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Smooth muscle cell spheroids as 3D model of phenotypic plasticity and matrix deposition revealed by 2D–3D proteomics

'Our findings show that VSMC 3D culture promotes the transition from contractile to synthetic VSMC phenotypes with ECM deposition, providing a translational in vitro system to investigate VSMC-mediated repair mechanisms and vascular remodeling in cardiovascular disease's.

Jul 28, 2026

WORC Update, Elvira Weber & Julia Hesse

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Abstract

Vascular smooth muscle cells (VSMCs) are regulators of vascular homeostasis and play a role in cardiovascular diseases, including atherosclerosis and aortic aneurysms. Together with endothelial cells (ECs), they control vascular remodeling by producing extracellular matrix (ECM), regulating the expression of adhesion molecules, and releasing signaling factors. Since changes in the ECM composition critically determine disease progression, models that reproduce authentic cell–ECM interactions are indispensable. Two-dimensional (2D) cultures fail to capture the mechanical integration of vascular cells within their ECM environment. We established a scaffold-free three-dimensional (3D) spheroid model of murine and human aortic VSMCs derived from heart failure patients to mimic their multicellular organization, cell plasticity, and interactions with the ECM. Using mass spectrometry, we provided to our knowledge the first comprehensive proteomic profiling of relevant 3D-cultured VSMCs in comparison to conventional 2D culture. VSMCs formed compact spheroids with a protein signature indicative of a synthetic phenotype, characterized by active ECM organization, adhesion, and energy homeostasis pathways. We demonstrated that the spheroid protocol can also be used to generate EC spheroids, offering opportunities to study VSMC-EC crosstalk. Our findings show that VSMC 3D culture promotes the transition from contractile to synthetic VSMC phenotypes with ECM deposition, providing a translational in vitro system to investigate VSMC-mediated repair mechanisms and vascular remodeling in cardiovascular diseases.

Hesse, J., Resch, U., Görtz, L. et al. Smooth muscle cell spheroids as 3D model of phenotypic plasticity and matrix deposition revealed by 2D–3D proteomics. Sci Rep 16, 23357 (2026). https://doi.org/10.1038/s41598-026-61834-7

Acknowledgements

We want to thank Susanne Pfeiler, Sarah Piel, and Norbert Gerdes (Division of Cardiology, Pulmonology and Vascular Medicine, Medical Faculty and University Hospital, Heinrich Heine University Düsseldorf, Germany), Katharina Scherschel (Institute of Neural and Sensory Physiology, Medical Faculty and University Hospital Düsseldorf, Heinrich Heine University Düsseldorf, Germany) as well as Maria Grandoch (Institute for Translational Pharmacology, Heinrich Heine University, 40225, Düsseldorf, Germany) for kindly providing support, materials and equipment.

Funding

Open Access funding enabled and organized by Projekt DEAL. This study was supported by “Christiane + Claudia Hempel-Stiftung für regenerative Medizin” to E.W. and H.A. Funding for this study was provided by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) project number 397484323 (TRR259/C04) to H.A.; and TRR259/B09, 468236352, and 384170921 (FOR2722/C01) to G.S.

Hug Aubin and Elvira Weber equally contributed to this work.

Authors and Affiliations

Research Group 3D Cardiovascular Regenerative Medicine and Tissue Engineering (CURE 3D), Department of Cardiac Surgery, Medical Faculty and University Hospital Düsseldorf, Heinrich Heine University Düsseldorf, Düsseldorf, Germany

Julia Hesse, Ulrike Resch, Lotte Görtz, Sarah Lang, Roya Batool, Sarah Saradar, Patricia Schoof, Kevin Gehlweiler, Daniela Zouikova, Pandora Jashnieh, Ricardo Fernandes Velosa, Nino Hotzel-Hacker, Fumitaka Suzuki, Samet Bayraktar, Melanie Cappallo, Vera Schmidt, Lara Ebbert, Margret H. Bülow, Nicole Kucharowski, Artur Lichtenberg, Hug Aubin & Elvira Weber
Department of Molecular Cardiology, Medical Faculty and University Hospital Düsseldorf, Heinrich Heine University Düsseldorf, Düsseldorf, Germany

Julia Hesse
CARID, Cardiovascular Research Institute Düsseldorf, Medical Faculty and University Hospital Düsseldorf, Heinrich Heine University Düsseldorf, Düsseldorf, Germany

Julia Hesse, Artur Lichtenberg, Hug Aubin & Elvira Weber
Centre of Physiology and Pharmacology, Department of Vascular Biology and Thrombosis Research, Medical University of Vienna, Vienna, Austria

Ulrike Resch
Gerontology Research Centre, Faculty of Sport and Health Sciences, University of Jyväskylä, Jyväskylä, Finland

Eija K. Laakkonen
Cologne Excellence Cluster on Cellular Stress Responses in Ageing-Associated Diseases (CECAD), University of Cologne, Cologne, Germany

Marcus Krüger & Gerhard Sengle
Department of Pediatrics and Adolescent Medicine, Faculty of Medicine and University Hospital Cologne, University of Cologne, Cologne, Germany

Gerhard Sengle
Center for Biochemistry, Faculty of Medicine, University Hospital of Cologne, Cologne, Germany

Gerhard Sengle
Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany

Gerhard Sengle
Cologne Center for Musculoskeletal Biomechanics (CCMB), Cologne, Germany

Gerhard Sengle
Medical Faculty, Institute for Clinical Biochemistry and Pathobiochemistry, German Diabetes Center, Leibniz Center for Diabetes Research at Heinrich Heine University Düsseldorf, Düsseldorf, Germany

Alexandra Chadt
German Center for Diabetes Research (DZD), Neuherberg, Munich, Germany

Alexandra Chadt

Contributions

J. Hesse: Conceptualization, formal analysis, data curation, visualization, writing—original draft, writing—review and editing. U. Resch: Investigation, methodology, data curation, writing—original draft, writing—review and editing. L. Goertz: Investigation, methodology. S. Lang: Investigation, methodology. R. Batool: Investigation. S. Saradar: Investigation, methodology. P. Schoof: Investigation, methodology. K. Gehlweiler: Investigation. D. Zouikova: Investigation. P. Jashnieh: Investigation. R. Fernandes Velosa: Investigation. N. Hotzel-Hacker: Writing-original draft. F. Suzuki: Writing-review and editing; data curation. S. Bayraktar: Formal analysis, data curation, writing—review and editing. M. Cappallo: Writing—review and editing. V. Schmidt: Investigation, writing—original draft, writing—review and editing. L. Ebbert: Investigation. M. Bülow: Investigation, writing—review and editing. N. Kucharowski: Investigation, writing—review and editing. E. Laakkonen: Conceptualization, writing—review and editing, M. Krüger: Conceptualization, methodology. G. Sengle: Writing-original draft; writing—review and editing; conzeptualisation. A. Chadt: Writing—review and editing. A. Lichtenberg: Resources, funding acquisition, writing—review and editing. H. Aubin: Conceptualization, resources, supervision, writing—review and editing, project administration. E. Weber: Methodology, investigation, writing—original draft, writing—review and editing, conceptualization, formal analysis, data curation, supervision, project administration.