Immune Mechanism Of Adult Asthma Exacerbation By Exposure To Atmospheric PM_2.5 And Its Polycyclic Aromatic Hydrocarbons

Objective:To study the effects of exposure to atmospheric PM_2.5and polycyclic aromatic hydrocarbons（PAHs）on the number of CD4⁺CD25⁺Treg cells,STAT5/Foxp3 signaling pathway and inflammatory cytokines in peripheral blood of patients with asthma,which provide scientific basis for exploring the mechanism of atmospheric PM_2.5and PAHs aggravating asthma.Methods:Nineteen asthmatic patients were selected and investigated from April to May 2019 in a Three-A hospital in Taiyuan city.Measurement of lung function and collection of nasal mucus and blood in asthmatic patients was carried out at the following periods,May（spring）,July（summer）and November（autumn）in 2019,and January（winter）in 2020.The samples were collected twice each season,each time for 3 days,twice intervals for 1week.Three days before the collection of biological samples,PM_2.5was collected at a fixed point,PM_2.5concentration was recorded at 9 monitoring stations in Taiyuan urban area,and indoor PM_2.5monitoring was carried out for typical residential buildings.Based on the daily PM_2.5concentration at nine environmental monitoring stations,inverse distance weighted（IDW）was used to estimate the outdoor concentration of PM_2.5.Indoor PM_2.5concentrations in residences can be detected using aerosol monitors.Finally,combined with the results of time-activity model to estimate individual PM_2.5exposure concentration.High-performance liquid chromatography（HPLC）was applied to determinate the content of polycyclic aromatic hydrocarbons in PM_2.5,including naphthalene,acenaphthylene,acenaphthene,fluorene,phenanthrene,anthracene,fluoranthene,pyrene,benzo[a]anthracene,chrysene,benzo[b]fluoranthene,benzo[k]fluoranthene,benzo[a]pyrene,indeno[1,2,3,-,c,d]pyrene,dibenzo[a,h]anthracene,dibenzo[a,h]anthr-acene,which were considered as individual PAHs exposure.The basic information of asthma patients was obtained through questionnaire survey;the time-activity model recorded the symptoms,medication and daily activities of asthma patients;determination of blood routine of asthmatic patients;using flow cytometry to detect the percentage of Treg cells in peripheral blood,the level of IL-17,IL-23,IL-10,TGF-βin plasm and IL-10,TGF-βin nasal were detected by enzyme-linked immunosorbent assay（ELISA）;the gene expression level of STAT5,Foxp3 in the peripheral blood were detected by real-time fluorescent quantitative polymerase chain reaction（q RT-PCR）;Spearman correlation analysis was performed to observe the correlation between PM_2.5and PAHs concentration,and to analyze the correlation between PM_2.5and PAHs exposure and the conditions of asthma patients;a mixed-effect model was used to analyze the association between PM_2.5and its PAHs exposure concentrations and the expression of biological indicators.Results:1.The concentrations of ambient PM_2.5and 14 kinds of polycyclic aromatic hydrocarbons（acenaphthene and acenaphthene undetected）varied with seasons.PM_2.5concentration was higher in autumn and winter and the differences between winter and spring,summer were statistically significant.The difference between autumn and summer was statistically significant.The highest concentration of most PAHs was in autumn,and the lowest concentration was in summer.There were significant differences in the concentration of anthracene,fluoranthene,pyrene,benzo[a]anthracene,benzo[a]pyrene,dibenzo[a,h]anthracene,indeno[1,2,3-c,d]pyrene between summer and autumn.2.PM_2.5was significant positive correlation with naphthalene,anthracene,Indeno[1,2,3-c,d]pyrene,dibenzo[a,h]anthracene,PM_2.5had the strongest correlation with dibenzo[a,h]anthracene.3.The condition of asthma patients varied with seasons,and the symptom in autumn was more serious than in spring.4.The expression of IL-17 and IL-23 in the peripheral blood of asthmatic patients showed seasonal differences,and the concentrations in autumn and winter were significantly higher than those in spring and summer.The percentage of Treg cells in peripheral blood in autumn was significantly lower than that in spring.TGF-β,IL-10 in nasal mucus and TGF-βin peripheral blood in spring,summer and autumn were significantly lower than that in winter.The plasma IL-10 concentration in autumn was significantly higher than that in spring.The level of STAT5 gene in peripheral blood in spring was significantly higher than that in summer,autumn and winter.There was no statistical difference in the expression of Foxp3 gene in peripheral blood among the four seasons.5.PM_2.5and its PAHs were significant positive correlation with the condition of asthma patients,and the PAHs were negatively correlated with lung function.The correlation showed lag effect and cumulative lag effect,mainly concentrated in 0-3 days（lag0-lag3 and lag01-lag03）,but part of the correlation could still extend to 9 days（lag09）.6.Mixed effect model showed that with the increase of PM_2.5concentration,IL-17significantly increased,the percentage of CD4⁺CD25⁺Treg cells and the expression of Foxp3 gene significantly decreased in the peripheral blood of asthmatic patients,and the lag effect and cumulative lag effect could extend to 10 days.7.With the increasing concentrations of naphthalene,acenaphthylene,acenaphthene,fluorene,phenanthrene,anthracene,fluoranthene,pyrene,benzo[a]anthracene,chrysene,benzo[b]fluoranthene,benzo[k]fluoranthene,benzo[a]pyrene,indeno[1,2,3,-,c,d]pyrene,dibenzo[a,h]anthracene,dibenzo[a,h]anthraceneh in ambient PM_2.5,IL-17 and IL-23expression in peripheral blood were significantly increased.However,CD4⁺CD25⁺Treg cells,TGF-β,STAT5,Foxp3 expression in peripheral blood and IL-10,TGF-βin nasal mucus were clearly decreased.The lag effect and cumulative lag effect are mainly concentrated in 0-4 days（lag0-lag4 and lag01-lag04）,but some of them can still extend to the 10th day（lag10 and lag010）.Conclusion:1.Exposure to PM_2.5is positive correlation with the expression of IL-17 and aggravation of asthma;PAHs in PM_2.5can decrease lung function,promote the secretion of inflammatory cytokines IL-17 and IL-23,then aggravate asthma.2.PM_2.5can reduce the expression of CD4⁺CD25⁺Treg cells and its specific marker of Foxp3,then aggravate asthma;PAHs in PM_2.5can reduce the expression of CD4⁺CD25⁺Treg cells through STAT5/Foxp3 signaling pathway,and inhibit the secretion of IL-10 and TGF-β,then aggravate asthma.3.PM_2.5and its PAHs showed lag effects and cumulative lag effect on asthma exacerbation,mainly concentrated in 0-3 days,but some of them can still extend to the10th day.