Protective Effect Of Streptococcus Xiaochunlingii E24 On PM2.5 Induced Respiratory Tract Injury And Its Effect On Respiratory Microflora In Mice

ObjectiveIn this study,streptococcus xiaochunlingii E24 was used to screen potential probiotic strains from the microflora in oropharynx of healthy children.These strains are from the dominant flora of the respiratory tract and are sensitive to common antibiotics.They have the characteristics of in vitro bacteriostasis and remain stable after freezing and resuscitation.It was identified as a new species of streptococcus,named streptococcus xiaochunlingii sp.nov.This topic cell model is established and the animal model of probiotics to atmospheric pollutants PM2.5(Particulate Matter 2.5)as a result of exposure to human bronchial epithelial cells and BALB/c mice respiratory damage protection related research.In vivo and in vitro experiments were conducted to explore the inflammatory response caused by exposure to the atmospheric pollutant PM2.5 and the changes in respiratory microflora.Methods1.Collect PM2.5 freeze-dried powder,prepare PM2.5 suspension with different concentrations(12.5,25,50 and 100?g/ml),and expose human bronchial epithelial16-HBE cells for 6,12,24 and 48h,respectively.Use CCK-8 kit to detect cell activity and determine PM2.5 exposure concentration and time.The levels of inflammatory cytokines IL-1?,IL-6 and TNF-?in cell culture supernatant were detected by ELISA after exposure to 16-HBE cells for 24h with 50 and 100mg/ml of PM2.5.The expression levels of Nrf2,IKK-?,P65,Cyclin A2,Cyclin D and other proteins after exposure to PM2.5 in 16-HBE cells were detected by Western-blot.2.BALB/c mice were randomly divided into the control group and the PM2.5exposure group,and exposed to PM2.5 suspension with a concentration of 750g/m~3 for4 hours a day for 4 weeks.Mouse serum was collected for the detection of levels of inflammatory factors IL-1?,IL-6 and TNF-?.3.Streptococcus xiaochunlingii E24 interfered with PM2.5 exposure to 16-HBE cells,and cell culture supernant was collected for inflammatory factor level detection.The expression levels of Nrf2,IKK and P65 after exposure to PM2.5 in 16-HBE cells were detected by extracting cell protein Western-blot.4.BALB/c mice were randomly divided into control group,PM2.5 exposure group,streptococcus xiaochunlingii E24 protection group and streptococcus xiaochunlingii E24 treatment group.Mice in the experimental group were exposed to PM2.5 for 4 hours a day for 15 days.The streptococcus xiaochunlingii E24 protection group was given E24 suspension for 8 days before exposure,and the streptococcus xiaochunlingii E24 treatment group was given E24 suspension for 8 days after exposure to PM2.5.Control animals were raised under similar conditions and exposed to normal saline for 4 hours per day.Mouse pharynx swabs were collected for analysis of high throughput sequencing results.The trachea of mice was collected and the pathological changes were observed.The levels of inflammatory factors in the serum of mice were evaluated to explore the effect of E24 on PM2.5 induced respiratory system injury.Results1.CCK-8 test results showed that PM2.5 could induce a decline in the activity of16-HBE cells,which decreased with the increase of PM2.5 concentration and time.ELISA results showed that PM2.5 exposure to 16-HBE cells could lead to increased levels of inflammatory cytokines.Western-blot results showed that PM2.5can lead to increased expression of cell cycle and inflammation-related proteins.2.Compared with mice in the control group,mice in the PM2.5 exposure group showed increased expression of serum inflammatory factors,and increased expression of tracheal inflammation-related proteins,indicating an obvious inflammatory response.3.Streptococcus xiaochunlingii E24 acted on 16-HBE cells for 9 and 12h,they could reduce the cell apotosis induced by PM2.5,and E24 had a certain protective effect on the cell inflammatory injury induced by PM2.5.4.The streptococcus xiaochunlingii E24 treatment group and streptococcus xiaochunlingii E24 protection in mice serum inflammatory factor expression level in mice PM2.5 exposure group showed a trend of decrease in mice,and Western-blot detection results showed that the E24 treatment group and E24 protection in mice trachea inflammation related protein expression in mice,the mice induced by the use of probiotics can reduce PM2.5 airway inflammation injury,and respiratory tract flora has certain regulation in mice.Conclusion1.PM2.5 exposure induced changes in cell cycle expression of 16-HBE and inflammatory damage to the airway of BALB/c mice.2.The treatment of streptococcus xiaochunlingii E24 had a protective effect on PM2.5 induced 16-HBE and respiratory tract injury in mice.The exposure of PM2.5 in the air changed the abundance and diversity of bacteria in the respiratory tract of mice,and the treatment of E24 had a certain regulating effect on the imbalance of bacteria induced by PM2.5.