In recent years, the oxidative stress (in a broad sense) has been the subject of many studies. It is caused by the overproduction of molecules commonly known as "free radicals", more broadly referred to as reactive oxygen species (ROS). ROS include, among others, hydroxyl radical (* OH), superoxide anion (O*ˉ), singlet oxygen and hydrogen peroxide (H2O2), which as an exception does not have an unpaired electron. These molecules are formed during the body's physiological processes, such as aerobic respiration or inflammation. They are involved in many cellular processes, including the secretion of hormones, the functioning of the immune system, muscle contractions, apoptosis, vascular tone regulation and the elimination of xenobiotics from the body (Czajka 2006).
Healthy organism has defenses which detoxify reactive oxygen species by complex antioxidant mechanisms. They are especially important when the production of ROS increases, which is a consequence of smoking, drinking alcohol, improper diet, excessive physical stress, exposure to environmental pollution or ionizing radiation. Excessive production of free radicals, due to their high reactivity, can have a very negative effect on the body. The consequences of the uncontrolled action of reactive oxygen species on cells include oxidation of cell membranes, modification of proteins, and changes in the structure of DNA that may cause mutations and, ultimately, lead to initiation of the neoplastic process. Such an intensified attack of free radicals on the body's structures, when the antioxidant defense mechanisms fail and the physiological concentrations of ROS are exceeded, is called oxidative stress (Czajka 2006, Valko et al. 2006, Halliwell 2007, Visconti et al. 2009).
Defense mechanisms maintain an adequate level of free radicals so that they do not interfere with the proper functioning of the body. The formation and action of reactive oxygen species is counteracted by both enzymatic and non-enzymatic components of the antioxidant defense. Non-enzymatic mechanisms (the so-called free radical scavengers) are considered to be supplementary elements, while antioxidant enzymes seem to play a major role in the whole process (Wielkoszyński et al. 2007).
Enzymatic defenses against ROS include a system of specialized enzymes that prevent and remove free radicals. These enzymes are related to each other, participating in a cascade of events aimed at neutralizing free radicals. The most important antioxidant enzymes include superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glutathione S-transferase (Wielkoszyński et al. 2007). These enzymes cooperate in the direct neutralization of free radicals, inhibition of lipid peroxidation, reactivation of non-enzymatic elements of antioxidant defense, repair of damaged molecules and destruction of structures that could not be repaired (Wielkoszyński et al. 2007).
Superoxide dismutase is the body's main defense mechanism against the toxic effects of peroxides. It catalyzes the decomposition of superoxide anions to hydrogen peroxide and molecular oxygen.
MnSOD is believed to be one of the most important enzymes in cell defense against oxidative stress. Mitochondria, whose DNA is highly susceptible to attack by free radicals, must be protected by an effective manganese superoxide dismutase mechanism. A disturbance in the enzyme's activity could expose the cell to an intensified attack of ROS, which could damage the genetic material, leading to mutations, energy deficit and, consequently, the initiation of carcinogenesis (Mruk et al. 2002, Skrzycki et al. 2005, Czajka 2006, Wielkoszyński et al. 2007).
It is believed that there is a relationship between the activity of superoxide dismutase and the development of neoplastic changes in humans. The lack or limited activity of this enzyme can lead to mutations. An increase in SOD activity is associated with cancer progression and malignant transformation of neoplastic cells. On the other hand, the increased activity of SOD, which reduces the concentration of the superoxide radical, is associated with the suppression of the neoplastic phenotype. Understanding the exact mechanisms of SOD activity in healthy people and cancer patients may result in new treatment and diagnostic options in the future (Mruk et al. 2002, Skrzycki et al. 2005).
Catalase is the main line of defense against highly reactive hydrogen peroxide and is involved in its decomposition into water and oxygen. The enzyme exhibits catalase activity at high concentrations of hydrogen peroxide, causing it to decompose. In turn, at a low concentration of H2O2, catalase shows peroxidase activity when participating in the oxidation of compounds such as methanol, ethanol, formates, nitrites or quinones (Putnam et al. 2000, Czajka et al. 2006, Ścibior et al. 2006).
An effective action of catalase is especially important in the metabolism of erythrocytes, which function at high oxygen concentrations and are therefore exposed to oxidative stress. Moreover, catalase, by converting hydrogen peroxide, does not generate additional free radicals, which protects cells against other reactive oxygen species. Oxygen from the decomposition of H2O2 can be further used in other metabolic processes (Kirkman et al. 1984, Ścibior et al. 2006).
Catalase, as an enzyme that protects cells against the toxic effects of hydrogen peroxide, is associated with mutagenesis, carcinogenesis, inflammation and protection against apoptosis. Low activity of catalase was found in patients with pneumonia, atherosclerosis, diabetes, neurodegenerative diseases, nephritis and cancer. It is believed that the reduction of the enzyme activity may be caused by prolonged exposure of patients' cells to oxidative stress. Particularly low values of catalase were observed in patients with cancer of the lung, gastrointestinal tract, kidney, breast or with leukemia (Ścibior et al. 2006).
Glutathione S-transferases are ubiquitous multifunctional enzymes that play a major role in cell detoxification (Krajka-Kuźniak 2007, Wielkoszyński et al. 2007).
The main function of glutathione S-transferase is related to the participation in the second phase of detoxification of xenobiotics. The enzyme protects cells by catalyzing the conjugation of glutathione with toxins, thereby neutralizing their electrophilic sites and producing a more water-soluble products. Glutathione conjugates are further metabolized to mercapturic acid and then excreted (Cotton et al. 2000, Strange et al. 2000). For example, highly toxic and carcinogenic lipid peroxidation products such as 4-hydroxy-2,3-nonenal and other carcinogens, anti-cancer drugs, pesticides and herbicides can be detoxified according to the above mechanism. GST can also deactivate oxidative stress products, such as quinones, hydroperoxides, α- and β-unsaturated carbonyls (Krajka-Kuźniak 2007).
GSTs are enzymes that reduce the harmfulness of xenobiotics, improve their solubility in water and, consequently, facilitate their excretion from the body. These enzymes are associated with susceptibility to diseases caused by toxic extracorporeal compounds, including cancer (Wielkoszyński et al. 2007).
Malondialdehyde as an indicator of oxidative stress
Reactive oxygen species participate in the free radical oxidation of unsaturated fatty acids in lipids, i.e. in the so-called lipid peroxidation (Gaweł et al. 2004).
The end products of lipid peroxidation can be low molecular weight three-carbon malondialdehyde (MDA), and other aldehydes and hydroxyaldehydes. MDA is one of the most mutagenic products of lipid peroxidation. It reacts with DNA to form premutagenic lesions (Przybyszewski et al. 2005, Krzystak et al. 2009, Kulbacka et al. 2009).
Elevated levels of free radicals boost lipid peroxidation and increase the production of MDA. It is believed that the content of malondialdehyde may be an indicator of increased oxidative stress and the body antioxidant status (Gaweł et al. 2004, Kulbacka et al. 2009). Elevated levels of MDA in the blood were found in patients with breast, colorectal or prostate cancer (Surapaneni et al. 2006).
Oxidative stress in prostate cancer
The etiopathogenesis of neoplastic diseases is still the subject of many scientific studies, including the analysis of the imbalance between oxidation and reduction (Kaya et al. 2017). It has long been known that oxidative stress, including that caused by environmental factors, increases the activity of antioxidants that contribute to intracellular redox homeostasis. When this action is disturbed, an organism without adequate defense may be exposed to damage, including changes in the genetic material resulting in carcenogenesis (Agarwal et al. 2006, Kaya et al. 2017).
Currently, the drastic increase in cancer incidence is a growing problem in developed and developing countries. It is related to factors such as environmental pollution, diet and smoking. Today, cancer is the leading cause of death in developed countries. Prostate cancer is the third most frequently diagnosed neoplastic disease, after lung cancer and colorectal cancer (Farhood et al. 2018, Religioni et al. 2020). According to data from 2016, prostate cancer is the most common cancer in men in Poland and the third cause of cancer-related death in men. The overall mortality rate due to this disease slightly exceeds the European average (according to data from 2013, the mortality rate in Poland was 12.4/100,000, and 12.1/100,000 in the entire EU) (Religioni et al. 2020).
As a part of the research on the prostate cancer etiopathogenesis (which has not yet been clearly explained), the mechanisms of inactivation and excretion of toxic xenobiotics and harmful substances produced by the body itself were investigated. To determine the importance of antioxidant mechanisms in prostate cancer, the activity of enzymes such as catalase (CAT), superoxide dismutase (SOD) and the concentration of glutathione S-transferase (GST) were tested (Agarwal et al. 2006, Arsova-Sarafinovska et al. 2009, Battisti et al. 2011, Freitas et al. 2012). Battisti et al. (2011) demonstrated reduced catalase activity and increased superoxide dismutase activity in patients with prostate cancer compared to healthy controls. Studies involving patients from Macedonia and Turkey (Arsova-Sarafinovska et al. 2009) showed decreased catalase activity, as in the previously cited study. However, the activity of superoxide dismutase was lower in patients with prostate cancer than in the control group. The analysis of the influence of oxidative stress on prostate cancer cells by Freitas et al. (2012) included the measurement of the concentration of glutathione S-transferase. It was shown that the concentration of GST in cells treated with hydrogen peroxide was significantly lower, which might indicate a relationship between the low level of glutathione S-transferase and the progression of neoplastic changes. So far, inconclusive data indicate an imbalance of antioxidants in patients with prostate cancer, supporting the hypothesis of the influence of oxidative stress on this type of cancer (Arsova-Sarafinovska et al. 2009, Battisti et al. 2011, Freitas et al. 2012).
The aim of the study was to analyze the activity of antioxidant enzymes (glutathione S-transferase, catalase, superoxide dismutase) in order to determine the role of detoxification mechanisms in prostate cancer. The concentration of malondialdehyde, which is an indicator of lipid peroxidation in cancer patients, was also tested.