F. F. Liu

Hanging drop: an in vitro air toxic exposure model using human lung cells in 2D and 3D structures.

Using benzene as a candidate air toxicant and A549 cells as an in vitro cell model, we have developed and validated a hanging drop (HD) air exposure system that mimics an air liquid interface exposure to the lung for periods of 1h to over 20 days. Dose response curves were highly reproducible for 2D cultures but more variable for 3D cultures. By comparing the HD exposure method with other classically used air exposure systems, we found that the HD exposure method is more sensitive, more reliable and cheaper to run than medium diffusion methods and the CULTEX(®) system. The concentration causing 50% of reduction of cell viability (EC50) for benzene, toluene, p-xylene, m-xylene and o-xylene to A549 cells for 1h exposure in the HD system were similar to previous in vitro static air exposure. Not only cell viability could be assessed but also sub lethal biological endpoints such as DNA damage and interleukin expressions. An advantage of the HD exposure system is that bioavailability and cell concentrations can be derived from published physicochemical properties using a four compartment mass balance model. The modelled cellular effect concentrations EC50cell for 1h exposure were very similar for benzene, toluene and three xylenes and ranged from 5 to 15 mmol/kgdry weight, which corresponds to the intracellular concentration of narcotic chemicals in many aquatic species, confirming the high sensitivity of this exposure method.

Biomarkers for the evaluation of population health status 16 years after the intervention of arsenic-contaminated groundwater in Xinjiang, China

The arsenicosis endemic area in the region of Kuitun and Chepaizi, Dzungaria district, Xinjiang, People Republic of China was the first identified arsenic endemic area in China where arsenic concentration of up to 850 μg/L in the groundwater was reported. An intervention was put in place in 1985 by government to provide an alternative water source at a centralized community level. Sixteen years on since the intervention, we evaluated the health status of 178 villagers from endemic and 179 villagers from control sites. Biomarkers in their urine, included arsenic, porphyrins and malondialdehyde (MDA) were measured and the prevalence of skin lesions was also assessed. The average urinary arsenic (117 ± 8.3 μg/g of creatinine) from the endemic-villages was significantly higher (p<0.001) than that of the controls (73.6 ± 3.2 μg/g of creatinine) while no significant difference was found in urinary porphyrins and malondialdehyde concentrations in the overall studies subjects from these two areas. However when the urinary arsenic was higher than 150 μg/g of creatinine, MDA and porphyrins were higher in the endemic-villagers compared to the controls. Fifty-one out of 178 people from the arsenic endemic area showed skin lesions related to arsenicosis but these were absent among villagers from the control site. Of particular concern, skin lesions related to arsenicosis were observed in 4 out of 9 subjects 16 years of age or younger who were from different villages and born after the completion of water intervention. Although sporadic exposure and/or voluntary drinking contaminated water were thought to be a contributor of arsenicosis after the water intervention, the contribution from other dietary arsenic intakes remain unclear.

Arsenicosis status and urinary malondialdehyde (MDA) in people exposed to arsenic contaminated-coal in China

The current arsenic exposure condition, arsenicosis prevalence, urinary arsenic and MDA (malondialdehyde) concentrations in people were studied. The study area, a village in Xing Ren County in Guizhou Province, PR China, is a coal-borne arsenicosis endemic area that was identified several decades ago. The residents in Xing Ren have been using coal containing high arsenic levels all their life. Urinary arsenic levels of villagers were 192.2+/-22 microg/g creatinine (n=113) in the coal-borne endemic area (Xing Ren county) and were significantly higher than 63.6+/-5.9 microg/g creatinine (n=30) in a neighbouring control site (a village in Xing Yi county). The urinary MDA concentrations of villagers from the endemic area were also significantly higher compared to those of the control area. There was a strong correlation between age and urinary arsenic and MDA concentrations in the endemic area of Xing Ren; urinary arsenic and MDA levels decreased with age. Fifty out of 113 (44.3%) villagers in the endemic area had arsenicosis symptoms and the prevalence in villagers older than 40 y was 100% in male (92.2% overall). Urinary MDA concentration was significantly higher in people with arsenicosis symptoms in the endemic areas. Oxidative stress (urinary MDA concentration) was strongly related to arsenic exposure but not to the age and smoking habit. Higher urinary arsenic and MDA levels in younger villagers from the endemic area suggest that they are having a higher exposure to coal-borne emitted arsenic because they spend more time indoor. There is an urgent need to develop proper intervention methods in the Guizhou endemic areas in order to reduce the risk to the local communities who are still using arsenic contaminated-coal.

BTEX in vitro exposure tool using human lung cells: trips and gains.

Cytotoxicity of benzene, toluene, ethylbenzene and xylenes (BTEX) to human lung cells was explored using three different exposure methods: Method 1 - in normal 96-well plates using DMSO as a carrier vehicle, we exposed (a) human lung carcinoma A549 cells, (b) A549 cells over-expressed with cytochrome P450 2E1 cells, and (c) normal lung fibroblast LL-24 cells to benzene, toluene, ethylbenzene and xylene individually and in a mixture which models car exhaust gases for between 1-88 h. We found that the order of the BTEX potency is benzeneCYP2E1 over-expressed A549 cells. A significant difference was found between inter-assay responses for all 24h exposures (P<0.005) suggesting a poor assay repeatability. No sign of potency increase was found from 6 to 72 h exposures. Method 2 - Using sealed vials to expose A549 cells to benzene, toluene and ethylbenzene, we observed a twenty-fold increase in their cytotoxicity, but also with no time-course effect. Method 3 - Using air exposed hanging-drop cell culture, we were able to see both an increase of demonstration of toxicity and a time-course effect from 1 to 12h exposure. We conclude that exposing cells in sealed and unsealed media using DMSO as a carrier vehicle was not suitable for BTEX exposure studies. Hanging-drop air exposure has more potential. It should be noted that if there are any changes in their exposure matrixes, its exposure mass distribution in cells could differ.

Porphyrins and malondialdehyde as early biomarkers in individuals exposed to arsenic-contaminated coal in Guizhou, PR China: Isee-687

Introduction Coal is widely used in PR China. Unfortunately, coal from some areas in Guizhou Province contains elevated levels of arsenic. This has caused arsenicosis in individuals who use arsenic-contaminated coal for the purposes of cooking, heating and drying food. Aims Porphyrins are produced during heme biosynthetic pathways. Malondialdehyde (MDA) has been used to evaluate lipid peroxidation and DNA damage caused by exogenous free radicals or endogenous reactive oxygen species (ROS). This study aims to measure porphyrins and MDA and investigate their potential as early biomarkers of chronic arsenic poisoning. Methods We used HPLC to analyse porphyrin excretion profiles in urine samples from 113 people who live in Xing Ren of Guizhou Province, a coal-borne arsenicosis endemic area and from 30 control samples from Xing Yi where arsenicosis is not prevalent. We analysed urinary MDA by GC-MS in 119 and 30 urine samples obtained from the endemic and control sites respectively. Results Urinary porphyrins and MDA were higher in all arsenic exposed individuals than those of controls. Correlation between the urinary arsenic levels and porphyrin concentrations demonstrated the effect of arsenic on heme biosynthesis resulting in increased porphyrin excretion. Greater increases of urinary arsenic and porphyrin profiles in women, children and older age groups who spend much of their time indoors suggest that they might be at a higher risk. MDA concentrations were significantly higher in young age and female exposed groups than the controls. This confirmed the existence of lipid peroxidation and oxidative stress induced by chronic arsenic exposure. Conclusions Porphyrins and MDA are potential early biomarkers of arsenicosis. Elevated levels of porphyrins and MDA could lead to adverse health effects in chronically arsenic-exposed individuals associated with both non-cancer and cancer end-points.