摘要
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.
摘要
Purpose To investigate the expression of Dkk3 and Cyclin D1 protein in human hepatocellular carcinoma (HCC) and clinical significance.Methods Immunohistochemistry was used to detect Dkk-3 and Cyclin D1 protein expression level in 80 cases of hepatocellular carcinoma and corresponding para-cancer tissue.Results The expression of Dkk-3 in hepatocellular carcinoma was significantly lower than those in corresponding para-cancer tissue (P ＜ 0.05) and the expression of Cyclin D1 in hepatocellular carcinoma was significantly higher than those in CoTesponding para-cancer tissue (P ＜ 0.05).The up-regulation of Cyclin D1 and the down-regulation of Dkk-3 proteins were correlated with pathologic differentiation degree (P ＜0.05).There was a significant inverse correlation between Dkk3 and Cyclin D1 expression (P =0.044,rs =-0.226).Conclusion The abnormal expression of Dkk-3 and Cyclin D1 gene in human hepatocellular carcinoma suggest that Dkk-3 and Cyclin D1 gene may play an important role in the development and progression of the cancer.The combination deteetion of the two biomarkers may provide valuable data for diagnosis and prognosis estimation of HCC.