Abstract:

ABSTRACT Objective: To investigate the characteristics of proximal tubule epithelial cells injury and the epigenetic mechanism of the injury induced by organic solvents. Methodology:: Human proximal tubule epithelial cells were treated with different concentrations of xylene. The NGAL in the culture medium was measured by ELISA, the apoptosis of tubular cells was detected by flow cytometry and the Caspase-3 activity was detected by kit. The Illumina Infinium HumanMethylation450 (450K) BeadChips were used to detect the methylation changes in HK-2 cells after xylene treatment. The data of Illumina chips were validated by pyro-sequencing method. Results: The secretion of NGAL was increased in dose-and time-dependent manner after xylene treatment. The NGAL was increased significantly (4.52±0.49 pg/ml vs 2.30± 0.11 pg/ml, p < 0.01) after 48 h treatment with 1.4 mmol/L xylene. At the same time, with increasing concentrations of xylene, the ratio of HK-2 cells apoptosis and Caspase-3 activity was significantly increased. Xylene treatment induced DNA methylation changes, indicating by 243 probes in HK-2 cells after xylene stimulation, with 109 probes displayed increased levels of methylation and 134 probes showed reduced levels of methylation. The methylation changes of 138 genes were indicated by these probes, including apoptosis related gene, e.g. Bax, FAF1, etc. The DNA methylation of FAF1 was decreased while the DNA methylation of BAX was increased. The pyrosequencing assays showed that Bax gene methylation level changes of xylene stimulation group was 7% compared to control group, and FAF1 gene was 6%, its methylation change trend was consistent with the results of the chip. Conclusion: Xylene stimulation can cause HK-2 cells injury and increased apoptosis. The changes of DNA methylation level of Bax, FAF1 might involve in xylene induced injury.