Study On Acute Lung Injury Induced In Rats By PM_2.5 Dustfall Exposure Through DNA Methylation And Its Protection Effects By Vitamin B Complex

Objective:To investigate the role of DNA methylation in PM_2.5.5 dustfall induced acute lung injury in rats,clarify the mechanism of vitamin B complex(folic acid,B₆ and B₁₂)to reduce lung injury,and provide theoretical basis for the application of B vitamins to prevent the health effects of PM_2.5.Methods:1.PM_2.5 dustfall samples are mainly taken from the traffic zone and industrial zone in Mianyang,Sichuan,provided by Southwest University of Science and Technology.Fifty-six male SD rats aged 8-10 weeks were randomly divided into 7 groups,which were negative control group,different concentrations?3.5,7.0,14.0 mg/ml?of vitamin B complex protection+PM_2.5exposure groups,different doses?0.4,2.0,10.0 mg/ml?of PM_2.5?provided by Southwest University of Science and Technology,Mianyang,China?exposure groups,each group of 8.Rats of vitamin B complex protection+PM_2.5 exposure groups were administered by gavage with 0.5 ml?3.5,7.0,14.0 mg/ml?vitamin B complex dissovled in pure water once a day for 28 days,and other groups were administered by gavage with pure water in the same way.After 28 days,rats of all groups were administered by non-exposed intratracheal instillation for three days,once a day,0.5 ml each time.Rats of vitamin B complex protection+PM_2.5 exposure groups were administered by non-exposed intratracheal instillation with 2.0 mg/ml PM_2.5 dustfall suspension.Rats of PM_2.5 exposure groups were administered by non-exposed intratracheal instillation with 0.4,2.0,10.0 mg/ml PM_2.5 dustfall suspension.Rats of negative control group were administered by non-exposed intratracheal instillation with0.9%saline.We collected the abdominal aortic artery serum,lung tissue and bronchoalveolar lavage fluid of rats 24 hours after the last exposure.The histopathological alterations of lung tissues were observed by HE staining.The balf of rats were collected to measure the content of TP,LDH,AKP,ACP,MDA,SOD,CAT,IL-4 and IFN-?,which were associated with lung injury.Lung tissue and serum were collected to measure the contents and mRNA expression of IL-4 and IFN-?.Quantitative real-time PCR and western blot were used to detect the expression levels of DNMT 1,DNMT 3a and DNMT 3b in lung tissues.The 5-mC%levels were detected by quantitation kit to evaluate genomic DNA methylation.Results:Compared with the negative control group,low,medium and high doses of PM_2.5 exposure could cause different degrees of pathological damage of lung tissue in rats.The histopathological alterations of rats in the low,medium and high concentrations of vitamin B complex protection+PM_2.5exposure groups were reduced with different degrees compared with 2.0 mg/ml PM_2.5 exposure group.Compared with the negative control group,the contents of LDH,AKP,ACP,TP and MDA in the balf of rats in the low,medium and high doses of PM_2.5 exposure groups significantly increased?P<0.05?,while the activity of SOD and CAT significantly decreased?P<0.05?,and the contents of TP and MDA increased with the increase of PM_2.5 doses?P<0.05?,the activity of CAT decreased with the increase of PM_2.5 doses?P<0.05?.Compared with2.0 mg/ml PM_2.5 exposure group,the contents of LDH,TP and MDA in the balf of rats in the medium and high concentrations of vitamin B complex protection+PM_2.5 exposure groups decreased significantly?P<0.05?,SOD and CAT activity increased significantly?P<0.05?,the contents of AKP and ACP decreased in the high concentration of vitamin B complex protection+PM_2.5exposure group?P<0.05?.The content and mRNA expression of IL-4 in the balf,serum and lung tissue of rats in the medium and high doses of PM_2.5 exposure groups significantly increased?P<0.05?,while the content and mRNA expression of IFN-?significantly decreased?P<0.05?,IL-4/IFN-?increased?P<0.05?compared with the negative control group.Compared with 2.0 mg/ml PM_2.5 exposure group,The content of IL-4 in the balf and serum of rats in the medium and high concentrations of vitamin B complex protection+PM_2.5exposure groups significantly decreased?P<0.05?,while the content and mRNA expression in the lung tissue of IFN-?significantly increased?P<0.05?,IL-4/IFN-?decreased?P<0.05?.The mRNA expression of IL-4 in lung tissue of rats in the low,medium and high concentrations of vitamin B complex protection+PM_2.5 exposure groups significantly decreased?P<0.05?.The mRNA and protein expression of DNMT 1 in lung tissue of rats in the medium and high doses of PM_2.5 exposure groups significantly decreased?P<0.05?compared with the negative control group.The mRNA and protein expression of DNMT 3b significantly increased?P<0.05?,and the genomic methylation rate?5-mC%?decreases in the rats lung tissue of low,medium and high doses of PM_2.5 exposure groups and decreased with the increase of PM_2.5 doses?P<0.05?.Compared with 2.0 mg/ml PM_2.5 exposure group,The medium and high concentrations of vitamin B complex protation could activate DNMT 1?P<0.05?,increase genomic methylation?5-mC%??P<0.05?,significantly.But vitamin B complex had no significant effect on mRNA and protein expression of DNMT 3a and 3b?P>0.05?.Conclusion:1.PM_2.5 dustfall induces inflammation and oxidative stress in the lungs of rats to cause acute lung damage,and DNA methylation is involved in this process.2.Vitamin B complex has a protective effect on DNA methylation abnormality induced by PM_2.5 dustfall in rats.These results help us further explore the protection mechanisms of vitamin B in lung damages induced by PM_2.5.