Pathological Mechanism of “Phlegm, Blood stasis, Toxin” in a Rabbit Model of Carotid Atherosclerosis Based on Gut Microbiota-host Metabolism Interactions

Background

In Traditional Chinese Medicine (TCM) theory, "phlegm, blood stasis and toxin" are the pathogenesis of carotid atherosclerosis (CAS). The rabbit carotid atherosclerosis (CAS), which is induced by high-cholesterol diet combined with carotid artery balloon injury, is a classic model for studying CAS. Many studies indicate that gut microbiota and host metabolic disorders are involved in the pathogenesis of rabbit CAS. However, the TCM pathological features and syndromes of this classic rabbit CAS model have not been reported.

Objective

To explore the pathogenesis of the rabbit CAS model and its TCM syndrome types from the perspective of "phlegm, blood stasis, and toxin".

Methods

Twelve male New Zealand white rabbits were randomly divided into NC group and CAS group according to their body weight, followed by feeding of basic feed and a 1% high cholesterol diet, respectively. After two weeks, the rabbits in the CAS group underwent common carotid artery (CCA) balloon injury, while the rabbits in the NC group underwent only CCA separation without balloon injury. The two groups received differential feed postoperatively for six more weeks, after which, changes in lipids, hemorheology, inflammation, oxidative stress, and CAS phenotypes were analyzed. In addition, the colon contents and serum were collected for 16S rRNA sequencing and ¹H-NMR metabonomic analysis.

Results

The CAS rabbits were observed to form noticeable abnormalities in lipid metabolism and blood rheology, a sharp increase in oxidative stress levels, excessive release of inflammatory factors and apparent CAS plaque formation. Furthermore, 10 specific gut microbiota (such as Akkermansia muciniphila, Barnesiellaceae and Faecalibacterium) and 14 characteristic metabolites (such as trimethylamine oxide, acetic acid and L-carnitine) were identified in the CAS rabbits, which were significantly related to the CAS phenotypes. The pathway function analysis showed that the gut microbiota and its metabolites mainly affected cholesterol metabolism, energy metabolism, inflammation and oxidative stress.

Conclusion

The rabbit CAS model conforms to the “phlegm, blood stasis and toxin damage” theory. The gut microbiota and host metabolic disorders of the CAS rabbits interact and promote internal and external toxins, aggravating the progression of CAS. Our study provided experimental evidence for the application of this model in the TCM-based research of CAS.