Role Of NLRP3/IL-1β/TGF-β1 Signal Axis In The Development Of Silicosis Pulmonary Fibrosis And The Antagonistic Effect Of Luteolin

Objective:To investigate the role of NLRP3/IL-1β/TGF-β1 signal axis in the occurrence and development of silicosis pulmonary fibrosis,and to explore the antagonistic effect of luteolin on silicosis pulmonary fibrosis with NLRP3/IL-1β/TGF-β1 signal axis as the target.Methods:SPF Wistar male rats were randomly divided into high,medium and low doses of SiO₂ dust exposure group and control group.In the dust-exposed groups,one-time non-exposed tracheal intubation was used to make the model,while in the control group,0.9%sodium chloride solution was sterilized by equal volume.Five rats were randomly executed 7,14,28 and 56 days after exposure SiO₂ dust.The degree of pulmonary alveolitis and pulmonary fibrosis were observed.The expressions of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in lung tissue were detected.SPF Wistar male rats were randomly divided into negative control group,model group and inhibitor group.At the same time of modeling,the inhibitor group was given 10mg/kg of NLRP3 inhibitor,and the model group and the control group were given an equal volume of sterilized 0.9%sodium chloride solution,and administered once a day.Five rats were randomly executed at 7,14,28 and 56 days after exposure SiO₂ dust.The degree of pulmonary alveolitis and pulmonary fibrosis were observed.The expressions of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in lung tissue were detected.Rat alveolar macrophages were divided into blank control group,dusting group,NLRP3 inhibitor group and luteolin group.50mg/L of dust was added to the dusting group,50mg/L of dust and 20μmol/L of NLRP3 inhibitor were added to the NLRP3 inhibitor group,and 50mg/L of whisk and 20μmol/L of luteolin were added to the luteolin group.The samples were collected at 12,24 and 48h.The cell growth was detected by MTT assay,and the expression of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in lung tissue was detected.SPF Wistar male rats were randomly divided into negative control group,model group,Ajuga Decumbens group,NLRP3 inhibitor group,Caspase-1 inhibitor group,IL-1beta inhibitor group and luteolin high,medium and low dose group.Each administration group was given a corresponding dose of the therapeutic drug,and the model group and the control group were intragastrically administered with an equal volume of sterilized 0.9%sodium chloride solution,and administered once a day.Five rats were randomly sacrificed at 7,14,and 28 days after modeling.The changes of lung tissue morphology,lung index,lung alveolitis and pulmonary fibrosis were observed.The expressions of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in lung tissue were detected.Results:The alveolitis score,pulmonary fibrosis score and the expression of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in the lungs of the dust-treated group were higher than those in the control group at 7,14,28d and 56d（P<0.01）;the expression levels of NLRP3,Caspase-1,IL-1β,IL-18 and TGF-β1 in low,medium and high concentrations of SiO₂dusting group were compared between 7,14 and 28d（P<0.05）,a time-effect relationship that increases with increasing dusting time.The scores of pulmonary fibrosis degree and the levels of IL-1β,IL-18 and TGF-β1 in lung tissue of rats had statistical significance only in the main effect of dust treatment（P<0.01）.The order from high to low was model group,inhibitor group and control group（P<0.01）.At 7,14,28 and 56 days after modeling,the alveolitis score and the relative expression levels of NLRP3 and Caspase-1 protein in lung tissues of the inhibition group were lower than those of the model group at the same time point（P<0.01）,but higher than those of the control group at the same time point（P<0.01）.The survival rate of dust-exposed macrophages in NLRP3 inhibitor group and luteolin group at three time points was higher than that in dust-exposed group（P<0.05）,and at 12and 24 hours luteolin group was higher than that in inhibitor group（P<0.05）.Compared with the dust-stained group,the expressions of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 were decreased in the inhibitor group and the luteolin group（P<0.05）.Compared with the inhibitor group,the levels of IL-1βin luteolin group decreased at all three time points,while the levels of IL-18 and TGF-β1 decreased only at 48 hours（P<0.05）,and the levels of NLRP3 and Caspase-1 decreased only at 24 hours（P<0.05）.Compared with other treatment groups,the high-dose luteolin group had advantages in improving the general condition of silicosis rats,reducing the severity of pulmonary index,alveolitis and pulmonary fibrosis and the expression of IL-1β,IL-18,TGF-β1,NLRP3 and Caspase-1 in lung tissue,especially at 14 and 28 days.Conclusions:NLRP3/IL-1β/TGF-β1 signal axis plays an important role in the occurrence and development of silicosis pulmonary fibrosis.Luteolin can inhibit the activation of inflammatory bodies in NLRP3,reduce the activation of Caspase-1,inhibit the expression of inflammatory mediators IL-1βand IL-18,and ultimately reduce the expression of TGF-β1,thereby reducing the inflammatory response and fibrosis of the lung.It is suggested that luteolin can inhibit silicosis pulmonary fibrosis by interfering with NLRP3/IL-1β/TGF-β1 signal axis.