Effect Of T?R? DNA Methylation In Pulmonary Fibrosis Induced By Beryllium Sulfate

Objective To investigate the changes of fibrosis-related factors collagen I,collagen III and?-SMA mRNA and protein in human lung adenocarcinoma A549 cells stimulated by beryllium sulfate?BeSO₄?,transforming growth factor-?type II receptor?T?R II?mRNA.and protein changes,and changes in DNA methylation.To explore the role of T?R II DNA methylation in BeSO₄-induced pulmonary fibrosis.Treatment of A549 cells with the methyltransferase enzyme inhibitor 5-azacytidine?AZC?further verified T?RII DNA methylation changes.Histone acetylation was observed in A549 cells using the histone acetylase inhibitor trichostatin A?TSA?.At the same time,the expression of fiber-related factors and T?R II was observed under the action of these two inhibitors.Methods An experimental model of cultured A549 cells was established and treated with BeSO₄ at a final concentration of 1.0,10.0 and 100.0?mol/L?solvent as complete medium,set to low,medium and high dose groups?,and AZC intervention group?final?The concentration of 10.0?mol/L barium sulfate and 10.0?mol/L AZC mixture treatment group and TSA intervention group?final concentration of 10.0?mol/L barium sulfate and 0.5?mol/L TSA mixture treatment group?.Cells were harvested after 24,48 and 72 h of treatment.Detection of mRNAs of collagen I?COL-I?,collagen III?COL-III?,?-smooth muscle actin??-SMA?and T?R II by fluorescence quantitative polymerase chain reaction?Quantitative Real-time PCR?Expression level;relative expression levels of COL-I,COL-III,?-SMA and T?R II proteins were detected by Western blot;T?RII DNA methylation level was detected by nested falling methylation-specific PCR.Results1.The mRNA and protein relative expression levels of collagen I,collagen III and?-SMA inA549 cells treated with different doses of BeSO4 increased with the increase of dose ?P<0.05?;low,medium,The mRNA and protein relative expression levels of collagen I,collagen III and?-SMA increased with the increase of exposure time in high dose group ?P<0.05?;after AZC and TSA intervention in A549 cells,24 At 48 and 72 h,the relative expression levels of the three indicators in the BeSO₄+AZC group and the BeSO₄+TSA group were lower than those in the middle dose group?P<0.05?.2.At 24,48 and 72 h,the relative expression levels of T?RII mRNA and protein in A549 cells in low,medium and high dose groups were lower than those in the control group ?P<0.05?.After AZC and TSA intervention in A549 cells,At 48 and 72 h,the relative expression levels of T?RII protein in BeSO4+AZC group and BeSO4+TSA group were higher than those in middle dose group?P<0.05?.3.Compared with the control group,T?RII DNA methylation level was higher in the 72h low dose group,48h and 72h medium and high dose groups than in the control group ?P<0.05?;low,medium and high dose groups T?RII DNA methylation level The dose increased with the increase of dose?P<0.05?.Under the action of AZC and TSA as the intervention factor of BeSO₄,the T?RII DNA methylation level was lower in BeSO₄+AZC group and BeSO₄+TSA group at three time points.Medium and high doses ?P<0.05?.Conclusion Under the stimulation of BeSO₄,fibrosis of A549 cells and increased methylation of T?RII DNA in cells may be an important epigenetic mechanism in the development and progression of A549 cell fibrosis induced by BeSO₄.