ABSTRACT
Background Natural pyrethrins have long been widely used in the fields of environmental and household hygiene. Studies have reported that natural pyrethrins have potential liver toxicity, but their specific mechanisms are still unclear yet. Objective To explore the effect of natural pyrethrins on DNA damage in human liver cells. Methods This study used human liver cell QSG7701 as an in vitro testing model. After exposure to DMSO and a series of concentrations of natural pyrethrins (5, 10, 20, and 40 μg·mL−1) for 6 and 24 h, reactive oxygen species (ROS) was detected by fluorescence microscopy using a fluorescence probe, thiobarbituric acid reactive substance (TBARS) by colorimetric method using a microplate reader, DNA damage by comet assay through observing DNA fragment migration under microscope, and phospho H2AX (γH2AX) and 8-oxoguanine (8-oxoG) by immunofluorescence assay using a laser confocal microscope. Results As the exposure concentration of natural pyrethrins increased, the fluorescence intensity of ROS significantly increased in a concentration-dependent manner. The differences in ROS between the 10 μg·mL−1 and above groups and the control group were statistically significant (P<0.01), and the ROS levels in the 20 μg·mL−1 and 40 μg·mL−1 treatment groups were 2.17 and 3.05 times higher than that in the control group respectively. The TBARS level increased in a concentration-dependent manner in natural pyrethrins treated cells (P<0.01), and the levels in the 20 μg·mL−1 and 40 μg·mL−1 treatment groups were 2.46 and 3.01 times higher than that in the control group respectively. The results of comet assay showed trailing formation of cellular DNA in each dose group; as the exposure concentration of natural pyrethrins increased, indicators such as tail DNA content (TDNA%), tail length (TL), tail moment (TM), and Olive tail moment (OTM) increased in a concentration-dependent manner. Compared with the control group, the differences in the indicators between the 20 μg·mL−1 and above groups and the control group were statistically significant (P<0.01), especially in the 40 μg·mL−1 treatment groups, where TDNA%, TL, TM, and OTM were (46.92 ± 3.52) %, (64.67± 4.16) μm, 30.96 ± 2.94, and 22.64 ± 3.89, respectively. The cellular immunofluorescence results showed that natural pyrethrins induced the formation of γH2AX and 8-oxoG, the fluorescence intensities of γH2AX and 8-oxoG increased in a concentration-dependent manner, and the differences between the 10 μg·mL−1 and above groups and the control group were statistically significant (P<0.01). Conclusion Natural pyrethrins could induce DNA damage in human liver cells, and ROS-mediated oxidative stress may play an important role in its liver cell genotoxicity.
ABSTRACT
Objective To establish bovine corneal opacity and permeability （BCOP） test， and determine its predictive ability for the eye irritation evaluation of cosmetics. Methods A total of ten reference chemicals were selected to establish the BCOP test. Then eye irritation of 16 routinely collected cosmetics in our laboratory was predicted. In vitro scores were calculated by the change in the opacity and sodium fluorescein permeability after exposure to the testing cosmetics， and subsequently compared with the historical data by Draize test. Results Reference chemicals with known irritation classification were correctly classified by the BCOP test， which was consistent with the classification of UN globally harmonized system of classification and labeling of chemicals. Moreover， the specificity of the BCOP test for the classification of non-irritating cosmetics samples was 80.0% （8/10）， and the sensitivity for weak to mild irritating cosmetics samples was 83.3% （5/6）. The BCOP test demonstrated an overall classification consistency of 81.3% （13/16） with in vivo test. Conclusion BCOP test may be independently used to identify chemicals with potential eye irritation and serious eye damage， suggesting it is significant for in vitro integrated test strategy for predicting eye irritation due to cosmetics.