A Workflow to Produce a Low-Cost In Vitro Platform for the Electric-Field Pacing of Cellularised 3D Porous Scaffolds.

Endogenous electrically mediated signaling is a key feature of most native tissues, the most notable examples being the nervous and the cardiac systems. Biomedical engineering often aims to harness and drive such activity in vitro, in bioreactors to study cell disease and differentiation, and often in three-dimensional (3D) formats with the help of biomaterials, with most of these approaches adopting scaffold-free self-assembling strategies to create 3D tissues. In essence, this is the casting of gels which self-assemble in response to factors such as temperature or pH and have capacity to harbor cells during this process without imparting toxicity. However, the use of materials that do not self-assemble but can support 3D encapsulation of cells (such as porous scaffolds) warrants consideration given the larger repertoire this would provide in terms of material physicochemical properties and microstructure. In this method and protocol paper, we detail and provide design codes and assembly instructions to cheaply create an electrical pacing bioreactor and a Rig for Stimulation of Sponge-like Scaffolds (R3S). This setup has also been engineered to simultaneously perform live optical imaging of the in vitro models. To showcase a pilot exploration of material physiochemistry (in this aspect material conductivity) and microstructure (isotropy versus anisotropy), we adopt isotropic and anisotropic porous scaffolds composed of collagen or poly(3,4-ethylene dioxythiophene):polystyrenesulfonate (PEDOT:PSS) for their contrasting conductivity properties yet similar in porosity and mechanical integrity. Electric field pacing of mouse C3H10 cells on anisotropic porous scaffolds placed in R3S led to increased metabolic activity and enhanced cell alignment. Furthermore, after 7 days electrical pacing drove C3H10 alignment regardless of material conductivity or anisotropy. This platform and its design, which we have shared, have wide suitability for the study of electrical pacing of cellularized scaffolds in 3D in vitro cultures.

MXene functionalized collagen biomaterials for cardiac tissue engineering driving iPSC-derived cardiomyocyte maturation.

Electroconductive biomaterials are gaining significant consideration for regeneration in tissues where electrical functionality is of crucial importance, such as myocardium, neural, musculoskeletal, and bone tissue. In this work, conductive biohybrid platforms were engineered by blending collagen type I and 2D MXene (Ti3C2Tx) and afterwards covalently crosslinking; to harness the biofunctionality of the protein component and the increased stiffness and enhanced electrical conductivity (matching and even surpassing native tissues) that two-dimensional titanium carbide provides. These MXene platforms were highly biocompatible and resulted in increased proliferation and cell spreading when seeded with fibroblasts. Conversely, they limited bacterial attachment (Staphylococcus aureus) and proliferation. When neonatal rat cardiomyocytes (nrCMs) were cultured on the substrates increased spreading and viability up to day 7 were studied when compared to control collagen substrates. Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) were seeded and stimulated using electric-field generation in a custom-made bioreactor. The combination of an electroconductive substrate with an external electrical field enhanced cell growth, and significantly increased cx43 expression. This in vitro study convincingly demonstrates the potential of this engineered conductive biohybrid platform for cardiac tissue regeneration.

Beyond Chemistry: Tailoring Stiffness and Microarchitecture to Engineer Highly Sensitive Biphasic Elastomeric Piezoresistive Sensors.

Carbon-based nanoparticles and conductive polymers are two classes of materials widely used in the production of three-dimensional (3D) piezoresistive sensors. One conductive polymer, poly(3,4-ethylenedioxythiophene):polystyrenesulfonate (PEDOT:PSS) has excellent stability and conductivity yet is limited in its application as a sensor, often existing upon a base, limiting its performance and potential. Despite much progress in the field of materials chemistry and polymer synthesis, one aspect we consider worthy of exploration is the impact that microstructure and stiffness may have on the sensitivity of 3D sensors. In this study, we report a strategy for fabricating biphasic electroactive sponges (EAS) that combine 3D porous PEDOT:PSS scaffolds possessing either an isotropic or anisotropic microarchitecture, infused with insulating elastomeric fillers of varying stiffness. When characterizing the electromechanical behavior of these EAS, a higher stiffness yields a higher strain gauge factor, with values as high as 387 for an isotropic microarchitecture infused with a stiff elastomer. The approach we describe is cost-effective and extremely versatile, by which one can fabricate piezoresistive sensors with adaptable sensitivity ranges and excellent high strain gauge factor with the underlying microarchitecture and insulant stiffness dictating this performance.

Structural crystallisation of crosslinked 3D PEDOT:PSS anisotropic porous biomaterials to generate highly conductive platforms for tissue engineering applications.

An emerging class of materials finding applications in biomaterials science - conductive polymers (CPs) - enables the achievement of smarter electrode coatings, piezoresistive components within biosensors, and scaffolds for tissue engineering. Despite their advances in recent years, there exist still some challenges which have yet to be addressed, such as long-term stability under physiological conditions, adequate long-term conductivity and optimal biocompatibility. Additionally, another hurdle to the use of these materials is their adaptation towards three-dimensional (3D) scaffolds, a feature that is usually achieved by virtue of applying CPs as a functionalised coating on a bulk material. Poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) is by far one of the most promising CPs in terms of its stability and conductivity, with the latter capable of being enhanced via a crystallisation treatment using sulphuric acid. In this work, we present a new generation of 3D electroconductive porous biomaterial scaffolds based on PEDOT:PSS crosslinked via glycidoxypropyltrimethoxysilane (GOPS) and subjected to sulphuric acid crystallisation. The resultant isotropic and anisotropic crystallised porous scaffolds exhibited, on an average, a 1000-fold increase in conductivity when compared with the untreated scaffolds. Moreover, we also document a precise control over the pore microarchitecture, size and anisotropy with high repeatability to achieve both isotropic and aligned scaffolds with mechanical and electrical anisotropy, while exhibiting adequate biocompatibility. These findings herald a new approach towards generating anisotropic porous biomaterial scaffolds with superior conductivity through a safe and scalable post-treatment.

The rationale and emergence of electroconductive biomaterial scaffolds in cardiac tissue engineering.

The human heart possesses minimal regenerative potential, which can often lead to chronic heart failure following myocardial infarction. Despite the successes of assistive support devices and pharmacological therapies, only a whole heart transplantation can sufficiently address heart failure. Engineered scaffolds, implantable patches, and injectable hydrogels are among the most promising solutions to restore cardiac function and coax regeneration; however, current biomaterials have yet to achieve ideal tissue regeneration and adequate integration due a mismatch of material physicochemical properties. Conductive fillers such as graphene, carbon nanotubes, metallic nanoparticles, and MXenes and conjugated polymers such as polyaniline, polypyrrole, and poly(3,4-ethylendioxythiophene) can possibly achieve optimal electrical conductivities for cardiac applications with appropriate suitability for tissue engineering approaches. Many studies have focused on the use of these materials in multiple fields, with promising effects on the regeneration of electrically active biological tissues such as orthopedic, neural, and cardiac tissue. In this review, we critically discuss the role of heart electrophysiology and the rationale toward the use of electroconductive biomaterials for cardiac tissue engineering. We present the emerging applications of these smart materials to create supportive platforms and discuss the crucial role that electrical stimulation has been shown to exert in maturation of cardiac progenitor cells.

PEDOT:PSS interfaces stabilised using a PEGylated crosslinker yield improved conductivity and biocompatibility.

The rapidly expanding fields of bioelectronics, and biological interfaces with electronic sensors and stimulators, are placing an increasing demand on candidate materials to serve as robust surfaces that are both biocompatible, stable and electroconductive. Amongst conductive polymers, poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) is a promising material in biomedical research due to its appropriate stability and high conductivity, however its intrinsic solubility requires a crosslinking process that can limit its conductivity and biocompatibility. Poly(ethylene glycol) is known to be a suitably anti-immunogenic moiety and its derivatives have been widely used for biomedical applications. In this study we investigate the application of poly(ethylene glycol)diglycidyl ether (PEGDE) as an effective crosslinker and conductive filler for PEDOT:PSS. From our interpretation of XPS analysis we hypothesise that the crosslinking reaction is occurring via the epoxy ring of PEGDE interacting with the sulfonic groups of excel PSS chains, which reaches a saturation at 3 w/v% PEGDE concentration. PEGDE crosslinked films did not disperse in aqueous environments, had enhanced electrical conductivity and imparted a significant degree of hydrophilicity to PEDOT:PSS films. This hydrophilicity and the presence of biocompatible PEGDE led to good cell viability and a significantly increased degree of cell spreading on PEDOT:PSS films. In comparison to widely reported chemical crosslinking via glycidoxy propyltrimethoxysilane (GOPS), this original crosslinking yields a highly hydrophilic 2D film substrate with increased electroconductive and biocompatibility properties, resulting in a next-generation formulation for bioengineering applications.