Caracterização do material particulado e avaliação do risco ocupacional e mecanismos moleculares associados à queima artesanal da castanha de caju

Brazil is among the largest cashew nut producers of the world. However, the roasting process is still carried out artisanally, especially in the Brazilian semiarid region. In face of this occupational problem, the aim of this study was to perform a physical-chemical characterization of the particulate matter (PM) emitted by the roasting of cashew nuts, as well as to determine the occupational risk and molecular mechanisms associated. The most evident PM characteristics were the prevalence of fine particles, typical biomass burning morphologies such as tar ball and the presence of the elements K, Cl, S, Ca and Fe. In addition, atmospheric modeling analyses suggest that these particles can reach neighboring regions of the emission source. Polycyclic aromatic hydrocarbons (PAHs) with carcinogenic potential, such as benzo[a]pyrene, dibenz[a,h]anthracene, benzo[a]anthracene, benzo[b]fluoranthene, chrysene, benzo[k]fluoranthene, indeno[1,2,3-c,d]pyrene and benzo[j]fluoranthene were the most abundant PAHs found in the two air monitoring campaigns. Among the identified oxy-PAH the benzanthrone (7H-benz[d,e]anthracen-7-one) had the highest concentration and the evaluation of lifetime cancer risk showed an increase of 12 to 37 cases of cancer for every 10,000 exposed people. Chemical analysis of roasted cashew nuts identified the PAHs: phenanthrene, benzo[g,h,i]perylene, pyrene and benzo[a]pyrene, besides the 3-pentadecilfenol allergen (urushiol analogue) as prevalent. Occupational exposure to PAHs was confirmed by the increase of urinary 1-hydroxypyrene levels and genotoxic effects were evidenced by the increase on micronuclei and nuclear bud frequency in exfoliated buccal mucosa cells among the exposed workers. Other biomarkers of effects such as karyorrhexis, pyknotic, karyolytic, condensed chromatin and binucleated cells also have their frequencies increased when compared to an unexposed control group. The investigation of the molecular mechanisms associated with the PM organic extract showed cytotoxicity in human lung cell lines (A549) at concentrations ≥ 4 nM BaPeq. Using non-cytotoxic doses the extract was able to activate proteins involved in the DNA damage response pathway (Chk1 and p53). Moreover, the specific contribution of the four most representative PAHs in the cashew nut roasting sample showed that benzo[a]pyrene was the most efficient to activate Chk1 and p53. Finally, the organic extract was able to increase persistently the mRNA expression involved in the PAHs metabolism (CYP1A1 and CYP1B1), inflammatory response (IL-8 and TNF-α) and cell cycle arrest (CDKN1A) for DNA repair (DDB2). The high PM concentrations and its biological effects associated warn of the serious harmful effects of artisanal cashew nut roasting and urgent actions should be taken to the sustainable development of this activity.

Caracterização do material particulado e avaliação do risco ocupacional e mecanismos moleculares associados à queima artesanal da castanha de caju

Brazil is among the largest cashew nut producers of the world. However, the roasting process is still carried out artisanally, especially in the Brazilian semiarid region. In face of this occupational problem, the aim of this study was to perform a physical-chemical characterization of the particulate matter (PM) emitted by the roasting of cashew nuts, as well as to determine the occupational risk and molecular mechanisms associated. The most evident PM characteristics were the prevalence of fine particles, typical biomass burning morphologies such as tar ball and the presence of the elements K, Cl, S, Ca and Fe. In addition, atmospheric modeling analyses suggest that these particles can reach neighboring regions of the emission source. Polycyclic aromatic hydrocarbons (PAHs) with carcinogenic potential, such as benzo[a]pyrene, dibenz[a,h]anthracene, benzo[a]anthracene, benzo[b]fluoranthene, chrysene, benzo[k]fluoranthene, indeno[1,2,3-c,d]pyrene and benzo[j]fluoranthene were the most abundant PAHs found in the two air monitoring campaigns. Among the identified oxy-PAH the benzanthrone (7H-benz[d,e]anthracen-7-one) had the highest concentration and the evaluation of lifetime cancer risk showed an increase of 12 to 37 cases of cancer for every 10,000 exposed people. Chemical analysis of roasted cashew nuts identified the PAHs: phenanthrene, benzo[g,h,i]perylene, pyrene and benzo[a]pyrene, besides the 3-pentadecilfenol allergen (urushiol analogue) as prevalent. Occupational exposure to PAHs was confirmed by the increase of urinary 1-hydroxypyrene levels and genotoxic effects were evidenced by the increase on micronuclei and nuclear bud frequency in exfoliated buccal mucosa cells among the exposed workers. Other biomarkers of effects such as karyorrhexis, pyknotic, karyolytic, condensed chromatin and binucleated cells also have their frequencies increased when compared to an unexposed control group. The investigation of the molecular mechanisms associated with the PM organic extract showed cytotoxicity in human lung cell lines (A549) at concentrations ≥ 4 nM BaPeq. Using non-cytotoxic doses the extract was able to activate proteins involved in the DNA damage response pathway (Chk1 and p53). Moreover, the specific contribution of the four most representative PAHs in the cashew nut roasting sample showed that benzo[a]pyrene was the most efficient to activate Chk1 and p53. Finally, the organic extract was able to increase persistently the mRNA expression involved in the PAHs metabolism (CYP1A1 and CYP1B1), inflammatory response (IL-8 and TNF-α) and cell cycle arrest (CDKN1A) for DNA repair (DDB2). The high PM concentrations and its biological effects associated warn of the serious harmful effects of artisanal cashew nut roasting and urgent actions should be taken to the sustainable development of this activity.

Mechanisms of Cdc42 Polarization in Yeast

Polarization is important for the function and morphology of many different cell types. The keys regulators of polarity in eukaryotes are the Rho-family GTPases. In the budding yeast Saccharomyces cerevisiae, which must polarize in order to bud and to mate, the master regulator is the highly conserved Rho GTPase, Cdc42. During polarity establishment, active Cdc42 accumulates at a site on the plasma membrane characterizing the “front” of the cell where the bud will emerge. The orientation of polarization is guided by upstream cues that dictate the site of Cdc42 clustering. However, in the absence of upstream cues, yeast can still polarize in a random direction during symmetry breaking. Symmetry breaking suggests cells possess an autocatalytic polarization mechanism that can amplify stochastic fluctuations of polarity proteins through a positive feedback mechanism.

Two different positive feedback mechanisms have been proposed to polarize Cdc42 in budding yeast. One model posits that Cdc42 activation must be localized to a site at the plasma membrane. Another model posits that Cdc42 delivery must be localized to a particular site at the plasma membrane. Although both mechanisms could work in parallel to polarize Cdc42, it is unclear which mechanism is critical to polarity establishment. We directly tested the predictions of the two positive feedback models using genetics and live microscopy. We found that localized Cdc42 activation is necessary for polarity establishment.

While this explains how active Cdc42 localizes to a particular site at the plasma membrane, it does not address how Cdc42 concentrates at that site. Several different mechanisms have been proposed to concentrate Cdc42. The GDI can extract Cdc42 from membranes and selective mobilize GDP-Cdc42 in the cytoplasm. It was proposed that selectively mobilizing GDP-Cdc42 in combination with local activation could locally concentrate total Cdc42 at the polarity site. Although the GDI is important for rapid Cdc42 accumulation at the polarity site, it is not essential to Cdc42 concentration. It was proposed that delivery of Cdc42 by actin-mediated vesicle can act as a backup pathway to concentrate Cdc42. However, we found no evidence for an actin-dependent concentrating pathway. Live microscopy experiments reveal that prenylated proteins are not restricted to membranes, and can enter the cytoplasm. We found that the GDI-independent concentrating pathway still requires Cdc42 to exchange between the plasma membrane and the cytoplasm, which is supported by computational modeling. In the absence of the GDI, we found that Cdc42 GAP became essential for polarization. We propose that the GAP limits GTP-Cdc42 leak into the cytoplasm, which would be prohibitive to Cdc42 polarization.

Body measurements and blood plasma lipid, PCB and OH-PCB content of polar bear (U. maritimus) mothers and cubs, Svalbard

The aim of this study was to examine the plasma concentrations and prevalence of polychlorinated biphenyls (PCBs) and hydroxylated PCB-metabolites (OH-PCBs) in polar bear (Ursus maritimus) mothers (n = 26) and their 4 months old cubs-of-the-year (n = 38) from Svalbard to gain insight into the mother-cub transfer, biotransformation and to evaluate the health risk associated with the exposure to these contaminants. As samplings were performed in 1997/1998 and 2008, we further investigated the differences in levels and pattern of PCBs between the two sampling years. The plasma concentrations of Sum(21)PCBs (1997/1998: 5710 ± 3090 ng/g lipid weight [lw], 2008: 2560±1500 ng/g lw) and Sum(6)OH-PCBs (1997/1998: 228 ± 60 ng/g wet weight [ww], 2008: 80 ± 38 ng/g ww) in mothers were significantly lower in 2008 compared to in 1997/1998. In cubs, the plasma concentrations of Sum(21)PCBs (1997/1998: 14680 ± 5350 ng/g lw, 2008: 6070 ± 2590 ng/g lw) and Sum(6)OH-PCBs (1997/1998: 98 ± 23 ng/g ww, 2008: 49 ± 21 ng/g ww) were also significantly lower in 2008 than in 1997/1998. Sum(21)PCBs in cubs was 2.7 ± 0.7 times higher than in their mothers. This is due to a significant maternal transfer of these contaminants. In contrast, Sum(6)OH-PCBs in cubs were approximately 0.53 ± 0.16 times the concentration in their mothers. This indicates a lower maternal transfer of OH-PCBs compared to PCBs. The majority of the metabolite/precursor-ratios were lower in cubs compared to mothers. This may indicate that cubs have a lower endogenous capacity to biotransform PCBs to OH-PCBs than polar bear mothers. Exposure to PCBs and OH-PCBs is a potential health risk for polar bears, and the levels of PCBs and OH-PCBs in cubs from 2008 were still above levels associated with health effects in humans and wildlife.