Proteoglycan 4 Modulates Osteogenic Smooth Muscle Cell Differentiation during Vascular Remodeling and Intimal Calcification

Cells, vol. 10, pp. 1276

Abstract

Calcification is a prominent feature of late-stage atherosclerosis, but the mechanisms driving this process are unclear. Using a biobank of carotid endarterectomies, we recently showed that Proteoglycan 4 (PRG4) is a key molecular signature of calcified plaques, expressed in smooth muscle cell (SMC) rich regions. Here, we aimed to unravel the PRG4 role in vascular remodeling and intimal calcification. PRG4 expression in human carotid endarterectomies correlated with calcification assessed by preoperative computed tomographies. PRG4 localized to SMCs in early intimal thickening, while in advanced lesions it was found in the extracellular matrix, surrounding macro-calcifications. In experimental models, Prg4 was upregulated in SMCs from partially ligated ApoE−/− mice and rat carotid intimal hyperplasia, correlating with osteogenic markers and TGFb1. Furthermore, PRG4 was enriched in cells positive for chondrogenic marker SOX9 and around plaque calcifications in ApoE−/− mice on warfarin. In vitro, PRG4 was induced in SMCs by IFNg, TGFb1 and calcifying medium, while SMC markers were repressed under calcifying conditions. Silencing experiments showed that PRG4 expression was driven by transcription factors SMAD3 and SOX9. Functionally, the addition of recombinant human PRG4 increased ectopic SMC calcification, while arresting cell migration and proliferation. Mechanistically, it suppressed endogenous PRG4, SMAD3 and SOX9, and restored SMC markers’ expression. PRG4 modulates SMC function and osteogenic phenotype during intimal remodeling and macro-calcification in response to TGFb1 signaling, SMAD3 and SOX9 activation. The effects of PRG4 on SMC phenotype and calcification suggest its role in atherosclerotic plaque stability, warranting further investigations.

Authors 19

1. Till Seime

   Karolinska Institutet

   Affiliation as printed

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

2. Asim Cengiz Akbulut

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, CARIM, Maastricht University, 6229 ER Maastricht, The Netherlands

3. Moritz Lindquist Liljeqvist

   Karolinska Institutet

   Affiliation as printed

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

4. Antti Siika

   Karolinska Institutet

   Affiliation as printed

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

5. Hong Hua Jin

   Karolinska Institutet

   Affiliation as printed

   Department of Medicine, Karolinska Institutet, 17164 Stockholm, Sweden

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

6. Greg Winski

   Karolinska Institutet

   Affiliation as printed

   Department of Medicine, Karolinska Institutet, 17164 Stockholm, Sweden

7. Rick H. van Gorp

   Maastricht University

   Affiliation as printed

   Department of Biochemistry, CARIM, Maastricht University, 6229 ER Maastricht, The Netherlands

8. Eva Karlöf

   Karolinska Institutet

   Affiliation as printed

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

9. Mariette Lengquist

   Karolinska Institutet

   Affiliation as printed

   Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

10. Andrew J. Buckler

    Karolinska Institutet

    Affiliation as printed

    Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

11. Malin Kronqvist

    Karolinska Institutet

    Affiliation as printed

    Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

12. Olivia J. Waring

    Maastricht University Medical Centre · Maastricht University

    Affiliation as printed

    Department of Pathology, CARIM, Maastricht University Medical Center, 6200 MD Maastricht, The Netherlands

13. Jan H.N. Lindeman Aachen Department of Surgery, Leiden University Medical Center

    Leiden University Medical Center

    Affiliation as printed

    Department of Surgery, Leiden University Medical Center, 2300 RC Leiden, The Netherlands

14. Erik A.L. Biessen

    Maastricht University Medical Centre · Maastricht University

    Affiliation as printed

    Department of Pathology, CARIM, Maastricht University Medical Center, 6200 MD Maastricht, The Netherlands

15. Lars Mäegdefessel

    TUM Klinikum · Karolinska Institutet · Technical University of Munich

    Affiliation as printed

    Department for Vascular and Endovascular Surgery, Klinikum rechts der Isar, Technische Universität München, 81679 Munich, Germany

    Department of Medicine, Karolinska Institutet, 17164 Stockholm, Sweden

16. Anton Razuvaev

    Karolinska Institutet

    Affiliation as printed

    Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

17. Leon J. Schurgers Aachen Institute of Experimental Medicine and Systems Biology

    Maastricht University · RWTH Aachen University

    Affiliation as printed

    Department of Biochemistry, CARIM, Maastricht University, 6229 ER Maastricht, The Netherlands

    Institute of Experimental Medicine and Systems Biology, RWTH Aachen University, 52062 Aachen, Germany

18. Ulf Hedin

    Karolinska Institutet

    Affiliation as printed

    Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden

19. Ljubica Perisic Matic corresponding

    Karolinska Institutet

    Affiliation as printed

    Vascular Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, 17164 Stockholm, Sweden