PREDICTIVE POWER OF OXIDATIVE STRESS BIOMARKERS IN RECURRENCE AND SURVIVAL IN ADVANCED CERVICAL CANCER.

The aim of our study was to measure the levels of 8-hydroxy-2-deoxyguanosine, malondialdehyde, and antioxidant enzymes in patients with locally advanced cervical cancer prior to treatment to determine how these evaluated biomarkers are associated with cervical cancer recurrence and to estimate their potential in further research and clinical use. MATERIALS AND METHODS: The study included 45 female patients with newly diagnosed advanced cervical cancer who underwent concomitant chemoradiotherapy. The blood and urine samples were collected prior to treatment, between December 2013 and April 2016, and subsequent laboratory analysis was performed. After the medium follow-up of 29 months, the patients were divided into 3 groups according to the time of disease recurrence. A statistical analysis was performed in order to evaluate the relationship between the previously measured biomarkers and recurrence. RESULTS: Taken individually, the parameters of oxidative stress did not reveal significant differences between the three groups in our study. Nevertheless, the catalase and glutathione S-transferase activities were the best predictors of the recurrence. Based on the activities of these two oxidative enzymes, it was possible to separate the group of patients without recurrence after follow-up from the other two groups of patients with recurrent disease. CONCLUSIONS: The parameters of oxidative stress have a certain predictive value on the outcome of patients with advanced cervical cancer after concomitant chemo-radiotherapy.

The effects of phthalate exposure on metabolic parameters in polycystic ovary syndrome.

BACKGROUND-AIM: Phthalates are known as endocrine disrupting chemicals which are present in wide-range of products. The objective of the study was to investigate whether phthalate exposure may attribute to the metabolic syndrome development in women with polycystic ovary syndrome (PCOS). METHOD: The cross-sectional study involved 60 women in reproductive age with confirmed PCOS. Anthropometric and biochemical measurements were examined together with detected levels of ten phthalate metabolites measured by GC-MS in morning urine samples. RESULTS: In this study at least one phthalate metabolite was detected in 51.7% of samples. Total phthalate metabolites urine concentrations were positively associated with BMI, waist circumference, waist-to-height-ratio (WtHR), leptin serum levels as well as lipid accumulation product (LAP) and visceral adiposity index (VAI). Mono-methyl-phthalate (MMP) levels was significantly correlated with WtHR, LAP and VAI. Additionally, total phthalate metabolites levels were significantly linked with fasting plasma glucose and HOMA index, whereas MMP concentrations were associated with fasting plasma glucose and insulin levels. Total cholesterol (TC) level was statistically significantly higher among PCOS women with detected phthalate metabolites compared to those without phthalates. The sum of all phthalates was correlated with LDL and triglyceride levels as well as TC/HDL. MMP concentrations were linked positively with TC, LDL and triglyceride levels as well as with TC/HDL. It is noteworthy that MMP concentrations were positively associated with testosterone serum levels while the total phthalate metabolites concentrations were also linked but with moderate significance. CONCLUSIONS: The increased phthalate metabolites concentrations may interfere with obesity, glucose and lipid impairment in PCOS women. Additionally, testosterone serum levels can be disrupted by MMP.

Accidental Use of Milk With an Increased Concentration of Aflatoxins Causes Significant DNA Damage in Hospital Workers Exposed to Ionizing Radiation.

The occupational exposure to ionizing radiation (Irad) or associated with mycotoxin-contaminated food may lead to genome damage and contribute to health risk. DNA damage in 80 blood samples of hospital workers occupationally exposed to low-doses of Irad was compared with 80 healthy controls. Among them, 40 participants accidentally consumed milk with increased concentration of Aflatoxin. All participants underwent the testing for micronuclei from blood, and 40 of them 8-OHdG from urine. The frequency of micronuclei (MN) was analyzed by cytokinesis-block peripheral blood lymphocytes and the level of urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) by ELISA. The Irad led to increased frequency of MN (p < 0.05) and 8-OHdG level at exposed hospital workers. The consumption of milk with increased concentration of aflatoxin probably raised MN frequency and 8-OHdG value. Higher consumption of aflatoxin-contaminated milk (≥2 L/monthly) caused significantly increased MN frequency and 8-OHdG value in comparison to lower milk intake (≤0.5 L/monthly). Also, confounding factors, such as age, gender, and smoking status of all participants were included in the study. The obtained results revealed an increased incidence of MN and 8-OHdG level among hospital workers exposed to low-doses of IRad and milk with increased aflatoxin concentration.

Can environmental pollutant bisphenol A increase metabolic risk in polycystic ovary syndrome?

BACKGROUND: Bisphenol A (BPA), a widespread industrial substance is recognized as endocrine disrupting chemical and therefore could be associated with polycystic ovary syndrome (PCOS) related metabolic disturbances. PATIENTS: In this study 29 women of reproductive age with diagnosed PCOS were enrolled. METHODS: BPA in urine samples was analyzed by gas chromatography-mass spectrometry. RESULTS: BPA was detected in urines of 48.28% participants. The waist-to-height ratio (WtHR) was statistically significant higher in PCOS BPA+ in comparison to PCOS BPA- women (p = 0.046). PCOS BPA+ women had 6.88 times (95%Cl 1.3481-35.0600, z = 2.319, p = 0.020) higher risk for waist circumference above 80 cm and 4.95 odds (95%Cl 1.0169-24.096, z = 1.981, p = 0.048) to have WtHR over 0.5 when compared to PCOS BPA-. Statistically significant positive association between BPA urine concentrations and insulin serum levels (p = 0.038) was obtained. BPA urine values were associated with elevated HOMA-IR values and reduced HDL levels with moderate significance (p = 0.079 and p = 0.061, respectively). Also, there was 3.75 times (95%Cl 0.7936-17.7203, z = 1.668, p = 0.095) higher risk for PCOS BPA+ women to have testosterone levels above reference values. CONCLUSION: The obtained results suggested that the BPA exposition in PCOS women was followed by increased metabolic risk through promotion of obesity, especially the visceral type, hyperinsulinemia, insulin resistance, dyslipidemia and elevated androgen levels.

Could phthalates exposure contribute to the development of metabolic syndrome and liver disease in humans?

In the study, 305 patients of both genders were enrolled and divided into three groups: obese (BMI > 30 kg/m2), patients who were diagnosed type 2 diabetes mellitus (T2DM), and control, normal weight healthy volunteers. At least one of ten different phthalate metabolites was determined in the urine samples of 49.84% all enrolled participants. In the obese subgroup, the sum of all urinary phthalate metabolites was positively associated with TG levels (p = 0.031) together with derived TC/HDL and TG/HDL ratios (p = 0.023 and 0.015), respectively. Urinary MEP concentration was positively correlated with the HOMA-IR in T2DM subgroup (p = 0.016) while in the control subgroup, log10MEP levels were negatively correlated with total cholesterol (p = 0.0051), and LDL serum levels (p = 0.0015), respectively. Also, in the control subgroup, positive linear correlations between urinary log10MEP levels and TyG and TYG-BMI values (p = 0.028 and p = 0.027), respectively, were determined. Urinary MEHP levels were associated with glucose serum levels (p = 0.02) in T2DM subgroup, while in the control HDL values were negatively associated with log10MEHP (p = 0.0035). Healthy volunteers exposed to phthalates had elevated AST levels in comparison to non-exposed ones (p = 0.023). In control subgroup, ALT and AST values were increased (p = 0.02 and p = 0.01, respectively) in MEP exposed while GGT levels were enhanced (p = 0.017) in MEHP exposed in comparison with non-exposed. Combined phthalates influence on glucose and lipid metabolism may increase the possibility for NAFLD and insulin resistance development among exposed individuals.

Hypomagnesemia in adults of northern Serbia: prevalence, nutritional risk factors, and associated comorbidities.

INTRODUCTION: Magnesium (Mg) deficiency is associated with numerous non-communicable diseases. The aim of the study was to estimate the prevalence of hypomagnesemia in the general adult population of Northern Serbia (NS), and to determine the level of Mg in drinking water. METHODS: This is a cross-sectional study with 5,122 adults from the general population. Serum level of Mg was determined by spectrophotometry, while the level of Mg in the drinking water was done by atomic absorption spectrophotometry. Standard laboratory methods were used to determine individual's blood lipid status and complete blood count. RESULTS: The prevalence of hypomagnesemia (Mg < 0.75 mmol/L) in the general population was 2.7%, while the prevalence of the subjects with high risk for the Mg deficiency (Mg 0.75-0.85 mmol/L) was 20.1%. The public water supply showed variable values between 17.3-35.3 mg/L. Age, systolic blood pressure, duration of diabetes mellitus, and glycemia increase with the decrease of Mg level. In addition, increased level of Mg was associated with higher red blood cell count and hemoglobin levels. CONCLUSIONS: The prevalence of hypomagnesemia in Northern Serbia is low (2.7%) and level of Mg in drinking water is not satisfactory. Serum level of Mg in the range 0.75-0.85 mmol/L is present in about 1/5 of the population. Glycemia, advanced age, gender, and smoking have a predictive role in hypomagnesemia occurrence. There is a significant correlation between serum level of Mg and lifestyle and dietary habits.

Health risk assessment for pediatric population associated with ethanol and selected residual solvents in herbal based products.

In this study, 48 herbal based products (41 for the pediatric population) were analyzed for the presence of ethanol and residual solvents. Ethanol was not detected in only 12% of the products designed for infants or toddlers aged under 2, and not quantified in only 5 of 14 'alcohol free' products. Actual content was higher than labeled in six out of 11 samples with specified ethanol quantity. WHO proposed requirement for ethanol content in products intended for use in children under the age of 6 (<0.5%) was not met by as many as 26 samples. Furthermore, calculated blood alcohol levels in children exceeded the relevant toxicological levels for nine samples following a single dose, and for one sample in case of accidental poisoning with the entire package. Regarding the residual solvents, acetone, 1-propanol and 1-butanol were not quantified, 2-propanol was found in two samples in low concentrations, whereas methanol intake via one of the samples exceeded the permitted level for children. The obtained results revealed a significant health concern for the pediatric population due to ethanol intake via herbal based products, calling for the establishment of strict guidelines for ethanol content and labeling.

Lipid Peroxidation, Antioxidative Defense and Level of 8-hydroxy-2-deoxyguanosine in Cervical Cancer Patients.

BACKGROUND: Oxidative stress has been associated with cervical cancer. Our aim was to examine lipid peroxidation and the extent of oxidative stress in women diagnosed with different stages of cervical cancer in order to evaluate its potential role in the evolution of cancer. METHODS: We measured the concentration of thiobarbituric acid reactive substances, activities of antioxidative enzymes and 8-hydroxy-2-deoxyguanosine in 153 subjects. Enzymatic activity as well as TBARS concentration were measured spectrophotometrically, while 8-OHdG was determined by gas chromatography-mass spectrometry. PPatients were categorized: group II H-SIL; group III FIGO Ia-Ib and group IV FIGO IIa-IV. RESULTS: Our results showed highly significant increase in the level of lipid peroxidation in group IV when com pared to the control group, group II and group III (p<0.001). Activity of superoxide dismutase was also significantly higher in group IV when compared to control group (p<0.01), group II (p<0.01) and group III (p<0.05). Activity of catalase was also significantly higher in group IV when compared to control group (p<0.005), group II (p<0.005) and group III (p<0.05). Activity of glutathione-S-transferase was also significantly higher in group IV when compared to control group (p<0.05), group II (p<0.05) and group III (p<0.05). Activities of glutathione peroxidase and glutathione reductase showed no significant differences among the groups. Level of 8-OHdG was significantly higher in group IV than in the other groups (p<0.01). CONCLUSIONS: It can be concluded that oxidative stress is possibly involved in the pathogenesis of cervical cancer, demonstrated by increased lipid peroxidation and an altered antioxidant defense system and higher levels of 8-OHdG.

Screening for Chronic Kidney Disease in Adult Males in Vojvodina: A Cross-sectional Study.

BACKGROUND: Chronic kidney disease (CKD) is one of the most significant global health problems accompanied by numerous complicatons, with constant increase in the number of affected people. This number is much higher in early phases of disease and patients are mostly asymptomatic, so early detection of CKD is crucial. The aim was examination of the prevalence of CKD in the general population of males in Vojvodina, based on estimated glomerular filtration rate (eGFR) and urine albumin/creatinine ratio (ACR), and exploring the determinants and awareness of CKD. METHODS: This cross-sectional study included 3060 male examinees from the general population, over 18 years of age, whose eGFR and ACR were calculated, first morning urine specimen examined, arterial blood pressure measured and body mass index calculated. Standard biochemistry methods determined creatinine, urea, uric acid and glucose serum concentrations as well as albumin and creatinine urine levels. RESULTS: Prevalence of CKD in the adult male population is 7.9%, highest in men over 65 years of age (46.7%), while in the other age groups it is 3.6-12.6%. The largest number of examinees with a positive CKD marker suffer from arterial hypertension (HTA) and diabetes mellitus (DM). Only 1.3% of examinees with eGFR<60 ml/min/1.73 m2 and/or ACR≥ 3 mg/mmol had been aware of positive CKD biomarkers. CONCLUSIONS: Obtained results show the prevalence of CKD in adult males is 7.9%, HTA and DM are the most important CKD risk factors and the level of CKD awareness is extremely low (1.3%) indicating the necessity for introduction of early stage disease recognition measures, including raising CKD awareness.

Possible influence of the environmental pollutant bisphenol A on the cardiometabolic risk factors.

Bisphenol A (BPA) is a ubiquitous environmental pollutant which is often associated with various health issues. In this study 103 healthy female volunteers in reproductive age from Serbian north province Vojvodina were enrolled and examined for the BPA exposure in the urine samples after 12 h of fasting. BPA was found in 35.92 % (37/103) of subjects. Statistically significant increment in waist circumference (p = 0.045) and waist-to-height ratio (p = 0.037) was observed among the BPA positive women in comparison with the women who had the same energetic balance and had not been exposed to BPA. Linear correlation was obtained between the BPA concentration in urine samples and body mass index (r2 = 0.35, p = 0.003) waist circumference (r2 = 0.21, p = 0.02) and waist-to-height ratio (r2 = 0.25, p = 0.01) among the obese. High energetic intake and reduced physical activity additionally pronounced BPA positive association with obesity. No statistically significant difference was observed in triglycerides, HDL and LDL cholesterol levels between the BPA exposed and BPA non-exposed female volunteers.

Cryptorchidism and pesticides: Is there a connection?

INTRODUCTION: The aim of our study was to compare the level of the most common organophosphate metabolite, dimethyl phosphate, in urine of women giving birth to both boys with cryptorchidism (study group), and healthy boys (control group), as well as to compare the level of dimethyl phosphate in our population with the results obtained in other populations. MATERIAL AND METHODS: After the ethical approval we included thirty women in both study and control groups. All newborns were born between 38 and 42weeks' gestation. Urine samples were taken on 3rd postpartal day. Gas chromatography with flame photometric detection was used to analyze dimethyl phosphate in urine following the method of Wu et al. Statistical analysis was done using Mann-Whitney test to compare the results in the two groups. RESULTS: Geometric mean of dimethyl phosphate in the study group was 7.18±8.26µg/L and the creatinine-corrected level was 5.63±5.95µg/L, and in the control group, the values are 7.98±6.75µg/L and 6.15±7.01µg/L, respectively. There was not a statistically significant difference in levels of dimethyl phosphate between these two groups (p=0.72786). Dimethyl phosphate levels obtained in similar studies are: 14.4µg/L in Israel, 3.7µg/L in Palestine, 10.3µg/L in Jerusalem, 1.60µg/L in Caribbean islands and 2.60µg/L in Canada. CONCLUSIONS: Pregnant women in our country are exposed to organophosphate pesticides, but a correlation between the exposure to organophosphate pesticides and cryptorchidism was not found. LEVEL OF EVIDENCE: I. TYPE OF STUDY: Prognostic study, prospective study.

Effects of pharmaceutical formulations containing thyme on carbon tetrachloride-induced liver injury in rats.

BACKGROUND: Herbal supplements are widely used in the treatment of various liver disases, but some of them may also induce liver injuries. Regarding the infuence of thyme and its constituents on the liver, conflicting results have been reported in the literature. The objective of this study was to examine the influence of two commonly used pharmaceutical formulations containing thyme (Thymus vulgaris L.), tincture and syrup, on carbon tetrachloride-induced acute liver injury in rats. METHODS: Chemical composition of investigated formulations of thyme was determined by gas chromatography and mass spectrometry. Activities of enzyme markers of hepatocellular damage in serum and antioxidant enzymes in the liver homogenates were measured using the kinetic spectrophotometric methods. Liver morphology was characterized by light microscopy using routine hematoxylin and eosin staining. RESULTS: Thymol was found to be predominant active constituent in both tincture and syrup. Investigated thyme preparations exerted antioxidant effects in liver by preventing carbon tetrachloride-induced increase of lipid peroxidation. Furthermore, co-treatment with thyme preparations reversed the activities of oxidative stress-related enzymes xanthine oxidase, catalase, peroxidase, glutathione peroxidase and glutathione reductase, towards normal values in the liver. Hepatotoxicity induced by carbon tetrachloride was reflected by a marked elevation of AST and ALT activities, and histopathologic alterations. Co-administration of thyme tincture resulted in unexpected exacerbation of AST and ALT values in serum, while thyme syrup managed to reduce activites of aminotransferases, in comparison to carbon tetrachloride-treated animals. CONCLUSIONS: Despite demonstrated antioxidant activity, mediated through both direct free radical scavenging and activation of antioxidant defense mechanisms, thyme preparations could not ameliorate liver injury in rats. Molecular mechanisms of diverse effects of thyme preparations on chemical-induced hepatotoxicity should be more in-depth investigated.

Do diethyl phthalate (DEP) and di-2-ethylhexyl phthalate (DEHP) influence the metabolic syndrome parameters? Pilot study.

The study objective was to determine if the healthy participants were exposed to diethyl phthalate (DEP) and di (2-ethylhexyl) phthalate (DEHP) and if this exposure could be linked to the development of metabolic syndrome. The study included 103 healthy volunteers of similar age with normal BMI values, waist circumference, total cholesterol, HDL, LDL, and triglycerides. DEP and DEHP were measured in the morning urine samples to detect monoethyl phthalate (MEP) and mono-2-ethylhexyl phthalate (MEHP). Two phthalate groups and a control group were formed. Both MEP group and control group had similar results. The correlations between MEP and the measured parameters were insignificant. The correlation between the MEHP group and the age was significantly negative, but between the MHEP group and the waist circumference the correlation was significantly positive. Lipids and lipoproteins were within the reference values and equal in both groups. The significant negative correlation was observed only between MEHP and HDL. Our population is exposed to DEP and DEHP. There was only a significant correlation between DEHP and the observed metabolic syndrome components. Its negative impact was higher as the participants were younger.

Estimation of in vivo and in vitro exposure to bisphenol A as food contaminant.

The goal of this cross-sectional study was to examine the occurrence of bisphenol A (BPA) in the morning spot urine taken from 145 female volunteers of various ages. Total urine BPA concentration was detected in 38.6% samples in the 0.92-70.96 µg/g Cr range. The majority of BPA + women belonged to the 25 + body mass index (BMI) group (54.5% were overweight and 43.4% were obese women). Occurrence of BPA in the urine samples was higher at 40 + ages. The maximum BPA concentration of 70.96 µg/g Cr was detected in the urine sample of an obese woman. It is known that BPA is highly toxic in vitro. In this study BPA impaired significantly the growth of all investigated cell lines, i.e. the EC50 values were reached at very low concentrations, in the range from 3.24 to 34.85 µg/mL. The obtained in vivo results suggest that a higher exposure to BPA could contribute to weight problems in women and the absence of the BPA in vitro selective toxicity studies indicates to its general toxic mode of action and raises awareness of the health risks associated with its ubiquitous presence in the environment.

Effects of different extraction methods and conditions on the phenolic composition of mate tea extracts.

A simple and rapid HPLC method for determination of chlorogenic acid (5-O-caffeoylquinic acid) in mate tea extracts was developed and validated. The chromatography used isocratic elution with a mobile phase of aqueous 1.5% acetic acid-methanol (85:15, v/v). The flow rate was 0.8 mL/min and detection by UV at 325 nm. The method showed good selectivity, accuracy, repeatability and robustness, with detection limit of 0.26 mg/L and recovery of 97.76%. The developed method was applied for the determination of chlorogenic acid in mate tea extracts obtained by ethanol extraction and liquid carbon dioxide extraction with ethanol as co-solvent. Different ethanol concentrations were used (40, 50 and 60%, v/v) and liquid CO2 extraction was performed at different pressures (50 and 100 bar) and constant temperature (27 ± 1 °C). Significant influence of extraction methods, conditions and solvent polarity on chlorogenic acid content, antioxidant activity and total phenolic and flavonoid content of mate tea extracts was established. The most efficient extraction solvent was liquid CO2 with aqueous ethanol (40%) as co-solvent using an extraction pressure of 100 bar.

Biomarkers of oxidative stress in amniotic fluid and complications in pregnancy.

OBJECTIVE: In this study, we tried to determine whether the activities of the primary antioxidant enzymes are detectable in amniotic fluid and whether they can be used as early biomarkers of complications in pregnancy such as pregnancy-induced hypertension (PIH), gestational diabetes mellitus (GDM), and bacterial vaginosis. METHODS: This was a prospective study in which amniotic fluid was taken between 16 and 19 week of gestation. In all, 161 pregnant women were divided into two groups: study group - patients with the treated local infection, PIH, and GDM, and control group - healthy pregnant women. Levels of reduced glutathione (GSH) and activities of supeoxide dismutase (SOD), glutathione peroxidase (GSH-Px), glutathione reductase (GSHR), glutathione S-transpherase (GST), xanthine oxidase (XOD) and lipid peroxidation (LP) were determined spectrophotometrically in amniotic fluid samples. RESULTS: Concentration of malondialdehyde varied greatly between investigated groups. XOD and SOD activities, though very low, were present in amniotic fluid samples. Also, enzymes of glutathione cycle and GSH concentrations were detectable and showed certain variations. CONCLUSION: Parameters of oxidative stress in amniotic fluid could be altered in certain pathological conditions. Their use as clinical biomarkers is limited due to great variations of amniotic fluid volume between patients which gives favor to hemolysate or serum of pregnant women.

Bacterial vaginosis and biomarkers of oxidative stress in amniotic fluid.

OBJECTIVE: In this study we tried to determine if the activities of the primary antioxidant enzymes are detectable in amniotic fluid and if they can be used as early biomarkers of complications in pregnancy connected with bacterial vaginosis. METHODS: This was a prospective study in which amniotic fluid was taken between 16 and 19 weeks of gestation. 161 pregnant women were divided into two groups: study group--patients with the treated local infection and control group--healthy pregnant women. Levels of reduced glutathione, and the activities of glutathione peroxidase, glutathione reductase, glutathione S-transferase, xanthine oxidase, superoxide dismutase and lipid peroxidation were determined spectrophotometrically in amniotic fluid samples. RESULTS: Concentration of malonyldialdehide (product of lipid peroxidation) varied greatly between investigated groups. Xanthine oxidase and superoxide dismutase activities, though very low, were present in amniotic fluid samples. Also, enzymes of glutathione cycle and reduced glutathione concentrations were detectable and showed certain variations. CONCLUSION: Although, biomarkers of antioxidant activity are present in the amniotic fluid, they are not different between women with and without bacterial vaginosis.