Role Of C/EBP? Hypermethylation In Diesel Engine Exhaust Exposure-induced Lung Inflammation

Objective:Exposure to diesel engine exhaust(DEE)impairs lung function.But the underlying mechanisms are still not fully understood.The aim of this study was to investigate the effects of long-term DEE exposure on lung inflammation and the underlying mechanisms.Method:Sprague-Dawley male rats were exposed to DEE with 3 mg/m3 of diesel exhaust particles(DEP)for 12 weeks.Then urine,blood,bronchoalveolar lavage fluid(BALF),and lung tissue were collected for determination.In vivo,to evaluate the pathological changes of lung tissue and the metabolites of polycyclic aromatic hydrocarbons(PAHs)including 2-hydro-xyphenanthrene(2-OHPh)and 9-OHPh,and oxidation index: 8-hydroxy-2 ?-deoxyguanosine(8-OHd G)and malondialdehyde(MDA).Inflammatory indexes in serum and BALF: IL-6,IL-8,TNF – ? and lung injury index: CCAAT / enhancer binding protein alpha(C / EBP ?),pulmonary surfactant protein D(SP-D).At the same time,the methylation level of C/EBP ? in lung tissue was detected by Methylation-specific polymerase chain reaction(MSP).The level of C/EBP ? methylation and the expression of C/EBP ? and CC16 were detected after the stimulation of C22,DEP and DAC in vitro;the expression of CC16 was evaluated by C/EBP ? si-RNA transfection;the expression of inflammatory factors was detected after the intervention of srm1650 b and CC16.Result:The results showed that the metabolites of polycyclic aromatic hydrocarbons(PAHs)2-OHPh and 9-OHPh,and 8-OHd G,and MDA level were higher in urine of DEE-exposed rats than control group.The level of pro-inflammatory cytokines IL-8,IL-6,and TNF-? was significantly higher in serum(1.8,3.5,and nearly 1.0-fold increase,respectively),BALF(2.2,3.8,and 2.0-fold increase,respectively)and lung tissues(3.5,4.3,and 2.4-fold increase,respectively)of DEE-exposed rats than control group.While the level of CC16 and SP-D with anti-inflammatory property was obviously lower in serum(reduction of 29% and 38%,respectively),BALF(reduction of 50% and 46%,respectively)and lung tissues(reduction of 50% and 55%,respectively)of DEE-exposed rats than control group.Exposure to DEE also resulted in significant increases in total WBC,neutrophil,eosinophil,and lymphocyte number in BALF.Airway inflammation and remolding were apparent in DEE group.The methylation level of C/EBP? promoter was markedly increased(about 3.2-fold increase),and its m RNA and protein expression were significantly decreased(about 62% and 68% decrease,respectively)in the lungs of DEE-exposed rats compared with the group.Further,cell experiments were performed to investigate the relationship between C/EBP? and CC16,and CC16 function under DEP conditions.The results showed that DEP inhibited CC16 expression via methylation of C/EBP? promoter,and the increase of CC16 level significantly relieved the pro-inflammatory effects caused by DEP exposure.Conclusion:In conclusion,our data indicated that long-term exposure to DEE can cause lung inflammation,at least in part via methylation of C/EBP? promoter,and inhibition of CC16 expression.