Mechanical modeling of the maturation process for tissue-engineered implants: Application to biohybrid heart valves.

The development of tissue-engineered cardiovascular implants can improve the lives of large segments of our society who suffer from cardiovascular diseases. Regenerative tissues are fabricated using a process called tissue maturation. Furthermore, it is highly challenging to produce cardiovascular regenerative implants with sufficient mechanical strength to withstand the loading conditions within the human body. Therefore, biohybrid implants for which the regenerative tissue is reinforced by standard reinforcement material (e.g. textile or 3d printed scaffold) can be an interesting solution. In silico models can significantly contribute to characterizing, designing, and optimizing biohybrid implants. The first step towards this goal is to develop a computational model for the maturation process of tissue-engineered implants. This paper focuses on the mechanical modeling of textile-reinforced tissue-engineered cardiovascular implants. First, an energy-based approach is proposed to compute the collagen evolution during the maturation process. Then, the concept of structural tensors is applied to model the anisotropic behavior of the extracellular matrix and the textile scaffold. Next, the newly developed material model is embedded into a special solid-shell finite element formulation with reduced integration. Finally, our framework is used to compute two structural problems: a pressurized shell construct and a tubular-shaped heart valve. The results show the ability of the model to predict collagen growth in response to the boundary conditions applied during the maturation process. Consequently, the model can predict the implant's mechanical response, such as the deformation and stresses of the implant.

3D printed pH-responsive tablets containing N-acetylglucosamine-loaded methylcellulose hydrogel for colon drug delivery applications.

The pH-responsive drug release approach in combination with three-dimensional (3D) printing for colon-specific oral drug administration can address the limitations of current treatments such as orally administered solid tablets. Such existing treatments fail to effectively deliver the right drug dosage to the colon. In order to achieve targeted drug release profiles, this work aimed at designing and producing 3D printed tablet shells using Eudragit® FS100 and polylactic acid (PLA) where the core was filled with 100 µl of N-acetylglucosamine (GlcNAc)-loaded methyl cellulose (MC) hydrogel. To meet the requirements of such tablets, the effects of polymer blending ratios and MC concentrations on physical, thermal, and material properties of various components of the tablets and most importantly in vitro drug release kinetics were investigated. The tablets with 80/20 wt% of Eudragit® FS100/PLA and the drug-loaded hydrogel with 30 mg/ml GlcNAc and 3% w/v MC showed the most promising results having the best printability, processability, and drug release kinetics besides being non-cytotoxic. Manufacturing of these tablets will be the first milestone in shifting from the conventional "one size fits all" approach to personalized medicine where different dosages and various combinations of drugs can be effectively delivered to the inflammation site.

Transformative Materials for Interfacial Drug Delivery.

Drug delivery systems (DDS) are designed to temporally and spatially control drug availability and activity. They assist in improving the balance between on-target therapeutic efficacy and off-target toxic side effects. DDS aid in overcoming biological barriers encountered by drug molecules upon applying them via various routes of administration. They are furthermore increasingly explored for modulating the interface between implanted (bio)medical materials and host tissue. Herein, an overview of the biological barriers and host-material interfaces encountered by DDS upon oral, intravenous, and local administration is provided, and material engineering advances at different time and space scales to exemplify how current and future DDS can contribute to improved disease treatment are highlighted.

Bioactive tri-component nanofibers from cellulose acetate/lignin//N-vanillidene-phenylthiazole copper-(II) complex for potential diaper dermatitis control.

Current research targets innovative medical textiles of nanofibrous nature and antibacterial activity to prevent diaper dermatitis. The work is based on electrospun nanofibers from cellulose acetate (CA) and lignin (Lig) polymers. A series of new copper complexes were synthesized and loaded to the CA/Lig solution mix then subjected to electrospinning, giving rise to the tricomponent bioactive mats CA/Lig/Cu-complex. The surface morphology of electrospun nanofiber mats was smooth and homogenous as the concentration of lignin increased in the mixture. The incorporation of lignin improved the electrospinnability of the cellulose acetate; however, it increased the fiber diameter. The water contact angle, absorption underload were significantly improved as lignin content increased. The incorporation of Cu-complex in electrospun CA and CA/Lig fiber mats occurred without any substantial change in the surface morphology, indicating well encapsulation of the complex. The electrospun mats were active against Pseudomonas aeruginosa, Acinetobacter baumannii, Staphylococcus epidermidis, and Streptococcus faecalis. The cytotoxicity, protein leakage, and biological results, together with the above studies, would advocate copper complex loaded CA/Lig nanofibers as a potential candidate for hygienic applications.

A potential future Fontan modification: preliminary in vitro data of a pressure-generating tube from engineered heart tissue.

OBJECTIVES: Univentricular malformations are severe cardiac lesions with limited therapeutic options and a poor long-term outcome. The staged surgical palliation (Fontan principle) results in a circulation in which venous return is conducted to the pulmonary arteries via passive laminar flow. We aimed to generate a contractile subpulmonary neo-ventricle from engineered heart tissue (EHT) to drive pulmonary flow actively. METHODS: A three-dimensional tubular EHT (1.8-cm length, 6-mm inner diameter, ca. 1-mm wall thickness) was created by casting human-induced pluripotent stem cell-derived cardiomyocytes (0.9 ml, 18 mio/ml) embedded in a fibrin-based hydrogel around a silicone tube. EHTs were cultured under continuous, pulsatile flow through the silicone tube for 23 days. RESULTS: The constructs started to beat macroscopically at days 8-14 and remained stable in size and shape over the whole culture period. Tubular EHTs showed a coherent beating pattern after 23 days in culture, and isovolumetric pressure measurements demonstrated a coherent pulsatile wave formation with an average frequency of 77 ± 5 beats/min and an average pressure of 0.2 mmHg. Histological analysis revealed cardiomyocytes mainly localized along the inner and outer curvature of the tubular wall with mainly longitudinal alignment. Cell density in the center of the tubular wall was lower. CONCLUSIONS: A simple tube-shaped contractile EHT was generated from human-induced pluripotent stem cells and developed a synchronous beating pattern. Further steps need to focus on optimizing support materials, flow rates and geometry to obtain a construct that creates sufficient pressures to support a directed and pulsatile blood flow.

Patient-Specific 3-Dimensional Model of Smooth Muscle Cell and Extracellular Matrix Dysfunction for the Study of Aortic Aneurysms.

INTRODUCTION: Abdominal aortic aneurysms (AAAs) are associated with overall high mortality in case of rupture. Since the pathophysiology is unclear, no adequate pharmacological therapy exists. Smooth muscle cells (SMCs) dysfunction and extracellular matrix (ECM) degradation have been proposed as underlying causes. We investigated SMC spatial organization and SMC-ECM interactions in our novel 3-dimensional (3D) vascular model. We validated our model for future use by comparing it to existing 2-dimensional (2D) cell culture. Our model can be used for translational studies of SMC and their role in AAA pathophysiology. MATERIALS AND METHODS: SMC isolated from the medial layer of were the aortic wall of controls and AAA patients seeded on electrospun poly-lactide-co-glycolide scaffolds and cultured for 5 weeks, after which endothelial cells (EC) are added. Cell morphology, orientation, mechanical properties and ECM production were quantified for validation and comparison between controls and patients. RESULTS: We show that cultured SMC proliferate into multiple layers after 5 weeks in culture and produce ECM proteins, mimicking their behavior in the medial aortic layer. EC attach to multilayered SMC, mimicking layer interactions. The novel SMC model exhibits viscoelastic properties comparable to biological vessels; cytoskeletal organization increases during the 5 weeks in culture; increased cytoskeletal alignment and decreased ECM production indicate different organization of AAA patients' cells compared with control. CONCLUSION: We present a valuable preclinical model of AAA constructed with patient specific cells with applications in both translational research and therapeutic developments. We observed SMC spatial reorganization in a time course of 5 weeks in our robust, patient-specific model of SMC-EC organization and ECM production.

Degradation resistance of PVDF mesh in vivo in comparison to PP mesh.

Mesh implant has been applied in hernia repair and urogynecological reconstruction. Polypropylene (PP) is now the most widely used material for non-resorbable mesh implants. A degradation phenomenon of PP mesh, which is apparent on the mesh surface as cracking, flaking and peeling, was discovered in the 1990's. This phenomenon of mesh implant has drawn attention because of mesh-related litigations. Polyvinylidene fluoride (PVDF), due to its high biocompatible performance, has been used since 2003 as an alternative material for non-resorbable mesh implants. Till now, no such degradation phenomenon of PVDF mesh has been reported, although limited study on PVDF mesh is available. In this paper, we researched the degradation of PVDF meshes taking the degradation of PP mesh as a reference. The meshes analysed in this study were received from a previous animal experiment. To expose the surface of explanted meshes, a tissue removing method with protease was used and the result of this cleaning process was tested by X-ray Photoelectron Spectroscopy (XPS). The morphological condition of the mesh surface was compared using Scanning Electron Microscopy (SEM) and the chemical condition concerning degradation was analysed through Fourier Transform Infrared Spectroscopy (FTIR). The surface condition of PVDF mesh after 3-, 6-, 12- and 24-month implantation was illustrated and compared with two types of PP meshes. XPS revealed an absence of nitrogen, confirming the successful removal of tissue residues using protease. SEM results presented no notable morphological surface change of the PVDF mesh and progressive surface cracking processes over time of both types of PP meshes. FTIR spectra of the implanted PVDF meshes had no considerable difference from the spectrum of the pristine mesh, while FTIR spectra of both types of PP meshes had extra chemical functional groups (carbonyl (CO) and hydroxyl (-OH) groups) increasing with implantation time, indicating progressive degradation. This study highlights the morphological and chemical stability of the PVDF mesh and demonstrates that the PVDF mesh is more resistant to degradation in comparison to the other two types of PP meshes.

Development of in vitro endothelialized drug-eluting stent using human peripheral blood-derived endothelial progenitor cells.

We propose in vitro endothelialization of drug-eluting stents (DES) to overcome late stent thrombosis by directly introducing late-outgrowth human endothelial progenitor cells (EPCs) at the target site utilizing abluminal DES. Isolated EPCs were confirmed as late-outgrowth EPCs by flow cytometric analysis. Abluminally paclitaxel-loaded stents were seeded with different cell concentrations and durations to determine optimal seeding conditions, in both uncrimped and crimped configurations. The seeding yield was determined by evaluating the percent coverage of the stent struts' area. The EPC-seeded DES were exposed to arterial shear stress to evaluate the effect of high shear stress on EPCs. To investigate how much paclitaxel elutes during the seeding procedure, a pharmacokinetic analysis was performed. Finally, to validate the proof of concept, EPC-seeded DES were placed on a fibrin matrix with and without smooth muscle cells (SMCs) and cultured for 3 days under perfusion. The seeding procedure resulted in 47% and 26% coverage of the stent surface in uncrimped and crimped conditions, respectively. After the optimal seeding, almost 99% of drug was still available. When EPC-seeded DES were placed on a fibrin matrix and cultured for 3 days, the EPCs confluently covered the stent surface and spread to the surrounding fibrin gel. When EPC-seeded DES were placed on SMC-containing fibrin layers, cells in contact with the struts died. EPCs can be successfully seeded onto DES without losing drug-eluting capability, and EPCs exhibit sufficient proliferative ability. EPC-seeded DES may combine early re-endothelialization ability with the antirestenotic effectiveness of DES.

[Delayed Occluding Membrane (DOM) - A New Concept to Prevent Spinal Cord Ischaemia During Endovascular Aortic Aneurysm Repair]. / Delayed Occluding Membrane (DOM) ­ ein neuer Ansatz zur Protektion der spinalen Ischämie bei endovaskulärem Aortenrepair.

INTRODUCTION: The risk of spinal cord ischemia is a relevant problem in in fields of open and endovascular thoracoabdominal aortic aneurysm repair (TAAA). Despite all efforts, no therapeutical concept exists, which enables a complete treatment of the TAAA without open branches or fenestrations, and reduces the risk for a spinal cord ischemia (SCI) to the minimum. In this article, we would like to present a new concept based on slow-occluding hydrogel-textile membrane, which could help to reduce the SCI risk during endovascular TAAA repair. CONCEPT: A hydrogel textile membrane is under development, which could be used a functional unit of endovascular stentprosthesis. If in contact with blood, glutathion induces swelling of the induces ongoing swelling of the membrane because of the triggered degradation of the crosslinker. Due to the resulting water uptake of the hydrogel textile membrane and mass increase of the gel, the swelling leads to a stabilization of the membrane. In vitro studies show, that the swelling of the hydrogel textile membrane should lead to a controlled decreasing flow into the aneurysm sac. After a pre-defined period, the membrane is occluded and the aneurysm sac perfusion stops. So, by using the hydrogel textile membrane, a complete treatment of the TAAA can be realized in one procedure without further re-intervention or pre-interventional measures. Furthermore, the risk of a SCI would be minimized. As this treatment concept is under development, only interim results are presented. CONCLUSION: The successful development and usage of a slow-occluding hydrogel textile membrane as a part of endovascular stentprosthesis could help to reduce the risk SCI during endovascular TAAA surgery.

Fluid-structure interaction simulation of artificial textile reinforced aortic heart valve: Validation with an in-vitro test.

Prosthetic heart valves deployed in the left heart (aortic and mitral) are subjected to harsh hemodynamical conditions. Most of the tissue engineered heart valves have been developed for the low pressure pulmonary position because of the difficulties in fabricating a mechanically strong valve, able to withstand the systemic circulation. This necessitates the use of reinforcing scaffolds, resulting in a tissue-engineered textile reinforced tubular aortic heart valve. Therefore, to better design these implants, material behaviour of the composite, valve kinematics and its hemodynamical response need to be evaluated. Experimental assessment can be immensely time consuming and expensive, paving way for numerical studies. In this work, the material properties obtained using the previously proposed multi-scale numerical method for textile composites was evaluated for its accuracy. An in silico immersed boundary (IB) fluid structure interaction (FSI) simulation emulating the in vitro experiment was set-up to evaluate and compare the geometric orifice area and flow rate for one beat cycle. Results from the in silico FSI simulation were found to be in good coherence with the in vitro test during the systolic phase, while mean deviation of approximately 9% was observed during the diastolic phase of a beat cycle. Merits and demerits of the in silico IB-FSI method for the presented case study has been discussed with the advantages outweighing the drawbacks, indicating the potential towards an effective use of this framework in the development and analysis of heart valves.

Development of a Polymer-Based Biodegradable Neurovascular Stent Prototype: A Preliminary In Vitro and In Vivo Study.

Biodegradable stents are not established in neurovascular interventions. In this study, mechanical, radiological, and histological characteristics of a stent prototype developed for neurovascular use are presented. The elasticity and brittleness of PLA 96/4, PLDL 70/30, PCL, and PLGA 85/15 and 10/90 polymers in in vitro experiments are first analyzed. After excluding the inapt polymers, degradability and mechanical characteristics of 78 PLGA 85/15 and PLGA 10/90 stent prototypes are analyzed. After excluding PLGA 10/90 stents because of rapid loss of mass PLGA 85/15 stents in porcine in vivo experiments are analyzed. Angiographic occlusion rates 7 d, 1 month, 3 months, and 6 months after stent implantation are assessed. Histological outcome measures are the presence of signs of inflammation, endothelialization, and the homogeneity of degradation after six months. One case of stent occlusion occurs within the first 7 d. There is a prominent foreign-body reaction with considerable mononuclear and minor granulocytic inflammation combined with incomplete fragmental degradation of the struts. It is possible to produce a stent prototype with dimensions that fit the typical size of carotid arteries. Major improvements concerning thrombogenicity, degradation, and inflammatory response are required to produce biodegradable stents that are suitable for neurovascular interventions.

Targeting In-Stent-Stenosis with RGD- and CXCL1-Coated Mini-Stents in Mice.

Atherosclerotic lesions that critically narrow the artery can necessitate an angioplasty and stent implantation. Long-term therapeutic effects, however, are limited by excessive arterial remodeling. We here employed a miniaturized nitinol-stent coated with star-shaped polyethylenglycole (star-PEG), and evaluated its bio-functionalization with RGD and CXCL1 for improving in-stent stenosis after implantation into carotid arteries of mice. Nitinol foils or stents (bare metal) were coated with star-PEG, and bio-functionalized with RGD, or RGD/CXCL1. Cell adhesion to star-PEG-coated nitinol foils was unaltered or reduced, whereas bio-functionalization with RGD but foremost RGD/CXCL1 increased adhesion of early angiogenic outgrowth cells (EOCs) and endothelial cells but not smooth muscle cells when compared with bare metal foils. Stimulation of cells with RGD/CXCL1 furthermore increased the proliferation of EOCs. In vivo, bio-functionalization with RGD/CXCL1 significantly reduced neointima formation and thrombus formation, and increased re-endothelialization in apoE-/- carotid arteries compared with bare-metal nitinol stents, star-PEG-coated stents, and stents bio-functionalized with RGD only. Bio-functionalization of star-PEG-coated nitinol-stents with RGD/CXCL1 reduced in-stent neointima formation. By supporting the adhesion and proliferation of endothelial progenitor cells, RGD/CXCL1 coating of stents may help to accelerate endothelial repair after stent implantation, and thus may harbor the potential to limit the complication of in-stent restenosis in clinical approaches.

Characterisation of cell-substrate interactions between Schwann cells and three-dimensional fibrin hydrogels containing orientated nanofibre topographical cues.

The generation of complex three-dimensional bioengineered scaffolds that are capable of mimicking the molecular and topographical cues of the extracellular matrix found in native tissues is a field of expanding research. The systematic development of such scaffolds requires the characterisation of cell behaviour in response to the individual components of the scaffold. In the present investigation, we studied cell-substrate interactions between purified populations of Schwann cells and three-dimensional fibrin hydrogel scaffolds, in the presence or absence of multiple layers of highly orientated electrospun polycaprolactone nanofibres. Embedded Schwann cells remained viable within the fibrin hydrogel for up to 7 days (the longest time studied); however, cell behaviour in the hydrogel was somewhat different to that observed on the two-dimensional fibrin substrate: Schwann cells failed to proliferate in the fibrin hydrogel, whereas cell numbers increased steadily on the two-dimensional fibrin substrate. Schwann cells within the fibrin hydrogel developed complex process branching patterns, but, when presented with orientated nanofibres, showed a strong tendency to redistribute themselves onto the nanofibres, where they extended long processes that followed the longitudinal orientation of the nanofibres. The process length along nanofibre-containing fibrin hydrogel reached near-maximal levels (for the present experimental conditions) as early as 1 day after culturing. The ability of this three-dimensional, extracellular matrix-mimicking scaffold to support Schwann cell survival and provide topographical cues for rapid process extension suggest that it may be an appropriate device design for the bridging of experimental lesions of the peripheral nervous system.

FMN-coated fluorescent USPIO for cell labeling and non-invasive MR imaging in tissue engineering.

Non-invasive magnetic resonance imaging (MRI) is gaining significant attention in the field of tissue engineering, since it can provide valuable information on in vitro production parameters and in vivo performance. It can e.g. be used to monitor the morphology, location and function of the regenerated tissue, the integrity, remodeling and resorption of the scaffold, and the fate of the implanted cells. Since cells are not visible using conventional MR techniques, ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles are routinely employed to label and monitor the cells embedded in tissue-engineered implants. We here set out to optimize cell labeling procedures with regard to labeling efficiency, biocompatibility and in vitro validation during bioreactor cultivation, using flavin mononucleotide (FMN)-coated fluorescent USPIO (FLUSPIO). Efficient FLUSPIO uptake is demonstrated in three different cell lines, applying relatively short incubation times and low labeling concentrations. FLUSPIO-labeled cells were successfully employed to visualize collagen scaffolds and tissue-engineered vascular grafts. Besides promoting safe and efficient cell uptake, an exquisite property of the non-polymeric FMN-coating is that it renders the USPIO fluorescent, providing a means for in vitro, in vivo and ex vivo validation via fluorescence microscopy and fluorescence reflectance imaging (FRI). FLUSPIO cell labeling is consequently considered to be a suitable tool for theranostic tissue engineering purposes.

Autologous valve replacement-CD133+ stem cell-plus-fibrin composite-based sprayed cell seeding for intraoperative heart valve tissue engineering.

OBJECTIVE: The development of biological valve prostheses with lifetime native-like performance and optimal host engraftment is an ultimate goal of heart valve tissue engineering. We describe a new concept for autologous graft coating based on a CD133(+)-stem-cells-plus-fibrin (SC+F) complex processed from bone marrow and peripheral blood of a single patient. METHODS: CD133(+)-SC (1 × 10(6) cells/mL) from human bone marrow and autologous fibrin (20 mg/mL) were administered simultaneously via spray administration using the novel Vivostat Co-Delivery System. During static cultivation, SC+F performance was monitored for 20 days after delivery and compared with controls. For dynamic testing SC+F-composite was sprayed on a decellularized porcine pulmonary valve and transferred to a bioreactor under pulsatile flow conditions for 7 days. RESULTS: Static cultivation of SC+F-composite induced significant improvements in stem cell proliferation as compared with controls. For dynamic testing, microscopic analyses on a smooth engineered heart valve surface detected homogenous distribution of stem cells. Ultrasonic analysis revealed native-like valve performance. Applied CD133(+) stem cells differentiated into endothelial-like cells positive for CD31 and vascular endothelial growth factor receptor 2 and engrafted the valve. However, occasional delamination was observed. CONCLUSION: SC+F serves as an excellent autologous matrix for intraoperative tissue engineering of valve prostheses promising optimal in vivo integration. However, stability remains an issue.