Flexible shape-memory scaffold for minimally invasive delivery of functional tissues.

Despite great progress in engineering functional tissues for organ repair, including the heart, an invasive surgical approach is still required for their implantation. Here, we designed an elastic and microfabricated scaffold using a biodegradable polymer (poly(octamethylene maleate (anhydride) citrate)) for functional tissue delivery via injection. The scaffold's shape memory was due to the microfabricated lattice design. Scaffolds and cardiac patches (1 cm × 1 cm) were delivered through an orifice as small as 1 mm, recovering their initial shape following injection without affecting cardiomyocyte viability and function. In a subcutaneous syngeneic rat model, injection of cardiac patches was equivalent to open surgery when comparing vascularization, macrophage recruitment and cell survival. The patches significantly improved cardiac function following myocardial infarction in a rat, compared with the untreated controls. Successful minimally invasive delivery of human cell-derived patches to the epicardium, aorta and liver in a large-animal (porcine) model was achieved.

A Conductive Polymer Hydrogel Supports Cell Electrical Signaling and Improves Cardiac Function After Implantation into Myocardial Infarct.

BACKGROUND: Efficient cardiac function requires synchronous ventricular contraction. After myocardial infarction, the nonconductive nature of scar tissue contributes to ventricular dysfunction by electrically uncoupling viable cardiomyocytes in the infarct region. Injection of a conductive biomaterial polymer that restores impulse propagation could synchronize contraction and restore ventricular function by electrically connecting isolated cardiomyocytes to intact tissue, allowing them to contribute to global heart function. METHODS AND RESULTS: We created a conductive polymer by grafting pyrrole to the clinically tested biomaterial chitosan to create a polypyrrole (PPy)-chitosan hydrogel. Cyclic voltammetry showed that PPy-chitosan had semiconductive properties lacking in chitosan alone. PPy-chitosan did not reduce cell attachment, metabolism, or proliferation in vitro. Neonatal rat cardiomyocytes plated on PPy-chitosan showed enhanced Ca(2+) signal conduction in comparison with chitosan alone. PPy-chitosan plating also improved electric coupling between skeletal muscles placed 25 mm apart in comparison with chitosan alone, demonstrating that PPy-chitosan can electrically connect contracting cells at a distance. In rats, injection of PPy-chitosan 1 week after myocardial infarction decreased the QRS interval and increased the transverse activation velocity in comparison with saline or chitosan, suggesting improved electric conduction. Optical mapping showed increased activation in the border zone of PPy-chitosan-treated rats. Echocardiography and pressure-volume analysis showed improvement in load-dependent (ejection fraction, fractional shortening) and load-independent (preload recruitable stroke work) indices of heart function 8 weeks after injection. CONCLUSIONS: We synthesized a biocompatible conductive biomaterial (PPy-chitosan) that enhances biological conduction in vitro and in vivo. Injection of PPy-chitosan better maintained heart function after myocardial infarction than a nonconductive polymer.

A Novel Conductive Polypyrrole-Chitosan Hydrogel Containing Human Endometrial Mesenchymal Stem Cell-Derived Exosomes Facilitated Sustained Release for Cardiac Repair.

Myocardial infarction (MI) results in cardiomyocyte necrosis and conductive system damage, leading to sudden cardiac death and heart failure. Studies have shown that conductive biomaterials can restore cardiac conduction, but cannot facilitate tissue regeneration. This study aims to add regenerative capabilities to the conductive biomaterial by incorporating human endometrial mesenchymal stem cell (hEMSC)-derived exosomes (hEMSC-Exo) into poly-pyrrole-chitosan (PPY-CHI), to yield an injectable hydrogel that can effectively treat MI. In vitro, PPY-CHI/hEMSC-Exo, compared to untreated controls, PPY-CHI, or hEMSC-Exo alone, alleviates H2O2-induced apoptosis and promotes tubule formation, while in vivo, PPY-CHI/hEMSC-Exo improves post-MI cardiac functioning, along with counteracting against ventricular remodeling and fibrosis. All these activities are facilitated via increased epidermal growth factor (EGF)/phosphoinositide 3-kinase (PI3K)/AKT signaling. Furthermore, the conductive properties of PPY-CHI/hEMSC-Exo are able to resynchronize cardiac electrical transmission to alleviate arrythmia. Overall, PPY-CHI/hEMSC-Exo synergistically combines the cardiac regenerative capabilities of hEMSC-Exo with the conductive properties of PPY-CHI to improve cardiac functioning, via promoting angiogenesis and inhibiting apoptosis, as well as resynchronizing electrical conduction, to ultimately enable more effective MI treatment. Therefore, incorporating exosomes into a conductive hydrogel provides dual benefits in terms of maintaining conductivity, along with facilitating long-term exosome release and sustained application of their beneficial effects.

A Polypyrrole-Polycarbonate Polyurethane Elastomer Alleviates Cardiac Arrhythmias via Improving Bio-Conductivity.

Myocardial fibrosis, resulting from myocardial infarction (MI), significantly alters cardiac electrophysiological properties. As fibrotic scar tissue forms, its resistance to incoming action potentials increases, leading to cardiac arrhythmia, and eventually sudden cardiac death or heart failure. Biomaterials are gaining increasing attention as an approach for addressing post-MI arrhythmias. The current study investigates the hypothesis that a bio-conductive epicardial patch can electrically synchronize isolated cardiomyocytes in vitro and rescue arrhythmic hearts in vivo. A new conceived biocompatible, conductive, and elastic polyurethane composite bio-membrane, referred to as polypyrrole-polycarbonate polyurethane (PPy-PCNU), is developed, in which solid-state conductive PPy nanoparticles are distributed throughout an electrospun aliphatic PCNU nanofiber patch in a controlled manner. Compared to PCNU alone, the resulting biocompatible patch demonstrates up to six times less impedance, with no conductivity loss over time, as well as being able to influence cellular alignment. Furthermore, PPy-PCNU promotes synchronous contraction of isolated neonatal rat cardiomyocytes and alleviates atrial fibrillation in rat hearts upon epicardial implantation. Taken together, epicardially-implanted PPy-PCNU could potentially serve as a novel alternative approach for the treatment of cardiac arrhythmias.

An electro-spun tri-component polymer biomaterial with optoelectronic properties for neuronal differentiation.

Optoelectronic biomaterials have recently emerged as a potential treatment option for neurodegenerative diseases, such as optic macular degeneration. Though initial works in the field have involved bulk heterojunctions mimicking solar panels with photovoltaics (PVs) and conductive polymers (CPs), recent developments have considered abandoning CPs in such systems. Here, we developed a simple antioxidant, biocompatible, and fibrous membrane heterojunction composed of photoactive polymer poly(3-hexylthiophene) (P3HT), polycaprolactone (PCL) and polypyrrole (PPY), to facilitate neurogenesis of PC-12 cells when photo-stimulated in vitro. The photoactive prototype, referred to as PCL-P3HT/PPY, was fabricated via polymerization of pyrrole on electro-spun PCL-P3HT nanofibers to form a membrane. Four experimental groups, namely PCL alone, PCL/PPY, PCL-P3HT and PCL-P3HT/PPY, were tested. In the absence of the CP, PCL-P3HT demonstrated lower cell survival due to increased intracellular reactive oxygen/nitrogen species production. PCL-P3HT/PPY rescued these cells by virtue of scavenging radicals, where the CP, PPY, acted as an antioxidant. Apart from having lower impedance, the material also enhanced neurogenesis of PC-12 cells when photo-stimulated, compared to the traditional PCL-P3HT. Lastly, the in vitro system with PC-12 was used to demonstrate the practicality of the material for potential use as a cellular patch in optic and nerve regeneration. This work demonstrated the importance of maintaining PV-CP heterojunctions while simultaneously providing an optoelectrical platform for neural and optical tissue engineering. STATEMENT OF SIGNIFICANCE: Regeneration and repair of injured nervous systems have always been a major clinical challenge. Stem cell therapy is a promising approach for nerve regeneration, and opto-electrical stimulation, which converts light into an electrical signal, has been shown to efficiently regulate stem cell behaviors with enhanced neurogenesis. We developed a micro-fibrous membrane, composed of photoactive polymer, P3HT, scaffold material PCL and conductive polymer PPY. Our heterojunction system improved cell survival via PPY quenching PCL-P3HT-generated cell-damaging reactive oxygen species. PPY also conducted electrons produced from light-stimulated P3HT to promote neurogenesis. This photoactive microfiber biomaterial has great potential as a highly biocompatible and efficient platform to wirelessly promote neurogenesis and survival. Our approach thus showed possibilities with respect to optical tissue engineering.

Compatibility and function of human induced pluripotent stem cell derived cardiomyocytes on an electrospun nanofibrous scaffold, generated from an ionomeric polyurethane composite.

Synthetic scaffolds are needed for generating organized neo-myocardium constructs to promote functional tissue repair. This study investigated the biocompatibility of an elastomeric electrospun degradable polar/hydrophobic/ionic polyurethane (D-PHI) composite scaffold with human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The composite material was electrospun to generate scaffolds, with nanofibres oriented in aligned or random directions. These features enabled the authors to evaluate the effect of characteristic elements which mimic that of the native extracellular matrix (alignment, chemical heterogeneity, and fiber topography) on hiPSC-CMs activity. The functional nature of the hiPSC-CM cultured on gelatin and Matrigel-coated scaffolds were assessed, investigating the influence of protein interactions with the synthetic substrate on subsequent cell phenotype. After 7 days of culture, high hiPSC-CM viability was observed on the scaffolds. The cells on the aligned scaffold were elongated and demonstrated aligned sarcomeres that oriented parallel to the direction of the fibers, while the cells on random scaffolds and a tissue culture polystyrene (TCPS) control did not exhibit such an organized morphology. The hiPSC-CMs cultured on the scaffolds and TCPS expressed similar levels of cardiac troponin-T, but there was a higher expression of ventricular myosin light chain-2 on the D-PHI composite scaffolds versus TCPS, indicating a higher proportion of hiPSC-CM exhibiting a ventricular cardiomyocyte like phenotype. Within 7 days, the hiPSC-CMs on aligned scaffolds and TCPS beat synchronously and had similar conductive velocities. These preliminary results show that aligned D-PHI elastomeric scaffolds allow hiPSC-CMs to demonstrate important cardiomyocytes characteristics, critical to enabling their future potential use for cardiac tissue regeneration.

Bio-Conductive Polymers for Treating Myocardial Conductive Defects: Long-Term Efficacy Study.

Following myocardial infarction (MI), the resulting fibrotic scar is nonconductive and leads to ventricular dysfunction via electrical uncoupling of the remaining viable cardiomyocytes. The uneven conductive properties between normal myocardium and scar tissue result in arrhythmia, yielding sudden cardiac death/heart failure. A conductive biopolymer, poly-3-amino-4-methoxybenzoic acid-gelatin (PAMB-G), is able to resynchronize myocardial contractions in vivo. Intravenous PAMB-G injections into mice show that it does not cause any acute toxicity, up to the maximum tolerated dose (1.6 mL kg-1 ), which includes the determined therapeutic dose (0.4 mL kg-1 ). There is also no short- or long-term toxicity when PAMB-G is injected into the myocardium of MI rats, with no significant changes in body weight, organ-brain ratio, hematologic, and histological parameters for up to 12 months post-injection. At the therapeutic dose, PAMB-G restores electrical conduction in infarcted rat hearts, resulting in lowered arrhythmia susceptibility and improved cardiac function. PAMB-G is also durable, as mass spectrometry detected the biopolymer for up to 12 months post-injection. PAMB-G did not impact reproductive organ function or offspring characteristics when given intravenously into healthy adult rats. Thus, PAMB-G is a nontoxic, durable, and conductive biomaterial that is able to improve cardiac function for up to 1 year post-implantation.

Toward Hierarchical Assembly of Aligned Cell Sheets into a Conical Cardiac Ventricle Using Microfabricated Elastomers.

Despite current efforts in organ-on-chip engineering to construct miniature cardiac models, they often lack some physiological aspects of the heart, including fiber orientation. This motivates the development of bioartificial left ventricle models that mimic the myofiber orientation of the native ventricle. Herein, an approach relying on microfabricated elastomers that enables hierarchical assembly of 2D aligned cell sheets into a functional conical cardiac ventricle is described. Soft lithography and injection molding techniques are used to fabricate micro-grooves on an elastomeric polymer scaffold with three different orientations ranging from -60° to +60°, each on a separate trapezoidal construct. The width of the micro-grooves is optimized to direct the majority of cells along the groove direction and while periodic breaks are used to promote cell-cell contact. The scaffold is wrapped around a central mandrel to obtain a conical-shaped left ventricle model inspired by the size of a human left ventricle 19 weeks post-gestation. Rectangular micro-scale holes are incorporated to alleviate oxygen diffusional limitations within the 3D scaffold. Cardiomyocytes within the 3D left ventricle constructs showed high viability in all layers after 7 days of cultivation. The hierarchically assembled left ventricle also provided functional readouts such as calcium transients and ejection fraction.

Epicardial delivery of a conductive membrane synchronizes conduction to reduce atrial fibrillation.

Conductive polymers have been investigated as a medium for the transmission of electrical signals in biological tissues, but their capacity to rewire cardiac tissue has not been evaluated. Myocardial tissue is unique in being able to generate an electrical potential at a fixed rate; this potential spreads rapidly among cells to trigger muscle contractions. Tissue injuries result in myocardial fibrosis and subsequent non-uniform conductivity, leading to arrhythmia. Atrial fibrillation (AF) is the most common sustained arrhythmia, associated with disruption of atrial electrical signaling, which can potentially be restored by the epicardial delivery of conductive polymers. In this work, poly-3-amino-4-methoxybenzoic acid, conjugated to gelatin, is fabricated as a membrane (PAMB-G) to support conductive velocities that are close to that of the myocardium. A cross-linked gelatin membrane (Gelatin) is used as a control. The as-fabricated PAMB-G has similar tensile elasticities, determined using the Young's modulus, as contracting myocardium; it can also transmit electrical signals to initiate cardiac cell and tissue excitation. Delivering PAMB-G onto the atrium of a rat AF model shortens AF duration and improves post-AF recovery for the duration of a 28-day-long study. Atrial tissue in the PAMB-G-implanted group has lower impedance, higher conduction velocity, and higher field potential amplitude than that in the Gelatin-implanted group. Therefore, the as-proposed PAMB-G is a suitable medium for restoring proper cardiac electrical signaling in AF hearts.

Polyethylenimine-mediated gene delivery into human bone marrow mesenchymal stem cells from patients.

Transplantation of mesenchymal stem cells (MSCs) derived from adult bone marrow has been proposed as a potential therapeutic approach for post-infarction left ventricular (LV) dysfunction. However, age-related functional decline of stem cells has restricted their clinical benefits after transplantation into the infarcted myocardium. The limitations imposed on patient cells could be addressed by genetic modification of stem cells. This study was designed to improve our understanding of genetic modification of human bone marrow derived mesenchymal stem cells (hMSCs) by polyethylenimine (PEI, branched with Mw 25 kD), one of non-viral vectors that show promise in stem cell genetic modification, in the context of cardiac regeneration for patients. We optimized the PEI-mediated reporter gene transfection into hMSCs, evaluated whether transfection efficiency is associated with gender or age of the cell donors, analysed the influence of cell cycle on transfection and investigated the transfer of therapeutic vascular endothelial growth factor gene (VEGF). hMSCs were isolated from patients with cardiovascular disease aged from 41 to 85 years. Optimization of gene delivery to hMSCs was carried out based on the particle size of the PEI/DNA complexes, N/P ratio of complexes, DNA dosage and cell viability. The highest efficiency with the cell viability near 60% was achieved at N/P ratio 2 and 6.0 µg DNA/cm(2) . The average transfection efficiency for all tested samples, middle-age group (<65 years), old-age group (>65 years), female group and male group was 4.32%, 3.85%, 4.52%, 4.14% and 4.38%, respectively. The transfection efficiency did not show any correlation either with the age or the gender of the donors. Statistically, there were two subpopulations in the donors; and transfection efficiency in each subpopulation was linearly related to the cell percentage in S phase. No significant phenotypic differences were observed between these two subpopulations. Furthermore, PEI-mediated therapeutic gene VEGF transfer could significantly enhance the expression level.

Injectable conductive hydrogel can reduce pacing threshold and enhance efficacy of cardiac pacemaker.

Background: Pacemaker implantation is currently used in patients with symptomatic bradycardia. Since a pacemaker is a lifetime therapeutic device, its energy consumption contributes to battery exhaustion, along with its voltage stimulation resulting in local fibrosis and greater resistance, which are all detrimental to patients. The possible resolution for those clinical issues is an injection of a conductive hydrogel, poly-3-amino-4-methoxybenzoic acid-gelatin (PAMB-G), to reduce the myocardial threshold voltage for pacemaker stimulation. Methods: PAMB-G is synthesized by covalently linking PAMB to gelatin, and its conductivity is measured using two-point resistivity. Rat hearts are injected with gelatin or PAMB-G, and pacing threshold is evaluated using electrocardiogram and cardiac optical mapping. Results: PAMB-G conductivity is 13 times greater than in gelatin. The ex vivo model shows that PAMB-G significantly enhances cardiac tissue stimulation. Injection of PAMB-G into the stimulating electrode location at the myocardium has a 4 times greater reduction of pacing threshold voltage, compared with electrode-only or gelatin-injected tissues. Multi-electrode array mapping reveals that the cardiac conduction velocity of PAMB-G group is significantly faster than the non- or gelatin-injection groups. PAMB-G also reduces pacing threshold voltage in an adenosine-induced atrial-ventricular block rat model. Conclusion: PAMB-G hydrogel reduces cardiac pacing threshold voltage, which is able to enhance pacemaker efficacy.

A conductive cell-delivery construct as a bioengineered patch that can improve electrical propagation and synchronize cardiomyocyte contraction for heart repair.

Cardiac tissue engineering is of particular importance in the combination of contracting cells with a biomaterial scaffold, which serves as a cell-delivery construct, to replace cardiomyocytes (CMs) that are lost as a result of an infarction, to restore heart function. However, most biomaterial scaffolds are nonconductive and may delay regional conduction, potentially causing arrhythmias. In this study, a conductive CM-delivery construct that consists of a gelatin-based gelfoam that is conjugated with a self-doped conductive polymer (poly-3-amino-4-methoxybenzoic acid, PAMB) is proposed as a cardiac patch (PAMB-Gel patch) to repair an infarcted heart. A nonconductive plain gelfoam (Gel patch) is used as a control. The electrical conductivity of the PAMB-Gel patch is approximately 30 times higher than that of the Gel patch; as a result, the conductive PAMB-Gel patch can substantially increase electrical conduction between distinct clusters of beating CMs, facilitating their synchronous contraction. In vivo epicardial implantation of the PAMB-Gel patch that is seeded with CMs (the bioengineered patch) in infarcted rat hearts can significantly enhance electrical activity in the fibrotic tissue, improving electrical impulse propagation and synchronizing CM contraction across the scar region, markedly reducing its susceptibility to cardiac arrhythmias. Echocardiography shows that the bioengineered conductive patch has an important role in the restoration of cardiac function, perhaps owing to the synergistic effects of its conductive construct and the synchronously beating CMs. These experimental results reveal that the as-proposed bioengineered conductive patch has great potential for repairing injured cardiac tissues.

A self-doping conductive polymer hydrogel that can restore electrical impulse propagation at myocardial infarct to prevent cardiac arrhythmia and preserve ventricular function.

Following myocardial infarction (MI), necrotic cardiomyocytes (CMs) are replaced by fibroblasts and collagen tissue, causing abnormal electrical signal propagation, desynchronizing cardiac contraction, resulting in cardiac arrhythmia. In this work, a conductive polymer, poly-3-amino-4-methoxybenzoic acid (PAMB), is synthesized and grafted onto non-conductive gelatin. The as-synthesized PAMB-G copolymer is self-doped in physiological pH environments, making it an electrically active material in biological tissues. This copolymer is cross-linked by carbodiimide to form an injectable conductive hydrogel (PAMB-G hydrogel). The un-grafted gelatin hydrogel is prepared in a similar manner as a control. Both test hydrogels not only provide an optimal matrix for CM adhesion and growth but also maintain CM morphology and functional proteins. The conductivity of PAMB-G hydrogel is ca. 12 times higher than that of gelatin hydrogel. Microelectrode array analyses reveal that a heart placed on the PAMB-G hydrogel has a higher field potential amplitude than that placed on the gelatin hydrogel and can pass current from one heart to excite another heart at a distance. The injection of PAMB-G hydrogel into the scar zone following an MI in a rat heart improves electrical impulse propagation over that in a heart that has been treated with gelatin hydrogel, and synchronizes heart contraction, leading to preservation of the ventricular function and reduction of cardiac arrhythmia, demonstrating its potential for use in treating MI.

Enhanced thoracic gene delivery by magnetic nanobead-mediated vector.

BACKGROUND: Systemic gene delivery is limited by the adverse hydrodynamic conditions on the collection of gene carrier particles to the specific area. In the present study, a magnetic field was employed to guide magnetic nanobead (MNB)/polymer/DNA complexes after systemic administration to the left side of the mouse thorax in order to induce localized gene expression. METHODS: Nonviral polymer (poly ethyleneimine, PEI) vector-gene complexes were conjugated to MNBs with the Sulfo-NHS-LC-Biotin linker. In vitro transfection efficacy of MNB/PEI/DNA was compared with PEI/DNA in three different cell lines as well as primary endothelial cells under magnetic field stimulation. In vivo, MNB/PEI/DNA complexes were injected into the tail vein of mice and an epicardial magnet was employed to attract the circulating MNB/PEI/DNA complexes. RESULTS: Endocytotic uptake of MNB/PEI/DNA complexes and intracellular gene release with nuclear translocation were observed in vitro, whereas the residues of MNB/PEI complexes were localized at the perinuclear region. Compared with PEI/DNA complexes alone, MNB/PEI/DNA complexes had a 36- to 85-fold higher transfection efficiency under the magnetic field. In vivo, the epicardial magnet effectively attracted MNB/PEI/DNA complexes in the left side of the thorax, resulting in strong reporter and therapeutic gene expression in the left lung and the heart. Gene expression in the heart was mainly within the endothelium. CONCLUSIONS: MNB-mediated gene delivery could comprise a promising method for gene delivery to the lung and the heart.

Bioactive coating of decellularized vascular grafts with a temperature-sensitive VEGF-conjugated hydrogel accelerates autologous endothelialization in vivo.

No ideal small-diameter vascular graft for widespread clinical application has yet been developed and current approaches still suffer from graft failure because of thrombosis or degeneration. Decellularized vascular grafts are a promising strategy as they preserve native vessel architecture while eliminating cell-based antigens and allow for autologous recellularization. In the present study, a functional in vivo rodent aortic transplantation model was used in order to evaluate the benefit of bioactive coating of decellularized vascular grafts with vascular endothelial growth factor (VEGF) conjugated to a temperature-sensitive aliphatic polyester hydrogel (HG). Luminal HG-VEGF coating persistence up to 4 weeks was confirmed in vivo by rhodamine-labelling. Doppler-sonography showed that the grafts were functional for up to 8 weeks in vivo. Histological and immunohistochemical analysis of the explanted grafts after 4 weeks and 8 weeks in vivo demonstrated significantly increased endothelium formation in the HG-VEGF group compared with the control group (luminal surface covered with single-layered endothelium, 4 weeks: 64.8 ± 7.6% vs. 40.4 ± 8.3%, p = 0.025) as well as enhanced media recellularization (absolute cell count, 8 weeks: 22.1 ± 13.0 vs. 3.2 ± 3.6, p = 0.0039). However, HG-VEGF coating also led to increased neo-intimal hyperplasia, resulting in a significantly increased intima-to-media ratio in the perianastomotic regions (intima-to-media ratio, 8 weeks: 1.61 ± 0.17 vs. 0.93 ± 0.09, p = 0.008; HG-VEGF vs. control). The findings indicate that HG-VEGF coating has potential for the development of engineered small-diameter artificial grafts, although further research is needed to prevent neo-intimal hyperplasia. Copyright © 2016 John Wiley & Sons, Ltd.

Preservation of conductive propagation after surgical repair of cardiac defects with a bio-engineered conductive patch.

BACKGROUND: Both stable and biodegradable biomaterials have been used to surgically repair congenital cardiac defects. However, neither type of biomaterial can conduct electrical activity. We evaluated the conductivity and efficacy of a newly synthesized conductive polypyrrole-chitosan (Ppy+Chi) gelfoam patch to support cardiomyocyte (CM) viability and function in vitro and to surgically repair a cardiac defect in vivo. METHODS: Ppy+Chi was incorporated into gelfoam (Gel) to form a 3-dimensional conductive patch. In vitro, patch characteristics were evaluated and biocompatibility and bioconductivity were investigated by culturing neonatal rat CMs on the patches. In vivo, a full-thickness right ventricular outflow tract defect was created in rats and the patches were implanted. Four weeks after patch repair, cardiac electrical activation and conduction velocity were evaluated using an optical mapping system. RESULTS: In vitro, the Ppy+Chi+Gel patch had a higher mean breaking stress than the Gel or Chi+Gel patches, and the highest conductivity. None of the patches altered cell growth. The Ca2+ transient velocity of CMs cultured on the Ppy+Chi+Gel patch was 2.5-fold higher than that of CMs cultured on the Gel or Chi+Gel patches. In vivo, optical mapping at 4 weeks post-implantation demonstrated that Ppy+Chi+Gel patch-implanted hearts had faster conduction velocities, as measured on the epicardial surface. Continuous electrocardiographic telemetry did not reveal any pathologic arrhythmias after patch implantation. Ex-vivo patch conductivity testing also revealed that the Ppy+Chi+Gel patch was more conductive than the Gel and Chi+Gel patches. CONCLUSIONS: The Ppy+Chi+Gel patch was biocompatible, safe and conductive, making it an attractive candidate for a new biomaterial platform for cardiac surgical repair to preserve synchronous ventricular contraction.

Polypyrrole-chitosan conductive biomaterial synchronizes cardiomyocyte contraction and improves myocardial electrical impulse propagation.

Background: The post-myocardial infarction (MI) scar interrupts electrical impulse propagation and delays regional contraction, which contributes to ventricular dysfunction. We investigated the potential of an injectable conductive biomaterial to restore scar tissue conductivity and re-establish synchronous ventricular contraction. Methods: A conductive biomaterial was generated by conjugating conductive polypyrrole (PPY) onto chitosan (CHI) backbones. Trypan blue staining of neonatal rat cardiomyocytes (CMs) cultured on biomaterials was used to evaluate the biocompatibility of the conductive biomaterials. Ca2+ imaging was used to visualize beating CMs. A cryoablation injury rat model was used to investigate the ability of PPY:CHI to improve cardiac electrical propagation in the injured heart in vivo. Electromyography was used to evaluate conductivity of scar tissue ex vivo. Results: Cell survival and morphology were similar between cells cultured on biomaterials-coated and uncoated-control dishes. PPY:CHI established synchronous contraction of two distinct clusters of spontaneously-beating CMs. Intramyocardial PPY:CHI injection into the cryoablation-induced injured region improved electrical impulse propagation across the scarred tissue and decreased the QRS interval, whereas saline- or CHI-injected hearts continued to have delayed propagation patterns and significantly reduced conduction velocity compared to healthy controls. Ex vivo evaluation found that scar tissue from PPY:CHI-treated rat hearts had higher signal amplitude compared to those from saline- or CHI-treated rat heart tissue. Conclusions: The PPY:CHI biomaterial is electrically conductive, biocompatible and injectable. It improved synchronous contraction between physically separated beating CM clusters in vitro. Intra-myocardial injection of PPY:CHI following cardiac injury improved electrical impulse propagation of scar tissue in vivo.

VEGF-loaded microsphere patch for local protein delivery to the ischemic heart.

BACKGROUND: Revascularization of the heart after myocardial infarction (MI) using growth factors delivered by hydrogel-based microspheres represents a promising therapeutic approach for cardiac regeneration. Microspheres have tuneable degradation properties and support the prolonged release of soluble factors. Cardiac patches provide mechanical restraint, preventing dilatation associated with ventricular remodelling. METHODS: We combined these approaches and produced a compacted calcium-alginate microsphere patch, restrained by a chitosan sheet, to deliver vascular endothelial growth factor (VEGF) to the heart after myocardial injury in rats. RESULTS: Microspheres had an average diameter of 3.2µm, were nonporous, and characterized by a smooth dimpled surface. Microsphere patches demonstrated prolonged in vitro release characteristics compared to non-compacted microspheres and VEGF supernatants obtained from patches maintained their bioactivity for the 5day duration of the study in vitro. In vivo, patches were assessed with magnetic resonance imaging following MI, and demonstrated 50% degradation 25.6days after implantation. Both VEGF(-) and VEGF(+) microsphere patch-treated hearts had better cardiac function than unpatched (chitosan sheet only) controls. However, VEGF(+) microsphere-patched hearts had thicker scars characterized by higher capillary density in the border zone than did those treated with VEGF(-) patches. VEGF was detected in the patches 4weeks post-implantation. CONCLUSION: The condensed microsphere patch represents a new therapeutic platform for cytokine delivery and could be used as an adjuvant to current biomaterial and cell-based therapies to promote localized angiogenesis in the infarcted heart. STATEMENT OF SIGNIFICANCE: Following a heart attack, a lack of blood flow to the heart results in loss of heart cells. Growth factors may facilitate growth of blood vessels and heart tissue repair and prevent the onset of heart failure. Determining a way to deliver these growth factors directly to the heart is vital. Here, we combined two biomaterial-based approaches to deliver vascular endothelial growth factor (VEGF) to rat hearts after heart attack: a microsphere for prolonged release of VEGF, and a cardiac patch for mechanical restraint to prevent heart dysfunction. The feasibility of this microsphere patch was demonstrated by surgically implanting it over the infarct region of the heart post-injury. VEGF-patched hearts had better blood vessel growth, tissue repair, and heart function.

Optimal biomaterial for creation of autologous cardiac grafts.

BACKGROUND: The optimal cardiac graft for the repair of congenital heart defects will be composed of autologous cells and will grow with the child. The biodegradable material should permit rapid cellular growth and delayed degradation with minimal inflammation. We compared a new material, epsilon-caprolactone-co-L-lactide sponge reinforced with knitted poly-L-lactide fabric (PCLA), to gelatin (GEL) and polyglycolic acid (PGA), which are previously evaluated materials. METHODS: Syngenic rat aortic smooth muscle cells (SMCs, 2x10(6)) were seeded onto GEL, PGA, and PCLA patches and cultured (n=11 per group). The DNA content in each patch was measured at 1, 2, and 3 weeks after seeding. Histological examination was performed 2 weeks after seeding. Cell-seeded patches were employed to replace a surgically created defect in the right ventricular outflow tract (RVOT) of rats (n=5 per group). Histology was studied at 8 weeks following implantation. RESULTS: In vitro studies showed that the DNA content increased significantly (P<0.05) in all patches between 1 and 3 weeks after seeding. Histology and staining SMCs for anti-alpha-smooth muscle actin (alphaSMA) revealed better growth of cells in the interstices of the grafts with GEL and PCLA than the PGA graft. In vivo studies demonstrated that seeded SMCs survived at least 8 weeks after the patch implantation in all groups. PCLA scaffolds were replaced by more cells with larger alphaSMA-positive areas and by more extracellular matrix with larger elastin-positive areas than with GEL and PGA. The patch did not thin and expanded significantly. The GEL and PGA patches thinned and expanded. All grafts had complete endothelialization on the endocardial surface. CONCLUSIONS: SMC-seeded biodegradable materials can be employed to repair the RVOT. The novel PCLA patches permitted better cellular penetration in vitro and did not thin or dilate in vivo and did not produce an inflammatory response. The cell-seeded PCLA patch may permit the construction of an autologous patch to repair congenital heart defects.

Histologic changes of nonbiodegradable and biodegradable biomaterials used to repair right ventricular heart defects in rats.

OBJECTIVES: Nonbiodegradable synthetic materials have been widely used to repair cardiac defects. Material-related failures, however, such as lack of growth, thrombosis, and infection, do occur. Because a biodegradable scaffold can be replaced by the patient's own cells and will be treated as a foreign body for a limited period, we compared four biodegradable materials (gelatin, polyglycolic acid (PGA), and copolymer made of epsilon-caprolactone and l-lactic acid reinforced with a poly-l-lactide knitted [KN-PCLA] or woven fabric [WV-PCLA]) with a nonbiodegradable polytetrafluoroethylene (PTFE) material. An animal heart model was tested that simulates the in vivo clinical condition to which a synthetic material would be used. METHODS: The five patches were used to repair transmural defects surgically created in the right ventricular outflow tracts of adult rat hearts (n = 5, each patch group). The PTFE patch group served as a control group. At 8 weeks after implantation, the biomaterials were excised. Patch size, patch thickness, infiltrated cell number, extracellular matrix composition, and patch degradation were evaluated. RESULTS: The PTFE patch itself did not change in size except for increasing in thickness because of fibroblast and collagen coverage of both its surfaces. Host cells did not migrate into the PTFE biomaterial. In contrast, cells migrated into the biodegrading gelatin, PGA, and KN-PCLA and WV-PCLA scaffolds. Cellular ingrowth per unit patch area was highest in the KN-PCLA patch. The KN-PCLA patch increased modestly in size and thinness. The WV-PCNA patch did not change in size or thickness. Fibroblasts and collagen were the dominant cellular infiltrate and extracellular matrix formed in the biodegrading scaffolds. The in vivo rates of biomaterial degradation, thinning, and expansion were material specific. All the subendocardial patch surfaces were covered with endothelial cells. No thrombi were seen. CONCLUSIONS: The unique, spongy matrix structure of the PCLA patch favored cell colonization relative to the other patches. The strong, durable outer poly-l-lactide fabric layers in these patches offered physical, biocompatible, and bioresorbable advantages relative to the other biodegradable materials studied. Host cells migrated into all the biomaterials. The cells secreted matrix and formed tissue, which was endothelialized on the endocardial surface. The biomaterial degradation rates and the tissue formation rates were material related. The PCLA grafts hold promise to become a suitable patch for surgical repair.