Generation of a Physiological Three-Layered Bioartificial Blood Vessel by Mechanical Stimulation

 

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Objectives: Vascular grafts are widely used in vascular and cardiac surgery. However, limited availability of autologous vessels and insufficient biocompatibility of synthetic grafts represent major limitations. Tissue engineered bioartificial grafts are a promising approach to overcome these problems. To correctly emulate the morphology and function of natural vessels, the three layers of the vascular wall, namely Tunica intima, media and adventitia, must be properly recreated in bioartificial vessels. This requires not only integrating the specific cell types for each layer, but also demands accurate cell alignment, morphology and phenotype specific for each layer. Therefore, sophisticated bioreactor techniques enabling physiological mechanical stimulation are of pivotal importance.

Methods: Three layered bioartificial vessels were generated based on fibrin matrices in a stepwise molding technique. For the Tunica media, adipogenic stem cells (ASC) were differentiated to smooth muscle cells (SMC) and integrated in a compacted tubular fibrin matrix. Subsequently, the Tunica adventitia containing human umbilical vein endothelial cells (HUVEC) and ASC in a low concentration fibrin matrix was molded around it. Finally, luminal seeding with HUVEC created the Tunica intima. The bioartificial vessels were exposed to physiological mechanical stimulation in a custom built bioreactor or incubated statically for 72 hours prior to immunofluorescence microscopy analysis.

Result: Mechanical stimulation under physiological conditions induced physiological cell alignment in each layer: Endothelial cells of the Tunica intima showed confluent coverage and longitudinal alignment. Cells in the Media showed circumferential alignment and expressed the SMC-markers αSMA and Calponin. HUVEC in the Adventitia formed tubes resembling the Vasa vasorum capillaries that were aligned in longitudinal direction. In the statically incubated controls, random orientation of the cells in each layer was observed.

Conclusion: To our knowledge, this is the first description of a completely cellularized three-layered bioartificial vessel including a capillarized adventitial layer generated in vitro. Fibrin proved to be an ideal matrix material for all three layers of the vascular wall. Most importantly, physiological mechanical stimulation induced accurate alignment and morphology in each layer and thus is pivotal for a physiological vessel wall architecture of bioartificial vascular grafts.

Publication History

Article published online:
19 February 2021

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