Identification of an Additional Metal-Binding Site in Human Dipeptidyl Peptidase III.

Dipeptidyl peptidase III (DPP III, EC 3.4.14.4) is a monozinc metalloexopeptidase that hydrolyzes dipeptides from the N-terminus of peptides consisting of three or more amino acids. Recently, DPP III has attracted great interest from scientists, and numerous studies have been conducted showing that it is involved in the regulation of various physiological processes. Since it is the only metalloenzyme among the dipeptidyl peptidases, we considered it important to study the process of binding and exchange of physiologically relevant metal dications in DPP III. Using fluorimetry, we measured the Kd values for the binding of Zn2+, Cu2+, and Co2+ to the catalytic site, and using isothermal titration calorimetry (ITC), we measured the Kd values for the binding of these metals to an additional binding site. The structure of the catalytic metal's binding site is known from previous studies, and in this work, the affinities for this site were calculated for Zn2+, Cu2+, Co2+, and Mn2+ using the QM approach. The structures of the additional binding sites for the Zn2+ and Cu2+ were also identified, and MD simulations showed that two Cu2+ ions bound to the catalytic and inhibitory sites exchanged less frequently than the Zn2+ ions bound to these sites.

Chemical Characterization and Antioxidant Potential of the Rowan (Sorbus aucuparia L.) Fruits from Alpine-Dinaric Region of Croatia§.

Research background: The rowan (Sorbus aucuparia L.) is a small tree in the Rosaceae family with characteristic orange-red fruits. The raw fruits can be used for making jams, juices and puree, while the dried fruits are used for teas. In folk medicine, they have been used to prevent scurvy and bleeding or as a diuretic and laxative. The aim of this study is to characterize the proximate chemical composition, antioxidant potential and macro- and trace elements of the rowan fruits for their potential use as a functional food. Experimental approach: The fruits were collected from 12 populations in the Alpine-Dinaric region of Croatia. After collection, the samples were transported to the laboratory, cut into small pieces, placed in plastic containers and stored at -20 °C until analysis. Proximate chemical composition, including ash, water, cellulose, crude fat and crude protein, was determined according to standard methods and total carbohydrates as non-structural carbohydrates. Total phenolic content (TPC) and antioxidant capacity (TAC) were also measured. For multielement analysis, fruits were cleaned from the dust, lyophilised, homogenised and acid-digested in a microwave system. Concentrations of elements were determined using the inductively coupled plasma mass spectrometry. Results and conclusions: The basic constituents in the analysed fruits were (in %): water 76.53, total carbohydrates 17.45, crude proteins 2.98, crude fats 1.49, cellulose 1.07 and ash 1.29. On average, the TPC was 932 mg/100 g and the TAC was (60.1±14.5) % and (4.1±1.2) mmol/100 g, determined by DPPH and FRAP assay, respectively. Mass fractions of elements decreased as follows (in mg/kg): K 2485>Ca 459>P 206, Mg 193>Na 6.29>Fe 3.68>Mn 3.58>Zn 1.11>Cu 0.731>Mo 0.098>Co 0.003>Se 0.001. Compared to the literature, the phenolic and element content of the rowan fruits is similar to that of blueberry and raspberry. The obtained results suggest that rowan fruits have valuable nutritional properties and could be useful for fortification in the food industry. Novelty and scientific contribution: The importance of the obtained results is reflected in filling in the gaps in the literature on the composition of elements, especially on the content of essential macro- and trace elements as well as the antioxidant potential of rowan fruits.

Heavy Metals and Essential Metals Are Associated with Cerebrospinal Fluid Biomarkers of Alzheimer's Disease.

Various metals have been associated with the pathogenesis of Alzheimer's disease (AD), principally heavy metals that are environmental pollutants (such as As, Cd, Hg, and Pb) and essential metals whose homeostasis is disturbed in AD (such as Cu, Fe, and Zn). Although there is evidence of the involvement of these metals in AD, further research is needed on their mechanisms of toxicity. To further assess the involvement of heavy and essential metals in AD pathogenesis, we compared cerebrospinal fluid (CSF) AD biomarkers to macro- and microelements measured in CSF and plasma. We tested if macro- and microelements' concentrations (heavy metals (As, Cd, Hg, Ni, Pb, and Tl), essential metals (Na, Mg, K, Ca, Fe, Co, Mn, Cu, Zn, and Mo), essential non-metals (B, P, S, and Se), and other non-essential metals (Al, Ba, Li, and Sr)) are associated with CSF AD biomarkers that reflect pathological changes in the AD brain (amyloid ß1-42, total tau, phosphorylated tau isoforms, NFL, S100B, VILIP-1, YKL-40, PAPP-A, and albumin). We used inductively coupled plasma mass spectroscopy (ICP-MS) to determine macro- and microelements in CSF and plasma, and enzyme-linked immunosorbent assays (ELISA) to determine protein biomarkers of AD in CSF. This study included 193 participants (124 with AD, 50 with mild cognitive impairment, and 19 healthy controls). Simple correlation, as well as machine learning algorithms (redescription mining and principal component analysis (PCA)), demonstrated that levels of heavy metals (As, Cd, Hg, Ni, Pb, and Tl), essential metals (Ca, Co, Cu, Fe, Mg, Mn, Mo, Na, K, and Zn), and essential non-metals (P, S, and Se) are positively associated with CSF phosphorylated tau isoforms, VILIP-1, S100B, NFL, and YKL-40 in AD.

Spatio-temporal monitoring of mercury and other stable metal(loid)s and radionuclides in a Croatian terrestrial ecosystem around a natural gas treatment plant.

The natural gas industry bears a certain contamination risk to human and biota due to, among others, mercury, arsenic, and naturally occurring radioactive material content in gas. We tracked multiple stabile metal(loid)s and radionuclides within the natural gas treatment plant Molve, Croatia, ecosystem during the last decade through a comprehensive monitoring of soil, earthworms, moss, livestock (blood, milk, hair, urine, and feces from cows), and wildlife animals (brain, muscle, liver, and kidney of European hare and pheasant). The level of mercury and other stable metal(loid)s has shown temporal variation, but without an obvious trend. The found spatial differences in soil and earthworms were based on the differing soil characteristics of the sampled locations and exceeded the maximal allowable concentration of arsenic and zinc for agricultural soil. The status of essential copper, selenium, and zinc in cows, hares, and pheasants inclined towards deficiency. The measured stable metal(loid) levels in soil and animal samples were generally in the same range of values reported in earlier decades from the same area or non-polluted areas across Europe. The consumption of local cow and game products (e.g., milk, meat) can be considered safe for human health, although game offal is advised to be avoided as a food item due to the low risk of lead and cadmium's adverse effects. Although the activity concentrations of some radionuclides in moss were higher than reported for pristine areas, transfer from soil to moss was assessed as average (except for lead-210). Radiological risk to human and biota around the gas treatment plant Molve was estimated as negligible.

Metal(loid) exposure assessment and biomarker responses in captive and free-ranging European brown bear (Ursus arctos).

We investigated the level of five non-essential metal(loid)s (As, Cd, Hg, Tl, Pb) and nine essential metals (Mg, Ca, Mn, Fe, Co, Cu, Zn, Se, Mo) in hair and blood components of captive and free-ranging European brown bear populations in Croatia and Poland. Metal(loid) associations with biomarkers of oxidative stress (superoxide dismutase, SOD; glutathione-peroxidase, GSH-Px; malondialdehyde, MDA) and metal exposure (metallothionein, MT) were estimated in this top predatory mammal. Lead was the most abundant non-essential metal(loid) in both blood and hair, with 4 of 35 individuals having blood levels over 100 µg/L. A positive association was found between Pb level and SOD activity in blood. Free-ranging bears had higher blood SOD activity, Mn, Zn and Cd levels, hair Co, Cd, Tl and Pb compared to captive individuals, while the opposite was true for Mg and hair Ca thereby reflecting habitat and diet differences. With increasing age, animals showed lower levels of SOD activity and certain essential metals. Females had higher SOD activity and blood levels of some essential metals than males. Hair showed a higher Fe and Co level when sampled during the growth phase and was not predictive of non-essential metal(loid) blood levels. The established metal(loid) baseline values will enable future risk assessment in both captive and wild European brown bear populations.

Sexual Maturity and Life Stage Influences Toxic Metal Accumulation in Croatian Brown Bears.

The influence of reproductive and (early) life stages on toxic metal levels was investigated in the brown bear (Ursus arctos), the largest mammalian predator species in Croatia. The purpose was to examine critical clusters in a population that might be at a higher risk of adverse health effects caused by metals as environmental contaminants. Levels of cadmium, mercury and lead in muscle, liver and kidney cortex of 325 male and 139 female bears, quantified by inductively coupled plasma mass spectrometry, were analysed according to distinct bear life stages (young: cub, yearling, subadult; mature: adult). Metal levels did not differ among sexes in young animals (< 4 years), except for mercury in muscles (higher in females), and adult females had higher cadmium and mercury. A trend of renal cadmium accumulation with age in immature male animals disappeared once they reached maturity, whereas for females this trend has only slowly declined in mature compared to immature bears. In early life stage (< 1 year), bear cubs had lower cadmium, comparable mercury, and higher lead in the kidneys than the bears of the following age category (yearlings). Due to a higher proportion of renal lead transfer from the mother to the cub compared with cadmium, it may be that the high burden of cadmium found in kidneys of older females has lower toxicological concern for their cubs than the lead content. Sex, reproductive, and life stages of bears were confirmed as important in assessing toxic metal burden.

Assessment of multiple anthropogenic contaminants and their potential genotoxicity in the aquatic environment of Plitvice Lakes National Park, Croatia.

In this study, the influence of anthropogenic pollution on the aquatic environment of Plitvice Lakes National Park (PLNP) was investigated during 2011-2012 using a combination of chemical and cytogenetic analyses. Four groups of major contaminants [(volatile organic compounds: benzene, toluene, ethylbenzene, and xylenes (BTEX); persistent organochlorine pollutants: organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs); major and trace elements; anthropogenic radionuclides (90Sr, 134Cs, and 137Cs)] were determined in three aquatic compartments (water, sediment, fish). Mass fractions of inorganic constituents in different compartments reflected the geological background of the area, indicating their origin from predominantly natural sources. Levels of volatile and persistent organic compounds in water and fish, respectively, were very low, at levels typical for remote pristine areas. Analysis of anthropogenic radionuclides in water and sediment revealed elevated activity concentrations of 137Cs in water, and measurable 134Cs in the upper sediment layers from April 2011, possibly as a consequence of the Fukushima nuclear accident in March 2011. The potential genotoxicity of river and lake water and lake sediment was assessed under laboratory conditions using the alkaline comet assay on human peripheral blood lymphocytes, and measured levels of primary DNA damage were within acceptable boundaries. The results showed that despite the protected status of the park, anthropogenic impact exists in both its terrestrial and aquatic components. Although contaminant levels were low, further monitoring is recommended to make sure that they will not rise and cause potentially hazardous anthropogenic impacts.

Trace elements concentrations association with schizophrenia symptoms; A cross-sectional study in Croatia.

BACKGROUND: Significant inconsistencies exist in findings on association of bio-elements (BE) concentrations and schizophrenia. Hypothesis of this research was that different concentrations of BE are associated with different psychopathological schizophrenia symptoms. SUBJECTS AND METHODS: This cross-sectional study was performed from 2014 to 2016 at Psychiatric Hospital "Sveti Ivan" and University Psychiatric Hospital "Vrapce", Zagreb, Croatia, on the consecutive sample of 67 patients diagnosed with schizophrenia. BE concentrations were measured by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) at the Institute for Medical Research and Occupational Health in Zagreb. Severity of schizophrenia symptoms was assessed on Brief Psychiatric Rating Scale (BPRS). RESULTS: After adjustment for all preplanned possible confounding variables, the first canonical correlation between BE and BPRS dimensions variates were statistically significant (Rc2=0.73; P=0.006). The first pair of canonical variates is defined by BPRS negative dimension (and marginally by positive symptoms and lack of resistance), and copper (Cu), lead (Pb), lithium (Li) and cobalt (Co) (marginally by cadmium (Cd) and nickel (Ni)). CONCLUSIONS: Concentrations of different BE are associated with different schizophrenia symptoms. Maximal correlation between BPRS and BE may be achieved with the weighted linear composite of negative schizophrenia symptoms and copper (Cu), lead (Pb), lithium (Li) and cobalt (Co).

Placental cadmium as an additional noninvasive bioindicator of active maternal tobacco smoking.

Tobacco smoke (TS) is a mixture of chemicals that is known to exert carcinogenic and endocrine-disrupting effects, as well as adverse effects on various systems. In TS nicotine is the major alkaloid and cadmium (Cd) the most abundant metal ion. The aim of this investigation was to assess exposure to Cd attributed to TS in healthy postpartum subjects (mean age 28 years) after term vaginal delivery in a clinical hospital by determining metal levels in maternal blood, placenta, and cord blood in relation to nicotine in maternal hair (12-cm-long samples). Two study groups were compared based upon self-reporting data: smokers (n = 32; continual cigarette smoking 3 months before and 9 months during pregnancy) and nonsmokers (n = 54; including passive smokers whose parameters did not differ from unexposed nonsmokers). In smokers compared to nonsmokers maternal hair nicotine concentrations increased approximately sevenfold, while Cd levels rose fourfold in maternal blood and up to twofold in placenta. Significant positive correlations were noted between maternal hair nicotine and placental Cd, maternal hair nicotine and maternal blood Cd, and placental Cd and maternal blood Cd. Levels of cord blood Cd were low in both study groups (<0.1 ng/ml). Data indicate that Cd in placenta may serve as a noninvasive bioindicator in addition to commonly used noninvasive hair nicotine in maternal TS assessment, especially in cases where unavailable or inappropriate (short or chemically treated) hair samples occur.

Mercury in Eisenia fetida and soil in the vicinity of a natural gas treatment plant in northern Croatia.

In the last two decades (1990-2012), as part of a mercury monitoring programme, earthworms and soils have been collected from four locations in the vicinity of a natural gas production and treatment plant near the village of Molve, Croatia. The aim of this study was to determine the concentration of mercury in the collected samples, monitor its changes over a longer period of time and determine the bioaccumulation of total mercury in earthworms (Eisenia fetida) from the soil. Total mercury concentrations in earthworms from the surroundings of four boreholes (Molve 9-12) ranged within 0.195-1.050, 0.129-1.0, 0.229-1.236 and 0.223-0.799 µg g-1 dry weight, while total mercury concentrations in different soil types at the same locations within 0.055-0.350, 0.035-0.250, 0.031-0.240 and 0.071-0.475 µg Hg g-1 of soil. The calculated mercury bioaccumulation factor ranged between 0.9 and 17.5. Mercury levels in soil and earthworms, as a tool for soil pollution assessment, suggested low mercury exposure and risks for human health in the monitored area.

The impact of multiple anthropogenic contaminants on the terrestrial environment of the Plitvice Lakes National Park, Croatia.

The anthropogenic impact on the terrestrial environment of the Plitvice Lakes National Park (PLNP) was investigated through the analysis of three groups of major contaminants (persistent organochlorine pollutants including 15 organochlorine pesticides (OCPs) and 17 polychlorinated biphenyls (PCBs), trace elements/heavy metals (6 major and 23 trace constituents), and anthropogenic radionuclides ((90)Sr, (134)Cs, and (137)Cs)) in three terrestrial compartments (soil, air, and bioindicators of air contamination) during 2011-2013. The correlation coefficients of element mass fractions with soil properties indicated that total Fe and Al minerals, soil organic matter (OM), and organic carbon (OC) content affected the mass fractions of most trace elements in the topsoils. The annual and spatial distributions of heavy metals in total deposited matter (TDM) indicated that the metals came from natural sources and long-range transfer of particulate matter. The PCB and OCP levels found in soil and conifer needles corresponded to global environmental pollution levels by persistent organic pollutants and represented the lower end of the mass fraction ranges reported in the relevant literature. Analyses of anthropogenic radionuclides in bioindicators (conifer needles, lichens, and mosses) showed low but measurable activity concentrations of (134)Cs (for the first time after the Chernobyl accident), which indicated origin from the March 2011 Fukushima Nuclear Power Plant accident. Our overall results indicated that human activity inside or near the PLNP had no significant impact either on contaminant spread by air or on their content in topsoils.

In female rats, ethylene glycol treatment elevates protein expression of hepatic and renal oxalate transporter sat-1 (Slc26a1) without inducing hyperoxaluria.

AIM: To investigate whether the sex-dependent expression of hepatic and renal oxalate transporter sat-1 (Slc26a1) changes in a rat model of ethylene glycol (EG)-induced hyperoxaluria. METHODS: Rats were given tap water (12 males and 12 females; controls) or EG (12 males and 12 females; 0.75% v/v in tap water) for one month. Oxaluric state was confirmed by biochemical parameters in blood plasma, urine, and tissues. Expression of sat-1 and rate-limiting enzymes of oxalate synthesis, alcohol dehydrogenase 1 (Adh1) and hydroxy-acid oxidase 1 (Hao1), was determined by immunocytochemistry (protein) and/or real time reverse transcription polymerase chain reaction (mRNA). RESULTS: EG-treated males had significantly higher (in µmol/L; mean±standard deviation) plasma (59.7±27.2 vs 12.9±4.1, P<0.001) and urine (3716±1726 vs 241±204, P<0.001) oxalate levels, and more abundant oxalate crystaluria than controls, while the liver and kidney sat-1 protein and mRNA expression did not differ significantly between these groups. EG-treated females, in comparison with controls had significantly higher (in µmol/L) serum oxalate levels (18.8±2.9 vs 11.6±4.9, P<0.001), unchanged urine oxalate levels, low oxalate crystaluria, and significantly higher expression (in relative fluorescence units) of the liver (1.59±0.61 vs 0.56±0.39, P=0.006) and kidney (1.77±0.42 vs 0.69±0.27, P<0.001) sat-1 protein, but not mRNA. The mRNA expression of Adh1 was female-dominant and that of Hao1 male-dominant, but both were unaffected by EG treatment. CONCLUSIONS: An increased expression of hepatic and renal oxalate transporting protein sat-1 in EG-treated female rats could protect from hyperoxaluria and oxalate urolithiasis.

Molecular field analysis of trophic relationships in soil-dwelling invertebrates to identify mercury, lead and cadmium transmission through forest ecosystems.

Contamination pathways in complex food chains in soil ecosystems can be difficult to elucidate. Molecular analysis of predator gut content can, however, rapidly reveal previously unidentified trophic interactions between invertebrates and thereby uncover pathways of pollutant spread. Here, we measured concentrations of the toxic metals lead, cadmium and mercury in carabid beetle predators and their prey. Invertebrates were sampled at one control and four heavy metal-polluted sites to reveal the impact of diet composition and seasonal variation in prey availability on metal burden in carabids and metal transfer pathways through forest ecosystems. This is the first report, to our knowledge, of carabid diet composition based on PCR analysis of gut contents at the forest community level, rather than in cultivated fields. Extensive screening using group- and species-specific primers revealed that carabids ate primarily earthworms and slugs, as well as smaller numbers of woodlice and springtails. Metal concentrations in carabids correlated with seasonal changes in diet. Mercury accumulated in beetle predators more than in their slug prey. As earthworms, slugs and carabid beetles are the major prey of many birds and mammals, prey-predator transfer and associated toxicity are major risks at mercury-contaminated sites. Carabids may be useful bioindicators for assessing the impact of pollutants on soil ecosystems, as long as species and seasonal factors are taken into account.

Cadmium in placenta- a valuable biomarker of exposure during pregnancy in biomedical research.

Rats were exposed to cadmium (Cd) during pregnancy orally (50 ppm Cd in drinking water from gestation day [GD] 1-20) or parenterally (5 mg/kg sc during GD 1-19 or on GD 16). Cadmium levels were measured in blood, placenta, and fetus at the end of exposure. Human studies comprised of two cohorts of healthy Croatian postpartum women in the continental versus coastal area; in these women, Cd levels were measured in maternal blood, placenta, and cord blood. Cadmium in rat placenta was 10-fold higher following parenteral versus oral exposure, with highest value after exposure in mid pregnancy. The values were 10-fold higher in placenta than blood, and the fetus had ≤10% of blood metal levels. Cadmium levels in human placenta noted increased values in smokers that were 10- to 20-fold higher than in maternal blood and in cord blood were very low (<0.1 ppb). In nonsmokers, different Cd levels in placenta and blood in coastal versus continental area were found due to dietary intake. Data showed that Cd in placenta is a reliable biomarker of exposure following oral and parenteral treatment during pregnancy in rats and in human biomonitoring that includes prenatal assessment. In addition, tobacco smoking is a prominent source of metal exposure. Cd levels in placenta are also a valuable biomarker of metal dietary exposure related to specific dietary habits and soil characteristics.

Selenium in brown bears (Ursus arctos) from Croatia: Relation to cadmium and mercury.

Muscle (n = 111), liver (n = 111), and kidney cortex (n = 101) samples from brown bears (Ursus arctos) were collected in the 2009 and 2010 hunting seasons in Croatia and analysed for selenium (Se), cadmium (Cd), and total mercury (Hg). The aim was to assess the levels of these elements according to age, sex, and season of collection, and to investigate possible Se/Cd and Se/Hg interactions. Median Se concentrations were 0.139 µg/g in muscle, 0.409 µg/g in liver and 1.75 µg/g wet mass in kidney cortex. Median Cd and Hg were 0.0078 and 0.0018 µg/g in muscle, 1.09 and 0.031 µg/g in liver, and 16.5 and 0.206 µg/g wet mass in the renal cortex, respectively. Se/Cd molar ratios were less than 1 in the kidney cortex, and close to or above 1 in liver and muscle, respectively. Toxic Cd and Hg correlated with Se in all of the studied tissues. Sex differences were found for all three elements (except Se in liver), with females having higher tissue concentration than males. Only Cd showed age-dependence. Bear samples collected in fall had higher Se in muscles, and Hg in muscles and liver compared to samples collected in spring. Element concentrations in brown bear tissues were within the range of previously reported studies. Bear meat is considered a rich source of Se, safe for consumption with regard to its Cd and Hg content. According to the molar ratio and correlation results, we assume that Se binding is not the primary detoxification pathway for Cd and Hg in brown bears.

The Impact of Long-Term Clinoptilolite Administration on the Concentration Profile of Metals in Rodent Organisms.

Heavy metals are dangerous systemic toxicants that can induce multiple organ damage, primarily by inducing oxidative stress and mitochondrial damage. Clinoptilolite is a highly porous natural mineral with a magnificent capacity to eliminate metals from living organisms, mainly by ion-exchange and adsorption, thus providing detoxifying, antioxidant and anti-inflammatory medicinal effects. The in vivo efficiency and safety of the oral administration of clinoptilolite in its activated forms, tribomechanically activated zeolite (TMAZ) and Panaceo-Micro-Activated (PMA) zeolite, as well as the impact on the metallic biodistribution, was examined in healthy female rats. Concentration profiles of Al, As, Cd, Co, Pb, Ni and Sr were measured in rat blood, serum, femur, liver, kidney, small and large intestine, and brain using inductively coupled plasma mass spectrometry (ICP-MS) after a 12-week administration period. Our results point to a beneficial effect of clinoptilolite materials on the concentration profile of metals in female rats supplemented with the corresponding natural clinoptilolite materials, TMAZ and PMA zeolite. The observed decrease of measured toxicants in the kidney, femur, and small and large intestine after three months of oral intake occurred concomitantly with their most likely transient release into the bloodstream (serum) indicative of a detoxification process.

Effects of Maternal Cigarette Smoking on Trace Element Levels and Steroidogenesis in the Maternal-Placental-Fetal Unit.

This study evaluates the interaction of toxic elements cadmium (Cd) and lead (Pb) due to exposure from cigarette smoking, essential elements, and steroidogenesis in the maternal-placental-fetal unit. In a cohort of 155 healthy, postpartum women with vaginal term deliveries in clinical hospitals in Zagreb, Croatia, samples of maternal blood/serum and urine, placental tissue, and umbilical cord blood/serum were collected at childbirth. The biomarkers determined were concentrations of Cd, Pb, iron (Fe), zinc (Zn), copper (Cu), and selenium (Se), and steroid hormones progesterone and estradiol in maternal and umbilical cord blood and the placenta. Three study groups were designated based on self-reported data on cigarette smoking habits and confirmed by urine cotinine levels: never smokers (n = 71), former smokers (n = 48), and active smokers (n = 36). Metal(loid)s, steroid hormones, urine cotinine, and creatinine levels were analyzed by ICP-MS, ELISA, GC-MS, and spectrophotometry. Cigarette smoking during pregnancy was associated with increased Cd levels in maternal, placental, and fetal compartments, Pb in the placenta, and with decreased Fe in the placenta. In active smokers, decreased progesterone and estradiol concentrations in cord blood serum were found, while sex steroid hormones did not change in either maternal serum or placenta. This study provides further evidence regarding toxic and essential metal(loid) interactions during prenatal life, and new data on sex steroid disruption in cord serum related to cigarette smoking. The results indicate that umbilical cord sex steroid levels may be a putative early marker of developmental origins of the future burden of disease related to harmful prenatal exposure to cigarette smoke.

The Effects of Chromium and Vanadium on Biomarkers of Carbohydrate and Lipid Metabolism in Workers Exposed to Coal Fly Ash.

Chromium (Cr) and vanadium (V) are micronutrients playing a role in carbohydrate and lipid metabolism but can be toxic at high concentrations, especially in specific forms. The study documents the effect of Cr and V concentrations on glucose and lipid metabolism in workers exposed to coal fly ash. We quantified selected metals (Cr, V) in the blood and serum of workers from a thermal power plant in Kosovo and compared them with the reference biological values. We determined fasting serum glucose and lipid profiles using a biochemical analyzer Synchron CX7 (Beckman Coulter). We quantified blood and serum Cr and V by inductively coupled plasma mass spectrometry. We also evaluated the association between carbohydrate and lipid metabolism biomarkers (glucose, cholesterol, and triglycerides) and co-exposure to coal fly ash. Power plant workers had significantly higher blood Cr and V levels (p < 0.0001) and significantly lower serum Cr and V levels (p < 0.0001) than the controls. We also found statistically significant (p < 0.0001) correlations between high blood Cr levels and low glucose/blood Cr ratios as well as between high serum Cr levels and low glucose/serum Cr ratios. Finally, in power plant workers, high blood V levels significantly correlated with low triglycerides/blood V and cholesterol/blood V ratios (p < 0.0001), while high serum V levels correlated with low cholesterol/serum V ratios (p = 0.005). Based on these findings, we concluded that the glucose/Cr, triglycerides/V and cholesterol/V ratios should be considered when evaluating carbohydrate and lipid metabolism disorders in occupationally-exposed workers.

Diurnal Salivary Cortisol in Relation to Body Composition and Heart Rate Variability in Young Adults.

Objective: Chronic stress has been related to impaired body composition (bone, muscle, adipose tissue), and disturbed autonomic nervous system (ANS), the latter regulated by hypothalamic-pituitary-adrenal (HPA) axis. Our objective was to investigate the relationship between salivary cortisol, body composition and heart rate variability (HRV-highly influenced by ANS), in a young student population. Methods: Body composition and HRV parameters were measured using two portable devices based on multi-frequency bioelectrical impedance and photoplethysmography. Body composition included measurement of bone, lean/muscle and adipose tissues. HRV included time domain and frequency domain indices. Salivary cortisol, immediately after awakening, 60 min post awakening and late night was collected and analyzed by ELISA. Results: Participants included n = 43 healthy university students (39 women, 4 men); 21.1 ± 1.3 years, BMI = 21.8 ± 3.4 kg/m2. Skeletal muscle was reduced in most of the participants, bone mass was at the lower normal range, while the fat tissue parameters were normal with only 7% participants being overweight. Cortisol and HRV parameters of sympathetic predominance (low frequency-LF and low frequency/high frequency ratio-LF/HF) were significantly associated with higher fat tissue parameters (fat mass-FM% and intramuscular adipose tissue-IMAT%) (p <0.001 and p = 0.035, respectively) and with lower skeletal muscle (p = 0.002) and bone mass (p = 0.025). Conclusions: The results point to a role of cortisol and ANS in the control of all three body composition compartments, suggesting that the stress can result in adverse effects on body composition, even in young, apparently healthy individuals.

The effects of maternal cigarette smoking on cadmium and lead levels, miRNA expression and biochemical parameters across the feto-placental unit.

Several miRNAs have been previously identified to be associated with cigarette smoke and/or the toxic metals cadmium (Cd) and lead (Pb). The aim of this study was to investigate the associations of maternal cigarette smoking with cadmium (Cd) and lead (Pb) levels, candidate miRNA expression and biochemical parameters across the feto-placental unit. miRNAs were isolated according to protocols provided by manufacturer from 72 healthy postpartum women using Qiagens' kits based on phenol/guanidine samples lysis and silica-membrane purification of total RNA. Candidate miRNAs (miR-1537, miR-190b, miR-16, miR-21, and miR-146a) were quantified by real-time PCR. Biochemical parameters were analyzed in plasma samples by standardized and harmonized enzymatic methods using appropriate calibrators, while CRP was determined by immunoturbidimetric method. Concentration of Cd and Pb in whole blood and placenta samples were measured by inductively coupled plasma mass spectroscopy. Cd levels in smokers were higher in all of the analyzed compartments of the feto-placental unit, Pb in maternal blood and placenta than non-smokers. Smokers also had a higher expression of miR-16 in maternal and miR-146a in cord plasma, and lower expression of miR-21 in the placenta in comparison to non-smokers. Urate concentrations in the maternal plasma of smokers were lower than this value in non-smokers. The study has demonstrated that maternal smoking was associated with toxic metals (Cd and Pb) levels, urate concentration and alteration of miRNA expression. Given that the effects of maternal smoking on miRNA expression are inadequate, all compartments of the feto-placental unit should be analyzed to obtain a complete picture. This paper is the first to report on the results of expression of cellular and circulating miRNAs simultaneously in maternal and fetal compartments and in the placenta.