| Objective:Trough studying the effect of Maxingshigan Decoction on model mice's lung index,pulmonary pathological changes,ultrastructure,gene and protein expression of TLR7/8 in lung,to investigate the Maxinshigan decoction's effect on lung inflammation of influenza A virus infected model mice,to elucidate its part mechanism relate Toll like receptor signal pathway,and finally to construct the theoretical basis for further clarity of its clinical effect of anti-influenza virus.Methods:Nasal inoculation with A-type influenza virus(mice lungs adapted strain A/PR/8/34)was adopted to establish animal model of infection.The experiment set up normal control group,model control group,antiviral granule group,oseltamivir group and Maxinshigan Decoction group.After giving the corresponding drug or saline by gavage for seven days,lung index of mice were observed and compared.The pulmonary pathological changes in mice were observed with HE staining under optical microscope,ultrastructure of lung of mice were observed under transmission electron microscopy,Western Blotting and RT-qPCR assay were applied to detect the expression level of gene and protein of TLR7/8 in mice lungs.Results:1.Effect of Maxinshigan Decoction on lung index of mice infected with A-type influenza virus led to lung index increased in model group,and there was a significant difference compared with that of the normal control group(P<0.01).Lung index decreased in antiviral granule group,oseltamivir group and Maxinshigan Decoction group,there were difference(antiviral granule group and Maxingshigan Decoction group,P<0.05)and significant variation(oseltamivir group,P<0.01)compared with that of the model group.2.Effect of Maxinshigan Decoction in the 1112 lung pathological changes of mice infected with influenza virusThe appearance of mice lungs in normal control group showed pink color,smooth surface,soft texture and no lesion;the result of pathological sections showed that mice alveolus in normal control group were in normal size,the interval was not broadened,alveolus and pulmonary interstitial were absent of inflammatory cell infiltration.The appearance of mice lungs in model group showed multiple lobi pulmonis infection,several lesion areas of consolidation,which were dark red or brown;the result of pathological sections showed that mice alveolus in model group had tissue and structure damage,connective tissue proliferation,and alveolus with a mass of inflammatory cells infiltration.After drug intervention in each group,observed mice lungs directly or with optical microscope,the result showed that the lesion extent and pathological changes of their lungs were relieved and improved,alveolus morphology was mostly intact,the number of inflammatory cells in alveolus and pulmonary interstitial were reduced in different degree.3.Effect of Maxinshigan Decoction on the pulmonary ultrastructure of influenza A virus infected miceCell structures of normal control group mice were intact,organelles-like mitochondria,endoplasmic reticulum,the Golgi complex,lamellar body and so on were abundant,nuclear membranes were clear enough.The pulmonary ultrastructures were damaged in model group,karyolemma and karyoplasm were inexplicit,the quantity of cell crgans and mitochondrial cristas became smaller or even disappear,vacuolated changes in endoplasmic reticulum showed up and there were plenty of viral particles in endochylema.After drug intervention in each group,damaged extent of mice pulmonary ultrastructures in oseltamivir group,Maxinshigan Decoction group and antiviral granule group were alleviated in different degree.4.The gene expression levels of TLR7/8 in lung of mice infected by influenza A virusThe results of real-time fluorescent quantitative PCR assay showed A-type influenza virus infection in mice led to TLR7/8 gene expression levels increased in lungs,and there was a significant difference compared with the normal control group(P<0.01).Compared with model group,the TLR7 gene expression levels in oseltamivir group,Maxinshigan Decoction group and antiviral granule group were improved,and the difference were significant(P<0.01),the TLR8 gene expression levels in oseltamivir group and antiviral granule group were improved with statistically significant difference(P<0.01).The mice lungs TLR8 gene expression level in Maxinshigan Decoction group had no significant difference compared with model group(P>0.05).5.The protein expression levels of TLR7/8 in lung of mice infected by influenza A virusThe results of western blotting assay showed A-type influenza virus infection in mice led to TLR7/8 protein expression levels increased in lungs,and there was a significant difference compared with the normal control group(P<0.01).Compared with model group,the TLR7 protein expression levels in oseltamivir group,antiviral granule group and Maxinshigan Decoction group were improved,and the difference were significant(P<0.01),the TLR8 protein expression levels in Maxinshigan Decoction group were improved with statistically significant difference(P<0.01).The mice lungs TLR8 protein expression level in antiviral granule group and oseltamivir group had no significant difference compared with model group(P>0.05).Conclusion:1.Maxinshigan Decoction can improve the lung inflammation and damaged lung ultrastructure of mice infected by influenza A virus and relieve pathological damage.2.Maxinshigan Decoction can promote TLR7 gene expression level of influenza A virus infected mice lungs.3.Maxinshigan Decoction has an obvious regulating effect on mice lungs protein expression of TLR7/8 in TLRs signal pathway.To sum up,Maxinshigan Decoction may play the role of against mice lungs pathological damage and anti influenza A virus though regulating gene and protein expression of TLR7/8 in TLRs-MyD88 signal pathway. |
PDF Full Text Request