Human Gut Microbiome Impacts Skeletal Muscle Mass Via Gut Microbial Synthesis Of The Short-Chain Fatty Acid Butyrate Among Healthy Menopausal Women

Background:Sarcopenia is defined as a progressive and systemic skeletal muscle disease,characterized by loss of skeletal muscle mass（SMM）and function with people of advanced age,increasing the risk of falls,fractures,disability and death.The prevalence of sarcopenia in 60 to 70-year-olds is in the range of 5-13%,while prevalence estimates increase to 11-50%for the population aged 80 years or older.On the other hand,loss of SMM is a more serious problem,with a prevalence of approximately 39%in elderly.It is urgent to find more effective therapeutic regimens to prevent and treat loss of SMM or sarcopenia.Gut microbiota-derived metabolites are implied in skeletal muscle,the gut-muscle axis was proposed to study this relationship.Developing interventions that target the gut-muscle axis may be a new target for preserving or alleviating age-related loss of SMM and function.Methods and purpose:Part Ⅰ:Spearman’s rank correlation test was used to explore the potential associations between gut microbial features（including gut metagenomics species and gut metabolic pathways）and SMM by leveraging shotgun metagenomic sequencing,serum short-chain fatty acids（SCFAs）,as well as available host SMM measurements among community-dwelling healthy menopausal women（N=482）.The potential associations between gut microbial features and SMM could provide a prerequisite for causality.Part Ⅱ:By leveraging whole-genome sequencing of the participants and combining publicly available summary statistics from genome-wide association analyses for human gut microbiome and SMM,we conducted metagenome-genome-wide association analysis,polygenic risk score,and Mendelian randomization（MR）analysis to further investigate an important finding of the Part Ⅰ whether there is a causal relationship between the increased capacity for gut microbial synthesis of the SCFA butyrate and SMM.Part Ⅲ:Based on the results provided by Part I and Part Ⅱ about the relationship between gut microbial features and SMM,F.prausnitzii may be the driver species for the positive association between gut microbial synthesis of the SCFA butyrate and SMM,suggesting a possible causal relation.Thus,we explored the molecular mechanism of F.prausnitzii affecting SMM and function through animal experiments in aging mice.Research result:Part Ⅰ:Gut microbial diversity was positively correlated with SMM.Based on the associations between gut microbial features and SMM,the capacity for gut microbial synthesis of the SCFA butyrate was positively associated with skeletal muscle index[Spearman correlation coefficient（SCC）=0.08,P=0.002]and serum butyrate（SCC=0.13,P=0.02）.Of interest was the finding that two main butyrate-producing bacterial species were both positively associated with the increased capacity for gut microbial synthesis of butyrate[F.prausnitzii(SCC=0.25,P=6.6×10^-7)and B.virosa（SCC=0.15,P=0.001）]and for skeletal muscle index[F.prausnitzii(SCC=0.16,P=6.2×10^-4)and B.virosa(SCC=0.17,P=2.4×10^-4)].From the results of the metagenomics association analysis,there was an implication that,in the study sample of Chinese menopausal women,gut microbiome may have clinical benefit to SMM attributable to gut microbial synthesis of SCFA butyrate.Part Ⅱ:One-sample MR results showed a causal effect between gut microbial synthesis of the SCFA butyrate and appendicular lean mass[β=0.04,95%confidence interval（CI）:0.029 to 0.051,P=0.003).Two-sample MR results further confirmed the causal effect between gut microbial synthesis of the SCFA butyrate and appendicular lean mass（β=0.06,95%CI:0 to 0.13,P=0.06）.Taken together,there may be a causal effect between gut microbial synthesis of the SCFA butyrate and SMM.Part Ⅲ:The F.prausnitzii-gavaged mice had higher SMM（including gastrocnemius and tibialis anterior）and anti-fatigue capacity compared to sham-gavaged controls after eight weeks challenge,indicating a healthy status.To reveal the effects of F.prausnitzii on SMM,metabolomic analysis based on liquid chromatography tandem mass spectrometry were utilized to detail the metabolic characterization of SMM after the gavage with F.prausnitzii.Metabolic pathways including oxidative phosphorylation,energy metabolism and amino acid metabolism were modulated.Of interest was the finding that levels of butyrate in muscle was significantly increased in the F.prausnitzii-gavaged mice.RNA sequencing of tibialis anterior muscle was performed to investigate the effects of F.prausnitzii on gene expression in muscle,there were significant differences in expression of genes encoding different cell signaling molecules that regulate SMM,including NF-kappa B signaling pathway,gamma-aminobutyric acid signaling pathway and chemical synaptic transmission.Conclusions:In conclusion,we confirmed the key role of gut microbial synthesis of the SCFA butyrate on host SMM by leveraging multi-omics data from 482 menopausal individuals,combining the results with publicly available summary statistics from genome-wide association analyses for human gut microbiome and SMM,as well as animal experiments in aging mice.Our results may help the future development of novel intervention approaches for preserving or preventing/alleviating loss of SMM.