Background/Purpose: Trimethylamine (TMA) generated by the gut microbiome is converted into trimethylamine N-oxide (TMAO) via the host enzyme FMO3. The TMA-FMO3-TMAO metaorganismal axis has been linked to chronic metabolic, renal, and cardiovascular diseases, and triggers myofibroblast reprogramming in mesenchymal cell types^1,2. However, despite growing evidence implicating gut dysbiosis in SSc³, the expression and pathogenic role in FMO3 in SSc has never been investigated.

Methods: The TMA-derived metabolite TMAO was measured in the circulation by LC tandem mass spectrometry in 200 adults with SSc, and 100 age-matched controls undergoing elective cardiac evaluation. FMO3 was evaluated in skin biopsies and fibroblasts via single cell RNA-seq (scRNA), immunohistochemistry and immunofluorescence. To investigate the pathogenic role of the TMAO-producing enzyme FMO3, C57BL/6 mice engineered with liver-specific gain-of-function of human FMO3 (hFMO3-TG)⁴ were used in models of fibrosis and inflammation.

Results: Serum TMAO was elevated in SSc patients compared to controls (median 3.31 µM [IQR: 2.18, 5.23] vs. 2.70 µM [IQR: 1.74, 4.11]; p=0.016), with highest levels among obese and male SSc patients. Remarkably, while FMO3 is expressed in the liver, we also detected FMO3 in skin. SSc patient-derived biopsies and explanted fibroblasts showed elevated FMO3 protein and mRNA expression colocalizing with collagen production. scRNA-seq of the skin showed the highest expression of FMO3 localized to CLDN1⁺ fibroblasts, a cellular subpopulation seen primarily in SSc biopsies and associated with extracellular matrix remodeling. Male mice engineered to express human FMO3 (hFMO3-TG) show a significant increase in circulating TMAO⁴. Bleomycin treatment caused significantly exaggerated fibro-inflammatory pathology in hFMO3-TG mice compared to identically treated wildtype mice.

Conclusion: Collectively, our results demonstrate that gut microbiome-derived vasculopathic/profibrotic TMAO is elevated in patients with SSc, and transgenic mice expressing human FMO3, the enzyme that generates TMAO, develop augmented fibrotic responses associated with elevated TMAO. This is the first study to identify a potential pathogenic role for FMO3 in SSc, linking gut dysbiosis to fibrosis and vasculopathy via a metaorganismal axis.

REFERENCES
1. Brown et al., 2017. Targeting of microbe-derived metabolites to improve human health: the next frontier for drug discovery. Journal of biological chemistry, 292(21), pp.8560-8568.
2. Kim et al., 2022. Gut microbe-derived metabolite trimethylamine N-oxide activates PERK to drive fibrogenic mesenchymal differentiation. Iscience, 25(7), p.104669.
3. Tan et al., 2023. Gut microbiome profiling in systemic sclerosis: a metagenomic approach. Clinical and Experimental Rheumatology.
4. Zhu et al., 2018. Flavin monooxygenase 3, the host hepatic enzyme in the metaorganismal trimethylamine N‐oxide‐generating pathway, modulates platelet responsiveness and thrombosis risk. Journal of Thrombosis and Haemostasis, 16(9), pp.1857-1872.

« Back to ACR Convergence 2023

ACR Meeting Abstracts - https://acrabstracts.org/abstract/flavin-containing-monooxygenase-fmo3-links-the-gut-and-fibrosis-in-systemic-sclerosis/