| PART ? HIGH-THROUGHPUT SEQUENCING-BASED GUT MICROBIOME COMPOSITION ANALYSIS IN INFANTS WITH BRONCHOPULMONARY DYSPLASIAObjective: This study investigated an association between bronchopulmonary dysplasia(BPD)and infants' gut microbiota.The study included determination of the composition of the gut microbiome associated with bronchopulmonary dysplasia.Methods: The study population consisted of 10 infants with BPD and 10 infants without BPD(Control).Premature infants admitted to the Department of Neonatology,Yuzhong District,Children's Hospital of Chongqing Medical University from March 2019 to November 2020 were selected as the research objects.Collected feces were analyzed by high-throughput sequencing.Results: At day 7,the intestinal flora of the BPD group and the Control group were not significantly different at the phylum and genus levels(P>0.05);At day 28,the BPD group had a significant increased Proteobacteria(P<0.01)and decreased Firmicutes at the phylum level compared to the Control group(P<0.01);at the genus level,Klebsiella was the main colonized bacteriain the BPD group(P<0.05),and Clostridium?sensu?stricto?1 was significantly decreased(P<0.05).Conclusion: The gut composition of bronchopulmonary dysplasia patients were as follows: Proteobacteriaincreased and Firmicutes decreased;at the genus level,the Klebsiella increased,and the Clostridium?sensu?stricto?1 decreased.The results suggest that antibiotic-induced intestinal dysbacteriosisis related to BPD development.PART ? ANTIBIOTIC-INDUCED INTESTINAL DYSBACTERIOSIS ON THE PATHOGENESIS OF BRONCHOPULMONARY DYSPLASIAObjective: According to the results of the part ?,antibioticsinduced intestinal dysbacteriosisaggravated bronchopulmonary dysplasia.Antibiotic-induced intestinal dysbacteriosis can interfere with the function of macrophages,leading to immune responses and increasing sensitivity to inflammation.These immune changes can be prevented by supplementing short-chain fatty acids.Therefore,this part intends to explore whether the intestinal dysbacteriosisparticipate in the pathogenesis of BPD by influencing the polarization of M1/M2 macrophages.Methods: Mice were randomly assigned to different cages,and were gavaged with antibiotics for 7 days to induce intestinal dysbacteriosis.In the case of the BPD model establishment,neonatal mice along with their mothers were exposed to hyperoxia(85%O2)for 14 consecutive days and were gavaged with butyrate.The study was divided into 5 groups:Saline/Air group,ABX/Air group,Saline/O2 group,ABX/O2 group,ABX/O2/SCFAs group.Percent of survivalin each group were observed,HE staining of lung tissue and alveolar count were obtained.,ELISA was used to detect the expression of cytokines IL-1?,IL-6,TNF-a,IL-10.Assay of commensal bacteria in the small intestine were performed by16 S r RNA.Results:The survival rate of mice in ABX/O2 group was significantly lower than that in Saline/O2 group.After the intervention of butyrate,the survival rate of mice in the ABX/O2/SCFAs group increased.HE staining: The alveolar structure of the mice treated with saline and antibiotics is complete;the number of alveoli in mice exposed to hyperoxia is reduced,interstitial congestion,edema and inflammatory cell infiltration;compared to Saline/O2,the alveolar wall is thicker and the number of alveoli reduce in the ABX/O2 group,the lung tissue is disordered.After the intervention of butyrate,compared to the mice whic exposed to hyperoxia and treated with antibiotics,the inflammation of the alveolar wall of the mice was reduced and the number of alveoli increased.The lungs of BPD micehad an elevated MLIand decreased RAC after exposed to hyperoxia.Moreover,lungs of mice in the ABX/O2 group had an elevated MLI and reduced RAC compared to the Saline/O2 group.Lungs of mice in the ABX/O2/SCFAs group had an reduced MLI and reduced elevated RAC compared to the ABX/O2 group.ELISA: The expression levels of IL-1?,IL-6 and TNF-? in the Saline/O2 group were significantly higher than those in the Saline/Air group(P<0.05,P<0.05,P<0.05).After antibiotics treatment,the expression levels of the above inflammatory cytokines increased(P<0.05,P<0.05,P<0.05).In addition,the expression level of IL-10 in the Saline/O2 group was significantly lower than that in the Saline/Air group(P<0.05).After antibiotics treatment,the expression levels of the cytokines were reduced(P<0.05).Compared to the ABX/O2 group,the expression levels of IL-1?,IL-6 and TNF-? were lower in ABX/O2/SCFAs group(P<0.01,P<0.05,P<0.05),and IL-10 expression levels were increased(P<0.05).16 Sr RNA: After antibiotics treatment,the proportion of proteobacteria increased significantly(P<0.01),Firmicutes and Bacteroides decreased significantly(P<0.01);At the genus level,the proportion of Citrobacter and unclassifiedf<sub>Enterobacteriaceae in the ABX group increased(P<0.01),the proportion of Bacteroides and norankf<sub>Muribaculaceae decreased(P<0.01).SCFAs: After antibiotics treatment,acetic acid(P<0.05),propionic acid(P<0.05)and butyric acid(P<0.01)in serum of newborn mice were significantly reduced.Conclusion: The composition of the gut microbiota of mice after antibiotics treatment was significantly changed and the serum short-chain fatty acids decreased,indicating that antibiotics treatment induced intestinal dysbacteriosis.Compared with Saline/O2,the ABX/O2 group had higher mortality,decreased alveolar number,increased IL-1?,IL-6,and TNF-? and decreased IL-10.It is suggested that antibiotics treatment promotes the BPD development,and supplementation of butyrate inhibits the effect of antibiotics on the onset of BPD.PART ? INTESTINAL DYSBACTERIOSIS AGGRAVATES BRONCHOPULMONARY DYSPLASIA BY PROMOTING THE IMBALANCE OF THE POLARIZATION OF M1/M2 MACROPHAGES IN THE LUNGSObjective: Bronchopulmonary dysplasia is a serious respiratory disease that occurs in premature infants.Bacterial colonization may drive an inflammatory response,disrupt lung development and then cause BPD.Therefore,this study intends to use antibiotics to induce intestinal dysbacteriosis to explore its impact on the pathogenesis of BPD,to study the role of intestinal flora in the occurrence and development of BPD,and to provide a new target for clinical treatment.Methods: The study was divided into 5 groups: Saline/Air group,ABX/Air group,Saline/O2 group,ABX/O2 group,ABX/O2/SCFAs group.Western blot and immunofluorescence were used to detect the protein expression of i NOS and Arg-1.Results: After exposed to hyperoxia,the expression of i NOS in the lung tissue of mice was significantly increased.After antibiotics treatment,the expression of i NOS in the ABX/O2 group increased,and the expression of i NOS in the ABX/O2/SCFAs group was lower than that of ABX/O2.In addition,after exposed to hyperoxia,the expression of Arg-1in the lung tissue of mice was significantly reduced.After antibiotics treatment,the expression of Arg-1 in the ABX/O2 group was reduced,and the expression of Arg-1 in the ABX/O2/SCFAs group was higher than that of ABX/O2.Conclusion: After antibiotics treatment induces intestinal dysbacteriosis,it promotes M1 polarization and inhibits M2 polarization,thereby aggravating BPD.Butyrate intervention inhibits M1 polarization and promotes M2 polarization,thereby reducing the inflammatory response in BPD mice caused by antibiotics. |
PDF Full Text Request