Intestinal Microbiota And Its Metabolites Are Correlated With Poor Prognosis In Ischemic Stroke

BackgroundEmerging evidences suggest that the gut-brain axis plays an important role in stroke.Significant dysbiosis of intestinal microbiota was found in large-artery atherosclerotic stroke patients,with an overload of Enterobacteriaceae and a lack of Bacteroides.Trimethylamine-N-oxide(TMAO),a gut microbiota-dependent metabolite,has been shown to be proatherogenic and prothrombotic,and corelate with an increased risk of cardiovascular events independent of traditional risk factors.Short-chain fatty acids(SCFAs),another gut microbiota-originating metabolic product,were shown to modulate the permeability of gut and blood-brain barrier,and modulate the brain functions through immune,endocrine,vagal and other humoral pathways.Until now,gut microbiome,plasma TMAO and fecal SCFAs levels among acute ischemic stroke(AIS)patients,as well as their potential associations with stroke outcomes,have not yet been fully evaluated.Here,we designed a prospective observational study of AIS patients and healthy individuals,upon the hypothesis that differential microbial members,TMAO and SCFAs levels existed in AIS,and they correlated with stroke prognosis.MethodsFinally,204 AIS patients and 108 healthy controls were included in the first part of this study;while 140 AIS patients and 92 healthy controls were recruited in the second part.Blood and fecal samples,demographic characteristics,history of risk factors and clinical features were collected.Stroke outcomes were assessed by two trained staff members blinded to the study design via structured follow-up telephone interviews.Plasma TMAO levels were quantified by stable isotope dilution liquid chromatography tandem mass spectrometry(6460 Series Triple Quadrupole LC/MS;Agilent).Serum concentrations of intestinal fatty acid binding protein(FABP),D-lactate,lipopolysaccharides(LPS)and LBP binding protein(LBP)levels were determined using commercially available enzyme-linked immunosorbent assay(ELISA)kits(Bioswamp,Myhalic Biotechnology Co.,Ltd.,Wuhan,China).After fecal DNA extraction,PCR amplification and sequencing of bacterial 16S rRNA genes,bioinformatics were obtained and then analyzed with the Quantitative Insights Into Microbial Ecology(QIIME 1.80).The concentrations of fecal SCFAs were detected by gas chromatography tandem mass spectrometry(5977B GC/MS,Agilent Technologies,Santa Clara,CA,USA).The associations with poor stroke outcomes were evaluated by univariate and multivariate logistic regression models.Receiver operating characteristic(ROC)curves were used to examine the prognostic ability of TMAO.ResultsTMAO levels of AIS patients were higher than that of healthy controls,but decreased with time since stroke onset.TMAO levels before or within 24 hours of AIS treatment were defined as the baseline levels.Elevated log2-transformed baseline TMAO levels but not those of afterwards were associated with increased risks of 90-day and 12-month major ischemic events and unfavorable functional outcomes after adjustments for the currently established risk factors.With a moderate predictive potential,baseline TMAO levels improved the prognostic accuracy of conventional risk factors,the NIHSS scores and the NT-proBNP levels.Gut microbial composition and fecal SCFAs levels distinguished AIS patients from healthy individuals.A lack of SCFAs-producing bacteria(Roseburia,Bacteroides,Lachnospiraceae,Faecalibacterium,Blautia and Anaerostipes),an overload of potentially undesirable bacteria(Lactobacillaceae,Akkermansia,Enterobacteriaceae and Porphyromonadaceae)and a low fecal SCFAs level defined dysbiosis of AIS patients.AIS patients with increased stroke severity exhibited significantly reduced SCFAs levels.Low log2-transformed SCFAs levels,especially acetate,were associated with an increased risk of 90-day poor functional outcomes even after strict adjustments.ConclusionsTMAO levels of AIS patients decreased with time since stroke onset.A lack of SCFAs-producing bacteria and a low fecal SCFAs level defined dysbiosis of AIS patients,especially those with increased stroke severity.Elevated TMAO and reduced SCFAs levels were associated with increased risks of major ischemic events and unfavorable functional outcomes even after adjustments for the currently established risk factors.With a moderate predictive potential,baseline TMAO levels improved the prognostic accuracy of conventional risk factors,the NIHSS scores and the NT-proBNP levels.Our data broaden the potential clinical utility of intestinal microbiota and its metabolites as independent prognostic markers and therapeutic targets.