Interfering in the ALK1 Pathway Results in Macrophage-Driven Outward Remodeling of Murine Vein Grafts

Alwin de Jong; Vincent Q Sier; Hendrika A B Peters; Natalia K M Schilder; J Wouter Jukema; Marie José T H Goumans; Paul H A Quax; Margreet R de Vries

doi:10.3389/fcvm.2021.784980

Interfering in the ALK1 Pathway Results in Macrophage-Driven Outward Remodeling of Murine Vein Grafts

Front Cardiovasc Med. 2022 Feb 3:8:784980. doi: 10.3389/fcvm.2021.784980. eCollection 2021.

Authors

Alwin de Jong^{1

2}, Vincent Q Sier^{1

2}, Hendrika A B Peters^{1

2}, Natalia K M Schilder^{1

2}, J Wouter Jukema³, Marie José T H Goumans⁴, Paul H A Quax^{1

2}, Margreet R de Vries^{1

2}

Affiliations

¹ Department of Surgery, Leiden University Medical Center, Leiden, Netherlands.
² Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.
³ Department of Cardiology, Leiden University Medical Center, Leiden, Netherlands.
⁴ Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, Netherlands.

Abstract

Aims: Vein grafts are frequently used to bypass coronary artery occlusions. Unfortunately, vein graft disease (VGD) causes impaired patency rates. ALK1 mediates signaling by TGF-β via TGFβR2 or BMP9/10 via BMPR2, which is an important pathway in fibrotic, inflammatory, and angiogenic processes in vascular diseases. The role of the TGF-β pathway in VGD is previously reported, however, the contribution of ALK1 signaling is not known. Therefore, we investigated ALK1 signaling in VGD in a mouse model for vein graft disease using either genetic or pharmacological inhibition of the Alk1 signaling.

Methods and results: Male ALK1 heterozygous (ALK1^+/-), control C57BL/6, as well as hypercholesterolemic ApoE3^*Leiden mice, underwent vein graft surgery. Histologic analyses of ALK1^+/- vein grafts demonstrated increased outward remodeling and macrophage accumulation after 28 days. In hypercholesterolemic ApoE3^*Leiden mice receiving weekly ALK1-Fc injections, ultrasound imaging showed 3-fold increased outward remodeling compared to controls treated with control-Fc, which was confirmed histologically. Moreover, ALK1-Fc treatment reduced collagen and smooth muscle cell accumulation, increased macrophages by 1.5-fold, and resulted in more plaque dissections. No difference was observed in intraplaque neovessel density. Flow cytometric analysis showed increased systemic levels of Ly6C^High monocytes in ALK1-Fc treated mice, supported by in vitro increased MCP-1 and IL-6 production of LPS-stimulated and ALK1-Fc-treated murine monocytes and macrophages.

Conclusion: Reduced ALK1 signaling in VGD promotes outward remodeling, increases macrophage influx, and promotes an unstable plaque phenotype.

Translational perspective: Vein graft disease (VGD) severely hampers patency rates of vein grafts, necessitating research of key disease-driving pathways like TGF-β. The three-dimensional nature of VGD together with the multitude of disease driving factors ask for a comprehensive approach. Here, we combined in vivo ultrasound imaging, histological analyses, and conventional in vitro analyses, identifying the ambiguous role of reduced ALK1 signaling in vein graft disease. Reduced ALK1 signaling promotes outward remodeling, increases macrophage influx, and promotes an unstable plaque phenotype in murine vein grafts. Characterization of in vivo vascular remodeling over time is imperative to monitor VGD development and identify new therapies.

Keywords: ALK1 signaling; macrophage; ultrasound; vascular remodeling; vein graft disease.