MSACL 2025 Abstract

Imidazole Propionate as a New Player in Atherosclerosis

Annalaura Mastrangelo (1), Iñaki Robles-Vera (1), Diego Mañanes (1), Miguel Galán (1), Marcos Femenía-Muiña (1), Rafael Barrero-Rodríguez (1), Eleftheria Papaioannou (2), Joaquín Amores-Iniesta (1), Ana Devesa (1), Manuel Lobo-González (1), Alba Carreras (3), Katharina R. Beck (3), Sophie Ivarsson (3), Georgios Georgiopoulos (4,5,6), Anders Gummesson (3,7), Manuel Rodrigo-Tapias (1,8), Sarai Martínez-Cano (1,8), Ivan Fernández-López (1), Alessia Ferrarini (1), Naohiro Inohara (9), Kimon Stamatelopoulos (4,10), Alberto Benguría (1), Danay Cibrian (1,11,12), Francisco Sánchez-Madrid (1,11,12), Ana Dopazo (1,11), Coral Barbas (13), Jesús Vázquez (1,12), Juan Antonio López (1,12), Alicia González-Martín (2), Gabriel Nuñez (9), Göran Bergström (3,14), Konstantinos Stellos (10,15), Fredrik Bäckhed (3,14), Valentín Fuster (1,16), Borja Ibañez (1,12,17), David Sancho (1)
(1) Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), Madrid, Spain. (2) Instituto de Investigaciones Biomédicas Sols-Morreale, Madrid, Spain. (3) University of Gothenburg, Gothenburg, Sweden. (4) National and Kapodistrian University of Athens, Athens, Greece. (5) King's College, London, UK. (6) University of Patras, Achaia, Greece. (7) Sahlgrenska University Hospital, Gothenburg, Sweden. (8) Inmunotek S.L., Alcalá de Henares, Spain. (9) University of Michigan, Ann Arbor, MI. (10) Newcastle University, Newcastle Upon Tyne, UK. (11) Hospital Universitario de La Princesa, Madrid, Spain. (12) CIBER de enfermedades cardiovasculares, Madrid, Spain, (13) Universidad San Pablo-CEU, Madrid, Spain. (14) Sahlgrenska University Hospital, Gothenburg, Sweden, (15) Heidelberg University, Mannheim, Germany. (16) The Zena and Michael A. Wiener Cardiovascular Institute, New York, NY. (17) Hospital Universitario Fundación Jiménez Díaz, Madrid, Spain.

Annalaura Mastrangelo, Five-year pharmacy degree and a doctorate in medicinal chemistry (Presenter) Spanish National Centre for Cardiovascular Research (CNIC)

Presenter Bio: Dr. Annalaura Mastrangelo holds a First-Class Honours M.Sc. in Pharmacy from the University of Modena and Reggio Emilia (2011) and an International PhD in Medicinal Chemistry from the University San Pablo CEU (2017).During her PhD at the Metabolomics and Bioanalysis Excellence Centre (CEMBIO), she gained expertise in metabolomics, analytical chemistry (MS, chromatography), and data science, focusing on obesity and its complications. In 2017, she joined the Spanish National Center for Cardiovascular Research (CNIC) as a postdoctoral fellow, investigating the role of metabolites in atherosclerosis and the immune response.

Dr. Mastrangelo is the author of a patent (PCT/EP2024/066950), one book chapter, and 17 publications in high-impact international journals (JCR), including five as first author and one as last author. Her work has been cited over 1309 times (H index=14, Scopus and Google Scholar). She has presented at 10 international conferences and participated in various communication and outreach activities. Her research focuses on applying metabolomics to develop personalized treatments and diagnostic tools for chronic diseases.

Relevant Financial Disclosures (within past 24 months, reported on Mar 26, 2025) No relevant financial relationship(s) to disclose.

INTRODUCTION:
Atherosclerosis is a complex multifactorial disease whose slow and asymptomatic development poses a challenge for its early detection and treatment. Current prevention strategies rely on traditional cardiovascular (CV) risk factor-based scores1, which fail to identify individuals at risk at early stages2. Recent research has questioned the notion of these traditional risk factors, emphasizing the need for alternative targets to improve therapy. This is particularly relevant for those subjects that suffer from CV events despite receiving appropriate lipid-lowering treatments3. Although alternative treatments are available, they are not yet established for atherosclerosis and are generally bound to established risk factors, emphasizing the need for alternative treatments to be explored. Accumulating evidence suggests the microbiota-host metabolism crosstalk as a contributor to CV disease (CVD)4. However, only a few gut microbiota-dependent metabolites, which correlate with late stages of the disease, have been described 5-7.

OBJECTIVE:
We aimed to identify microbial metabolites that could contribute to atherosclerosis progression, potentially serving as early biomarker of the disease and alternative targets for therapy.

METHODS:
Our discovery strategy employed untargeted metabolomics to analyse plasma samples from an apolipoprotein E deficient mouse (ApoE-/- mice). High-cholesterol diets and broad-spectrum antibiotics were used to shape the microbiota. Upon identifying imidazole propionate (ImP) as a microbial metabolite linked to atherosclerosis in mice, we evaluated its relevance in humans using targeted metabolomics. Metabolites of the ImP biosynthetic pathway were quantified in two cohorts: a discovery cohort (PESA-CNIC, N=400) and a validation cohort (N=1844) by a LC-MS validated method. 16S rRNA gene sequencing was also performed to assess microbiota composition and its relationship with ImP in both animal and human models. Additionally, correlation and logistic regression analyses were performed to explore the connection between plasma ImP and cardiovascular risk factors. Next, the role of ImP in the disease was studied by supplementing the metabolite in the drinking water of atheroprone murine models (ApoE-/- and LDLr-/- mice) fed a chow diet. Single-cell RNA sequencing was employed to examine the local effects of ImP supplementation in the aorta. Bulk RNA sequencing and phosphoproteomics were used to assess the metabolic effects of ImP in target cells following in vitro stimulation. Finally, in vitro cell assays identified the receptor through which ImP is sensed, and in vivo studies were conducted to assess the effects of its antagonist on the induction and progression of atherosclerosis.

RESULTS:
We discovered that microbially produced ImP is associated with, and causal for, atherosclerosis. Specifically, ImP is linked to atherosclerosis in mice and in two independent human cohorts, and induces atherosclerosis without altering the cholesterol levels in the bloodstream. We found that ImP-induced atherosclerosis was associated with increased inflammation and activation of both the innate and adaptive immune response, both systemically and locally in the aorta. We further identified the Imidazoline-1 receptor (I1R) as the receptor through which ImP mediates its effect. Notably, the blockade of the ImP/I1R axis inhibited the development of atherosclerosis induced both by ImP and high cholesterol diet in mice. This effect is independent of changes in cholesterol concentration but dependent on mTOR inhibition in macrophages.

CONCLUSION: or DISCUSSION:
The use of complementary omics techniques, along with in vitro and in vivo models, has allowed us to identify a novel role for the microbially produced ImP, previously linked to type 2 diabetes, cardiometabolic disease in people with HIV, heart failure and all-causes mortality 6,8-10. Our study goes beyond these associations, providing evidence of ImP’s causal role in atherosclerosis and demonstrating the blocking of its sensing axis as a therapeutic target for the disease. This showcases ImP as a new element in CVD management, both as biomarker for primary prevention and as therapeutic target for personalized medicine.

REFERENCES:
1. SCORE2 working group and ESC Cardiovascular risk collaboration. SCORE2 risk prediction algorithms: new models to estimate 10-year risk of cardiovascular disease in Europe. Eur. Heart J., 2021. 42(25): p. 2439–2454.
2. Fernández-Friera, L. et al. Prevalence, vascular distribution, and multiterritorial extent of subclinical atherosclerosis in a middle-aged cohort. Circulation, 2015. 131(24): p. 2104–2113.
3. Kaasenbrood, L. et al. Distribution of estimated 10-year risk of recurrent vascular events and residual risk in a secondary prevention population. Circulation, 2016. 134(19): p. 1419–1429.
4. Witkowski, M., Weeks, T. L. & Hazen, S. L. Gut microbiota and cardiovascular disease. Circ. Res., 2020. 127(4): p. 553–570.
5. Nemet, I. et al. A cardiovascular disease-linked gut microbial metabolite acts via adrenergic receptors. Cell, 2020. 180(5): p. 862-877.
6. Molinaro, A. et al. Microbially produced imidazole propionate is associated with heart failure and mortality. JACC Heart Fail., 2023. 11(7): p.810-821.
7. Zhu, W. et al. Gut microbial metabolite TMAO enhances platelet hyperreactivity and thrombosis risk. Cell, 2016. 165(1): p. 111–124.
8. Koh, A. et al. Microbially produced imidazole propionate impairs insulin signaling through mTORC1. Cell, 2018. 175(4): p. 947-961.
9. Wang, Z. et al. Gut microbiota, circulating inflammatory markers and metabolites, and carotid artery atherosclerosis in HIV infection. Microbiome, 2023. 11(1): p. 119.
10. Trøseid, M. et al. Gut microbiota alterations and circulating imidazole propionate levels are associated with obstructive coronary artery disease in people with HIV. J. Infect. Dis., 2024. 229(3): p.898-907.