Effects Of Guizhimahuanggebantang On The TLR7 And RLH Pathway In Influenza Virus Infected Mouse Lungs In A Cold Environment

ObjectiveThis study was to investigate the effects of the traditional Chinese medicine Guizhimahuanggebantang on controlling influenza-A virus（IAV）infection and improving inflammation in mouse lungs and to explore its anti-viral mechanism.This experiment based on the environment-body-pathogen interactions of the three conditions,Preliminary elucidation of the mechanism of TCM treatment of influenza using different desease methods and provide the basis for the principle of treating"different diseases".Method60 female SPF C57BL/6 mice（6°C-wild type（6°C-WT））,60 female SPF TLR7^-/-mice(6°C-TLR7^-/-)and 60 female SPF MAVS^-/-mice(6°C-MAVS^-/-)were maintained incold environments.A control group of 60 female SPF C57BL/6 mice（20°C-WT）were maintained in a normal environment.Mice were divided randomly into six groups with ten animals in each group:normal control（“NC”）;IAV control（“VC”）;IAV+oseltamivir（“OSE”）;IAV+yinqiaosan（“YQS”）;IAV+guizhimahuanggebantang（“GZMHGBT”）;IAV+xinjiaxiangruyin（“XJXRY”）.On the eleventh day,the mice model of influenza was established by nasal drops.After 24 hours,the mice were treated with oseltamivir,yinqiaosan,guizhimahuanggebantang,and xinjiaxiangruyin and was administered（0.2 mL/day,i.g.）to each mouse 24 h after virus challenge for 5 days（day 12 to day 16）.Daily observation of the mouse’s general survival status,weight and so on.Six days after infection（day 17）,mice were anesthetized and sacrificed.The spleen and lung were harvested.The proportion of Th1/Th2 and Th17/Treg in splenic immune cells was detected by flow cytometry.The lung inflammation was observed in lung tissue.Real-time quantitative polymerase chain reaction was used to measure mRNA expression of TLR7,MyD88,RIG-I,MAVS,NF-κBp65 in the TLR7 and RLH signaling pathway and virus replication in lungs.Western blotting was used to measure expression levels of TLR7,MyD88,RIG-I,MAVS,and NF-κB p65 proteins.resultGuizhimahuanggebantang can reduce lung inflammation of C57BL/6 mice in normal and cold environment.Flow cytometry results showed that oseltamivir and three groups of Chinese herbs could down-regulate the Th1/Th2 and Th17/Treg ratios.RT-PCR and Western blot analysis showed that guizhimahuanggebantang could down-regulate the expression of TLR7,MyD88,RIG-I,MAVS,NF-κBp65 in TLR7and RLH signaling pathways in the lung of C57BL/6 mice.ConclusionGuizhimahuanggebantang had a good therapeutic effect on mice infected with IAV,especially in the cold environment.It could reduce lung inflammation in mice significantly and elicit an anti-influenza effect by down-regulating expression of the key factors in TLR7and RLH signaling pathway.