The Study Of The Intervention Effect Of Qingfei Paidu Decoction On NLRP3 Signaling Pathway Of Coronavirus Infection Model

ObjectiveTo observe the inhibitory effect of Qingfei Paidu Decoction on the inflammation caused by coronavirus infection,and to study its mechanism based on NLRP3 pathway.MethodsFirstly,BNL CL.2 cells were infected with coronavirus-murine hepatitis virus-to construct in vitro MHV-infected cell model.Microscopic observation,AO/EB staining was used to detect the effect of Qingfei Paidu Decoction on MHV-infected cell model.The IC50 concentration of Qingfei Paidu Decoction on MHV-infected cell model was detected by MTT assay,Quantitative Real-time PCR was used to detect the intracellular antiviral effect and the effect of intracellular pro-inflammatory factor IL-1β mRNA of Qingfei Paidu Decoction.Then,Kunming mice was infected with coronavirus-murine hepatitis virus to construct MHV-infected mouse model.We observed the effects of different dosage and duration of Qingfei Paidu Decoction on body weight,diet and water intake of MHV-infected mouse model.HE staining was used to detect the effects of different doses and duration of Qingfei Paidu Decoction on the pathological injury of liver and lung tissue in MHV-infected mouse models.Quantitative Real-time PCR was used to detect the effects of different doses and duration of Qingfei Paidu Decoction on NLRP3,ASC,caspase-1,IL-1β,IL-18,NF-κB mRNA and the relative amount of MHV virus in liver and lung.Western Blot assay was used to determine the effects of different doses and duration of Qingfei Paidu Decoction on NLRP3,ASC,caspase-1,IL-1β,IL-18,NF-κB protein in liver and lung.ResultsThe therapeutic effect of Qingfei Paidu Decoction on MHV-infected cell model was studied by microscope observation,MTT assay and AO/EB staining.AO/EB staining showed that 0.030625mg/mL of Qingfei Paidu Decoction still had therapeutic effect MHV-infected cell model.The CC50 and IC50 of Qingfei Paidu Decoction on BNL CL.2 cells were 8.951 mg/mL and 0.535 mg/mL respectively by MTT assay.The treatment index was 16.7308.Quantitative Real-time PCR showed that Qingfei Paidu Decoction had mild antiviral effect in vitro,and could reduce the mRNA expression level of IL-1β in BNL cell infected model.It was observed that Qingfei Paidu Decoction could reduce the decrease of diet and water intake of MHV-infected mouse model and reduce the influence of MHV virus on body weight of mouse model.HE staining showed that Qingfei Paidu Decoction could reduce the pathological injury of liver and lung tissue in MHV-infected mouse model with different dosage and duration of administration.Quantitative Real-time PCR showed that Qingfei Paidu Decoction could slightly inhibit the replication of MHV virus in mice with different dosage and duration.Quantitative Real-time PCR and Western Blot assay showed that NLRP3,ASC,caspase-1,IL-1 β,IL-18,NF-κ B mRNA and protein expression in liver and lung tissues of MHV-infected mice could be reduced at different doses and duration of Qingfei Paidu Decoction.ConclusionQingfei Paidu Decoction has therapeutic effect on the infection of mouse hepatitis virus,the alternative coronavirus of SARS-Co V-2,both in vitro and in vivo.It can improve the survival rate of BNL cell infection model,reduce the loss trend of weight gain in MHV infection model,reduce the decline of diet and water intake,and reduce the pathological damage in MHV infection model.Qingfei Paidu Decoction has antiviral effect in vitro and in vitro.Qingfei Paidu Decoction may inhibit the expression of NLRP3 protein,ASC protein and caspase-1 protein,and lead to the down-regulation of the downstream signaling molecule IL-1β and inflammatory response-related protein NF-κB.Thus,it can inhibit the inflammatory response of the body,avoid the excessive inflammatory stress of the body,reduce the pathological damage of the body,and finally play a therapeutic effect on the body.