Based on the theory of “the heart is connected with the small intestine” in traditional Chinese medicine，this study explored the mechanism of gut microbiota driving the TMA/FMO3/TMAO pathway to affect platelet aggregation in acute coronary syndrome（ACS）with syndrome of phlegm and blood stasis rats.

Forty-five SPF healthy SD rats were randomly divided into control group（CON），ACSgroup and ACSwith syndrome of phlegm and blood stasis group（TYACS），with 15 rats in each group.Eight-channel multi-lead electrocardiogram was used to record the changes of leadⅡelectrocardiogram of the standard limb of rats.The serum lipid levels of rats were detected by automatic biochemical analyzer.HE staining was used to observe the structural changes of myocardial tissue.The maximum aggregation rate of platelet arachidonic acid（AA）and adenosine diphosphate（ADP）were detected by photoelectric turbidity method.LC/MS technology was used for detecting the levels of trimethylamine（TMA）and trimethylamine N-oxide（TMAO）in the plasma.Flavin containing monooxygenase 3（FMO3）gene and protein expressions in liver were detected by RT-PCR and Western blot.Spearman correlation analysis was used to investigate the correlation between TMAO and the maximum aggregation rate of platelet AA and ADP.Metagenomic technology was used to detect the characteristics of gut microbiome in rats.

The ECG changes of rats in the ACS group and the TYACS group were consistent with ACS，and the serum lipid levels of rats in the TYACSgroup were consistent with the dietary imbalance of the syndrome of phlegm and blood stasis.HE staining showed that the myocardium of the CON group was arranged neatly and the cytoplasm was abundant.In the ACS group and the TYACSgroup，the inflammatory cell infiltration in the ischemic area，the loss of myocardial nucleus，the vacuolar degeneration of myocardial cells and the disorder of myocardial organization were observed.The maximum aggregation rate of platelet AA and ADPwere significantly increased in the ACS and TYACSgroups（P<0.01）.LC/MS showed that the levels of TMA and TMAO in ACS group and TMA in TYACS group were showed an upward trend，and the level of TMAO in TYACSgroup was significantly increased（P<0.001）.FMO3 mRNA and protein in liver of the ACS and TYACS groups were significantly increased（P<0.01）.Maximum aggregation rate of platelet AA and ADP were positively correlated with TMAO level.Metagenomic analysis revealed that the characteristics of gut microbiome in the ACSand TYACS groups were significantly changed.The species markers significantly different in the CON group was Bacteroidetes，in the ACSgroup were Firmicutes and Fusobacteria，in the TYACS group were Proteobacteria，Actinobacteria and Spirochaetes.KEGG enrichment analysis showed that the bacteria were most enriched in AA metabolism，riboflavin metabolism and other pathways in ACSwith syndrome of phlegm and blood stasis rats，and the differences were the most obvious.ACS rats were especially related to AA metabolism，insulin resistance，riboflavin metabolism and other pathways.

The gut microbiome of rats with ACS and ACSwith syndrome of phlegm and blood stasis had significant characteristic changes.The platelet aggregation process of model rats may be related to a variety of different gut microbiota driving the TMA/FMO3/TMAO pathway，participating in AA metabolism，riboflavin metabolism，etc.This finding may provide a theoretical basis for exploring new methods to improve platelet aggregation function and prevent ACSby regulating flora forest，and enrich the theory of “the heart is connected with the small intestine” in traditional Chinese medicine.