The saphenous vein is the conduit of choice in bypass graft procedures. Unfortunately, haemodynamic stress associated with the arterial environment of the bypass vein graft leads to the development of intimal hyperplasia (IH), an excessive cell growth and collagen deposition that results in restenosis and secondary occlusion of the graft. IH processes involve endothelial cells injury, inflammation, vascular smooth muscle cells (VSMC) dysfunction and phenotypic modulation and extracellular matrix deposition. However, there are potential mechanisms that are still unaddressed. Here, we aim to establish an ex vivo model of graft IH by static human vein culture.

Discarded segments of great saphenous vein from patients undergoing lower limb bypass surgery were cut in 5 mm segments and randomly immediately preserved (D0) or maintained in cell culture for 7 days (D7). Immunohistochemical staining and molecular analyses were performed to identify the regulatory pathways involved in neointima formation.

As assessed by VGEL staining, veins placed in cell culture developed IH. Contractile markers (MYH11, MYOCD, Calponin) and the anti-apoptotic marker (Bcl2) were significantly downregulated, while the pro-apoptosis marker (Bax) and proliferative marker (PCNA) were upregulated in vein culture (D7) as compared to D0 in vitro. Furthermore, vein culture (D7) resulted in overexpression of matrix remodeling markers (MMP2, MMP9, Cathepsin K), inflammatory cytokines (IL6, IL1β, CCL5, CXCL2), profibrotic VSMC markers (YAP, HSP47) and osteochondrogenic VSMC markers (RunX2, SOX9, Osteopontin).

We have successfully established an in vitro model of graft IH. Our results show that human vein culture recreates key features of VSMC apoptosis, dedifferentiation, proliferation, extracellular matrix deposition and a phenotypic switch from contractile to profibrotic and osteochondrogenic VSMC in the development of IH. This novel model allows analysis of vascular remodeling and reprogramming of cells and the understanding of mechanisms underlying the dysregulated function of VSMC in IH.