Reconstruction of a hybrid vascular graft hierarchically layered with three cell types

Hybrid grafts with hierarchically incorporated endothelial cells (ECs), smooth muscle cells (SMCs), and fibroblasts (FCs), were developed, and the cellular behavior and ultrastructure of the regenerated extracellular matrix (ECM) of the grafts were investigated in vivo. Two models of hybrid vascular grafts were constructed on knitted Dacron grafts (internal diameter, 4 mm; length, 6 cm) in vitro. Model I grafts were hierarchically structured with a confluent monolayer of ECs, a middle layer of SMCs, and an outer layer of FCs. Model II consisted of a confluent EC monolayer and a homogeneously mixed layer composed of SMCs and FCs prelabeled with fluorescent lipids. The designed artificial ECM was a mixed gel of type I collagen and dermatan sulfate. Both grafts were implanted bilaterally in the carotid arteries of 11 dogs for up to 12 weeks without anticoagulant. Regardless of model type, patent grafts (n = 18 of 22 implanted grafts) were completely endothelialized. In model II grafts at 4 weeks, SMCs migrated to the subendothelial layer, whereas labeled FCs migrated to an outer layer. Observation of model I grafts at 12 weeks showed that most of the collagen fibers in the luminal surface layer were longitudinally oriented, but those in deeper layers tended to orient circumferentially, and that newly generated elastic tissues were composed of many concentric laminae. SMCs of both grafts at 12 weeks were predominantly of the contractile phenotype. Thus, hybrid grafts incorporating intimal-, medial-, and adventitial-type layers can enhance the process of arterial wall reconstruction.