Developmental and reproductive toxicity evaluation of toluene vapor in the rat II. Developmental toxicity

The developmental toxicity of toluene was evaluated via whole body inhalation exposure, in pregnant Sprague Dawley rats exposed to toluene (99.9% pure) from gestation day (GD) 6–15 inclusive, 6 h/day, at concentrations of 0, 250, 750, 1500 and 3000 ppm (0, 938, 2812, 5625 and 11250 mg/m³). Doses were selected from a preliminary study performed over a range of concentrations from 0 to 5000 ppm, in which maternal and fetal toxicity were observed at 2000 ppm and above. This study has been cited in various regulatory documents and is presented here to allow greater accessibility to results and conclusions.

Toluene induced clinical signs in pregnant dams (ataxia, hyper-responsivity, increased water intake, decreased food consumption) at 3000 ppm, ataxia and hyper-responsivity at 1500 ppm, and reduced maternal body weight gain at 1500 during the exposure period only and at 3000 ppm from initiation of exposure to GD20. At Caesarean section on GD20, no adverse effects on implantation, number and viability of fetuses, or fetal sex distribution were observed. Litter weight and mean fetal weight was reduced at 3000 ppm and mean fetal weight was reduced at 1500 ppm. Instances of reduced or unossified skeletal elements occurred at the same dose levels. Mean fetal weight was also reduced at 250 ppm but not at 750 ppm. Extensive statistical analysis of fetal body weight data support the conclusion that there is no toxicologically significant dose-related effect on fetal body weight at or below 750 ppm. Low incidences (≤2.5%) of various malformations occurred in the 250, 1500, and 3000 ppm groups; there was no increase in the incidence of specific or total malformations with increased exposure and thus these were not attributed to toluene.

In this Toluene study, the maternal toxicity NOAEL was 750 ppm with a defined maternal and developmental toxicity LOAEL of 1500 ppm.     

气相色谱法测定甲苯含量扩展不确定度的简化评定法

目的介绍气相色谱法测定工作场所空气中甲苯的结果扩展不确定度的简化评定方法。推广实验室测量结果不确定度的应用。方法确定和计算测定过程各不确定度分量,最后整体合成。结果气相色谱法测工作场所空气中甲苯的扩展不确定度为1．9μg／ml。结论该法评定过程合理,思路简捷,适用于实验室气相法测定扩展不确定度的日常应用。     

Gene profiles of a human bronchial epithelial cell line after in vitro exposure to respiratory (non-)sensitizing chemicals: Identification of discriminating genetic markers and pathway analysis

Respiratory sensitization is a concern for occupational and environmental health in consumer product development. Despite international regulatory requirements there is no established protocol for the identification of chemical respiratory sensitizers. New tests should be based on mechanistic understanding and should be preferentially restricted to in vitro assays. The major goal of this study was to investigate the alterations in gene expression of human bronchial epithelial (BEAS-2B) cells after exposure to respiratory sensitizers and respiratory non-sensitizing chemicals, and to identify genes that are able to discriminate between both groups of chemicals. BEAS-2B cells were exposed during 6, 10, and 24 h to the respiratory sensitizers ammonium hexachloroplatinate IV, hexamethylene diisocyanate, and trimellitic anhydride, the irritants acrolein and methyl salicylate, and the skin sensitizer 1-chloro-2,4-dinitrobenzene. Overall changes in gene expression were evaluated using Agilent Whole Human Genome 4× 44K oligonucleotide arrays. Fisher Linear Discriminant Analysis was used to obtain a ranking of genes that reflects their potential to discriminate between respiratory sensitizing and respiratory non-sensitizing chemicals. The 10 most discriminative genes were BC042064, A_24_P229834, DOCK11, THC2544911, DLGAP4, NINJ1, PFKM, FLJ10986, IL28RA, and CASP9. Based on the differentially expressed genes, pathway analysis was used to identify possible underlying mechanisms of respiratory sensitization. We demonstrated that in bronchial epithelial cells the canonical PTEN signaling pathway is probably the most specific pathway in the context of respiratory sensitization. Results are indicative that the BEAS-2B cell line can be used as an alternative cell model to screen chemical compounds for their respiratory sensitizing potential.     

Time course of the ethanol-like discriminative stimulus effects of abused inhalants in mice

Abused solvents have effects similar to those of abused depressant drugs. This experiment evaluated the time course of the discriminative stimulus effects of toluene and 1,1,1-trichloroethane (TRI). Mice were trained to discriminate between i.p. injections of ethanol (EtOH; 1.25 g/kg) and saline in a two-lever operant task in which responding was under the control of a fixed-ratio 20 schedule. After 20-min inhalation exposures to toluene (500-6000 ppm) or TRI (1000-12,000 ppm), stimulus generalization was examined at 0, 5, 10, 20, and 40 min post-exposure. Ethanol doses ≥ 0.25 g/kg produced increases in EtOH-lever responding with full substitution occurring immediately after testing for doses between 1.25 and 2.5 g/kg. Toluene and TRI produced increased EtOH-lever responding at 0-10 min post-exposure with some EtOH-lever responding occurring up to 20-min post-exposure. Response rates were not decreased for any concentration of toluene or TRI immediately following inhalant exposure but several concentrations elevated rates from 5 to 40 min post-exposure. These results confirm and extend previous studies and show these solvents produce similar effects in EtOH-lever responding but with potency differences. The time-dependent differences in EtOH-lever responding suggest that as solvents are cleared from the body, the EtOH-like subjective effects also fade.     

Does early life toluene exposure alter the expression of NMDA receptor subunits and signal transduction pathway in infant mouse hippocampus?

We aim to investigate the critical window of susceptibility to toluene exposure during brain development and the effects of fetal and neonatal toluene exposure on the expression of N-methyl-d-aspartate (NMDA) receptor subunits and related transduction pathway in infant mice hippocampus. Pregnant mice (GD 14), male offspring (postnatal day; PND 2) or PND 8 were exposed to either a filtered air control (0 ppm), or 5, or 50 ppm of toluene for 6 h per day for 5 consecutive days. On PND 21, the expression levels of NMDA receptor subunits, cyclic AMP responsive element binding protein (CREB)-1, calcium/calmodulin-dependent protein kinase (CaMK)-IV, and apoptotic related genes (Bax, Bcl) mRNAs in the hippocampus were estimated using quantitative real-time RT-PCR and immunohistochemical analyses. NR2B, CaMKIV and CREB1 mRNAs increased significantly in the hippocampus of mice exposed to 50 ppm toluene on PND 2–6. In contrast, almost all memory function-related gene mRNAs and proapoptotic and anti-apoptotic ratio increased significantly in mice exposed to 5 or 50 ppm toluene on PND 8–12. However, mice exposed to toluene on GD 14–18 showed no significant change. Increased active caspase-3 immunoreactive cells were found in hippocampal CA1 area of PND 21 male mice exposed to 5 ppm toluene during PND 8–12. Our results suggest that late postnatal period may be a vulnerable and critical period to toluene exposure. Then, we have also examined the effect of toluene exposure in brain development on learning ability in young adult mice and found that poor spatial learning performance in PND 49 male mice exposed to 5 ppm toluene during critical period. This is the first study to show that the early toluene exposure induces persistent of the alteration of memory function-related genes in infant mice and memory deficit in later life via modulating the synaptic morphology and function.     

Self-collected urine sampling to study the kinetics of urinary toluene (and <Emphasis Type="Italic">o</Emphasis>-cresol) and define the best sampling time for biomonitoring

Purpose  To study the excretion kinetics of urinary toluene, TOL-U, and o-cresol, o-C, following occupational exposure to toluene in order to define the best time for sample collection, to apply a non-invasive approach based on self-collected urine sampling. Methods  Five rotogravure printing workers exposed to uncontrolled levels of toluene collected spot urine samples over three consecutive working days and the following day of rest. In each sample TOL-U and o-C were measured and kinetics of excretion evaluated. Results  Toluene exposure ranged from 48.3 to 75.3 mg/m³; TOL-U and o-C ranged from 1.4 to 34.6 μg/L and from 0.013 to 1.012 mg/L. A time course trend was obtained: TOL-U and o-C increased during the shift and peaked at the end of exposure and up to 2 h later, respectively; afterwards they rapidly decreased following apparent first order kinetics. Considering TOL-U, the elimination half-life for the first fast phase was 79 (±35 standard error) min, and for the second slow phase was 1,320 (±1,162) min. For o-C the elimination half-life for the first fast phase was 231 (±48) min. Considering a toluene uptake of 86%, TOL-U and o-C excreted in urine were about 0.0067 and 0.18% of the up taken. Conclusion  Our results support the use of end shift TOL-U as a short term biomarker of occupational exposure to toluene and show the feasibility of self-collected urine sampling to investigate the elimination kinetics of industrial toxics in humans.     

孕期二苯基甲烷二异氰酸酯暴露对子代雄性小鼠性成熟后睾丸组织能量代谢功能的影响

目的探讨孕期二苯基甲烷二异氰酸酯(MDI)暴露对子代雄性小鼠性成熟后睾丸组织能量代谢功能的影响。方法将清洁级健康昆明种孕小鼠16只按体重随机分为高剂量(1/8LD50)、中剂量(1/16LD50)、低剂量(1/32LD50)MDI染毒组和溶剂对照组(玉米油),每组4只。于妊娠第14天开始,每日灌胃染毒,灌胃剂量为5ml/kg,持续至母鼠自然分娩。记录孕鼠染毒期间体重增量和分娩时间。待仔鼠8周龄(性成熟)时,分离双侧睾丸,称重,并计算睾丸的脏器系数。并检测睾丸组织中琥珀酸脱氢酶(SDH)、Na-K-ATP酶、Ca-Mg-ATP酶和总ATP酶的活力以及蛋白的含量。结果各组孕鼠分娩时间和体重增量间比较,差异均无统计学意义(P>0.05)。与高剂量MDI染毒组比较,中、低剂量MDI染毒组和溶剂对照组睾丸系数较高,差异均有统计学意义(P<0.01或P<0.05)。与溶剂对照组比较,高、中、低剂量MDI染毒组SDH、Na-K-ATPase、Ca-Mg-ATPase以及总ATPase酶活力均下降,差异均有统计学意义(P<0.01或P<0.05);且随着染毒剂量的增高,SDH、Na-K-ATPase、Ca-Mg-ATPase...     

Reduced exposure to organic solvents by use of water-based paint systems in car repair shops

Objective The objective of this study was to determine to what extent the substitution of solvent-based paint by water-based paint has reduced potential exposure to organic solvents for spray painters in car repair shops.Methods Full-shift personal air sampling (n=79) was carried out over 3 consecutive days in eight car repair shops. Blood samples on the Monday morning (n=26) and at the end of the shift on the Wednesday (n=26), were analysed for organic solvents by headspace techniques.Results Toluene was the organic solvent detected at the highest geometric mean concentration in air samples when solvent-based paint systems were used (0.8 ppm), whereas xylene was found at the highest level when water-based systems were used (0.25 ppm). Toluene, isopropanol, acetone and butyl acetate were detected at higher concentrations when solvent-based paint was used than when water-based paint was employed. The additive factor, based on Norwegian limit values, was three-times higher for the painters using solvent-based paint (0.15) than for those using water-based paint (0.05). On Wednesday after shift the geometric mean of toluene in blood was significantly higher for the painters using solvent-based paint (0.044 g/ml) than for the painters using water-based paint (0.007 g/ml). There was a significant correlation between toluene in personal air samples and toluene in blood samples taken at the end of the shift on the same day.Conclusions When solvent-based paint systems were used the additive factor for organic solvent exposure was three-times higher than when water-based systems were employed. The exposure levels of the organic solvents were well below the Norwegian limit values. The significant correlation between the toluene concentration in air and blood samples indicated that the uptake of organic solvents was correspondingly reduced. At the levels of organic solvents presently described the risk of acute and chronic health effects caused by organic solvents is low.     

Behavioral effects of toluene in rats selectively bred for infantile vocalization rate

Glue sniffing is epidemic among children living in poverty in Latin America. Previous research has shown that abused inhalants such as toluene share pharmacological properties with anxiolytic drugs, and that personality factors such as degree of anxiety have been proposed to modulate the effects of these drugs. To study this interaction in an animal model, rats selectively bred for high (High) or low (Low) rates of distress calls after maternal separation (ultrasonic vocalizations, USVs) were used to investigate toluene's acute and long-term effects on two measures of anxiety behavior. At ten days of age, neonatal subjects were administered toluene (1 g/kg i.p.) and USVs were recorded. The subjects were retested as juveniles on an elevated plus maze to examine sequela of earlier toluene exposure. Acute toluene administration reduced USVs relative to control groups in neonates of both lines, indicating anxiolysis. As expected, Lows had reduced USVs relative to Highs. At 28 days of age, Highs spent more time in the open arms of the elevated plus maze than Lows. However, prior neonatal toluene exposure blocked this reversal of behavioral phenotype. This suggests that early toluene exposure compromised a compensatory process occurring during this developmental period, which may have been maternally mediated. These results have implications for the effects of early drug exposure on plasticity in the developing nervous system.