Inorganic Arsenic-mediated Up Regulation Of TUG1 Promoted Apoptosis In Human Bronchial Epithelial By Activating The P53 Signaling Pathway

ObjectiveArsenic and its compounds have been classified as human carcinogens by the International Agency for Research on Cancer and the U.S.Environmental Protection Agency.In this study,we analyzed the distribution pattern of arsenic in different urinary arsenic compounds by detecting the content and morphology of arsenic elements in the urine of arsenic-exposed people and analyzing the expression level of TUG1 gene in their bodies.To investigate the changes of TUG1 in human normal lung epithelial cell line 16 HBE after exposure to different concentrations of arsenic,and the effects of arsenic metabolites on TUG1 gene expression.To observe the apoptosis of 16 HBE cells induced by inorganic arsenic after TUG1 knockdown using Rfect;to further explore the mechanism of action in TUG1 on proliferation and apoptosis of 16 HBE cells,and to provide theoretical support for the molecular mechanism of arsenic carcinogenesis and about the function of TUG1.MethodsSeventy-four arsenic-exposed workers at a highly contaminated arsenic plant in Yunnan Province were selected as the arsenic-exposed group,and 25 people who had lived at the site for more than one year were selected as the control group at a location ≥10 km away from the plant,using a questionnaire in which participants provided an additional 10.0 m L of morning urine sample and 5.0 m L of venous blood.The levels of i As and its metabolites MMA and DMA in the urine samples of the study subjects were measured by hydride generation-ultralow temperature trapping-atomic absorption spectrophotometry.Peripheral blood RNA was extracted for reverse transcription,and TUG1 m RNA expression levels in the arsenic-exposed population were detected using q RT-PCR.The correlation between TUG1 gene expression and arsenic exposure was further analyzed,and the statistical characteristics of the relationship between arsenic exposure and TUG1 gene expression were derived.Specific silencing fragments were designed for TUG1 to transfect cells to reduce intracellular TUG1 expression in 16 HBE cells,and the viability and apoptosis of transfected cells were observed by Hoechst/PI two-color assay as well as MTS assay.Western Blotting assay was performed to determine the apoptotic gene expression after low expression of TUG1 and to explore its mechanism in the proliferation and apoptosis process in vivo.Results1.i As,MMA,and DMA concentrations as well as i As% and MMA% were significantly elevated in the arsenic-exposed group,while DMA%,PMI,and SMI were lower than those in the control group.2.TUG1 m RNA expression in peripheral blood lymphocytes was significantly elevated in the arsenic-exposed population compared with the control group.And TUG1 m RNA expression was found to be positively correlated with i As,MMA,DMA and t As levels in urine in the study population.3.Cell staining experiments revealed that TUG1 m RNA expression was significantly increased in arsenic-stained 16 HBE cells in a dose-dependent manner.And the metabolites of arsenic,MMA and DMA,were significantly lower than i As in inducing TUG1 m RNA expression.4.cell viability of 16 HBE cells was significantly reduced after TUG1 silencing,and the degree of reduction in cell viability was more significant after arsenic treatment than that of cells without arsenic treatment.apoptosis of 16 HBE cells was significantly increased after TUG1 silencing.5.knocking down the expression of TUG1,the protein expression of MDM2,Fas,BCL2,Caspase7 and p53 were found to be increased in 16 HBE cells by Western blotting assay,and p53 phosphorylation sites(Ser392)was also increased.Conclusion1.The concentrations of arsenic and its compounds were significantly higher in the occupational arsenic-exposed population,and the expression of TUG1 was higher in the arsenic-exposed population than in the control group.2.Arsenic exposure induced TUG1 expression in 16 HBE cells,while the arsenic metabolites MMA and DMA induced a significantly lower abnormal expression of TUG1 than the same concentration of inorganic arsenic.3.Silencing of TUG1 significantly inhibited cell viability and promoted apoptosis in16 HBE cells.Low expression of TUG1 enhanced phosphorylation of the p53(Ser392)site,the apoptotic pathway mainly by the p53 pathway,upregulated protein expression of FAS,MDM2,and BCL2,and activated Caspase7 to promote apoptosis.