Carbon Content Of Macrophages And Its Application In Evaluating Human Exposure To Carbon Particles

[Objective]Currently,it is not possible to accurately evaluate individual particulate exposure level based on macroscopic population or particle metabolite measurements,but macrophage carbon content as an internal exposure biomarker can accurately evaluate individual particulate exposure level.In order to verify the effectiveness of macrophage carbon content as an internal dose biomarker,this study conducted research from two aspects of methodology and population epidemiology:firstly,to explore and establish a new preservation method of freshly induced sputum and a quantitative method of macrophage carbon content in induced sputum,and to standardize the preservation,treatment and quantitative methods.Then,through the population epidemiological study,to explore the levels and differences of macrophage carbon in different populations,and to explore the relationship chain of particulate matter external exposure-biomarkers-health effects,and finally verify the role of macrophage carbon as biomarkers.[Methods]1.Through the selection of water pump worker without special occupational harmful factors exposure as the general population to verify the methodology of carbon content in induced sputum and study the influencing factors and give the general population reference value.Subjects inhaled 4.5%hypertonic saline and coughed up sputum.About 20-30mL Saccomanno solution was added to the sputum and shook well and stored in a cool place away from light.Then standardized staining was carried out to make smear,and the pictures of macrophages were obtained by microscope and analyzed quantitatively by Image J software.The influence factors of CCAM are analyzed by generalized linear model.2.Workers in a black carbon plant and a diesel engine plant were selected to further verify the effectiveness of macrophage carbon as an internal exposure marker.Induced sputum was collected by ultrasonic atomization for carbon quantification of macrophages and classified count of inflammatory cells in sputum.Venous blood was drawn to measure the level of inflammatory factors,and micronucleus test was used to detect gene instability.Poisson regression analyzed the influencing factors of macrophage carbon and inflammation levels and explored the relationship between external exposure-macrophage carbon and biological effects.[Results]1.The sputum preserved with Saccomanno's fixative had fewer impurities intact cell morphology,clear nucleus and cytoplasm in the visual field,the nuclei are bluish-purple,and the cytoplasm is light pink or light blue and cells were easy to distinguish.The nucleus is round or kidney-shaped on one side,and the black carbon particles in the cytoplasm are clearly visible.Most of them are clustered and distributed in small clumps,which can be used to quantify the carbon content of airway macrophage successfully.2.The range of carbon content of macrophages in general population is as follows(M(P₂₅?P₇₅):0.83%?0.63%?1.34%?,for male is 0.79%?0.59%?1.27%?,and female is 0.95%?0.75%?1.34%?.3.The results of environmental exposure assessment showed that the PM_2.5concentration of carbon black?CB?exposed population and diesel exhaust particles?DEP?exposed population was 637.4 mg/m³ and 430.8 mg/m³,respectively,which was much higher than that of the control group?130.0 mg/m³?,about 5 times and 3 times higher than that of the control group.The carbon content levels of macrophages in CB exposed population and DEP exposed population(PCOC Q3,M(P₂₅?P₇₅))is 8.80%?3.91%?14.44%?and 4.31%?3.08%?7.61%?)were generally higher than those in their respective control groups?1.42%?1.19%?1.84%?and 1.72%?1.18%,2.21%??.Also,the proportion of macrophages?82.22%and 81.38%?was higher than that in their respective control groups.4.Poisson regression analysis showed that exposure to carbon black and diesel exhaust could increase the carbon content in macrophages:?5.20?95%CI:3.78-7.16?and 2.99?95%CI:2.24-3.99??.5.The results of Poisson regression showed that the CCAM level increased by one quartile?IQR,0.71?and the proportion of macrophages increased by 2.73%in the carbon black group and the carbon black control group?FR:1.02?95%CI:1.01-1.05??.6.The increase of macrophage carbon is related to the level of pulmonary inflammatory cells,the levels of TNF-?,IL-1?,IL-6 and micronucleus.[Conclusion]1.This research method is convenient,reliable,and efficient,and can be used to determine the carbon content of large-scale macrophages based on induced sputum.2.Elevated carbon content of airway macrophages was detected in those exposed to pure carbon particles of carbon black,and those exposed to carbon particles mixed with diesel exhaust,which can be used as an internal dose marker for exposure to carbon particles.3.Macrophages are one of the target cells of particulate matter.We found that there is a dose-response relationship between the carbon content of airway macrophages and the body's inflammatory level and gene instability level.Therefore,they can also be used as a dose marker for the biological effect of carbon particulate exposure.