Study On Mechanism Of Liver And Kidney Inflammation Caused By Fine Particulate Matter In Subway Platform In Mice

Objective:With the continuous increase and extension of global urban rail transit mileage,the subway has become the preferred public transportation mode for people’s daily travel,greatly relieving the pressure of urban surface transportation.Due to the high concentration of fine particles matter(PM_2.5)in subway stations,which are rich in toxic and harmful components such as heavy metals,their exposure can adversely affect human heart and lung,but the effect on extrapulmonary organs is unknown.In recent years,it has been found that adaptive immune cells and group 2 innate lymphoid cells（ILC2）are involved in mediating a variety of liver and kidney diseases,but the relevant mechanism of action is not completely clear.In innate immunity,interleukin（IL）-33activates ILC2 to drive the early immune response against environmental insults.Therefore,we used Rag1 knockout mice(Rag1^-/-)lacking T and B cells and set up IL-33treatment group to investigate the role of T cells,B cells and ILC2 in subway PM_2.5-induced liver and kidney inflammatory injury,which may provide a theoretical basis for the prevention and treatment of air pollutant-induced liver and kidney injury in subway stations in the future,and sound the alarm for subway passengers to strengthen their health precaution awareness.Methods:In this study,BALB/c mice were used in control,low,medium and high dose subway PM_2.5groups;wild type（WT）C57BL/6 mice and Rag1^-/-mice were used in control,subway PM_2.5-treated and IL-33-treated groups.The mice were intratracheally administrated subway PM_2.5suspensions of various concentrations or IL-33 solution（0.05 ml/time）through a polyethylene tube under anesthesia with 4%halothane 4 times at one-week interval.One day after the last intratracheal administration,the mice from all groups were killed by exsanguination under deep anesthesia by intraperitoneal（i.p.）injection of pentobarbital.Blood was collected from the heart,serum was separated,and liver and kidney tissues were collected.The level of aspartate aminotransferase（AST）,alanine aminotransferase（ALT）and blood urea nitrogen（BUN）in murine serum was determined using automatic biochemical analyzer.And the level of creatinine（CRE）was determined using Creatinine Assay Kit.Gross samples of liver and kidney tissues were observed and weighed for organ weight.HE staining was used to observe the structural changes and the degree of inflammatory cell infiltration in liver and kidney tissues.Transmission electron microscopy（TEM）was used to observe the ultrastructure of liver and kidney tissue.Immunohistochemistry assay（IHC）was performed to detect the expression of macrophages and neutrophils in liver and kidney tissues.RT-q PCR and Western Blot assays were performed to detect the m RNA and protein expressions of TLR2,TLR4,My D88,NF-κB,COX-2,TNF-α,IL-6 and MCP-1 in liver and kidney tissues of BALB/c mice,and the m RNA and protein expressions of COX-2,TNF-α,IL-6and MCP-1 were detected in liver and kidney tissues of WT C57BL/6 and Rag1^-/-mice.The experimental results adopt SPSS25.0 to conduct statistical analysis.Results:1.After treatment with different doses of subway PM_2.5,the body weight and organ coefficients of mice decreased significantly with increasing doses of dyeing（P<0.05）.2.The serum ALT and AST amounts in all treatment groups were remarkably elevated in a dose-dependent relationship,the serum CRE and BUN amounts were also enhanced in the high dose group（P<0.05）.3.HE staining results showed that there was no significant change in the control group,while the structure disorder and inflammatory cell infiltration of the liver and kidney tissues of mice in the PM_2.5treatment group were increased.4.TEM observed subway PM_2.5in liver and kidney tissues of mice in the high-dose group.5.The results of IHC experiment showed that macrophages and neutrophils infiltration in a dose-dependent relationship in the hepatic and renal areas.6.Subway PM_2.5enhanced hepatic m RNA levels of TLR2,TLR4,My D88,NF-κB,COX-2,TNF-α,IL-6 and MCP-1（P<0.01）.Correspondingly,the protein expression levels of TLR2,TLR4,NF-κB,TNF-αand IL-6 in liver tissue of mice in all the three exposure groups were elevated in a dose-dependent relationship（P<0.05）.7.With the increase of exposure dose,the m RNA and protein expression levels of all the above inflammatory factors in kidney tissue of mice in each treatment group were increased in turn.8.The liver coefficients of Rag1^-/-mice were significantly increased after treatment with subway PM_2.5and IL-33 compared to WT mice（P<0.01）,and the kidney coefficients of mice were not significantly changed compared to their control group.9.Compared with its control group,the serum ALT amounts in Rag1^-/-mice were significantly increased（P<0.05）,but the AST,CRE and BUN amounts were not significantly changed.10.The results of HE staining and IHC experiments showed that the degree of macrophage and neutrophil infiltration in the liver and kidney tissues of Rag1^-/-mice was significantly reduced.11.Compared with WT mice,the m RNA and protein expression levels of COX-2,TNF-α,IL-6 and MCP-1 were significantly reduced in the liver tissues of Rag1^-/-mice（P<0.05）,and the m RNA and protein expression levels of TNF-α,IL-6 and MCP-1 were significantly reduced in the kidney tissues of Rag1^-/-mice（P<0.05）.Conclusion:1.Subway PM_2.5could enter liver and kidney after tracheal infusion,leading to inflammatory cell infiltration in liver and kidney tissues and triggering subacute liver and kidney inflammation in mice.2.The TLR/My D88/NF-κB signaling pathway may mediate subway PM_2.5-induced inflammatory injury in mice liver and kidney.3.Subway PM_2.5-induced inflammatory in mice liver and kidney was mainly dependent on T and B cells,while the role of ILC2 was not obvious in this process.