The levels of 12 cytokines and growth factors in tears: hyperthyreosis vs euthyreosis.

PURPOSE: Simultaneous analyses of the contents and ratios of 12 cytokines and growth factors in single samples of human tears were performed, and the results were compared between a group of healthy subjects and a group of patients with Graves' hyperthyreosis (GH) without thyroid-associated orbitopathy (TAO). METHODS: Determinations and concentration measurements of interleukins (IL-2, IL4, IL-6, IL-8, IL-10, IL-1α, and IL-1ß) interferon (IFN-γ), tumor necrosis factor (TNF-α), monocyte chemoattractant protein (MCP-1), vascular endothelial growth factor (VEGF), and epidermal growth factor (EGF) were performed with single tear samples from 21 patients with hyperthyreosis and 22 healthy subjects. The analyses were performed using a Randox microchip with an Evidence Biochip Array Analyzer. RESULTS: We found significant differences between the healthy donor group and the hyperthyreosis group in the levels of IL-6, IL-10, VEGF, IL-1α, and MCP-1. The concentration of IL-6 was considerably higher in the hyperthyreosis group, IL-10 was higher in the healthy donor group, and VEGF and MPC-1 were higher in the hyperthyreosis group. The IL-8 and IFN-γ levels were higher in the hyperthyreosis group. The ratios of all of the cytokines to anti-inflammatory IL-10 were significantly elevated in the hyperthyreosis group. CONCLUSION: There are clear differences in the levels of cytokines and growth factors in the tears of healthy subjects and patients with GH without TAO. Tear cytokine changes and related dysfunctional tear syndrome (DTS) could be an early sign of occult TAO in Graves' hyperthyreosis patients.

Urine high-sensitive troponin I in children cannot offer an applicable alternative to serum.

Introduction: In children, congenital heart defects represent the primary cause of increased serum troponin I. The elimination process of cardiac troponin I from the bloodstream and the factors influencing this process remain unknown. The objective of this study was to explore the role of troponin I as an indicator of cardiac damage in children both in serum and urine, a concept previously investigated in adults. Methods: Our prospective study involved 70 children under 24 months of age. The first group underwent ventricular septal defect repair, while the second group involved children who had undergone partial cavopulmonary anastomosis. For these groups, urine and serum troponin I were assessed on four occasions. The third group, consisting of healthy children, underwent a single measurement of urine troponin I. Results: Serum troponin I values exhibited an expected elevation in the early postoperative period, followed by a return to lower levels. Significantly higher concentrations of serum troponin I were observed in the first group of children (p < 0.05). A positive correlation was found between troponin I in the first three measurements and cardiopulmonary bypass and aortic cross-clamping time. There was no discernible increase in urine troponin I directly related to myocardial damage; troponin I couldn't be detected in most urine samples. Discussion: The inability to detect troponin I in urine remains unexplained. Potential explanatory factors may include the isoelectric point of troponin I, elevated urinary concentrations of salts and urea, variations in urine acidity (different pH levels), and a relatively low protein concentration in urine.

Inflammation, haemostatic disturbance, and obesity: possible link to pathogenesis of diabetic retinopathy in type 2 diabetes.

PURPOSE: The pathogenesis of diabetic retinopathy (DR) is insufficiently understood but may possibly involve chronic, low-grade inflammation. The aim of this cross-sectional study was to investigate the relationship between inflammatory and haemostatic markers, other markers of endothelial dysfunction and anthropometric parameters, and their association with DR in patients with type 2 diabetes. METHODS: According to the DR status patients were divided into three groups: no retinopathy, mild/moderate nonproliferative (NPDR), and severe NPDR/proliferative retinopathy (PDR). RESULTS: The groups did not differ in the levels of inflammatory and haemostatic markers, other markers of endothelial dysfunction, and anthropometric parameters. After dividing the patients according to the level of obesity (defined by BMI, WC, and WHR) into three groups ANOVA showed the differences in C-reactive protein according to the WC (P = 0.0265) and in fibrinogen according to the WHR (P = 0.0102) as well as in total cholesterol (P = 0.0109) and triglycerides (P = 0.0133) according to the BMI. Logistic regression analyses showed that diabetes duration and prolonged poor glycemic control are the main predictors of retinopathy in patients with type 2 diabetes. CONCLUSION: Interrelations between obesity, inflammation, haemostatic disturbance, and other risk factors may possibly play an important additional role in endothelial dysfunction involved in the pathogenesis of diabetic retinopathy.

Monitoring of nitrates in drinking water from agricultural and residential areas of Podravina and Prigorje (Croatia).

Nitrates are the most common chemical pollutant of groundwater in agricultural and suburban areas. Croatia must comply with the Nitrate Directive (91/676/EEC) whose aim is to reduce water pollution by nitrates originating from agriculture and to prevent further pollution. Podravina and Prigorje are the areas with a relatively high degree of agricultural activity. Therefore, the aim of this study was, by monitoring nitrates, to determine the distribution of nitrates in two different areas, Podravina and Prigorje (Croatia), to determine sources of contamination as well as annual and seasonal trends. The nitrate concentrations were measured in 30 wells (N = 382 samples) in Prigorje and in 19 wells (N = 174 samples) in Podravina from 2002 to 2007. In Podravina, the nitrate content was 24.9 mg/l and 6% of the samples were above the maximum available value (MAV), and in Prigorje the content was 53.9 mg/l and 38% of the samples above MAV. The wells were classified as correct, occasionally incorrect and incorrect. In the group of occasionally incorrect and incorrect wells, the point sources were within 10 m of the well. There is no statistically significant difference over the years or seasons within the year, but the interaction between locations and years was significant. Nitrate concentrations' trend was not significant during the monitoring. These results are a prerequisite for the adjustment of Croatian standards to those of the EU and will contribute to the implementation of the Nitrate Directive and the Directives on Environmental Protection in Croatia and the EU.

Influence of Clinically Active Graves' Ophthalmopathy on Spherical Equivalent and Visual Acuity.

Purpose: Clinical experience regarding the fluctuations of the refractive error of the eye during the different stages of Graves' ophthalmopathy observed through outpatient clinic frequent check-ups points towards an underestimated and often overlooked problem. Published data about it are sparse. The clinical manifestations of Graves' ophthalmopathy can be understood from the perspective of "compartment syndrome" and literature implies how such changes can affect the refractive error and consequently, the visual acuity. The purpose of the study was to explore how the clinical activity score of Graves' ophthalmopathy affects refractive error and visual acuity. Patients and Methods: The study was prospective and observational, including 60 eyes of 30 patients with clinically active Graves' ophthalmopathy. All the patients were monitored and evaluated over a period of 36 months by the clinical activity score, spherical equivalent and visual acuity. All the observed parameters were statistically analyzed. Results: The mean values of spherical equivalent and visual acuity throughout the observed period showed continuous fluctuation. Repeated measure analysis of variance showed statistically significant differences in visual acuity and spherical equivalent over the observed period. There was a statistically significant positive correlation between visual acuity and clinical activity score. The correlation between spherical equivalent and clinical activity score was also positive but not statistically significant. Conclusion: A decrease in the clinical activity score is either the result of a spontaneously resolving course of Graves' ophthalmopathy or a consequence of treatment, so lowering in fluctuation of refractive error and improved visual acuity may be associated with a reduction in orbital inflammation.

Adiponectin has different mechanisms in type 1 and type 2 diabetes with C-peptide link.

PURPOSE: Adiponectin (ApN) is considered to be responsible for reduction of inflammation and is known to be included in lipid metabolism. This study was designed to assess the role of adiponectin in patients with type 1 and type 2 diabetes and to determine parameters important in the prediction of adiponectin. METHODS: Adiponectin, high sensitive C-reactive protein, fibrinogen, homocysteine, C-peptide, and lipid panel in addition to clinical and laboratory parameters important for the definition of diabetes, obesity and the metabolic syndrome were measured in 118 patients. RESULTS: The best model (R2=0.989) for predicting adiponectin in type 1 diabetes included fibrinogen, white blood cell count, uric acid and triglycerides. In type 2 diabetes the best model (R2=0.751) included C-peptide, white blood cell count, systolic blood pressure, fasting blood glucose, glycated hemoglobin and high-density lipoprotein cholesterol. ANOVA showed among-group differences in adiponectin (P=0.028), body mass index (P < 0.001), fasting blood glucose (P < 0.001) and high-density lipoprotein cholesterol (P =0.012) according to the type of diabetes. Between-group differences were also observed in adiponectin (P =0.033) and high-density lipoprotein cholesterol (P =0.009) according to sex. Adiponectin correlated (P < 0.05) with body mass index, C-peptide, pulse pressure and high-density lipoprotein cholesterol. CONCLUSION: Adiponectin levels were higher in type 1 diabetes. The association between C-peptide and adiponectin is probably one of the reasons for their different respective levels in different types of diabetes. Interrelations between adiponectin and inflammation, dyslipidemia, C-peptide levels and sex appear to be important for complex adiponectin modulation and action.

Genotoxic effects of anaesthetics in operating theatre personnel evaluated by the comet assay and micronucleus test.

Genetic damage induced by anaesthetic gases in occupationally exposed populations was investigated using the comet assay and micronucleus test. The study included two groups of subjects: 50 operating theatre medical workers (anaesthesiologists, technicians and nurses) and 50 control subjects corresponding in sex, age and smoking habit. The exposed group revealed an increase in genome damage in both tests. In the comet assay, exposure to anaesthetics was a highly significant predictor of the tail length for technicians, while sex proved to be significant predictor of tail moment for women in exposed group. Micronucleus frequency increased significantly, showing threefold increase in exposed groups (RR>3.029). Univariate analysis showed significant influence of duration of exposure, while multivariate analysis showed age to be significant predictor of micronucleus frequency. The obtained results call for further, targeted investigation of exposure risk.

Developmental and transplacental genotoxicology: fluconazole.

Over the last 40 years mankind has been facing new types of radiochemical environmental settings with every decade. During the last decade, biomonitoring was additionally focused on assessing associations between environmental exposure(s) and both early and late biological effects in children. Despite efforts to control and avoid child exposure to genotoxic agents the incidence of childhood cancers is increasing. Some cancers in adulthood may be the consequence of a multi-step process which starts with intrauterine and childhood exposure. This highlights the importance of a comprehensive interpretation of multiple health effects, especially considering recent studies suggesting that most health disorders are related to DNA changes. When exposed to genotoxic agents, a developing organism (fetus or child) is constantly being forced to reorganize into new equilibriums in order to adjust to a xenobiotic environment. In addition, the influence of sex hormones on radiochemical sensitivity is still unknown. For this reason special attention should be paid to puberty. The results of recent studies on animal models and follow up studies on children after nuclear accidents show long-lasting cytogenetic damage even after low dose exposures and their transgenerational persistance. To evaluate age-related difference and transplacental genotoxic potency fluconazole (FC) was investigated by in vivo micronucleus (MN) assay in adult mice, young mice and in transplacentally exposed newborn pups. Compared to the baseline values, FC caused no detectable genome damage in adult animals, but there was a significant increase in MN frequency in young animals and in newborn pups. Our study thus exemplifies an age-related chemosensitivity, and argues that cancer-promoting disturbances of complex prenatal developmental mechanisms and maturation during childhood require a new approach using systems biology.

Metallothionein 2A gene polymorphism and trace elements in mother-newborn pairs in the Croatian population.

The main source of exposure for all essential and toxic elements in the general population is diet. In smokers, the main route for cadmium (Cd) and lead (Pb) intake is the inhalation of tobacco smoke. Besides gender, age, nutrition, lifestyle, and physiological conditions such as pregnancy, specific genetic characteristics also influence individual element uptake. Metallothionein MT2 is a cysteine-rich low-weight protein found ubiquitously throughout the body. Specific gene polymorphism may influence MT2 expression and subsequent binding, transfer and organ accumulation of metals, though data on these influences are lacking, especially in human mother-newborn pairs. The objective of this study was to determine selected toxic (Cd, Pb, Hg) and essential (Fe, Zn, Cu, Se) elements in maternal blood, placenta, and cord blood (by ICP-MS), and MT2 levels in maternal serum (by ELISA) in relation to maternal MT2A -5A/G (rs28366003) polymorphism (by RFLP-PCR and electrophoresis). Study participants were healthy postpartum women in Croatia (n=268, mean age 29 years) with term vaginal childbirth in a maternity ward assigned into two study groups by self-reporting about their smoking habit (by questionnaire). Smokers vs. non-smokers had increased levels of Cd and Pb in all measured samples, Fe and Cu in cord blood, Zn in placenta, and MT2 in maternal serum. Among subjects with AG/GG genotype, placental Fe was significantly lower only among non-smokers, while MT2 levels in serum were lower, though not significantly, regardless of maternal smoking habit. There was no impact of MT2A -5A/G SNP on any element in maternal or cord blood. In conclusion, the results confirmed maternal smoking-related increases in Cd and Pb levels in the maternal-placental-foetal unit. They also provided additional data on concomitant metal concentrations in representative samples of maternal blood, placenta, and cord blood, as well as increased cord blood Fe and Cu, placental Zn, and maternal serum MT2 in smokers. New evidence is that MT2A -5A/G SNP was associated with decreased placental Fe levels in non-smokers. For a final conclusion on the influence of the MT2A -5A/G polymorphism on toxic and essential element levels in mother-newborn pairs, further research would require a larger number of participants divided across subgroups defined by the main source of particular toxic metal exposure (such as specific food intake, cigarette smoking, air pollution and/or occupational exposure).

Comparison of genomic damage caused by 5-nitrofurantoin in young and adult mice using the in vivo micronucleus assay.

The antibiotic 5-nitrofurantoin (5-NF) has been used widely for the treatment of urosepsis in children during the last 20 years. Recent experimentation suggests the need for reevaluating its genotoxic potential. Because of possible differences in the metabolism and clearance of 5-NF in young and adult animals, we conducted a study to determine whether micronuclei caused by 5-NF were age-related. The in vivo micronucleus (MN) assay was performed on 3- and 8-week-old mice given single intraperitoneal injections of 5, 10, and 50 mg/kg 5-NF. Blood samples from the tail vein were taken before injection (baseline) and at 48, 96, 168, and 336 hr (2 weeks) after the treatment. One thousand reticulocytes were analyzed for micronuclei from each animal. Compared to similar baseline values for young and adult mice, 5-NF caused a significant increase in MN frequency in both age groups. The mean MN frequency in the young animals was higher than in the adult animals for each dose and sampling time. MN frequencies remained significantly elevated in young animals even 2 weeks after exposure to 5-NF. The results of the study confirm the genotoxic potential of 5-NF in young and adult animals, and indicate that young animals are more sensitive to the genotoxic effects of 5-NF than adult mice and that the response in young mice persists for a significantly longer time. These findings may be related to poorly developed mechanisms of xenobiotic detoxification and renal elimination in young animals.

Inverse Levels of Adiponectin in Type 1 and Type 2 Diabetes Are in Accordance with the State of Albuminuria.

Aims. To investigate the behaviour of adiponectin (ApN) in patients with type 1 and type 2 diabetic nephropathy. Methods. ApN and inflammatory and other markers of the metabolic syndrome were compared across diabetes types, albumin excretion rate (AER), and creatinine clearance (CrCl) categories in 219 type 1 and type 2 diabetic patients. Results. Significant differences among ApN levels according to AER were found in both types of diabetes (F = 8.45, df = 2, P < 0.001). With the progression of albuminuria, ApN increased in type 1 and decreased in type 2 diabetes. Patients with decreased CrCl had higher ApN levels than those with normal CrCl in either type of diabetes (F = 12.7, df = 1, P < 0.001). The best model for ApN (R (2) = 0.9002) obtained from stepwise regression in type 1 diabetes included CrCl, BMI, WBC, CRP, and age, while in type 2 diabetes (R (2) = 0.2882) it included ppPG, LDL, and UA. Conclusion. ApN behaved differently in relation to albuminuria, increasing with its progression in type 1 diabetes and decreasing in type 2 diabetes. It was however increased in the subgroups with decreased CrCl in both types of diabetes. Albuminuria seems to be more important than renal insufficiency in the definition of ApN levels in type 1 and type 2 diabetes.

Polymorphisms in DNA repair genes: link with biomarkers of the CBMN cytome assay in hospital workers chronically exposed to low doses of ionising radiation.

Individual sensitivity to ionising radiation (IR) is the result of interaction between exposure, DNA damage, and its repair, which is why polymorphisms in DNA repair genes could play an important role. We examined the association between DNA damage, expressed as micronuclei (MNi), nuclear buds (NBs), and nucleoplasmic bridges (NPBs) and single nucleotide polymorphisms in selected DNA repair genes (APE1, hOGG1, XRCC1, XRCC3, XPD, PARP1, MGMT genes; representative of the different DNA repair pathways operating in mammals) in 77 hospital workers chronically exposed to low doses of IR, and 70 matched controls. A significantly higher MNi frequency was found in the exposed group (16.2±10.4 vs. 11.5±9.4; P=0.003) and the effect appeared to be independent from the principal confounding factor. Exposed individuals with hOGG1, XRCC1, PARP1, and MGMT wild-type alleles or APEX1, as well as XPD (rs13181) heterozygous showed a significantly higher MNi frequency than controls with the same genotypes. Genetic polymorphism analysis and cytogenetic dosimetry have proven to be a powerful tool complementary to physical dosimetry in regular health surveillance programmes.

Impact of coffee and other selected factors on general mortality and mortality due to cardiovascular disease in Croatia.

In Croatia, the mortality rate is higher than that in the countries of the European Union (EU), and consumption of coffee is moderate compared to the EU countries. The study examined the effects of coffee consumption on all-cause (general) mortality, mortality due to cardiovascular disease, and survival. Analyses were based on data obtained from an epidemiological longitudinal study started in 1969 with follow-ups in 1972, including 1,571 men and 1,793 women aged 35-59 years, and in 1982, including 1,093 men and 1,330 women. The sample was age- and gender-stratified and included urban and rural populations from three coastal and three continental regions of Croatia. During the observation period from spring 1972 to the end of 1999, 568 men and 382 women died. In total, 254 men and 181 women died due to cardiovascular disease. The sample was classified in 4 groups: non-drinkers, consumption of coffee sometimes, regularly 1-2 cup(s), and regularly more than 2 cups per day. Apart from coffee, the effects of diastolic blood pressure, smoking habit, well-being, stomach ulcer, and resident status were analyzed. Data on general mortality and mortality due to cardiovascular disease were also analyzed. The influence of region and the effects of diastolic pressure and smoking habit on general mortality and cardiovascular disease-associated mortality were confirmed in both the sexes. No significant effects of coffee consumption on general mortality and mortality due to cardiovascular disease were found among men. Positive effects of coffee on general mortality (p = 0.0089) but not on cardiovascular disease-associated mortality were observed among women. Women who regularly drank coffee 1-2 cup(s) per day had a significantly lower risk of all-cause death adjusted for age, region, smoking, diastolic blood pressure, feeling of well-being, and history of stomach ulcer (relative risk = 0.631; p = 0.0033; confidence interval: 0.464-0.857). The role of coffee consumption on mortality was less relevant than other variables. However, it cannot be completely neglected in women.

Smoking habits, signs of chronic diseases and survival in inland and coastal regions of Croatia: a follow-up study.

Aim of the study was to estimate, the relationship between survival, smoking habits, and the results of medical examinations in inland and coastal regions of Croatia. Age and sex stratified sample of general population (1,571 men and 1,793 women, 37-56 years old in 1972; followed in 1982: N = 1,090 men; 1,325 women and/or 1972-1999 controlling vital status). Relative risks and 95% confidence limits were estimated using Cox regression in the model with time dependent covariates, separately by sex. In all regions, the proportion of male smokers decreased between 1972 and 1982. The proportion of female smokers increased, differently in urban and rural regions. During the follow-up between 1972 and 1999, 568 deaths were recorded among men and 382 among women. In men, in addition to age, significant hazards of death were the number of smoked cigarettes per day, body mass index, sedative intake, vital lung capacity (FVC), 100FEV1/VC, systolic blood pressure, electrocardiogram, history of heart attack, and region. In women, in addition to age, significant predictors were the number of smoked cigarettes per day, systolic blood pressure, electrocardiogram, history of heart disease (excluding coronary diseases), and region. Survival relative risk increased with each additional smoked pack of cigarettes by 2.4% in women and 1.3% in men. Regional differences vs. smoking habit were observed. These data emphasize the need for prevention of smoking.

Effect of summer ozone concentrations on the lung function of walkers in the Medvednica Mountain Nature Park, Croatia.

The study examines the influence of naturally elevated ozone concentrations and some meteorological variables on the lung function of untrained volunteers walking in an unpolluted mountain area. Forty male participants between 18 and 70 years (smokers and nonsmokers) walked at the top of Medvednica Mountain near Zagreb, Croatia, at approximately 1000 m above sea level (a.s.l.) and engaged in other recreational activities for at least 1 hour. Forced vital capacity (FVC) and forced expiratory volume in 1 second (FEV(1)) of the participants were measured at arrival and before they left the mountain. Regression analysis showed a statistically significant influence of environmental variables on lung function tests. However, all variables together explained less than 35% variability of FVC and 41% variability of FEV(1). The results suggest that short-term exposure to ozone may affect lung function tests, but no more than temperature and humidity. FEV(1) was found to be more sensitive to ozone than FVC. Smoking habit also had an important role in subject's sensitivity to ozone.

Chromosome aberrations in peripheral blood lymphocytes of Croatian hospital staff occupationally exposed to low levels of ionising radiation.

Medical staff is an occupational group exposed to different agents suspected to induce genetic damage. Among them ionising radiation is the most studied. Cytogenetic analysis of human chromosomes in peripheral lymphocytes allows direct detection of mutation in somatic cells. This study investigated the cytogenetic effects of low-level ionising x-radiation in 48-hour peripheral blood lymphocyte cultures sampled from 765 hospital staff occupationally exposed to several agents known or suspected to induce chromosome damage and compared them with 200 control subjects. The exposed subjects were divided in eight (8) groups according to their specialties and job titles. The exposed groups manifested an increase in all types of chromosome aberrations. Acentric fragments were the most frequent chromosome-type aberration. Dicentric chromosomes were statistically significant only in urologists/gynaecologists. Age and smoking significantly influenced the incidence of dicentrics in the exposed groups. The frequency of ring chromosomes was low in all exposed groups (range: 0-2), and none were found in the control group. These findings indicate the importance of periodic medical checkups of hospital staff occupationally exposed to low doses of ionising radiation. The purpose is to create an individual cytogenetic register, where changes could evidence individual risks.