Study On Intestinal Mucosal Barrier Damage Markers And GutMicrobiota In Patients With Insomnia Disorder

ObjectiveBased on our previous findings of altered diversity and abundance of intestinal flora in patients with insomnia disorder,this study examined the intestinal mucosal barrier function and intestinal flora of patients with insomnia disorder,to assess whether patients with insomnia disorder have impaired intestinal mucosal barrier.To reveal the relationship between intestinal flora and intestinal mucosal barrier in patients with insomnia disorder,and to screen for possible biomarkers of intestinal flora in patients with insomnia disorder.Materials and MethodsAccording to the diagnostic criteria of insomnia disorder in The Diagnostic and Statistical Manual of Mental Disorders-5,general condition questionnaire,clinical interview,subjective mood and sleep related scales screening and PSG,a total of 53 cases in the insomnia disorder group and 38 cases in the healthy control group were finally included.Serum concentrations of Diamine oxidase(DAO),D-lactic acid(D-LA),intestinal fatty acid binding protein(I-FABP)and bacterial endotoxin(ET)were measured by ELISA and the 16 S rDNA V4 region of feces was sequenced by Illumina Nova Seq PE250 by 16 S rDNA sequencing technology.Spearman correlation analysis was used to analyze the correlation between intestinal microflora and intestinal mucosal markers.The random forest algorithm was used to screen out the biomarkers.Data analysis was performed using SPSS 26.0 and R language.Results1.Compared to healthy controls,the insomnia disorder group had significantly higher scores on the subjective mood and sleep scales(such as GAD-7,PHQ-9,HAMA,HAMD,PSQI and ISI)(p<0.05).Nocturnal PSG suggested that patients with insomnia disorder had significantly more bedtime,total waking time,sleep latency and wake after sleep onset,but lower sleep efficiency.And their sleep structure was changed(increasing the duration and proportion of N1 phase and decreasing the duration of N2 phase)(p<0.05).2.Compared to healthy controls,serum concentrations of DAO(3.34±0.47)ng/m L,D-LA(6.74±1.16)μmol/L,I-FABP(193.88±29.18)pg/m L and ET(2.96±0.42)pg/m L were significantly lower in the insomnia disorder group(P<0.001).3.There were significant differences in species diversity,composition,function,and co-occurrence network of the intestinal flora between the two groups(P<0.05).There was no difference on the alpha diversity indices Chao1,ACE,Simpson and Shannon between the groups(P>0.05).PCo A and PERMANOVA analyses revealed significant differences in beta diversity(P<0.05).The relative abundance of p＿Acidobacteria,p＿Proteobacteria and p＿Firmicutes were increased while the p＿Bacteroidota was decreased.Using the Spar CC algorithm to calculate the species interaction matrix,the MD index was-0.4296,indicating that the taxon richness of the insomnia disorder group was generally reduced.The ROC curve was constructed based on the random forest algorithm and the AUC value was calculated to be 0.62.Finally,biomarkers such as s＿metagenome,s＿[Clostridium]＿spiroforme,g＿Lachnoclostridium,and g＿Agathobacter were screened out.4.The results of Mantel test analysis showed that there was a certain correlation between intestinal flora and intestinal mucosal barrier.The relative abundance matrix of species in the insomnia disorder group was positively correlated with DAO(r=0.135,P=0.029)and ET(r=0.123,P=0.02).The biomarkers of intestinal flora were closely related to clinical indicators in insomnia disorder group.ET was negatively correlated with Monoglobus(r=-0.27,P=0.029);D-LA was negatively correlated with CAG-352(r=-0.31,P=0.012)and so on.ConclusionPatients with insomnia disorder have disturbed sleep structure and are more likely to experience anxiety and depression.Patients with insomnia disorder have impaired intestinal mucosal barrier function and significant changes in intestinal flora species diversity,structure,function and co-occurrence network.There is a close correlation between the intestinal mucosal barrier function and intestinal flora.The biomarkers s＿metagenome,s＿[Clostridium]＿spiroforme,g＿Lachnoclostridium,g＿Agathobacter,could effectively distinguish the two groups.The results of this study provides some reference for the clinical diagnosis of insomnia disorder.