The fusion of tissue spheroids attached to pre-stretched electrospun polyurethane scaffolds.

Effective cell invasion into thick electrospun biomimetic scaffolds is an unsolved problem. One possible strategy to biofabricate tissue constructs of desirable thickness and material properties without the need for cell invasion is to use thin (<2 µm) porous electrospun meshes and self-assembling (capable of tissue fusion) tissue spheroids as building blocks. Pre-stretched electrospun meshes remained taut in cell culture and were able to support tissue spheroids with minimal deformation. We hypothesize that elastic electrospun scaffolds could be used as temporal support templates for rapid self-assembly of cell spheroids into higher order tissue structures, such as engineered vascular tissue. The aim of this study was to investigate how the attachment of tissue spheroids to pre-stretched polyurethane scaffolds may interfere with the tissue fusion process. Tissue spheroids attached, spread, and fused after being placed on pre-stretched polyurethane electrospun matrices and formed tissue constructs. Efforts to eliminate hole defects with fibrogenic tissue growth factor-ß resulted in the increased synthesis of collagen and periostin and a dramatic reduction in hole size and number. In control experiments, tissue spheroids fuse on a non-adhesive hydrogel and form continuous tissue constructs without holes. Our data demonstrate that tissue spheroids attached to thin stretched elastic electrospun scaffolds have an interrupted tissue fusion process. The resulting tissue-engineered construct phenotype is a direct outcome of the delicate balance of the competing physical forces operating during the tissue fusion process at the interface of the pre-stretched elastic scaffold and the attached tissue spheroids. We have shown that with appropriate treatments, this process can be modulated, and thus, a thin pre-stretched elastic polyurethane electrospun scaffold could serve as a supporting template for rapid biofabrication of thick tissue-engineered constructs without the need for cell invasion.

Age-related analysis of structural, biochemical and mechanical properties of the porcine mitral heart valve leaflets.

The aim of this study was to assess structural and biochemical differences in the extracellular matrix of the fetal and adult porcine mitral heart valves in relation to their mechanical characteristics. Using tensile tests it was demonstrated that the material properties of porcine mitral heart valves progressively change with age. The collagen content of the adult heart valve, as estimated by hydroxyproline assay, increases three times as compared with fetal heart valves. Transmission electron microscopy demonstrated that the diameter of collagen fibrils increased in adult heart valves compared with fetal heart valves. The level of collagen cross-linking is lower in the fetal heart valve than the adult heart valve. The reported age differences in the material properties of fetal and adult porcine heart valves were associated with increases in collagen content, the diameter of collagen fibrils and the level of collagen cross-linking. These data lay a foundation for systematic elucidation of the structural determinants of material properties of heart valves during embryonic and postnatal valvulogenesis. They are also essential to define the desirable level of tissue maturation in heart valve tissue engineering.

Comparison of radial deformability of stent posts of different aortic bioprostheses.

OBJECTIVES: Little is known about the stent deformability required for optimal stented heart valve bioprosthesis design. Therefore, two bioprosthetic valves with known good long-term clinical results were tested. The strain in the radial direction of the stent posts of these valves was compared with contemporary bioprosthetic valves and a native porcine aortic root. METHODS: Medtronic Intact and Carpentier-Edwards Standard (CES), and four contemporary bioprostheses, including one self-expanding prosthesis, were tested with three sonomicrometry probes per valve fixed at commissure attachment points. The mean values from 2400 data points from three measurements of the interprobe distances were used to calculate the radius of the circle circumscribed around the three probes. Changes in the radius of the aortic root at pressures 70-90 and 120-140 mmHg (pressure during diastole and systole) and that of the stent posts at 70-90 and 0-10 mmHg (transvalvular pressure gradient during diastole and systole) were compared. RESULTS: An increase in radius by 7.3 ± 2.6, 8.7 ± 0.0 and 3.9 ± 0.0% for the porcine aortic root, CES and Intact valves, respectively, was observed during transition from diastolic to systolic pressure and less for contemporary bioprostheses-mean 2.5 ± 0.9%, lowest 1.2 ± 0.0. CONCLUSIONS: The results indicate that the radial deformability of bioprosthetic valve stent posts can be as low as 1.2% for xenoaortic and 3.0% for xenopericardial prostheses with no compromise of valve durability. Although these results suggest that valve stent post-deformability might not be of critical importance, a concrete answer to the question of the significance of stent deformability for valve durability can be obtained only by acquiring long-term follow-up results for valve prostheses with rigid stents.

St Jude Epic heart valve bioprostheses versus native human and porcine aortic valves - comparison of mechanical properties.

OBJECTIVES: The major problem with heart valve bioprostheses made from chemically treated porcine aortic valves is their limited longevity caused by gradual deterioration, which has a causal link with valve tissue mechanical properties. To our best knowledge, there are no published studies on the mechanical properties of modern, commercially available bioprostheses comparing them to native human valves. The objective of this study is to determine the mechanical properties of St Jude Epic bioprostheses and to compare them with native human and porcine aortic valves. METHODS: Leaflets from eight porcine aortic valves and six Epic bioprostheses were analyzed using uni-axial tensile tests in radial and circumferential directions. Mechanical properties of human valves have been previously published by our group. Results are represented as mean values+/-S.D. RESULTS: Circumferential direction. Modulus of elasticity of Epic bioprostheses in circumferential direction at the level of stress 1.0 MPa is 101.99+/-58.24 MPa, 42.3+/-4.96 MPa for native porcine and 15.34+/-3.84 MPa for human aortic valves. Ultimate stress is highest for Epic bioprostheses 5.77+/-1.94 MPa, human valves have ultimate stress of 1.74+/-0.29 MPa and porcine 1.58+/-0.26 MPa. Ultimate strain in circumferential direction is highest for human valves 18.35+/-7.61% followed by 7.26+/-0.69% for porcine valves and 5.95+/-1.54% for Epic bioprostheses. Radial direction. Modulus of elasticity in radial direction is 9.18+/-1.81 MPa for Epic bioprostheses, 5.33+/-0.61 MPa for native porcine, and 1.98+/-0.15 MPa for human aortic valve leaflets. In the radial direction ultimate stress is highest for Epic bioprostheses 0.7+/-0.21 MPa followed by native porcine valves 0.55+/-0.11 MPa and 0.32+/-0.04 MPa for human valves. For human valves ultimate strain is 23.92+/-4.87%, for native porcine valves 8.57+/-0.8% and 7.92+/-1.74% for Epic bioprostheses. CONCLUSIONS: Epic bioprostheses have non-linear stress-strain behavior similar to native valve tissue, but they are significantly stiffer and hence less elastic compared to native porcine and human aortic valves. These differences in mechanical properties may cause variations in stress distribution within leaflets of the bioprosthetic valves and accelerate their deterioration.

Rapid biofabrication of tubular tissue constructs by centrifugal casting in a decellularized natural scaffold with laser-machined micropores.

Centrifugal casting allows rapid biofabrication of tubular tissue constructs by suspending living cells in an in situ cross-linkable hydrogel. We hypothesize that introduction of laser-machined micropores into a decellularized natural scaffold will facilitate cell seeding by centrifugal casting and increase hydrogel retention, without compromising the biomechanical properties of the scaffold. Micropores with diameters of 50, 100, and 200 mum were machined at different linear densities in decellularized small intestine submucosa (SIS) planar sheets and tubular SIS scaffolds using an argon laser. The ultimate stress and ultimate strain values for SIS sheets with laser-machined micropores with diameter 50 mum and distance between holes as low as 714 mum were not significantly different from unmachined control SIS specimens. Centrifugal casting of GFP-labeled cells suspended in an in situ cross-linkable hyaluronan-based hydrogel resulted in scaffold recellularization with a high density of viable cells inside the laser-machined micropores. Perfusion tests demonstrated the retention of the cells encapsulated within the HA hydrogel in the microholes. Thus, an SIS scaffold with appropriately sized microholes can be loaded with hydrogel encapsulated cells by centrifugal casting to give a mechanically robust construct that retains the cell-seeded hydrogel, permitting rapid biofabrication of tubular tissue construct in a "bioreactor-free" fashion.

Cardiovascular tissue engineering I. Perfusion bioreactors: a review.

Tissue engineering is a fast-evolving field of biomedical science and technology with future promise to manufacture living tissues and organs for replacement, repair, and regeneration of diseased organs. Owing to the specific role of hemodynamics in the development, maintenance, and functioning of the cardiovascular system, bioreactors are a fundamental of cardiovascular tissue engineering. The development of perfusion bioreactor technology for cardiovascular tissue engineering is a direct sequence of previous historic successes in extracorporeal circulation techniques. Bioreactors provide a fluidic environment for tissue engineered tissue and organs, and guarantee their viability, maturation, biomonitoring, testing, storage, and transportation. There are different types of bioreactors and they vary greatly in their size, complexity, and functional capabilities. Although progress in design and functional properties of perfusion bioreactors for tissue engineered blood vessels, heart valves, and myocardial patches is obvious, there are some challenges and insufficiently addressed issues, and room for bioreactor design improvement and performance optimization. These challenges include creating a triple perfusion bioreactor for vascularized tubular tissue engineered cardiac construct; designing and manufacturing fluidics-based perfused minibioreactors; incorporation of systematic mathematical modeling and computer simulation based on computational fluid dynamics into the bioreactor designing process; and development of automatic systems of hydrodynamic regime control. Designing and engineering of built-in noninvasive biomonitoring systems is another important challenge. The optimal and most efficient perfusion and conditioning regime, which accelerates tissue maturation of tissue-engineered constructs also remains to be determined. This is a first article in a series of reviews on critical elements of cardiovascular tissue engineering technology describing the current status, unsolved problems, and challenges of bioreactor technology in cardiovascular tissue engineering and outlining future trends and developments.

Tannic acid mimicking dendrimers as small intestine submucosa stabilizing nanomordants.

Chemical stabilization resulting in increased resistance to proteolytic degradation is one of the approaches in prevention of post-implantational aneurysm development in decellularized natural vascular scaffolds. Recently, tannic acid (TA) and tannic acid mimicking dendrimers (TAMD) have been suggested as potential stabilization agents for collagen and elastin. The aim of this work was to determine the stabilizing effects of TAMD on decellularized natural scaffolds. Vascular scaffolds fabricated from small intestine submucosa (SIS) and SIS plane sheets (Cook Biotech Inc.) were used. The biomechanical properties of the SIS vascular graft segments treated with TA and TAMD were tested. The effect of TAMD treatment on resistance to proteolytic degradation was evaluated by measuring biomechanical properties of TAMD stabilized and non-stabilized SIS specimens after incubation in collagenase solution. It was shown that treatment with TA as well as with TAMD increased the strength of tubular SIS as well as their resistance to proteolytic biodegradation manifested by preservation of biomechanical properties after collagenase treatment. Transmission electron microscopy demonstrated that treatment with TAMD increased the periodical pattern typical of collagen fiber ultrastructure as a result of the "mordant" effect. The possible collagen cross-linking effect of TAMD on SIS was investigated by differential scanning calorimetry (DSC). The treatment with TAMD induced a small, but detectable cross-linking effect, suggesting that TAMD do not establish extensive covalent cross links within the extracellular matrix but rather interact with collagen, thus rendering SIS scaffolds more resistant to proteolytic degradation.

Comparison of biomechanical and structural properties between human aortic and pulmonary valve.

OBJECTIVE: Pulmonary valve autografts have been reported as clinically effective for replacement of diseased aortic valve (Ross procedure). Published data about pulmonary valve mechanical and structural suitability as a long-term substitute for aortic valve are limited. The aim of this study was to compare aortic and pulmonary valve properties. METHODS: Experimental studies of biomechanical properties and structure of aortic and pulmonary valves were carried out on pathologically unchanged human heart valves, collected from 11 cadaveric hearts. Biomechanical properties of 84 specimens (all valve elements: cusps, fibrous ring, commissures, sinotubular junction, sinuses) were investigated using uniaxial tensile tests. Ultrastructure was studied using transmission and scanning electron microscopy. RESULTS: Ultimate stress in circumferential direction for pulmonary valve cusps is higher than for aortic valve (2.78+/-1.05 and 1.74+/-0.29 MPa, respectively). Ultimate stress in radial direction for pulmonary and aortic cusps is practically the same (0.29+/-0.06 and 0.32+/-0.04 MPa, respectively). In ultrastructural study, different layout and density in each construction element are determined. The aortic and pulmonary valves have common ultrastructural properties. CONCLUSIONS: Mechanical differences between aortic and pulmonary valve are minimal. Ultrastructural studies show that the aortic and pulmonary valves have similar structural elements and architecture. This investigation suggests that the pulmonary valve can be considered mechanically and structurally suitable for use as an aortic valve replacement.

Biomechanical properties of oesophagus wall under loading.

In this investigation, firstly, the biomechanical properties of different parts of oesophagus were determined. Oesophagus stress and strain are the greatest in the cervical part for all age groups. The human oesophagus deforms unevenly, depending on the direction of load in relation to the organ's axis, it exhibits anisotropical behaviour. With the age the values of mechanical parameters of the oesophagus wall reduce, in particular beginning from 45 years of age, but the modulus of elasticity increases. Biomechanical properties of the oesophagus depend on the architecture of its structure. By loading the organ in the circumferential direction, microfibrilae rupture and deformation of the muscular fibres occurs. With increase of load, collagenous fibres straighten and microruptures in collagenous fibrilae occur. With stretching of oesophagus longitudinally, collagenous fibres partially preserve their wavy and helical configuration. Therefore, higher resistance of the oesophageal wall occurs in the longitudinal direction.