On the shape and structure of the murine pulmonary heart valve.

Murine animal models are an established standard in translational research and provides a potential platform for studying heart valve disease. To date, studies on heart valve disease using murine models have been hindered by a lack of appropriate methodologies due to their small scale. In the present study, we developed a multi-scale, imaging-based approach to extract the functional structure and geometry for the murine heart valve. We chose the pulmonary valve (PV) to study, due to its importance in congenital heart valve disease. Excised pulmonary outflow tracts from eleven 1-year old C57BL/6J mice were fixed at 10, 20, and 30 mmHg to simulate physiological loading. Micro-computed tomography was used to reconstruct the 3D organ-level PV geometry, which was then spatially correlated with serial en-face scanning electron microscopy imaging to quantify local collagen fiber distributions. From the acquired volume renderings, we obtained the geometric descriptors of the murine PV under increasing transvalvular pressures, which demonstrated remarkable consistency. Results to date suggest that the preferred collagen orientation was predominantly in the circumferential direction, as in larger mammalian valves. The present study represents a first step in establishing organ-level murine models for the study of heart valve disease.

Age related extracellular matrix and interstitial cell phenotype in pulmonary valves.

Heart valve disease is a common manifestation of cardiovascular disease and is a significant cause of cardiovascular morbidity and mortality worldwide. The pulmonary valve (PV) is of primary concern because of its involvement in common congenital heart defects, and the PV is usually the site for prosthetic replacement following a Ross operation. Although effects of age on valve matrix components and mechanical properties for aortic and mitral valves have been studied, very little is known about the age-related alterations that occur in the PV. In this study, we isolated PV leaflets from porcine hearts in different age groups (~ 4-6 months, denoted as young versus ~ 2 years, denoted as adult) and studied the effects of age on PV leaflet thickness, extracellular matrix components, and mechanical properties. We also conducted proteomics and RNA sequencing to investigate the global changes of PV leaflets and passage zero PV interstitial cells in their protein and gene levels. We found that the size, thickness, elastic modulus, and ultimate stress in both the radial and circumferential directions and the collagen of PV leaflets increased from young to adult age, while the ultimate strain and amount of glycosaminoglycans decreased when age increased. Young and adult PV had both similar and distinct protein and gene expression patterns that are related to their inherent physiological properties. These findings are important for us to better understand the physiological microenvironments of PV leaflet and valve cells for correctively engineering age-specific heart valve tissues.

The interaction of laser radiation with tissue in the aspect of generating the process of decellularization in the preparation of animal origin autologous tissue.

PURPOSE: The aim of the work was to create an appropriate substrate for organ transplantation using bioactive tissue-based scaffold populated by cells of the graft recipient. The purpose of the modeling was to investigate the mechanical effects of wave loading of aortic and pulmonary tissue material. METHODS: The biological properties of tissues of aortic and pulmonary valves were modified by the process of decellularization. The host cells were removed by various physical methods with focus on minimal degradation of the extracellular matrix. Thus, the decellularization process was controlled by histological methods. The tissue decellularization process was simulated by finite element modelling. RESULTS: The mechanical results represented by a displacement at the center of the sample were coherent and the heterogeneity of the distribution of the caves on the surface of the samples was confirmed, both by experiment and in the simulation by the alternate occurrence of local minima and maxima. The latter results from the uneven removal of cells from the effect of the wave causing decellularization were also predicted by the numerical model. Laser radiation had a destructive effect on the components of the extracellular matrix (e.g., collagen and elastic fibers), mainly depending on the fluence and number of pulses in a single exposure. CONCLUSIONS: The differences between the valve tissue materials were shown, and the impact of the process of decellularization on the properties of the tissues was analyzed. It should be emphasized that due to low absorption and high scattering, laser radiation can deeply penetrate the tissue, which allows for effective decellularization process in the entire volume of irradiated tissue.

Analyzing valve interstitial cell mechanics and geometry with spatial statistics.

Understanding cell geometric and mechanical properties is crucial to understanding how cells sense and respond to their local environment. Moreover, changes to cell mechanical properties under varied micro-environmental conditions can both influence and indicate fundamental changes to cell behavior. Atomic Force Microscopy (AFM) is a well established, powerful tool to capture geometric and mechanical properties of cells. We have previously demonstrated substantial functional and behavioral differences between aortic and pulmonary valve interstitial cells (VIC) using AFM and subsequent models of VIC mechanical response. In the present work, we extend these studies by demonstrating that to best interpret the spatially distributed AFM data, the use of spatial statistics is required. Spatial statistics includes formal techniques to analyze spatially distributed data, and has been used successfully in the analysis of geographic data. Thus, spatially mapped AFM studies of cell geometry and mechanics are analogous to more traditional forms of geospatial data. We are able to compare the spatial autocorrelation of stiffness in aortic and pulmonary valve interstitial cells, and more accurately capture cell geometry from height recordings. Specifically, we showed that pulmonary valve interstitial cells display higher levels of spatial autocorrelation of stiffness than aortic valve interstitial cells. This suggests that aortic VICs form different stress fiber structures than their pulmonary counterparts, in addition to being more highly expressed and stiffer on average. Thus, the addition of spatial statistics can contribute to our fundamental understanding of the differences between cell types. Moving forward, we anticipate that this work will be meaningful to enhance direct analysis of experimental data and for constructing high fidelity computational of VICs and other cell models.

Rate-dependency of the mechanical behaviour of semilunar heart valves under biaxial deformation.

This paper presents an experimental investigation and evidence of rate-dependency in the planar mechanical behaviour of semilunar heart valves. Samples of porcine aortic and pulmonary valves were subjected to biaxial deformations across 1000-fold stretch rate, ranging from λ̇=0.001 to 1 s-1. The experimental campaign encompassed protocols covering (i) tests on samples without preconditioning, (ii) preconditioning immediately followed by tensile tests; and (iii) tensile tests at different rates performed on the same preconditioned specimen. Our results indicate that under all employed loading protocols, heart valve samples exhibit a marked rate-dependency in their deformation behaviour. This rate-dependency is reflected in stress-stretch curves and the calculated ensuing gradients, where samples typically show stiffening with increased rate. These results underpin one conclusive outcome: the in-plane mechanical behaviour of semilunar valves is rate-dependent (p<0.05 for Cauchy stress levels ≥50 kPa). This outcome implies that the rate of deformation for characterising the mechanical behaviour of semilunar heart valves may not be chosen arbitrarily low, and models that incorporate rate-effects may be more appropriate for better capturing the mechanical behaviour of heart valves. STATEMENT OF SIGNIFICANCE: This study presents for the first time a comprehensive set of results and evidence of rate-dependency in the mechanical behaviour of semilunar heart valves under biaxial deformation. Our results challenge the widely-applied assumption in the bulk of the existing literature, where an implicit rate-independency is assumed in both experimental and modelling propositions related to the biomechanics of the aortic and pulmonary valves. This study therefore creates a solid platform for future research in heart valve biomechanics with two important implications. First, experimental campaigns have to be carried out at high stretch rates; ideally as close to the physiological rate as possible. Second, new continuum/computational models are required to address the rate-dependent mechanical behaviour of the semilunar valves.

A geometric model for the human pulmonary valve in its fully open case.

The human pulmonary valve, one of the key cardiac structures, plays an important role in circulatory system. However, there are few mathematical models to accurately simulate it. In this paper, we establish a geometric model of the normal human pulmonary valve from a mathematical perspective in the fully opening case. Based on the statistical data of the human pulmonary valves, we assume that the motions of the three cusps are symmetrical in the cardiac cycle. Thus, we first propose that each cusp is a part of the cylindrical shell according to its structure and physiological feature. The parameters for the pulmonary valve cusps in three-dimensional space are obtained by the fitting functions. We verify the accuracy of our results by comparing the areas of the pulmonary valve and pulmonary valve flap.

Computational modeling guides tissue-engineered heart valve design for long-term in vivo performance in a translational sheep model.

Valvular heart disease is a major cause of morbidity and mortality worldwide. Current heart valve prostheses have considerable clinical limitations due to their artificial, nonliving nature without regenerative capacity. To overcome these limitations, heart valve tissue engineering (TE) aiming to develop living, native-like heart valves with self-repair, remodeling, and regeneration capacity has been suggested as next-generation technology. A major roadblock to clinically relevant, safe, and robust TE solutions has been the high complexity and variability inherent to bioengineering approaches that rely on cell-driven tissue remodeling. For heart valve TE, this has limited long-term performance in vivo because of uncontrolled tissue remodeling phenomena, such as valve leaflet shortening, which often translates into valve failure regardless of the bioengineering methodology used to develop the implant. We tested the hypothesis that integration of a computationally inspired heart valve design into our TE methodologies could guide tissue remodeling toward long-term functionality in tissue-engineered heart valves (TEHVs). In a clinically and regulatory relevant sheep model, TEHVs implanted as pulmonary valve replacements using minimally invasive techniques were monitored for 1 year via multimodal in vivo imaging and comprehensive tissue remodeling assessments. TEHVs exhibited good preserved long-term in vivo performance and remodeling comparable to native heart valves, as predicted by and consistent with computational modeling. TEHV failure could be predicted for nonphysiological pressure loading. Beyond previous studies, this work suggests the relevance of an integrated in silico, in vitro, and in vivo bioengineering approach as a basis for the safe and efficient clinical translation of TEHVs.

In vitro biomechanical and hydrodynamic characterisation of decellularised human pulmonary and aortic roots.

BACKGROUND AND PURPOSE OF THE STUDY: The use of decellularised biological heart valves in the replacement of damaged heart valves offers a promising solution to reduce the degradation issues associated with existing cryopreserved allografts. The purpose of this study was to assess the effect of low concentration sodium dodecyl sulphate decellularisation on the in vitro biomechanical and hydrodynamic properties of cryopreserved human aortic and pulmonary roots. METHOD: The biomechanical and hydrodynamic properties of cryopreserved decellularised human aortic and pulmonary roots were fully characterised and compared to cellular human aortic and pulmonary roots in an unpaired study. Following review of these results, a further study was performed to investigate the influence of a specific processing step during the decellularisation protocol ('scraping') in a paired comparison, and to improve the method of the closed valve competency test by incorporating a more physiological boundary condition. RESULTS: The majority of the biomechanical and hydrodynamic characteristics of the decellularised aortic and pulmonary roots were similar compared to their cellular counterparts. However, several differences were noted, particularly in the functional biomechanical parameters of the pulmonary roots. However, in the subsequent paired comparison of pulmonary roots with and without decellularisation, and when a more appropriate physiological test model was used, the functional biomechanical parameters for the decellularised pulmonary roots were similar to the cellular roots. CONCLUSION: Overall, the results demonstrated that the decellularised roots would be a potential choice for clinical application in heart valve replacement.

In vivo performance of freeze-dried decellularized pulmonary heart valve allo- and xenografts orthotopically implanted into juvenile sheep.

The decellularization of biological tissues decreases immunogenicity, allows repopulation with cells, and may lead to improved long-term performance after implantation. Freeze drying these tissues would ensure off-the-shelf availability, save storage costs, and facilitates easy transport. This study evaluates the in vivo performance of freeze-dried decellularized heart valves in juvenile sheep. TritonX-100 and sodium dodecylsulfate decellularized ovine and porcine pulmonary valves (PV) were freeze-dried in a lyoprotectant sucrose solution. After rehydration for 24 h, valves were implanted into the PV position in sheep as allografts (fdOPV) and xenografts (fdPPV), while fresh dezellularized ovine grafts (frOPV) were implanted as controls. Functional assessment was performed by transesophageal echocardiography at implantation and at explantation six months later. Explanted grafts were analysed histologically to assess the matrix, and immunofluorescence stains were used to identify the repopulating cells. Although the graft diameters and orifice areas increased, good function was maintained, except for one insufficient, strongly deteriorated frOPV. Cells which were positive for either endothelial or interstitial markers were found in all grafts. In fdPPV, immune-reactive cells were also found. Our findings suggest that freeze-drying does not alter the early hemodynamic performance and repopulation potential of decellularized grafts in vivo, even in the challenging xenogeneic situation. Despite evidence of an immunological reaction for the xenogenic valves, good early functionalities were achieved. STATEMENT OF SIGNIFICANCE: Decellularized allogeneic heart valves show excellent results as evident from large animal experiments and clinical trials. However, a long-term storing method is needed for an optimal use of this limited resource in the clinical setting, where an optimized matching of graft and recipient is requested. As demonstrated in this study, freeze-dried and freshly decellularized grafts reveal equally good results after implantation in the juvenile sheep concerning function and repopulation with recipients' cells. Thus, freeze-drying arises as a promising method to extend the shelf-life of valvular grafts compared to those stored in antibiotic-solution as currently practised.

Decellularization of human donor aortic and pulmonary valved conduits using low concentration sodium dodecyl sulfate.

The clinical use of decellularized cardiac valve allografts is increasing. Long-term data will be required to determine whether they outperform conventional cryopreserved allografts. Valves decellularized using different processes may show varied long-term outcomes. It is therefore important to understand the effects of specific decellularization technologies on the characteristics of donor heart valves. Human cryopreserved aortic and pulmonary valved conduits were decellularized using hypotonic buffer, 0.1% (w/v) sodium dodecyl sulfate and nuclease digestion. The decellularized tissues were compared to cellular cryopreserved valve tissues using histology, immunohistochemistry, quantitation of total deoxyribose nucleic acid, collagen and glycosaminoglycan content, in vitro cytotoxicity assays, uniaxial tensile testing and subcutaneous implantation in mice. The decellularized tissues showed no histological evidence of cells or cell remnants and >97% deoxyribose nucleic acid removal in all regions (arterial wall, muscle, leaflet and junction). The decellularized tissues retained collagen IV and von Willebrand factor staining with some loss of fibronectin, laminin and chondroitin sulfate staining. There was an absence of major histocompatibility complex Class I staining in decellularized pulmonary valve tissues, with only residual staining in isolated areas of decellularized aortic valve tissues. The collagen content of the tissues was not decreased following decellularization however the glycosaminoglycan content was reduced. Only moderate changes in the maximum load to failure of the tissues were recorded postdecellularization. The decellularized tissues were noncytotoxic in vitro, and were biocompatible in vivo in a mouse subcutaneous implant model. The decellularization process will now be translated into a good manufacturing practices-compatible process for donor cryopreserved valves with a view to future clinical use. Copyright © 2016 The Authors Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

Implantation of a modified stented bovine pulmonary valve in a beating heart sheep model.

PURPOSE: The aim of this research was to assess the performance of a modified bovine stent valve implanted transventricularly in the pulmonary position in sheep with a 3-month follow-up period. MATERIALS AND METHODS: Seven modified pulmonary bovine stent valves were transventricularly implanted in the pulmonary position into seven sheep using a delivery system. Stent valve performance was investigated and evaluated hemodynamically, angiographically, and with echocardiograms before, immediately after, and 3 months following implantation. Macroscopic, histologic, and radiographic examinations were performed on the explanted graft at 3 months. RESULTS: The modified stent valves were all deployed and implanted successfully in the pulmonary position in seven sheep. Angiographic, echocardiographic, hemodynamic, and macroscopic analyses confirmed firm anchoring of the stents in the target position in the early and 3-month follow-up period. All modified stent valves showed satisfactory function, except one moderate stenosis (32 mmHg gradient) with mild regurgitation that was discovered at 3 months. All seven valves were free of any calcification and thrombus formation at postmortem macroscopic examination, which was confirmed by histologic and radiographic examination. All stents were intact without any fracture at microscopic or radiographic examination. CONCLUSIONS: Transventricular implantation of a modified nitinol pulmonary valve stent showed good structural and functional outcomes without stent fracture or migration.

The composition and biomechanical properties of human cryopreserved aortas, pulmonary trunks, and aortic and pulmonary cusps.

Human cryopreserved allografts of pulmonary and aortic heart valves, aortas and pulmonary trunks are used for valve replacement. However, it is unknown how the composition of these allografts relate to their mechanical properties. Our aims were to correlate the histological compositions and passive mechanical properties of aortic and pulmonary valves and to observe the microcracks of aortas and pulmonary trunks. The following parameters were quantified: ultimate stress; ultimate strain; Young's modulus of elasticity; valve cusp wall thickness; pulmonary and aortic intima-media thickness; area fraction of elastin, collagen and calcification; and length density of elastic fibres. The propagation of experimentally induced microcracks avoided elastic fibres. Ultimate strain was negatively correlated with the area fraction of calcification (r=-0.4) in aortas. Ultimate stress (r=0.27) and Young's modulus in small deformation (r=0.29) and in large deformation (r=0.32) correlated with wall thickness in valve cusps. Young's modulus (r=0.34) and ultimate strain (r=0.31) correlated with intima-media thickness. Ultimate strain correlated with the area fraction of elastin (r=-0.40) and collagen in the arteries (r=0.31). As conventional histology does not fully explain the mechanical properties of cryopreserved grafts, both morphological and biomechanical tests should be used complementarily when characterizing the ageing of the grafts.

Combined Bone Marrow-Derived Mesenchymal Stromal Cell Therapy and One-Way Endobronchial Valve Placement in Patients with Pulmonary Emphysema: A Phase I Clinical Trial.

One-way endobronchial valves (EBV) insertion to reduce pulmonary air trapping has been used as therapy for chronic obstructive pulmonary disease (COPD) patients. However, local inflammation may result and can contribute to worsening of clinical status in these patients. We hypothesized that combined EBV insertion and intrabronchial administration of mesenchymal stromal cells (MSCs) would decrease the inflammatory process, thus mitigating EBV complications in severe COPD patients. This initial study sought to investigate the safety of this approach. For this purpose, a phase I, prospective, patient-blinded, randomized, placebo-controlled design was used. Heterogeneous advanced emphysema (Global Initiative for Chronic Lung Disease [GOLD] III or IV) patients randomly received either allogeneic bone marrow-derived MSCs (108 cells, EBV+MSC) or 0.9% saline solution (EBV) (n = 5 per group), bronchoscopically, just before insertion of one-way EBVs. Patients were evaluated 1, 7, 30, and 90 days after therapy. All patients completed the study protocol and 90-day follow-up. MSC delivery did not result in acute administration-related toxicity, serious adverse events, or death. No significant between-group differences were observed in overall number of adverse events, frequency of COPD exacerbations, or worsening of disease. Additionally, there were no significant differences in blood tests, lung function, or radiological outcomes. However, quality-of-life indicators were higher in EBV + MSC compared with EBV. EBV + MSC patients presented decreased levels of circulating C-reactive protein at 30 and 90 days, as well as BODE (Body mass index, airway Obstruction, Dyspnea, and Exercise index) and MMRC (Modified Medical Research Council) scores. Thus, combined use of EBV and MSCs appears to be safe in patients with severe COPD, providing a basis for subsequent investigations using MSCs as concomitant therapy. Stem Cells Translational Medicine 2017;6:962-969.

Effects of combined cryopreservation and decellularization on the biomechanical, structural and biochemical properties of porcine pulmonary heart valves.

UNLABELLED: Non-fixed, decellularized allogeneic heart valve scaffolds seem to be the best choice for heart valve replacement, their availability, however, is quite limited. Cryopreservation could prolong their shelf-life, allowing for their ideal match to a recipient. In this study, porcine pulmonary valves were decellularized using detergents, either prior or after cryopreservation, and analyzed. Mechanical integrity was analyzed by uniaxial tensile testing, histoarchitecture by histological staining, and composition by DNA, collagen (hydroxyproline) and GAG (chondroitin sulfate) quantification. Residual sodium dodecyl sulfate (SDS) in the scaffold was quantified by applying a methylene blue activation assay (MBAS). Cryopreserved decellularized scaffolds (DC) and scaffolds that were decellularized after cryopreservation (CD) were compared to fresh valves (F), cryopreserved native valves (C), and decellularized only scaffolds (D). The E-modulus and tensile strength of decellularized (D) tissue showed no significant difference compared to DC and CD. The decellularization resulted in an overall reduction of DNA and GAG, with DC containing the lowest amount of GAGs. The DNA content in the valvular wall of the CD group was higher than in the D and DC groups. CD valves showed slightly more residual SDS than DC valves, which might be harmful to recipient cells. In conclusion, cryopreservation after decellularization was shown to be preferable over cryopreservation before decellularization. However, in vivo testing would be necessary to determine whether these differences are significant in biocompatibility or immunogenicity of the scaffolds. STATEMENT OF SIGNIFICANCE: Absence of adverse effects on biomechanical stability of acellular heart valve grafts by cryopreservation, neither before nor after decellularization, allows the identification of best matching patients in a less time pressure dictated process, and therefore to an optimized use of a very limited, but best-suited heart valve prosthesis.

Lesión en el lóbulo inferior derecho pulmonar por catéter Shaldon / Right lower pulmonary lobe injury due to a Shaldon catheter

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Engineering based assessment for a shape design of a pediatric ePTFE pulmonary conduit valve.

The authors examined the hemodynamic characteristics of expanded polytetrafluoroethylene (ePTFE) pulmonary valved conduits quantitatively by our originally developed pediatric pulmonary mechanical circulatory system, in order to suggest the optimal shape design. The system consisted of pneumatically driven right atrium and ventricle model, a pulmonary valve chamber, and elastic pulmonary compliance model with peripheral vascular resistance units, a venous reservoir. We employed two different types of ePTFE valve and evaluated the relationship between the leaflets motion and hemodynamic characteristics by using a high-speed video camera. As a result, we successfully reproduced hemodynamic simulations in our pediatric pulmonary mock system. We confirmed that the presence of bulging sinuses in the pulmonary valved conduit reduced the transvalvular energy loss and increased the valve opening area during systolic period. Our engineering-based in vitro analysis could be useful for proposing a shape design optimization of sophisticated pediatric ePTFE pulmonary valve.

Material modeling of cardiac valve tissue: Experiments, constitutive analysis and numerical investigation.

A key element of the cardiac cycle of the human heart is the opening and closing of the four valves. However, the material properties of the leaflet tissues, which fundamentally contribute to determine the mechanical response of the valves, are still an open field of research. The main contribution of the present study is to provide a complete experimental data set for porcine heart valve samples spanning all valve and leaflet types under tensile loading. The tests show a fair degree of reproducibility and are clearly indicative of a number of fundamental tissue properties, including a progressively stiffening response with increasing elongation. We then propose a simple anisotropic constitutive model, which is fitted to the experimental data set, showing a reasonable interspecimen variability. Furthermore, we present a dynamic finite element analysis of the aortic valve to show the direct usability of the obtained material parameters in computational simulations.

On the bending properties of porcine mitral, tricuspid, aortic, and pulmonary valve leaflets.

The atrioventricular valve leaflets (mitral and tricuspid) are different from the semilunar valve leaflets (aortic and pulmonary) in layered structure, ultrastructural constitution and organization, and leaflet thickness. These differences warrant a comparative look at the bending properties of the four types of leaflets. We found that the moment-curvature relationships in atrioventricular valves were stiffer than in semilunar valves, and the moment-curvature relationships of the left-side valve leaflets were stiffer than their morphological analog of the right side. These trends were supported by the moment-curvature curves and the flexural rigidity analysis (EI value decreased from mitral, tricuspid, aortic, to pulmonary leaflets). However, after taking away the geometric effect (moment of inertia I), the instantaneous effective bending modulus E showed a reversed trend. The overall trend of flexural rigidity (EI: mitral > tricuspid > aortic > pulmonary) might be correlated with the thickness variations among the four types of leaflets (thickness: mitral > tricuspid > aortic > pulmonary). The overall trend of the instantaneous effective bending modulus (E: mitral < tricuspid < aortic < pulmonary) might be correlated to the layered fibrous ultrastructures of the four types of leaflets, of which the fibers in mitral and tricuspid leaflets were less aligned, and the fibers in aortic and pulmonary leaflets were highly aligned. We also found that, for all types of leaflets, moment-curvature relationships are stiffer in against-curvature (AC) bending than in with-curvature bending (WC), which implies that leaflets tend to flex toward their natural curvature and comply with blood flow. Lastly, we observed that the leaflets were stiffer in circumferential bending compared with radial bending, likely reflecting the physiological motion of the leaflets, i.e., more bending moment and movement were experienced in radial direction than circumferential direction.

Hemodynamic Characterization of a Mouse Model for Investigating the Cellular and Molecular Mechanisms of Neotissue Formation in Tissue-Engineered Heart Valves.

Decellularized allograft heart valves have been used as tissue-engineered heart valve (TEHV) scaffolds with promising results; however, little is known about the cellular mechanisms underlying TEHV neotissue formation. To better understand this phenomenon, we developed a murine model of decellularized pulmonary heart valve transplantation using a hemodynamically unloaded heart transplant model. Furthermore, because the hemodynamics of blood flow through a heart valve may influence morphology and subsequent function, we describe a modified loaded heterotopic heart transplant model that led to an increase in blood flow through the pulmonary valve. We report host cell infiltration and endothelialization of implanted decellularized pulmonary valves (dPV) and provide an experimental approach for the study of TEHVs using mouse models.