Background:
IDO1 (indoleamine 2,3-dioxygenase 1) is the rate-limiting enzyme for tryptophan metabolism. IDO1 malfunction is involved in the pathogenesis of atherosclerosis. Vascular smooth muscle cells (VSMCs) with an osteogenic phenotype promote calcification and features of plaque instability. However, it remains unclear whether aberrant IDO1-regulated tryptophan metabolism causes VSMCs osteogenic reprogramming and calcification.
Methods:
We generated global
Apoe
(
apolipoprotein E
) and
Ido1
double knockout mice, and
Apoe
knockout mice with specific deletion of IDO1 in VSMCs or macrophages. Arterial intimal calcification was evaluated by a Western diet–induced atherosclerotic calcification model.
Results:
Global deficiency of IDO1 boosted calcific lesion formation without sex bias in vivo. Conditional IDO1 loss of function in VSMCs rather than macrophages promoted calcific lesion development and the abundance of RUNX2 (runt-related transcription factor 2). In contrast, administration of kynurenine via intraperitoneal injection markedly delayed the progression of intimal calcification in parallel with decreased RUNX2 expression in both
Apoe
−/−
and
Apoe
−/−
Ido1
−/−
mice. We found that IDO1 deletion restrained RUNX2 from proteasomal degradation, which resulted in enhanced osteogenic reprogramming of VSMCs. Kynurenine administration downregulated RUNX2 in an aryl hydrocarbon receptor–dependent manner. Kynurenine acted as the endogenous ligand of aryl hydrocarbon receptor, controlled resultant interactions between cullin 4B and aryl hydrocarbon receptor to form an E3 ubiquitin ligase that bound with RUNX2, and subsequently promoted ubiquitin-mediated instability of RUNX2 in VSMCs. Serum samples from patients with coronary artery calcification had impaired IDO1 activity and decreased kynurenine catabolites compared with those without calcification.
Conclusions:
Kynurenine, an IDO1-mediated tryptophan metabolism main product, promotes RUNX2 ubiquitination and subsequently leads to its proteasomal degradation via an aryl hydrocarbon receptor–dependent nongenomic pathway. Insufficient kynurenine exerts the deleterious role of IDO1 ablation in promoting RUNX2-mediated VSMCs osteogenic reprogramming and calcification in vivo.
Gut microbial phenylalanine, tyrosine, and tryptophan metabolites are closely linked to various diseases. Monitoring the alterations of the related metabolites is vital to facilitate the understanding of pathophysiology of diseases. Herein, a rapid and sensitive assay based on LC–tandem mass spectrometry has been developed to analyze 20 gut microbial metabolites derived from phenylalanine, tyrosine, and tryptophan in rat serum, urine, and faeces. These microbial‐derived metabolites were separated on a phenyl‐hexyl column and simultaneously determined in a single run of 8 min. The detection limit for analytes ranged between 1.08 and 32.4 ng/mL. All calibration curves exhibited good linear relationships (R2 ≥ 0.9982). Intra‐ and inter‐assay precision values were below 15% and accuracies ranged from 85% to 115% for all analytes. The selectivity, matrix effect, and recovery of this method were all satisfactory. The validated method was successfully applied to characterize the alterations of these metabolites in type 2 diabetes mellitus rat. In general, the developed assay is suitable for high‐throughput monitoring of gut microbial phenylalanine, tyrosine, and tryptophan metabolites and provides a useful approach for exploring the mechanisms of microbial‐derived metabolites in diseases.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.