionase, as well as increased GSH consumption in reactions with ROS, produced in too large quantities .
An important enzyme involved in the defense of the body against oxidative stress is CAT. CAT reacts with hydrogen peroxide (H2O2) to form water and molecular oxygen, as well as compounds such as methanol, ethanol, formic acid, phenol, etc. CAT thus protects the body against the effects of hydrogen peroxide produced in cells and is one of the most efficient enzymes in the fight against oxidative stress . This study analyzed differences in the levels of antioxidant enzymes in particular age groups. A statistically significant increase in CAT activity with age was demonstrated. The highest activity was observed in the 46–55 age group. These results also seem to be confirmed by multivariate regression analysis, which showed an increase in CAT activity with increasing age in healthy volunteers (an average increase of 0.0057 U/mgHb per year). These data support the generally accepted hypothesis regarding the increase in the activity of CAT in an aging organism . The increase in the activity of this enzyme may also be related to the activity of GST. There was a statistically significant (p = 0.047) decline in its activity among the elderly. These enzymes, due to their function to reduce peroxides, can complement one another. Therefore, when one of these enzymes increases, the growth of the other is inhibited .
Another important enzyme in antioxidant defense is GPx. This enzyme reduces both inorganic peroxides, e.g. H2O2, as well as organic peroxides (ROOH) to form selenic acid as an intermediate [9, 10]. Peroxidase has a greater affinity for hydrogen peroxide than CAT, therefore it performs a more important function in most physiological situations, when the amount of hydrogen peroxide formed is not too high. Insufficient CAT activity is therefore compensated by an increase in GPx activity and, conversely, reduced peroxidase activity is compensated by an increase in CAT activity [3, 11]. In the current studies reported here, no increased GPx activity was observed, in any age range, as well as in dependence from BMI. This may be related to the greater activity of CAT, which, despite its lower affinity for H2O2, performs this reaction with greater efficiency. This is evidenced by the increase in the activity of CAT with age. However, this does not confirm that peroxidase is more active in physiological situations.
Olędzki et al. compared the activity of SOD and CAT in erythrocytes taken from young healthy people (aged 20–29) and older individuals (> 60 years of age). They reported reduced SOD activity in the older patients. The study also showed statistically significant differences in the activity of SOD between the group of the youngest and the oldest women. According to the study reported here, the SOD activity decreases by an average of 0.0033 U/mgHb per year. In the case of CAT, decreased activity in the elderly was observed in relation to younger people, which is in opposition to the data presented here. Based on these results, it was found that the antioxidant defense level of erythrocytes, during their more than 100-day duration, is not stable in both young and old people and that it decreases in the physiological aging process .
The reduction of SOD activity and the increase in CAT activity among aging women can be explained by enzyme inactivation by excess hydrogen peroxide, as well as by glycation of SOD molecules or reactions with lipid peroxidation products, the intensity of which increases with age . This study also analyzed antioxidative enzyme activities and GSH concentration in relation to the BMI of healthy volunteers. A statistically significant increase in the activity of SOD was found in people who were underweight and a significant decrease in people with the first degree of obesity. This enzyme is an antioxidant that catalyzes the superoxide anion radical dismutation reaction for hydrogen peroxide and molecular oxygen, thus contributing to effective defense against oxidative stress [7, 13].
Karolkiewicz and colleagues evaluated the relationship between body mass and insulin resistance parameters, as well as the relationship between body weight and oxidative stress markers in older women. The population studied consisted of 34 women aged 60–90 who were divided into three subgroups based on their BMI: normal weight, overweight and obese. The total antioxidative status (TAS), the concentration of substances reacting with thiobarbituric acid and the level of protein were measured in the plasma C-reactive protein (CRP). However, the concentration of GSH and GPx activity were determined in haemolysate of red blood cells. The results did not reveal any significant differences between the three groups of women surveyed in relation to antioxidant status parameters. There was also no disturbed balance between oxidants and antioxidants . However, the relationship in the case of SOD has not been studied, therefore it is unknown what the reaction would have been with this enzyme. The current study reported here also did not show statistical significance with respect to GSH concentration and GPx activity.
The impact of diet and increased plant sterol supply on the parameters of oxidative stress in the group of obese women was examined previously by Stelmach-Mardas and co-workers. The study covered 101 women with a BMI > 30 kg / m². They were divided into two groups: the study group (60 women) and the control group (41 women). Anthropometric measurements were made, such as body weight, height, waist and hip circumference. Parameters were calculated, including the value of the body mass index and waist-hip index and percentage of adipose tissue. The lipid profile, oxidative stress parameters (malondialdehyde (MDA), oxidized protein, hydroxydioxides, CAT, SOD) were measured by enzymatic-colorimetric methods. The results showed statistically significant (p < 0.05) differences between the examined groups in relation to oxidative stress parameters after supplementing the diet with plant sterols.
In this study group, a decrease in the amount of hydroxydioxides (p = 0.0011) and a tendency to lower the activity of malondialdehyde (p = 0.0018), oxidized protein, and most importantly SOD (p = 0.0004) . Xiao-Liao et al. conducted a study for SOD in a group of 136 young and middle-aged men. The men were divided into three groups based on BMI: group I - obese (43 people), group II - overweight (46 people) and a control group with normal weight (47 people). Statistical analysis was performed, the results of which, in relation to oxidative stress parameters, showed a significant decrease in the activity of MDA and SOD in overweight and obese people . With regard to our current study, it can be assumed that anthropometric features are also strongly related to the activity of individual antioxidant enzymes. It is true that healthy volunteers, before and during the study, did not have to be on a special diet, as was the case in the study by Stelmach-Mardas et al., although SOD activity was similar. The decrease in the activity of SOD in the case of people with I degree of obesity, and its increase in underweight individuals has also been demonstrated.
Obesity is defined as the body mass exceeding the upper limit of physiological needs caused by excessive fat accumulation [16, 17]. The human body has brown and white adipose tissue. White adipose tissue contains fibroblasts, adipocyte, and macrophages, which are characterized by an obvious heterogeneity associated with their location, e.g. subcutaneous or visceral. In addition, white adipose tissue is not only the tissue that stores energy in the body, but also has an endocrine, paracrine, and autocrine function [16, 18]. Bioactive substances secreted by adipose tissues are called adipocytes or adipocytokines (including leptin and adiponectin) [16, 19]. The latter two can increase energy consumption, insulin sensitivity, and fatty acid oxidation, while leptin can suppress appetite and fat aggregation. In addition, adipokines increase the production of reactive oxygen species and cause oxidative stress respectively. Therefore, obesity correlates significantly with the growth of oxidative stress markers [16, 20].
Of course, diet should be taken as a factor having a significant impact on the increase or decrease in the activity of individual antioxidant enzymes and GSH concentration. A diet rich in fat reduces the activity and concentration of antioxidants. On the contrary, eating a large amount of fruits and vegetables increases the activity and concentration of antioxidants . In this case, attention should be paid to the characteristics of the women surveyed in Table 3. Almost 70% of women limited fat intake, and the majority did not limit consumption of fruit and vegetables in their diet. This applies mainly to older people, because the diet changes with age and health status, which often determines the consumption of proper foods in the elderly.
Epidemiological studies indicate that fruit and vegetables have a protective effect against diseases typical of old age, such as, joint degeneration, cardiovascular disease, stroke, or various types of cancer. The benefits of a diet rich in fruits and vegetables may also result from the avoidance of ingredients of animal origin less desirable for this age, such as saturated fats, oxidized cholesterol, etc., and may result from the intake of various antioxidant compounds, such as vitamin C and major carotenoids and dietary polyphenols [10, 21, 22].
The relationship between the change in the activity of antioxidant enzymes and physical activity is not completely clear. Current research indicates that the systems of antioxidant enzymes undergo significant changes in response to acute and chronic exercise. Acute exercise may increase the activity of some antioxidant enzymes in various tissues. A small effect of physical activity on hepatic enzymes or cardiac muscle has been shown, but changes in the activity of these enzymes in skeletal muscles have been observed, especially in the case of GPx . In our own study, there were no differences in the activity of antioxidant enzymes and GSH concentration in dependence from physical activity, which may be due to the fact that there was minimal physical activity in the characteristics of the study group. More than half of the women surveyed did not show any physical activity, and the part who led an active lifestyle only spent a small number of hours exercising each week.
In the literature, a small number of studies indicate the relationship between antioxidant enzyme activity and cigarette smoking, however these studies are predominantly in pregnant women. In this case, it has been shown that smoking cigarettes during pregnancy and lactation results in disruption of the antioxidant balance in the woman's milk. It is also found that cigarette smoking reduces the total antioxidative capacity in colostrum and this is directly related to the number of cigarettes smoked by pregnant women [9, 23, 24]. As the literature presents, tobacco smoke contains large amounts of reactive oxygen species, which, as we know, intensify oxidative stress and cause of various pathological changes in the human body. In the serum of people who smoke, there is a significant increase in the products of oxidative damage DNA, proteins, and lipids, and at the same time a significant decrease in the activity of antioxidants. It is assumed that in one portion of inhaled tobacco smoke there are as many as 1015 ROS molecules.
These molecules include mainly semiquinone (QH˙) radicals, but also oxygen radicals such as the hydroxyl radical, superoxide anion radical, or hydroperoxide radical, and molecules that do not belong to free oxygen radicals but are easily transformed
in these forms. These include mainly hydrogen peroxide, which is a precursor of the reactive hydroxyl radical. In the human body, the concentration of hydrogen peroxide is very low, mainly due to the reductive function of CAT and GPx. Oxygen radicals react very easily with the molecules present in cigarette smoke. These molecules include, for example, hydrocarbons, and as a result of their reaction, alkoxy radicals (RO˙) or alkyl radicals (R˙) . Based on these data, it should be assumed that in smokers, there should be a significant increase in the activity of antioxidant enzymes, which is adequate to increase the level of reactive oxygen species contained in tobacco smoke. In our study, 57% of women smoke cigarettes. However, there were no significant differences in enzyme activity and GSH concentrations in dependence from smoking cigarettes. It does not confirm, therefore, the reports of other scientists on the increase in oxidative stress as a result of smoking cigarettes. It may be associated with a well-functioning antioxidant system in the group of women surveyed, especially considering most of them were women in the 20–35 age group.