Study On The Mechanism Of Inhibition Of NETosis-induced Asthma Inflammation By Penicilazaphilone C

Objective: Penicilazaphilone C(PAC)is a nitrogen ketone compound.It has been reported to have a variety of biological activities,mainly antibacterial,antifungal,antiviral,anti-inflammatory,antioxidant and cytotoxic.A fundamental feature of asthma is chronic inflammation of the airways,and despite medical advances in the treatment of asthma,its prevalence continues to increase,and given the high heterogeneity of the disease,more than 50% of patients are not fully controlled,the treatment of asthma remains a great challenge.Studies have shown that neutrophil extracellular traps(NETs)and their formation process(NETosis)have a certain degree of influence on asthma.The purpose of this study was to preliminarily explore the effect of Penicilazaphilone C on the inflammatory response of asthmatic mice,and the possible mechanism of the formation of NETosis in the development of asthma.Methods: In vitro,NETs were induced in neutrophils of healthy mice with phorbol 12-myristate-13-acetate(PMA).Four groups of different doses of Penicilazaphilone C(10mg/kg,20mg/kg,30mg/kg,40mg/kg)and blank control groups were set up to quantify the level of double-stranded DNA in the neutrophil culture supernatant to understand inhibitory effect and determination of optimal effective dose.In vivo,a neutrophilic asthma mouse model was established by inducing lipopolysaccharide(LPS),and 30 mice were randomly divided into 5groups: control group,asthma group,dexamethasone group,DNase I group and PAC group.24 hours after modeling,the levels of cytokines(IL-4,IL-6,IL-17 and TNF-α)in bronchoalveolar lavage fluid(BALF)were measured.The number of inflammatory cells in BALF was measured by cell counter.The changes of airway resistance in mice were recorded.Lung tissues were stained with HE staining,PAS staining and immunofluorescence.Measurement of reactive oxygen species(ROS)in neutrophils.Citrullination of histone H3 and histones in lung tissue by western blotting.NETs in peripheral blood of mice in each group were qualitatively and quantitatively detected.Results: In vitro,40mg/kg Penicilazaphilone C could significantly inhibit the formation of NETs.In vivo,compared with the control group,the NETs in the BALF,peripheral blood and lung tissue of mice in the asthma group were significantly increased,and there were a large number of inflammatory factors and inflammatory cells in the BALF,with a large proportion of IL-17 and neutrophils,and the lung tissue was significantly damaged.The levels of inflammatory factors and inflammatory cells in BALF in PAC group were significantly lower than those in asthma group.Compared with the asthma group,the pathological changes of the lung tissue in the PAC group were significantly decreased,and the levels of ROS,citrullination of histone H3 and NETs were also significantly decreased.Conclusion: The neutrophilic asthma mouse model was successfully established by LPS.Penicilazaphilone C can significantly alleviate inflammation in asthmatic mice,and the mechanism is related to the inhibition of ROS,citrullination of histone H3 and the formation of NETosis.