Anisotropic architecture and electrical stimulation enhance neuron cell behaviour on a tough graphene embedded PVA: alginate fibrous scaffold.

Tough scaffolds comprised of aligned and conductive fibers are promising for peripheral nerve regeneration due to their unique mechanical and electrical properties. Several studies have confirmed that electrical stimulation can control the axonal extension in vitro. However, the stimulatory effects of scaffold architecture and electrical stimulation have not yet been investigated in detail. Here, we assessed a comparison between aligned and random fibers made of graphene (Gr) embedded sodium alginate (SA) polyvinyl alcohol (PVA) (Gr-AP scaffolds) for peripheral nerve engineering. The effects of applied electrical stimulation and orientation of the fabricated fibers on the in vitro attachment, alignment, and proliferation of PC12 cells (a rat neuronal cell line) were investigated. The results revealed that the aligned fibrous Gr-AP scaffolds closely mimicked the anisotropic structure of the native sciatic nerve. Aligned fibrous Gr-AP scaffolds significantly improved mechanical properties as well as cell-scaffold integration compared to random fibrous scaffolds. In addition, electrical stimulation significantly improved PC12 cell proliferation. In summary, our findings revealed that aligned fibrous Gr-AP scaffolds offered superior mechanical characteristics and structural properties that enhanced neural cell-substrate interactions, resulting in a promising construct for nerve tissue regeneration.

Elastomeric Fibrous Hybrid Scaffold Supports In Vitro and In Vivo Tissue Formation.

Biomimetic materials with biomechanical properties resembling those of native tissues while providing an environment for cell growth and tissue formation, are vital for tissue engineering (TE). Mechanical anisotropy is an important property of native cardiovascular tissues and directly influences tissue function. This study reports fabrication of anisotropic cell-seeded constructs while retaining control over the construct's architecture and distribution of cells. Newly synthesized poly-4-hydroxybutyrate (P4HB) is fabricated with a dry spinning technique to create anelastomeric fibrous scaffold that allows control of fiber diameter, porosity, and rate ofdegradation. To allow cell and tissue ingrowth, hybrid scaffolds with mesenchymalstem cells (MSCs) encapsulated in a photocrosslinkable hydrogel were developed. Culturing the cellularized scaffolds in a cyclic stretch/flexure bioreactor resulted in tissue formation and confirmed the scaffold's performance under mechanical stimulation. In vivo experiments showed that the hybrid scaffold is capable of withstanding physiological pressures when implanted as a patch in the pulmonary artery. Aligned tissue formation occurred on the scaffold luminal surface without macroscopic thrombus formation. This combination of a novel, anisotropic fibrous scaffold and a tunable native-like hydrogel for cellular encapsulation promoted formation of 3D tissue and provides a biologically functional composite scaffold for soft-tissue engineering applications.

Design and testing of a cyclic stretch and flexure bioreactor for evaluating engineered heart valve tissues based on poly(glycerol sebacate) scaffolds.

Cyclic flexure and stretch are essential to the function of semilunar heart valves and have demonstrated utility in mechanically conditioning tissue-engineered heart valves. In this study, a cyclic stretch and flexure bioreactor was designed and tested in the context of the bioresorbable elastomer poly(glycerol sebacate). Solid poly(glycerol sebacate) membranes were subjected to cyclic stretch, and micromolded poly(glycerol sebacate) scaffolds seeded with porcine aortic valvular interstitial cells were subjected to cyclic stretch and flexure. The results demonstrated significant effects of cyclic stretch on poly(glycerol sebacate) mechanical properties, including significant decreases in effective stiffness versus controls. In valvular interstitial cell-seeded scaffolds, cyclic stretch elicited significant increases in DNA and collagen content that paralleled maintenance of effective stiffness. This work provides a basis for investigating the roles of mechanical loading in the formation of tissue-engineered heart valves based on elastomeric scaffolds.

Tri-layered elastomeric scaffolds for engineering heart valve leaflets.

Tissue engineered heart valves (TEHVs) that can grow and remodel have the potential to serve as permanent replacements of the current non-viable prosthetic valves particularly for pediatric patients. A major challenge in designing functional TEHVs is to mimic both structural and anisotropic mechanical characteristics of the native valve leaflets. To establish a more biomimetic model of TEHV, we fabricated tri-layered scaffolds by combining electrospinning and microfabrication techniques. These constructs were fabricated by assembling microfabricated poly(glycerol sebacate) (PGS) and fibrous PGS/poly(caprolactone) (PCL) electrospun sheets to develop elastic scaffolds with tunable anisotropic mechanical properties similar to the mechanical characteristics of the native heart valves. The engineered scaffolds supported the growth of valvular interstitial cells (VICs) and mesenchymal stem cells (MSCs) within the 3D structure and promoted the deposition of heart valve extracellular matrix (ECM). MSCs were also organized and aligned along the anisotropic axes of the engineered tri-layered scaffolds. In addition, the fabricated constructs opened and closed properly in an ex vivo model of porcine heart valve leaflet tissue replacement. The engineered tri-layered scaffolds have the potential for successful translation towards TEHV replacements.

Tough and flexible CNT-polymeric hybrid scaffolds for engineering cardiac constructs.

In the past few years, a considerable amount of effort has been devoted toward the development of biomimetic scaffolds for cardiac tissue engineering. However, most of the previous scaffolds have been electrically insulating or lacked the structural and mechanical robustness to engineer cardiac tissue constructs with suitable electrophysiological functions. Here, we developed tough and flexible hybrid scaffolds with enhanced electrical properties composed of carbon nanotubes (CNTs) embedded aligned poly(glycerol sebacate):gelatin (PG) electrospun nanofibers. Incorporation of varying concentrations of CNTs from 0 to 1.5% within the PG nanofibrous scaffolds (CNT-PG scaffolds) notably enhanced fiber alignment and improved the electrical conductivity and toughness of the scaffolds while maintaining the viability, retention, alignment, and contractile activities of cardiomyocytes (CMs) seeded on the scaffolds. The resulting CNT-PG scaffolds resulted in stronger spontaneous and synchronous beating behavior (3.5-fold lower excitation threshold and 2.8-fold higher maximum capture rate) compared to those cultured on PG scaffold. Overall, our findings demonstrated that aligned CNT-PG scaffold exhibited superior mechanical properties with enhanced CM beating properties. It is envisioned that the proposed hybrid scaffolds can be useful for generating cardiac tissue constructs with improved organization and maturation.

Fiber-reinforced hydrogel scaffolds for heart valve tissue engineering.

Heart valve-related disorders are among the major causes of death worldwide. Although prosthetic valves are widely used to treat this pathology, current prosthetic grafts cannot grow with the patient while maintaining normal valve mechanical and hemodynamic properties. Tissue engineering may provide a possible solution to this issue through using biodegradable scaffolds and patients' own cells. Despite their similarity to heart valve tissue, most hydrogel scaffolds are not mechanically suitable for the dynamic stresses of the heart valve microenvironment. In this study, we integrated electrospun poly(glycerol sebacate) (PGS)-poly(ɛ-caprolactone) (PCL) microfiber scaffolds, which possess enhanced mechanical properties for heart valve engineering, within a hybrid hydrogel made from methacrylated hyaluronic acid and methacrylated gelatin. Sheep mitral valvular interstitial cells were encapsulated in the hydrogel and evaluated in hydrogel-only, PGS-PCL scaffold-only, and composite scaffold conditions. Although the cellular viability and metabolic activity were similar among all scaffold types, the presence of the hydrogel improved the three-dimensional distribution of mitral valvular interstitial cells. As seen by similar values in both the Young's modulus and the ultimate tensile strength between the PGS-PCL scaffolds and the composites, microfibrous scaffolds preserved their mechanical properties in the presence of the hydrogels. Compared to electrospun or hydrogel scaffolds alone, this combined system may provide a more suitable three-dimensional structure for generating scaffolds for heart valve tissue engineering.

Electrospun PGS:PCL microfibers align human valvular interstitial cells and provide tunable scaffold anisotropy.

Tissue engineered heart valves (TEHV) can be useful in the repair of congenital or acquired valvular diseases due to their potential for growth and remodeling. The development of biomimetic scaffolds is a major challenge in heart valve tissue engineering. One of the most important structural characteristics of mature heart valve leaflets is their intrinsic anisotropy, which is derived from the microstructure of aligned collagen fibers in the extracellular matrix (ECM). In the present study, a directional electrospinning technique is used to fabricate fibrous poly(glycerol sebacate):poly(caprolactone) (PGS:PCL) scaffolds containing aligned fibers, which resemble native heart valve leaflet ECM networks. In addition, the anisotropic mechanical characteristics of fabricated scaffolds are tuned by changing the ratio of PGS:PCL to mimic the native heart valve's mechanical properties. Primary human valvular interstitial cells (VICs) attach and align along the anisotropic axes of all PGS:PCL scaffolds with various mechanical properties. The cells are also biochemically active in producing heart-valve-associated collagen, vimentin, and smooth muscle actin as determined by gene expression. The fibrous PGS:PCL scaffolds seeded with human VICs mimick the structure and mechanical properties of native valve leaflet tissues and would potentially be suitable for the replacement of heart valves in diverse patient populations.

PGS:Gelatin nanofibrous scaffolds with tunable mechanical and structural properties for engineering cardiac tissues.

A significant challenge in cardiac tissue engineering is the development of biomimetic grafts that can potentially promote myocardial repair and regeneration. A number of approaches have used engineered scaffolds to mimic the architecture of the native myocardium tissue and precisely regulate cardiac cell functions. However, previous attempts have not been able to simultaneously recapitulate chemical, mechanical, and structural properties of the myocardial extracellular matrix (ECM). In this study, we utilized an electrospinning approach to fabricate elastomeric biodegradable poly(glycerol sebacate) (PGS):gelatin nanofibrous scaffolds with a wide range of chemical composition, stiffness and anisotropy. Our findings demonstrated that through incorporation of PGS, it is possible to create nanofibrous scaffolds with well-defined anisotropy that mimic the left ventricular myocardium architecture. Furthermore, we studied attachment, proliferation, differentiation and alignment of neonatal rat cardiac fibroblast cells (CFs) as well as protein expression, alignment, and contractile function of cardiomyocyte (CMs) on PGS:gelatin scaffolds with variable amount of PGS. Notably, aligned nanofibrous scaffold, consisting of 33 wt. % PGS, induced optimal synchronous contractions of CMs while significantly enhanced cellular alignment. Overall, our study suggests that the aligned nanofibrous PGS:gelatin scaffold support cardiac cell organization, phenotype and contraction and could potentially be used to develop clinically relevant constructs for cardiac tissue engineering.

Valvular interstitial cell seeded poly(glycerol sebacate) scaffolds: toward a biomimetic in vitro model for heart valve tissue engineering.

Tissue engineered replacement heart valves may be capable of overcoming the lack of growth potential intrinsic to current non-viable prosthetics, and thus could potentially serve as permanent replacements in the surgical repair of pediatric valvular lesions. However, the evaluation of candidate combinations of cells and scaffolds lacks a biomimetic in vitro model with broadly tunable, anisotropic and elastomeric structural-mechanical properties. Toward establishing such an in vitro model, in the current study, porcine aortic and pulmonary valvular interstitial cells (i.e. biomimetic cells) were cultivated on anisotropic, micromolded poly(glycerol sebacate) scaffolds (i.e. biomimetic scaffolds). Following 14 and 28 days of static culture, cell-seeded scaffolds and unseeded controls were assessed for their mechanical properties, and cell-seeded scaffolds were further characterized by confocal fluorescence and scanning electron microscopy, and by collagen and DNA assays. Poly(glycerol sebacate) micromolding yielded scaffolds with anisotropic stiffnesses resembling those of native valvular tissues in the low stress-strain ranges characteristic of physiologic valvular function. Scaffold anisotropy was largely retained upon cultivation with valvular interstitial cells; while the mechanical properties of unseeded scaffolds progressively diminished, cell-seeded scaffolds either retained or exceeded initial mechanical properties. Retention of mechanical properties in cell-seeded scaffolds paralleled the accretion of collagen, which increased significantly from 14 to 28 days. This study demonstrates that valvular interstitial cells can be cultivated on anisotropic poly(glycerol sebacate) scaffolds to yield biomimetic in vitro models with which clinically relevant cells and future scaffold designs can be evaluated.

Laser microfabricated poly(glycerol sebacate) scaffolds for heart valve tissue engineering.

Microfabricated poly(glycerol sebacate) (PGS) scaffolds may be applicable to tissue engineering heart valve leaflets by virtue of their controllable microstructure, stiffness, and elasticity. In this study, PGS scaffolds were computationally designed and microfabricated by laser ablation to match the anisotropy and peak tangent moduli of native bovine aortic heart valve leaflets. Finite element simulations predicted PGS curing conditions, scaffold pore shape, and strut width capable of matching the scaffold effective stiffnesses to the leaflet peak tangent moduli. On the basis of simulation predicted effective stiffnesses of 1.041 and 0.208 MPa for the scaffold preferred (PD) and orthogonal, cross-preferred (XD) material directions, scaffolds with diamond-shaped pores were microfabricated by laser ablation of PGS cured 12 h at 160°C. Effective stiffnesses measured for the scaffold PD (0.83 ± 0.13 MPa) and XD (0.21 ± 0.03 MPa) were similar to both predicted values and peak tangent moduli measured for bovine aortic valve leaflets in the circumferential (1.00 ± 0.16 MPa) and radial (0.26 ± 0.03 MPa) directions. Scaffolds cultivated with fibroblasts for 3 weeks accumulated collagen (736 ± 193 µg/g wet weight) and DNA (17 ± 4 µg/g wet weight). This study provides a basis for the computational design of biomimetic microfabricated PGS scaffolds for tissue-engineered heart valves.

2D Computational Fluid Dynamic Modeling of Human Ventricle System Based on Fluid-Solid Interaction and Pulsatile Flow.

Many diseases are related to cerebrospinal fluid (CSF) hydrodynamics. Therefore, understanding the hydrodynamics of CSF flow and intracranial pressure is helpful for obtaining deeper knowledge of pathological processes and providing better treatments. Furthermore, engineering a reliable computational method is promising approach for fabricating in vitro models which is essential for inventing generic medicines. A Fluid-Solid Interaction (FSI)model was constructed to simulate CSF flow. An important problem in modeling the CSF flow is the diastolic back flow. In this article, using both rigid and flexible conditions for ventricular system allowed us to evaluate the effect of surrounding brain tissue. Our model assumed an elastic wall for the ventricles and a pulsatile CSF input as its boundary conditions. A comparison of the results and the experimental data was done. The flexible model gave better results because it could reproduce the diastolic back flow mentioned in clinical research studies. The previous rigid models have ignored the brain parenchyma interaction with CSF and so had not reported the back flow during the diastolic time. In this computational fluid dynamic (CFD) analysis, the CSF pressure and flow velocity in different areas were concordant with the experimental data.

Finite element analysis of blunt foreign body impact on the cornea after PRK and LASIK.

PURPOSE: To investigate the effect of blunt foreign body impact on a human cornea after photorefractive keratectomy (PRK) and LASIK using a simulation model. METHODS: Computational simulations were performed using a finite element analysis program (LS-Dyna, Livermore Software Technology Corp). The blunt foreign body was set to impact at the center of the corneal surface models (after PRK and LASIK) with thicknesses of 500, 450, 400, 350, and 300 µm. Corneal rupture was assumed to occur at a peak stress of 9.45 MPa and at a strain of 18%. The foreign body projectile was blunt in shape, made from aluminum, contained plastic-kinematic properties, and had a density of 2700 kg/m(3). RESULTS: The projectile was launched at the center of the cornea with velocities ranging from 20 to 60 m/s. The threshold of impact velocities creating rupture in corneal thicknesses of 500, 450, 400, 350, and 300 µm were 33, 32.8, 30.7, 27.9, and 22.8 m/s, respectively, in the PRK model. In the LASIK model, the thresholds creating rupture in the stromal bed of the corneas with thicknesses of 500, 450, 400, 350, and 300 µm were 40, 38.1, 35.6, 31.5, and 26.7 m/s, respectively. The 110-µm corneal flap in the LASIK model ruptured at all velocities. CONCLUSIONS: Ruptures occurred at lower velocities in the PRK cornea model than in the corneal stromal bed of the LASIK model following blunt foreign body impact.

Mathematical modeling of CSF pulsatile hydrodynamics based on fluid-solid interaction.

Intracranial pressure (ICP) is derived from cerebral blood pressure and cerebrospinal fluid (CSF) circulatory dynamics and can be affected in the course of many diseases. Computer analysis of the ICP time pattern plays a crucial role in the diagnosis and treatment of those diseases. This study proposes the application of Linninger et al.'s [IEEE Trans. Biomed. Eng., vol. 52, no. 4, pp. 557-565, Apr. 2005] fluid-solid interaction model of CSF hydrodynamic in ventricular system based on our clinical data from a group of patients with brain parenchyma tumor. The clinical experiments include the arterial blood pressure (ABP), venous blood pressure, and ICP in the subarachnoid space (SAS). These data were used as inputs to the model that predicts the intracranial dynamic phenomena. In addition, the model has been modified by considering CSF pulsatile production rate as the major factor of CSF motion. The approximations of ventricle enlargement, CSF pressure distribution in the ventricular system and CSF velocity magnitude in the aqueduct and foramina were obtained in this study. The observation of reversal flow in the CSF flow pattern due to brain tissue compression is another finding in our investigation. Based on the experimental results, no existence of large transmural pressure differences were found in the brain system. The measured pressure drop in the ventricular system was less than 5 Pa. Moreover, the CSF flow pattern, ICP distribution, and velocity magnitude were in good agreement with the published models and CINE (phase-contrast magnetic resonance imaging) experiments, respectively.