Study On The Mechanism Of Xuanbai Chengqi Jiawei On H1N1 Infection Of A549 Cells Based On TLR3/NF-κB Signaling Pathwa

Purpose:To construct a model of H1N1 infection with A549 influenza virus,to test the efficacy of Xuanbai Chengqi Tangwei against influenza virus at different time points,to study the effect of Xuanbai Chengqi Tangwei on the TLR3/NF-κB pathway of the virus model and its related factors TNF-αand IL-6,and to elucidate the mechanism of Xuanbai Chengqi Tangwei in the treatment of A549 influenza virus model.Materials and methods:First,A549 cell culture and chicken embryo enlargement were carried out,suitable influenza virus strains were screened according to influenza hemagglutination experiments,and half of H1N1 tissue infection（TCID50）was detected.Half of the cytotoxic concentration（TC50）and half of the effective concentration of the drug（IC50）were calculated by cytotoxicity experiment and drug antiviral index experiment,and then the influenza virus treatment index（TI）of the drug was calculated.Indirect immunofluorescence experiments were used to detect the expression of influenza A virus nucleoprotein（NP）in the normal group,model group,traditional Chinese medicine group and western medicine group in the three time points of the A549 influenza virus model at 24h,48h and 72h,and the corresponding different groups at each time point.Western blot was used to detect the changes of Toll receptor 3（TLR3）and nuclear factor k B（NF-κB）in the normal group,model group,traditional Chinese medicine group,western medicine group and western medicine group in each time point of A549 influenza virus model at 24h,48h and 72h.ELISA was used to detect the changes of tumor necrosis geneα（TNF-α）and interleukin 6（IL-6）in the supernatant of different time points and different groups of the A549 influenza virus model.Results:1.H1N1 half of the tissue infection volume（TCID50）is 10^-3.2.Influenza virus treatment index（TI）of drugs:Xuanbai Chengqi Tangjia flavor TI=4.94,oseltamivir phosphate TI=6.19.3.Indirect immunofluorescence experimental results:in the three time points of 24h,48h and72h,compared with the normal group,the model group and the drug group had the expression of influenza virus nucleoprotein;NP was reduced in both the drug group and the model group.4.Western blot experimental results:in the three time points of 24h,48h and 72h,the expression of TLR3 and NF-κB in the model group was increased compared with the normal group（P<0.05）;Compared with the model group,TLR3 and NF-κB were reduced in the drug group（P<0.05）.There were statistically significant differences between the Chinese medicine group and the Western medicine group（P<0.05）.5.Enzyme-linked immunosorbent assay（ELISA）experimental results:In the three time points of 24h,48h and 72h,the expression of TNF-αand IL-6 in the A549 supernatant of the model group and the drug group increased compared with the normal group（P<0.05）;Compared with the model group,TNF-αand IL-6 were reduced in the drug group（P<0.05）.There were differences between the Chinese medicine group and the Western medicine group（P<0.05）.Conclusion:1.Successfully constructed a model of H1N1 infection with A549 influenza virus.2.Xuanbai Chengqi soup flavor treatment of influenza virus is effective.3.Xuanbai Chengqi soup flavor may be by inhibiting the TLR3/NF-κB pathway,thereby reducing the release of related factors TNF-αand IL-6 to fight influenza viruses.