The Effect Of NLRP3 In Chronic Rhinosinusitis Without Nasal Polyps

Objective:To define the role of the NLRP3 in chronic sinusitis without nasal polyps(chronic rhinosinusitis without nasal polyps,CRSs NP).Methods:In this study,the C57BL/6J mouse model of CRSs NP was prepared with Streptococcus pneumoniae bacterial solution,the mice were randomly divided into three groups:surgical implantation group(Streptococcus pneumoniae fluid expansion sponge implantation group);the surgical non-implantation group(without expansion expansion implantation)and the control group(non-surgical group).Observe the inflammatory reaction of nasal sinus mucosa tissue in each group by HE staining,and detect NLRP3 protein expession of tissue in each group by Western blot.In vitro experiment,human nasal epithelial cell line(HNEPC)was selected,and LPS was used to induce the formation of inflammatory cell model.The optimal concentration of LPS was selected through CCK-8 experiment.Cells were randomly divided into two groups,inflammatory induction group and control group.The cells were randomly divided into two groups,inflammatory induction and control groups,the expression levels of NLRP3,IL-1β and IL-18 genes were determined by RT-PCR experiments;the protein expression level of NLRP3 in both groups was determined through the Western blot and the protein expression levels of IL-1β and IL-18 were tested through ELISA.Results:(1)HE staining results showed that chronic inflammation was seen in the sinuses and nasal mucosa of the surgical implanted group,with uneven distribution of mucosal epithelium,necrosis and shedding of epithelial cells,hyperplasia of submucosal glands and fibrous tissue,and lymphocytes and plasma cell infiltration.The mucosa epithelium of nasal sinuses and nasal cavity in the non-implantation group was the same as that in the normal group,without obvious infiltration of submucosal lymphocytes and monocytes,and without gland hyperplasia;in the control group,the nasal sinus and nasal cavity mucosa epithelial tissue structure is regular and orderly arranged,no obvious inflammatory cell infiltration is found in the submucosa,and a small amount of goblet cells are occasionally seen.(2)Western blot results showed that the expression of NLRP3 protein in the tissue of the operating group was significantly higher than that of the control group,and comparing the two groups of data can find that they have significant differencest(P<0.05).(3)CCK-8 experiment screened the optimal LPS induction concentration,and found that the OD value of each group increased with the increase of time;when cultured for 6h,12 h and 24 h,the OD of LPS at 1ng/m L and 10ng/m L was significantly higher than that of other groups(P<0.05);when cultured for 48 hours,there was no statistical difference between the 1ng/m L and 10ng/m L groups and the control group(P>0.05).At 24 h,the proliferation rate of cells with LPS concentration of 10ng/m L was significantly higher than that of 1ng/m L,and comparing the two groups of data can find that they have significant differencest(P<0.05);At 6h and 12 h,there was no significant difference between the 10ng/m L group and the 1ng/m L group(P>0.05).(4)RT-PCR detected inflammasome and inflammatory factor gene expression under LPS induction,and found significantly higher NLRP3,IL-1β and IL-18 gene expression in the LPS induction group at 4d than in the control group,with a statistically significant difference(P <0.05).(5)Western blot to detect NLRP3 protein expression under LPS induction,the results found that compared with the control group,the LPS-induced group has a higher level of NLRP3 in HNEPC,and comparing the two groups of data can find that they have significant differencest(P<0.05).(6)ELISA for IL-1β and IL-18 protein expression under LPS induction found that the expression of IL-1β and IL-18 protein in the supernatant of HNEp C culture in the LPS induction group was significantly higher than the control group,which was statistically significant(P <0.05).Conclusion: NLRP3 and its downstream factors IL-1βand IL-18 play important roles in the development of chronic sinusitis without nasal polyps.