Itaconate and citrate releasing polymer attenuates foreign body response in biofabricated cardiac patches.

Application of cardiac patches to the heart surface can be undertaken to provide support and facilitate regeneration of the damaged cardiac tissue following ischemic injury. Biomaterial composition is an important consideration in the design of cardiac patch materials as it governs host response to ultimately prevent the undesirable fibrotic response. Here, we investigate a novel patch material, poly (itaconate-co-citrate-co-octanediol) (PICO), in the context of cardiac implantation. Citric acid (CA) and itaconic acid (ITA), the molecular components of PICO, provided a level of protection for cardiac cells during ischemic reperfusion injury in vitro. Biofabricated PICO patches were shown to degrade in accelerated and hydrolytic conditions, with CA and ITA being released upon degradation. Furthermore, the host response to PICO patches after implantation on rat epicardium in vivo was explored and compared to two biocompatible cardiac patch materials, poly (octamethylene (anhydride) citrate) (POMaC) and poly (ethylene glycol) diacrylate (PEGDA). PICO patches resulted in less macrophage infiltration and lower foreign body giant cell reaction compared to the other materials, with corresponding reduction in smooth muscle actin-positive vessel infiltration into the implant region. Overall, this work demonstrates that PICO patches release CA and ITA upon degradation, both of which demonstrate cardioprotective effects on cardiac cells after ischemic injury, and that PICO patches generate a reduced inflammatory response upon implantation to the heart compared to other materials, signifying promise for use in cardiac patch applications.

Elastomeric Polyesters in Cardiovascular Tissue Engineering and Organs-on-a-Chip.

Cardiovascular tissue constructs provide unique design requirements due to their functional responses to substrate mechanical properties and cyclic stretching behavior of cardiac tissue that requires the use of durable elastic materials. Given the diversity of polyester synthesis approaches, an opportunity exists to develop a new class of biocompatible, elastic, and immunomodulatory cardiovascular polymers. Furthermore, elastomeric polyester materials have the capability to provide tailored biomechanical synergy with native tissue and hence reduce inflammatory response in vivo and better support tissue maturation in vitro. In this review, we highlight underlying chemistry and design strategies of polyester elastomers optimized for cardiac tissue scaffolds. The major advantages of these materials such as their tunable elasticity, desirable biodegradation, and potential for incorporation of bioactive compounds are further expanded. Unique fabrication methods using polyester materials such as micromolding, 3D stamping, electrospinning, laser ablation, and 3D printing are discussed. Moreover, applications of these biomaterials in cardiovascular organ-on-a-chip devices and patches are analyzed. Finally, we outline unaddressed challenges in the field that need further study to enable the impactful translation of soft polyesters to clinical applications.

Transfer learning in a biomaterial fibrosis model identifies in vivo senescence heterogeneity and contributions to vascularization and matrix production across species and diverse pathologies.

Cellular senescence is a state of permanent growth arrest that plays an important role in wound healing, tissue fibrosis, and tumor suppression. Despite senescent cells' (SnCs) pathological role and therapeutic interest, their phenotype in vivo remains poorly defined. Here, we developed an in vivo-derived senescence signature (SenSig) using a foreign body response-driven fibrosis model in a p16-CreERT2;Ai14 reporter mouse. We identified pericytes and "cartilage-like" fibroblasts as senescent and defined cell type-specific senescence-associated secretory phenotypes (SASPs). Transfer learning and senescence scoring identified these two SnC populations along with endothelial and epithelial SnCs in new and publicly available murine and human data single-cell RNA sequencing (scRNAseq) datasets from diverse pathologies. Signaling analysis uncovered crosstalk between SnCs and myeloid cells via an IL34-CSF1R-TGFßR signaling axis, contributing to tissue balance of vascularization and matrix production. Overall, our study provides a senescence signature and a computational approach that may be broadly applied to identify SnC transcriptional profiles and SASP factors in wound healing, aging, and other pathologies.

Harnessing conserved signaling and metabolic pathways to enhance the maturation of functional engineered tissues.

The development of induced-pluripotent stem cell (iPSC)-derived cell types offers promise for basic science, drug testing, disease modeling, personalized medicine, and translatable cell therapies across many tissue types. However, in practice many iPSC-derived cells have presented as immature in physiological function, and despite efforts to recapitulate adult maturity, most have yet to meet the necessary benchmarks for the intended tissues. Here, we summarize the available state of knowledge surrounding the physiological mechanisms underlying cell maturation in several key tissues. Common signaling consolidators, as well as potential synergies between critical signaling pathways are explored. Finally, current practices in physiologically relevant tissue engineering and experimental design are critically examined, with the goal of integrating greater decision paradigms and frameworks towards achieving efficient maturation strategies, which in turn may produce higher-valued iPSC-derived tissues.

Biomaterials direct functional B cell response in a material-specific manner.

B cells are an adaptive immune target of biomaterials development in vaccine research but, despite their role in wound healing, have not been extensively studied in regenerative medicine. To probe the role of B cells in biomaterial scaffold response, we evaluated the B cell response to biomaterial materials implanted in a muscle wound using a biological extracellular matrix (ECM), as a reference for a naturally derived material, and synthetic polyester polycaprolactone (PCL), as a reference for a synthetic material. In the local muscle tissue, small numbers of B cells are present in response to tissue injury and biomaterial implantation. The ECM materials induced mature B cells in lymph nodes and antigen presentation in the spleen. The synthetic PCL implants resulted in prolonged B cell presence in the wound and induced an antigen-presenting phenotype. In summary, the adaptive B cell immune response to biomaterial induces local, regional, and systemic immunological changes.

A well plate-based multiplexed platform for incorporation of organoids into an organ-on-a-chip system with a perfusable vasculature.

Owing to their high spatiotemporal precision and adaptability to different host cells, organ-on-a-chip systems are showing great promise in drug discovery, developmental biology studies and disease modeling. However, many current micro-engineered biomimetic systems are limited in technological application because of culture media mixing that does not allow direct incorporation of techniques from stem cell biology, such as organoids. Here, we describe a detailed alternative method to cultivate millimeter-scale functional vascularized tissues on a biofabricated platform, termed 'integrated vasculature for assessing dynamic events', that enables facile incorporation of organoid technology. Utilizing the 3D stamping technique with a synthetic polymeric elastomer, a scaffold termed 'AngioTube' is generated with a central microchannel that has the mechanical stability to support a perfusable vascular system and the self-assembly of various parenchymal tissues. We demonstrate an increase in user familiarity and content analysis by situating the scaffold on a footprint of a 96-well plate. Uniquely, the platform can be used for facile connection of two or more tissue compartments in series through a common vasculature. Built-in micropores enable the studies of cell invasion involved in both angiogenesis and metastasis. We describe how this protocol can be applied to create both vascularized cardiac and hepatic tissues, metastatic breast cancer tissue and personalized pancreatic cancer tissue through incorporation of patient-derived organoids. Platform assembly to populating the scaffold with cells of interest into perfusable functional vascularized tissue will require 12-14 d and an additional 4 d if pre-polymer and master molds are needed.

An Organ-on-a-Chip System to Study Anaerobic Bacteria in Intestinal Health and Disease.

A state-of-the-art organ-on-a-chip system enabled co-culture of highly oxygen-sensitive Faecalibacterium prausnitzii with human intestinal epithelial cells, recapitulating a critical role of the bacteria in the maintenance of anti-inflammatory phenotype of gut epithelium mediated by bacteria-secreted butyrate.

Enhancing senior high school student engagement and academic performance using an inclusive and scalable inquiry-based program.

The multi-disciplinary nature of science, technology, engineering, and math (STEM) careers often renders difficulty for high school students navigating from classroom knowledge to post-secondary pursuits. Discrepancies between the knowledge-based high school learning approach and the experiential approach of future studies leaves some students disillusioned by STEM. We present Discovery, a term-long inquiry-focused learning model delivered by STEM graduate students in collaboration with high school teachers, in the context of biomedical engineering. Entire classes of high school STEM students representing diverse cultural and socioeconomic backgrounds engaged in iterative, problem-based learning designed to emphasize critical thinking concomitantly within the secondary school and university environments. Assessment of grades and survey data suggested positive impact of this learning model on students' STEM interests and engagement, notably in under-performing cohorts, as well as repeating cohorts that engage in the program on more than one occasion. Discovery presents a scalable platform that stimulates persistence in STEM learning, providing valuable learning opportunities and capturing cohorts of students that might otherwise be under-engaged in STEM.

Elastic Biomaterial Scaffold with Spatially Varying Adhesive Design.

In order to secure biomaterials to tissue surfaces, sutures or glues are commonly used. Of interest is the development of a biomaterial patch for applications in tissue engineering and regeneration that incorporates an adhesive component to simplify patch application and ensure sufficient adhesion. A separate region dedicated to fulfilling the specific requirements of an application such as mechanical support or tissue delivery is also desirable. Here, the design and fabrication of a unique patch are presented with distinct regions for adhesion and function, resulting in a biomaterial patch resembling the Band-Aid. The adhesive region contains a novel polymer, synthesized to incorporate a molecule capable of adhesion to tissue, dopamine. The desired polymer composition for patch development is selected based on chemical assessment and evaluation of key physical properties such as swelling and elastic modulus, which are tailored for use in soft tissue applications. The selected polymer formulation, referred to as the adhesive patch (AP) polymer, demonstrates negligible cytotoxicity and improves adhesive capability to rat cardiac tissue compared to currently used patch materials. Finally, the AP polymer is used in the patch, designed to possess distinct adhesive and nonadhesive domains, presenting a novel design for the next generation of biomaterials.

Recapitulating pancreatic tumor microenvironment through synergistic use of patient organoids and organ-on-a-chip vasculature.

Tumor progression relies heavily on the interaction between the neoplastic epithelial cells and their surrounding stromal partners. This cell cross-talk affects stromal development, and ultimately the heterogeneity impacts drug efflux and efficacy. To mimic this evolving paradigm, we have micro-engineered a three-dimensional (3D) vascularized pancreatic adenocarcinoma tissue in a tri-culture system composed of patient derived pancreatic organoids, primary human fibroblasts and endothelial cells on a perfusable InVADE platform situated in a 96-well plate. Uniquely, through synergistic engineering we combined the benefits of cellular fidelity of patient tumor derived organoids with the addressability of a plastic organ-on-a-chip platform. Validation of this platform included demonstrating the growth of pancreatic tumor organoids by monitoring the change in metabolic activity of the tissue. Investigation of tumor microenvironmental behavior highlighted the role of fibroblasts in symbiosis with patient organoid cells, resulting in a six-fold increase of collagen deposition and a corresponding increase in tissue stiffness in comparison to fibroblast free controls. The value of a perfusable vascular network was evident in drug screening, as perfusion of gemcitabine into a stiffened matrix did not show the dose-dependent effects on tumor viability as those under static conditions. These findings demonstrate the importance of studying the dynamic synergistic relationship between patient cells with stromal fibroblasts, in a 3D perfused vascular network, to accurately understand and recapitulate the tumor microenvironment.

3D Printing of Vascular Tubes Using Bioelastomer Prepolymers by Freeform Reversible Embedding.

Bioelastomers have been extensively used in tissue engineering applications because of favorable mechanical stability, tunable properties, and chemical versatility. As these materials generally possess low elastic modulus and relatively long gelation time, it is challenging to 3D print them using traditional techniques. Instead, the field of 3D printing has focused preferentially on hydrogels and rigid polyester materials. To develop a versatile approach for 3D printing of elastomers, we used freeform reversible embedding of suspended prepolymers. A family of novel fast photocrosslinakble bioelastomer prepolymers were synthesized from dimethyl itaconate, 1,8-octanediol, and triethyl citrate. Tensile testing confirmed their elastic properties with Young's moduli in the range of 11-53 kPa. These materials supported cultivation of viable cells and enabled adhesion and proliferation of human umbilical vein endothelial cells. Tubular structures were created by embedding the 3D printed microtubes within a secondary hydrogel that served as a temporary support. Upon photocrosslinking and porogen leaching, the polymers were permeable to small molecules (TRITC-dextran). The polymer microtubes were assembled on the 96-well plates custom made by hot-embossing, as a tool to connect multiple organs-on-a-chip. The endothelialization of the tubes was performed to confirm that these microtubes can be utilized as vascular tubes to support parenchymal tissues seeded on them.

Biowire Model of Interstitial and Focal Cardiac Fibrosis.

Myocardial fibrosis is a severe global health problem due to its prevalence in all forms of cardiac diseases and direct role in causing heart failure. The discovery of efficient antifibrotic compounds has been hampered due to the lack of a physiologically relevant disease model. Herein, we present a disease model of human myocardial fibrosis and use it to establish a compound screening system. In the Biowire II platform, cardiac tissues are suspended between a pair of poly(octamethylene maleate (anhydride) citrate) (POMaC) wires. Noninvasive functional readouts are realized on the basis of the deflection of the intrinsically fluorescent polymer. The disease model is constructed to recapitulate contractile, biomechanical, and electrophysiological complexities of fibrotic myocardium. Additionally, we constructed a heteropolar integrated model with fibrotic and healthy cardiac tissues coupled together. The integrated model captures the regional heterogeneity of scar lesion, border zone, and adjacent healthy myocardium. Finally, we demonstrate the utility of the system for the evaluation of antifibrotic compounds. The high-fidelity in vitro model system combined with convenient functional readouts could potentially facilitate the development of precision medicine strategies for cardiac fibrosis modeling and establish a pipeline for preclinical compound screening.

One-Pot Synthesis of Unsaturated Polyester Bioelastomer with Controllable Material Curing for Microscale Designs.

Synthetic polyester elastomeric constructs have become increasingly important for a range of healthcare applications, due to tunable soft elastic properties that mimic those of human tissues. A number of these constructs require intricate mechanical design to achieve a tunable material with controllable curing. Here, the synthesis and characterization of poly(itaconate-co-citrate-co-octanediol) (PICO) is presented, which exhibits tunable formation of elastomeric networks through radical crosslinking of itaconate in the polymer backbone of viscous polyester gels. Through variation of reaction times and monomer molar composition, materials with modulation of a wide range of elasticity (36-1476 kPa) are generated, indicating the tunability of materials to specific elastomeric constructs. This correlated with measured rapid and controllable gelation times. As a proof of principle, scaffold support for cardiac tissue patches is developed, which presents visible tissue organization and viability with appropriate elastomeric support from PICO materials. These formulations present potential application in a range of healthcare applications with requirement for elastomeric support with controllable, rapid gelation under mild conditions.

A Platform for Generation of Chamber-Specific Cardiac Tissues and Disease Modeling.

Tissue engineering using cardiomyocytes derived from human pluripotent stem cells holds a promise to revolutionize drug discovery, but only if limitations related to cardiac chamber specification and platform versatility can be overcome. We describe here a scalable tissue-cultivation platform that is cell source agnostic and enables drug testing under electrical pacing. The plastic platform enabled on-line noninvasive recording of passive tension, active force, contractile dynamics, and Ca2+ transients, as well as endpoint assessments of action potentials and conduction velocity. By combining directed cell differentiation with electrical field conditioning, we engineered electrophysiologically distinct atrial and ventricular tissues with chamber-specific drug responses and gene expression. We report, for the first time, engineering of heteropolar cardiac tissues containing distinct atrial and ventricular ends, and we demonstrate their spatially confined responses to serotonin and ranolazine. Uniquely, electrical conditioning for up to 8 months enabled modeling of polygenic left ventricular hypertrophy starting from patient cells.

Macrophage Polarization with Angiopoietin-1 Peptide QHREDGS.

Currently, clinics are faced with the difficult task of treating non-healing wounds. While the treatment regimen varies between different patients and wounds, a non-healing wound can be a significant detriment to a patient's quality of life, thus highlighting the need for more effective treatments. The immune system is heavily involved in regulating the wound healing process, with delayed wounds often being plagued by prolonged inflammation. In this study, we uncover the interaction between an angiopoietin-1 mimetic peptide, QHREDGS (glutamine-histidine-arginine-glutamic acid-aspartic acid-glycine-serine), immobilized to a collagen-chitosan hydrogel, and murine bone marrow derived macrophages. When macrophages were cultured in the presence of the QHREDGS peptide conjugated to a hydrogel, both proinflammatory and anti-inflammatory cytokines were produced, in contrast to the application of soluble peptide which elicited minimal cytokine secretion. This indicates a unique macrophage polarization with covalently immobilized peptide hydrogels, which may be beneficial in the context of the wound microenvironment. The QHREDGS peptide hydrogel was further optimized to be easily delivered to a wound within a clinical setting.

Microfabrication of AngioChip, a biodegradable polymer scaffold with microfluidic vasculature.

Microengineered biomimetic systems for organ-on-a-chip or tissue engineering purposes often fail as a result of an inability to recapitulate the in vivo environment, specifically the presence of a well-defined vascular system. To address this limitation, we developed an alternative method to cultivate three-dimensional (3D) tissues by incorporating a microfabricated scaffold, termed AngioChip, with a built-in perfusable vascular network. Here, we provide a detailed protocol for fabricating the AngioChip scaffold, populating it with endothelial cells and parenchymal tissues, and applying it in organ-on-a-chip drug testing in vitro and surgical vascular anastomosis in vivo. The fabrication of the AngioChip scaffold is achieved by a 3D stamping technique, in which an intricate microchannel network can be embedded within a 3D scaffold. To develop a vascularized tissue, endothelial cells are cultured in the lumen of the AngioChip network, and parenchymal cells are encapsulated in hydrogels that are amenable to remodeling around the vascular network to form functional tissues. Together, these steps yield a functional, vascularized network in vitro over a 14-d period. Finally, we demonstrate the functionality of AngioChip-vascularized hepatic and cardiac tissues, and describe direct surgical anastomosis of the AngioChip vascular network on the hind limb of a Lewis rat model.

Method for the Fabrication of Elastomeric Polyester Scaffolds for Tissue Engineering and Minimally Invasive Delivery.

Using the methods described herein, we have demonstrated how scaffolds can be designed for a number of applications including tissue engineering, biomedical devices and injectable tissues. Details on the methods of polymerization and physical and chemical characterization of poly(octamethylene maleate (anhydride) citrate (POMaC) are described. Two POMaC polymer recipes with different monomer ratios of maleic anhydride and citric acid were synthesized and compared. Mechanical testing was performed on scaffolds of two distinct anisotropic designs to show how scaffold design influences the apparent elasticity in the long and short axis. POMaC scaffolds of various patterns and geometries were fabricated to demonstrate: (1) scaffold function can be determined by scaffold design (e.g., inherent shape-memory or self-assembling tubular structures), and (2) the soft lithography approach to fabricating biodegradable elastomers described here can be used to suit a number of different potential applications.

Review: Multimodal bioactive material approaches for wound healing.

Wound healing is a highly complex process of tissue repair that relies on the synergistic effect of a number of different cells, cytokines, enzymes, and growth factors. A deregulation in this process can lead to the formation of a non-healing chronic ulcer. Current treatment options, such as collagen wound dressings, are unable to meet the demand set by the wound environment. Therefore, a multifaceted bioactive dressing is needed to elicit a targeted affect. Wound healing strategies seek to develop a targeted effect through the delivery of a bioactive molecule to the wound by a hydrogel or a polymeric scaffold. This review examines current biomaterial and small molecule-based approaches that seek to develop a bioactive material for targeted wound therapy and accepted wound healing models for testing material efficacy.