Effect And Mechanism Of Protease-activated Receptor 2 On Bronchopulmonary Dysplasia In Neonatal Rats

Research background and objectives:Bronchopulmonary dysplasia(BPD)is a common lung disease in premature infants.Its pathogenesis is not clear,and there is no specific prevention and treatment method.protease activated receptor-2(PAR2)is an important pathogenic factor in the pathogenesis of a variety of lung diseases,which can drive inflammatory response and promote lung injury.This study intends to explore the lung PAR2 in high oxygen induced BPD rats model,the expression and its function mechanism in the lung tissue,the pathogenesis of BPD for premature babies and prevention strategies to provide new ideas.Method:Forty-eight neonatal SD rats at gestational age of 20-22 days were randomly divided into air group and high oxygen group,with 24 rats in each group.Neonatal SD rats in the air group were fed in continuous air condition after birth,while those in the high oxygen group were fed in a closed oxygen chamber with continuous oxygen concentration >60% within 6h after birth.Six rats were collected from the two groups at day 1(postnatal 1,P1),day 4(postnatal 4,P4),day 7(postnatal 7,P7)and day 10(postnatal 10,P10),respectively,and lung tissues were collected under anesthesia.The pathological changes of lung tissues were compared between the two groups,and the radiative alveolar count,alveolar septum thickness and collagen fiber hyperplasia were measured.Meanwhile,the expressions of PAR2 and IL-18 in lung tissues were detected by Western-blot,and the expressions of PAR2,P38 MAPK and NF-κB in lung tissues were detected by immunohistochemical method.Newborn rats were randomly divided into Air group(Air),Hyperoxia group(Hyperoxia),Hyperoxia group +PAR2 agomir(Hyperoxia+ Sligry-NH2),hyperoxia group +PAR2 antagomir(Hyperoxia+ Fslri-NH2)and hyperoxia group +P38MAPKantagomir(Hyperoxia+SB203580)5 groups,a total of 90 patients,PAR2 agonists and inhibitors,P38 MAPK inhibitors were injected intrabitoneally,after 7consecutive days of injection,H&E staining was used to understand lung tissue morphological changes,Masson staining was used to understand pulmonary fibrosis.The positive expression area of NF-κB in lung tissues was detected by immunofluorescence assay.The m RNA expression of PAR2,P38 MAPK and NF-κB was detected by q RT-PCR.The gene expressions of chemokines(CCL2,CCL7,CXCL1,CXCL2)and inflammatory factors(IL-6,IL-17 a,IL-18,TNF-α)in lung tissue were quantified by q RT-PCR.Results:1.With the increase of age,the growth and development of rats in the hyperoxic group were delayed,the body weight of P4 and P7 was significantly lower than that of the air group(p<0.01),and the body weight of P10 was significantly decreased(p<0.001).Pathological results showed that compared with the air group,P4 in the hyperoxic group showed alveolar retardation(decreased radial alveolar count,p<0.001)and inflammatory cell infiltration.P7 showed thickening of alveolar septa(thickening of alveolar septa,p<0.001),hyperplasia of collagen fibers(increased area of lung collagen fibers,p<0.001),and disturbance of alveolar structure.The expressions of PAR2,P38 MAPK,NF-κB and IL-18 in lung tissues of P4,P7 and P10 in hyperoxia group were increased(p<0.05).2.The collagen fiber staining area of Hyperoxia group and Hyperoxia+SLIGRL-NH2 group was higher than that of Air group,and the collagen fiber staining area of the Hyperoxia+ Sligrl-NH2 group was significantly lower than that of the Hyperoxia+ Sligrl-NH2 group after using PAR2 and P38 MAPK inhibitors(p<0.05).3.The expressions of chemokines and inflammatory cytokines in Hyperoxia group and Hyperoxia+SLIGRL-NH2 group were higher than those in Air group,and the expressions of chemokines and inflammatory cytokines in high oxygen group were significantly lower after the use of PAR2 and P38 MAPK inhibitors(p<0.05).4.The positive expression area of NF-κB in Hyperoxia group and Hyperoxia+SLIGRL-NH2 group,and the gene expressions of PAR2 and P38 MAPK in Hyperoxia group were higher than those in Air group,and the expressions of PAR2 and P38 MAPK inhibitors in Hyperoxia group were lower than those in AIR group(p<0.05).Conclusion:1.Hyperoxic environment promotes lung PAR2 overexpression and induces BPD lung injury.2.Overexpression of PAR2 can promote cascade effect of pulmonary inflammation,pulmonary growth retardation and pulmonary fibrosis,and inhibition of overexpression of PAR2 has protective effect on BPD,which may be a potential target for BPD treatment.3.The P38 MAPK/NF-ΚB signaling pathway may be an important pathway for PAR2 to deign the inflammatory reaction of BPD and pulmonary fibrosis.