Rachel Picada Bulcão

Biomarkers of occupational exposure to air pollution, inflammation and oxidative damage in taxi drivers

Exposure to environmental pollutants has been recognised as a risk factor for cardiovascular events. 1-hydroxypyrene (1-OHP) is a biomarker of exposure to polycyclic aromatic hydrocarbons (PAHs) from traffic-related air pollution. Experimental studies indicate that PAH exposure could be associated with inflammation and atherogenesis. Thus, the purpose of this study was to evaluate whether the biomarker of PAH exposure is associated with biomarkers of inflammation and oxidative stress and if these effects modulate the risk of developing cardiovascular diseases in workers exposed to air pollution. This study included 60 subjects, comprising 39 taxi drivers and 21 non-occupationally exposed persons. Environmental PM2.5 and benzo[a]pyrene (BaP) levels, in addition to biomarkers of exposure and oxidative damage, were determined. Inflammatory cytokines (IL-1β, IL-6, IL-10, TNF-α, IFN-γ and hs-CRP) and serum levels of oxidised LDL (ox-LDL), auto-antibodies (ox-LDL-Ab) and homocysteine (Hcy) were also evaluated. PM2.5 and BaP exhibited averages of 12.4±6.9 μg m(-3) and 1.0±0.6 ng m(-3), respectively. Urinary 1-OHP levels were increased in taxi drivers compared to the non-occupationally exposed subjects (p<0.05) and were positively correlated with pro-inflammatory cytokines and negatively correlated with antioxidants. Furthermore, taxi drivers had elevated pro-inflammatory cytokines, biomarkers of oxidative damage, and ox-LDL, ox-LDL-Ab and Hcy levels, although antioxidant enzymes were decreased compared to the non-occupationally exposed subjects (p<0.05). In summary, our findings indicate that taxi drivers showed major exposure to pollutants, such as PAHs, in relation to non-occupationally exposed subjects. This finding was associated with higher inflammatory biomarkers and Hcy, which represent important predictors for cardiovascular events. These data suggest a contribution of PAHs to cardiovascular diseases upon occupational exposure.

Effects of low-level exposure to xenobiotics present in paints on oxidative stress in workers

Paints are composed of an extensive variety of hazardous substances, such as organic solvents and heavy metals. Biomonitoring is an essential tool for assessing the risk to occupational health. Thus, this study analyzed the levels of biomarkers of exposure for toluene, xylene, styrene, ethylbenzene, and lead, as well as the oxidative stress biomarker alterations in painters of an industry. Lipid peroxidation biomarker (MDA), delta-aminolevulinate dehydratase (ALA-D), nonprotein thyol groups, superoxide dismutase and catalase (CAT) were analyzed in exposed and nonexposed subjects. We estimated which of the paint constituents have the greatest influence on the changes in the biomarkers of oxidative stress in this case of co-exposure. The results demonstrated that despite the fact that all the biomarkers of exposure were below the biological exposure limits, the MDA levels and antioxidant enzyme activities were increased, while nonprotein thyol groups and ALA-D levels were decreased in painters when compared with nonexposed subjects. After statistic test, toluene could be suggested as the principal factor responsible for increased lipid peroxidation and inhibition of ALA-D enzyme; however, further studies on the inhibition of ALA-D enzyme by toluene are necessary.

Evaluation of genotoxicity and oxidative damage in painters exposed to low levels of toluene

Toluene is an organic solvent used in numerous processes and products, including industrial paints. Toluene neurotoxicity and reproductive toxicity are well recognized; however, its genotoxicity is still under discussion, and toluene is not classified as a carcinogenic solvent. Using the comet assay and the micronucleus test for detection of possible genotoxic effects of toluene, we monitored industrial painters from Rio Grande do Sul, Brazil. The putative involvement of oxidative stress in genetic damage and the influences of age, smoking, alcohol consumption, and exposure time were also assessed. Although all biomarkers of toluene exposure were below the biological exposure limits, painters presented significantly higher DNA damage (comet assay) than the control group; however, in the micronucleus assay, no significant difference was observed. Painters also showed alterations in hepatic enzymes and albumin levels, as well as oxidative damage, suggesting the involvement of oxidative stress. According to multiple linear regression analysis, blood toluene levels may account for the increased DNA damage in painters. In summary, this study showed that low levels of toluene exposure can cause genetic damage, and this is related to oxidative stress, age, and time of exposure.

Acute and Subchronic Toxicity Evaluation of Poly(ɛ-Caprolactone) Lipid-Core Nanocapsules in Rats

Owing to concerns over the effects of the physicochemical properties of nanoparticles and their interaction with biological systems, further investigation is required. We investigated, for the first time, the toxicity of lipid-core nanocapsules (LNCs) containing a polymeric wall of poly(ε-caprolactone) and a coating of polysorbate 80 used as drug delivery devices (~245nm) in Wistar rats after single- and repeated-dose treatments. The suspensions were prepared by interfacial deposition of the polymer and were physicochemically characterized. Toxicological effects were determined after single doses of 18.03, 36.06, and 72.12 × 10(12) LNC/kg and repeated doses of 6.01, 12.02, and 18.03 × 10(12) LNC/kg for 28 days by ip administration. The results for both the treatments showed no mortality or permanent body weight changes during the experiments. A granulomatous foreign body reaction was observed in the liver and spleen of higher dose groups in acute and subchronic treatments. Most of the hepatotoxicity and nephrotoxicity markers were within the reference values and/or were similar to the control group. However, a slight alteration in the hematologic parameters was observed in both the studies. Thus, to verify a possible methodological influence, we performed an in vitro test to confirm such influence. These findings are in agreement with earlier reports regarding no appreciable toxicity of biodegradable polymeric nanoparticles, indicating that LNC might be a safe candidate for drug delivery system. Furthermore, the results presented in this study are important for health risk assessment and to implement strategies for testing biodegradable polymeric nanoparticles.

Genotoxicity and oxidative stress in gasoline station attendants

We evaluated genotoxic effects of exposure to low levels of benzene, a class I human carcinogen, among gasoline station attendants (GSA). Oxidative stress and the protective effects of antioxidants on DNA damage were also analyzed. Although exposures were below ACGIH (American Conference of Governmental Industrial Hygienists) limits, the GSA group presented higher DNA damage indices and micronucleus frequencies, increased oxidative protein damage, and decreased antioxidant capacity relative to the control group. Duration of benzene exposure was correlated with DNA and protein damage. The biomarkers evaluated in this work may provide early signals of damage in subjects occupationally exposed to benzene.

Blood thioredoxin reductase activity, oxidative stress and hematological parameters in painters and battery workers: relationship with lead and cadmium levels in blood

Oxidative stress has been shown to be involved in lead and cadmium toxicity. We recently showed that the activity of the antioxidant enzyme thioredoxin reductase (TrxR) is increased in the kidneys of lead‐exposed rats. The present study evaluated the blood cadmium and blood lead levels (BLLs) and their relationship with hematological and oxidative stress parameters, including blood TrxR activity in 50 painters, 23 battery workers and 36 control subjects. Erythrocyte δ‐aminolevulinate dehydratase (δ‐ALA‐D) activity and its reactivation index were measured as biomarkers of lead effects. BLLs increased in painters, but were even higher in the battery workers group. In turn, blood cadmium levels increased only in the painters group, whose levels were higher than the recommended limit. δ‐ALA‐D activity was inhibited only in battery workers, whereas the δ‐ALA‐D reactivation index increased in both exposed groups; both parameters were correlated to BLLs (r = −0.59 and 0.84, P < 0.05), whereas the reactivation index was also correlated to blood cadmium levels (r = 0.27, P < 0.05). The changes in oxidative stress and hematological parameters were distinctively associated with either BLLs or blood cadmium levels, except glutathione‐S‐transferase activity, which was correlated with both lead (r = 0.34) and cadmium (r = 0.47; P < 0.05). However, TrxR activity did not correlate with any of the metals evaluated. In conclusion, blood TrxR activity does not seem to be a good parameter to evaluate oxidative stress in lead‐ and cadmium‐exposed populations. However, lead‐associated changes in biochemical and hematological parameters at low BLLs underlie the necessity of re‐evaluating the recommended health‐based limits in occupational exposure to this metal. Copyright

The relationships between exogenous and endogenous antioxidants with the lipid profile and oxidative damage in hemodialysis patients

BackgroundWe sought to investigate the relationships among the plasma levels of carotenoids, tocopherols, endogenous antioxidants, oxidative damage and lipid profiles and their possible effects on the cardiovascular risk associated with hemodialysis (HD) patients.MethodsThe study groups were divided into HD and healthy subjects. Plasma carotenoid, tocopherol and malondialdehyde (MDA) levels, as well as erythrocyte reduced glutathione (GSH), were measured by HPLC. Blood antioxidant enzymes, kidney function biomarkers and the lipid profiles were analyzed by spectrophotometric methods.ResultsPlasma lycopene levels and blood glutathione peroxidase (GPx) activity were significantly decreased in HD patients compared with healthy subjects. Total cholesterol, low-density lipoprotein cholesterol (LDL-c), creatinine, urea, MDA, GSH, superoxide dismutase (SOD) and catalase (CAT) were significantly increased in HD (p < 0.05). Lycopene levels were correlated with MDA (r = -0.50; p < 0.01), LDL-c (r = -0.38; p = 0.01) levels, the LDL-c/HDL-c index (r = -0.33; p = 0.03) and GPx activity (r = 0.30; p = 0.03). Regression models showed that lycopene levels were correlated with LDL-c (β estimated = -31.59; p = 0.04), while gender was correlated with the TC/HDL-c index and triglycerides. Age did not present a correlation with the parameters evaluated. GPx activity was negatively correlated with MDA levels and with the LDL-c/HDL-c and CT/HDL-c indexes.ConclusionsLycopene may represent an additional factor that contributes to reduced lipid peroxidation and atherogenesis in hemodialysis patients.

The plasma retinol levels as pro-oxidant/oxidant agents in haemodialysis patients

BACKGROUND Oxidative stress is a process involved in haemodialysis-related pathologies such as cerebrovascular diseases. Retinol is the major circulating form of vitamin A and it is elevated in haemodialysis (HD) patients. It is known that these patients present anaemia that is not totally responsive to erythropoietin. The aim of this study was to evaluate the influence of plasma retinol levels on oxidative stress biomarkers, especially on delta-aminolevulinate dehydratase. METHODS Plasma retinol and malondialdehyde (MDA) levels were quantified by HPLC-UV/VIS; blood activities of catalase (CAT), superoxide dismutase (SOD) and delta-aminolevulinate dehydratase (ALA-D) were analysed by spectrophotometric methods, in HD patients (n = 29) and healthy subjects (n = 20). RESULTS The MDA and retinol levels, SOD and CAT activities were significantly increased in HD patients. ALA-D activity was significantly decreased. Retinol levels were correlated with MDA levels (r = 0.68), CAT (r = 0.39), SOD (r = 0.40) and ALA-D (r = -0.55). A partial correlation between retinol levels with ALA-D (r = 0.43), SOD (r = 0.30) and CAT (r = 0.36) activity was found, utilizing MDA levels as co-variable. CONCLUSION Higher retinol levels may be associated with the increase of SOD and CAT activities, but this increase was not sufficient to prevent the lipid peroxidation and ALA-D thiolic group oxidation. In this manner, our results could suggest that high retinol levels contribute as an additional factor to the oxidative tissue damage.

Quantification of reduced glutathione by HPLC‐UV in erythrocytes of hemodialysis patients

Reduced glutathione (GSH) is a well-known multifunctional antioxidant. Its depletion is linked to a number of pathologies, such as renal insufficiency. Feasible methodologies in clinical chemistry are vital. Therefore a methodology for GSH quantification was optimized and validated by HPLC-UV. Important aspects such as acid deproteinization and GSH stability were established. The erythrocytes were hemolyzed, deproteinized, derivatized with 5,5-dithio-bis (2-nitrobenzoic) acid and analyzed using HPLC, on an RP18 gradient elution, lambda=330 nm. The method was applied to hemodialysis patients (n=75) compared with healthy subjects (n=40). The assay was linear from 0.5 to 3.0 mm (r2>0.99). The intra- and inter-run reproducibilities were obtained with CV%<10%. The accuracy (bias %) ranged from 1.32 to -6.38%, and the recovery was >94%. The derivatized sample was stable for 60 days at -20 degrees C. The GSH levels in hemodialysis patients showed a significant increase compared with healthy subjects (p<0.05) and an inverse correlation with age (r=-0.286; p=0.013) was found. This method used UV detection, reduction of the phosphate concentration in the mobile phase and effective protein removal with trichloroacetic acid. The method proved to be reproducible, precise, accurate and stable. Thus, it can be suggested for routine laboratory tests for the verification of physiopathological conditions.

In vivo toxicological evaluation of polymeric nanocapsules after intradermal administration

Polymeric nanocarriers have shown great promise as delivery systems. An alternative strategy has been to explore new delivery routes, such as intradermal (i.d.), that can be used for vaccines and patch-based drug delivery. Despite their many advantages, there are few toxicity studies, especially in vivo. We report a safety assessment of biodegradable poly(ɛ-caprolactone) lipid-core nanocapsules (LNC) with a mean size of 245±10nm following single and repeated intradermal injections to Wistar rats. Suspensions were prepared by interfacial deposition of polymer. The animals (n=6/group) received a single-dose of saline solution (1.2ml/kg) or LNC (7.2×10(12)LNC/kg), or repeated-doses of two controls, saline solution or Tween 80 (0.9ml/kg), or three different concentrations of LNC (1.8, 3.6, and 5.4×10(12)LNC/kg) for 28 consecutive days. Clinical and physiological signs and mortality were observed. Samples of urine, blood, and tissue were used to perform toxicological evaluation. There were no clinical signs of toxicity or mortality, but there was a slight decrease in the relative body weights in the Tween 80-treated group (p<0.01) after repeated administration. No histopathological alterations were observed in tissues or significant changes in blood and urinary biomarkers for tissue damage. Mild alterations in white blood cells count with increases in granulocytes in the Tween-80 group (p<0.05) were found. Genotoxicity was evaluated through the comet assay, and no statistical difference was observed among the groups. Therefore, we conclude that, under the conditions of these experiments, biodegradable LNC did not present appreciable toxicity after 28 consecutive days of intradermal administration and is promising for its future application in vaccines and patch-based devices for enhancing the delivery of drugs.