M Maria Stekelenburg

Dynamic Straining Combined with Fibrin Gel Cell Seeding Improves Strength of Tissue-Engineered Small-Diameter Vascular Grafts

Vascular tissue engineering represents a promising approach for the development of living small-diameter vascular grafts that can be used for replacement therapy. The culture of strong human tissue-engineered (TE) vascular grafts has required long culture times, up to several months, whether or not combined with gene therapy. This article describes the culture of strong, genetically unmodified, human TE vascular grafts in 4 weeks Small-diameter vascular grafts were engineered using a fast-degrading polyglycolic acid scaffold coated with poly-4-hydroxybutyrate combined with fibrin gel and seeded with myofibroblasts isolated from discarded saphenous veins from patients undergoing coronary bypass surgery. The TE grafts were subjected to dynamic strain conditions. After 28 d of in vitro culture, the grafts demonstrated burst pressures of 903 +/- 123 mmHg. Comparison with native vessels (intact human left internal mammary arteries (LIMAs) and saphenous veins) showed no significant differences in the amount of DNA, whereas the TE vessels contained approximately 50% of the native collagen content. In the physiological pressure range, up to 300 mmHg, the mechanical properties of the TE vessels were comparable to the LIMA. In this study, we showed that dynamic conditioning combined with fibrin gel cell seeding enhances the mechanical properties of small-diameter TE grafts. These grafts might provide a promising alternative to currently used vascular replacements.

The influence of endothelial cells on the ECM composition of 3D engineered cardiovascular constructs

Tissue engineering of small diameter (<5 mm) blood vessels is a promising approach to develop viable alternatives for autologous vascular grafts. Development of a functional, adherent, shear resisting endothelial cell (EC) layer is one of the major issues limiting the successful application of these tissue engineered grafts. The goal of the present study was to create a confluent EC layer on a rectangular 3D cardiovascular construct using human venous cells and to determine the influence of this layer on the extracellular matrix composition and mechanical properties of the constructs. Rectangular cardiovascular constructs were created by seeding myofibroblasts (MFs) on poly(glycolic acid) poly‐4‐hydroxybutyrate scaffolds using fibrin gel. After 3 or 4 weeks, ECs were seeded and co‐cultured using EGM‐2 medium for 2 or 1 week, respectively. A confluent EC layer could be created and maintained for up to 2 weeks. The EGM‐2 medium lowered the collagen production by MFs, resulting in weaker constructs, especially in the 2 week cultured constructs. Co‐culturing with ECs slightly reduced the collagen content, but had no additional affect on the mechanical performance. A confluent endothelial layer was created on 3D human cardiovascular constructs. The layer was co‐cultured for 1 and 2 weeks. Although, the collagen production of the MFs was slightly lowered, co‐culturing ECs for 1 week results in constructs with good mechanical properties and a confluent EC layer. Copyright

Remodeling of the collagen fiber architecture due to compaction in small vessels under tissue engineered conditions

Mechanical loading protocols in tissue engineering (TE) aim to improve the deposition of a properly organized collagen fiber network. In addition to collagen remodeling, these conditioning protocols can result in tissue compaction. Tissue compaction is beneficial to tissue collagen alignment, yet it may lead to a loss of functionality of the TE construct due to changes in geometry after culture. Here, a mathematical model is presented to relate the changes in collagen architecture to the local compaction within a TE small blood vessel, assuming that under static conditions, compaction is the main factor responsible for collagen fiber organization. An existing structurally based model is extended to incorporate volumetric tissue compaction. Subsequently, the model is applied to describe the collagen architecture of TE constructs under either strain based or stress based stimulus functions. Our computations indicate that stress based simulations result in a helical collagen fiber distribution along the vessel wall. The helix pitch angle increases from a circumferential direction in the inner wall, over about 45 deg in the middle vessel layer, to a longitudinal direction in the outer wall. These results are consistent with experimental data from TE small diameter blood vessels. In addition, our results suggest a stress dependent remodeling of the collagen, suggesting that cell traction is responsible for collagen orientation. These findings may be of value to design improved mechanical conditioning protocols to optimize the collagen architecture in engineered tissues.

Living Heart Valve and Small-Diameter Artery Substitutes - An Emerging Field for Intellectual Property Development

Cardiovascular diseases, such as heart valve dysfunction and coronary artery stenosis, are next to cancer the leading cause of death in the US. Treatments involve replacement of the heart valve or bypassing the obstructed coronary artery with a small-diameter vascular graft. The major limitation of currently used replacements is their inability to grow, adapt and repair in the patient. Considering the increasing age of the population and the subsequent increase in cardiovascular disease incidence, efforts to improve existing replacements and unraveling novel types of replacements are of paramount importance. Cardiovascular tissue engineering represents a rapid evolving field of research, providing living heart valve and small-diameter vascular substitutes with the ability to grow, adapt and repair after implantation. Various tissue engineering approaches are being employed, based on in vivo and/or in vitro tissue formation. This review provides an overview of the current heart valve and small-diameter vascular replacements and presents the status and future developments within the various tissue engineering approaches. The potential of tissue engineering for the development of living heart valve and small-diameter vascular substitutes is reflected in the numerous patents related to this emerging field of research.

Remodeling and Compaction in Tissue Engineered Small Vessels

Small-diameter vessels tissue engineering (TE) seeks to provide viable replacements to the native ones. The approach involves seeding autologous cells onto a biodegradable scaffold material and delivering the suitable environmental factors (biochemical and biomechanical stimuli) during culture via a conditioning protocol. Still, these replacements appear to lack the native mechanical integrity. TE protocols should then be improved and optimized.Copyright

3D Coculture of Human Endothelial Cells and Myofibroblasts for Vascular Tissue Engineering

Cardiovascular disease is still the number one cause of death in the industrialized world. Diseased small diameter blood vessels are frequently replaced by native grafts. However, these vessels have a limited life time [1], for example the patency at 10 year after coronary artery bypass grafting of saphenous vein grafts is 57% [2]. Tissue engineering (TE) of small diameter blood vessels seems a promising approach to overcome these shortcomings or address the increasing need for substitutes during follow up surgery. Mechanical conditioning of myofibroblast (MFs) seeded constructs appears to be beneficial for functional tissue properties, such as cell proliferation, ECM production and mechanical strength [3,4]. Without a functional endothelial cell (ECs) layer however, patency may be compromised by thrombogenecity. Construction of an EC layer might on the other hand affect the tissue composition during culture, as was shown for bovine ECs, which influenced proliferation and ECM production of smooth muscle cells [5].Copyright