A cardiac extracellular matrix-based bilayer vascular graft with controlled microstructures for the reconstruction of small-diameter blood vessels

Despite recent progress, challenges with small-diameter vascular grafts, including mechanical strength, intimal hyperplasia, thrombosis, and poor endothelialization, remain unresolved. The present study reports a novel bilayer vascular graft designed to mimic the anatomical features of small-diameter blood vessels. The electrospun graft consists of a dense micro/nanofibrous inner layer of cardiac extracellular matrix (cECM), polycaprolactone (PCL) loaded with heparin (P-cECM-H), and a super porous and micro-fibrous PCL outer layer. Liquid chromatography-mass spectrometry (LC-MS/MS) proteome analysis of the cECM revealed that it is enriched with several bioactive proteins related to angiogenesis, wound regeneration, cell migration, etc. The porosities of the two layers are tailored according to endothelial and smooth muscle cell biology. The graft exhibited excellent mechanical properties, and the heparinized P-cECM inner layer improved hemocompatibility and anticoagulation efficacy. A significant increase in endothelial cell proliferation was noted in the P-cECM-H group after 7 days compared with the control group (p < 0.05). The bilayer graft maintained 100 % patency after 10 weeks of rat abdominal aorta implantation. Histological evaluation revealed smooth muscle cell infiltration inside the highly porous outer layer and neointima regeneration in the inner layer with a complete endothelial lining. RNA sequencing (RNA-Seq) analysis further confirmed smooth muscle formation and endothelial layer formation. The gene expression data also suggested that the hypoxia-inducible factor-1 (HIF-) and vascular endothelial growth factor (VEGF) signaling pathways are involved in endothelial layer remodeling. These promising results indicate that cECM could be a key material for vascular tissue regeneration.