Tropoelastin Improves Post-Infarct Cardiac Function.

BACKGROUND: Myocardial infarction (MI) is among the leading causes of death worldwide. Following MI, necrotic cardiomyocytes are replaced by a stiff collagen-rich scar. Compared to collagen, the extracellular matrix protein elastin has high elasticity and may have more favorable properties within the cardiac scar. We sought to improve post-MI healing by introducing tropoelastin, the soluble subunit of elastin, to alter scar mechanics early after MI. METHODS AND RESULTS: We developed an ultrasound-guided direct intramyocardial injection method to administer tropoelastin directly into the left ventricular anterior wall of rats subjected to induced MI. Experimental groups included shams and infarcted rats injected with either PBS vehicle control or tropoelastin. Compared to vehicle treated controls, echocardiography assessments showed tropoelastin significantly improved left ventricular ejection fraction (64.7±4.4% versus 46.0±3.1% control) and reduced left ventricular dyssynchrony (11.4±3.5 ms versus 31.1±5.8 ms control) 28 days post-MI. Additionally, tropoelastin reduced post-MI scar size (8.9±1.5% versus 20.9±2.7% control) and increased scar elastin (22±5.8% versus 6.2±1.5% control) as determined by histological assessments. RNA sequencing (RNAseq) analyses of rat infarcts showed that tropoelastin injection increased genes associated with elastic fiber formation 7 days post-MI and reduced genes associated with immune response 11 days post-MI. To show translational relevance, we performed immunohistochemical analyses on human ischemic heart disease cardiac samples and showed an increase in tropoelastin within fibrotic areas. Using RNA-seq we also demonstrated the tropoelastin gene ELN is upregulated in human ischemic heart disease and during human cardiac fibroblast-myofibroblast differentiation. Furthermore, we showed by immunocytochemistry that human cardiac fibroblast synthesize increased elastin in direct response to tropoelastin treatment. CONCLUSIONS: We demonstrate for the first time that purified human tropoelastin can significantly repair the infarcted heart in a rodent model of MI and that human cardiac fibroblast synthesize elastin. Since human cardiac fibroblasts are primarily responsible for post-MI scar synthesis, our findings suggest exciting future clinical translation options designed to therapeutically manipulate this synthesis.

Engineering antibacterial bioceramics: Design principles and mechanisms of action.

The urgency to address skeletal abnormalities and diseases through innovative approaches has led to a significant interdisciplinary convergence of engineering, 3D printing, and design in developing individualised bioceramic bioscaffolds. This review explores into the recent advancements and future trajectory of non-antibiotic antibacterial bioceramics in bone tissue engineering, an importance given the escalating challenges of orthopaedic infections, antibiotic resistance, and emergent pathogens. Initially, the review provides an in-depth exploration of the complex interactions among bacteria, immune cells, and bioceramics in clinical contexts, highlighting the multifaceted nature of infection dynamics, including protein adsorption, immunological responses, bacterial adherence, and endotoxin release. Then, focus on the next-generation bioceramics designed to offer multifunctionality, especially in delivering antibacterial properties independent of traditional antibiotics. A key highlight of this study is the exploration of smart antibacterial bioceramics, marking a revolutionary stride in medical implant technology. The review also aims to guide the ongoing development and clinical adoption of bioceramic materials, focusing on their dual capabilities in promoting bone regeneration and exhibiting antibacterial properties. These next-generation bioceramics represent a paradigm shift in medical implant technology, offering multifunctional benefits that transcend traditional approaches.

Unraveling the influence of channel size and shape in 3D printed ceramic scaffolds on osteogenesis.

Bone has the capacity to regenerate itself for relatively small defects; however, this regenerative capacity is diminished in critical-size bone defects. The development of synthetic materials has risen as a distinct strategy to address this challenge. Effective synthetic materials to have emerged in recent years are bioceramic implants, which are biocompatible and highly bioactive. Yet nothing suitable for the repair of large bone defects has made the transition from laboratory to clinic. The clinical success of bioceramics has been shown to depend not only on the scaffold's intrinsic material properties but also on its internal porous geometry. This study aimed to systematically explore the implications of varying channel size, shape, and curvature in tissue scaffolds on in vivo bone regeneration outcomes. 3D printed bioceramic scaffolds with varying channel sizes (0.3 mm to 1.5 mm), shapes (circular vs rectangular), and curvatures (concave vs convex) were implanted in rabbit femoral defects for 8 weeks, followed by histological evaluation. We demonstrated that circular channel sizes of around 0.9 mm diameter significantly enhanced bone formation, compared to channel with diameters of 0.3 mm and 1.5 mm. Interestingly, varying channel shapes (rectangular vs circular) had no significant effect on the volume of newly formed bone. Furthermore, the present study systematically demonstrated the beneficial effect of concave surfaces on bone tissue growth in vivo, reinforcing previous in silico and in vitro findings. This study demonstrates that optimizing architectural configurations within ceramic scaffolds is crucial in enhancing bone regeneration outcomes. STATEMENT OF SIGNIFICANCE: Despite the explosion of work on developing synthetic scaffolds to repair bone defects, the amount of new bone formed by scaffolds in vivo remains suboptimal. Recent studies have illuminated the pivotal role of scaffolds' internal architecture in osteogenesis. However, these investigations have mostly remained confined to in silico and in vitro experiments. Among the in vivo studies conducted, there has been a lack of systematic analysis of individual architectural features. Herein, we utilized bioceramic 3D printing to conduct a systematic exploration of the effects of channel size, shape, and curvature on bone formation in vivo. Our results demonstrate the significant influence of channel size and curvature on in vivo outcomes. These findings provide invaluable insights into the design of more effective bone scaffolds.

In situ formation of osteochondral interfaces through "bone-ink" printing in tailored microgel suspensions.

Osteochondral tissue has a complex hierarchical structure spanning subchondral bone to articular cartilage. Biomaterials approaches to mimic and repair these interfaces have had limited success, largely due to challenges in fabricating composite hard-soft interfaces with living cells. Biofabrication approaches have emerged as attractive methods to form osteochondral analogues through additive assembly of hard and soft components. We have developed a unique printing platform that is able to integrate soft and hard materials concurrently through freeform printing of mineralized constructs within tunable microgel suspensions containing living cells. A library of microgels based on gelatin were prepared, where the stiffness of the microgels and a liquid "filler" phase can be tuned for bioprinting while simultaneously directing differentiation. Tuning microgel stiffness and filler content differentially directs chondrogenesis and osteogenesis within the same construct, demonstrating how this technique can be used to fabricate osteochondral interfaces in a single step. Printing of a rapidly setting calcium phosphate cement, so called "bone-ink" within a cell laden suspension bath further guides differentiation, where the cells adjacent to the nucleated hydroxyapatite phase undergo osteogenesis with cells in the surrounding medium undergoing chondrogenesis. In this way, bone analogues with hierarchical structure can be formed within cell-laden gradient soft matrices to yield multiphasic osteochondral constructs. This technique provides a versatile one-pot biofabrication approach without harsh post-processing which will aid efforts in bone disease modelling and tissue engineering. STATEMENT OF SIGNIFICANCE: This paper demonstrates the first example of a biofabrication approach to rapidly form osteochondral constructs in a single step under physiological conditions. Key to this advance is a tunable suspension of extracellular matrix microgels that are packed together with stem cells, providing a unique and modular scaffolding for guiding the simultaneous formation of bone and cartilage tissue. The physical properties of the suspension allow direct writing of a ceramic "bone-ink", resulting in an ordered structure of microscale hydrogels, living cells, and bone mimics in a single step. This platform reveals a simple approach to making complex skeletal tissue for disease modelling, with the possibility of repairing and replacing bone-cartilage interfaces in the clinic using a patient's own cells.

Electrostatic and Covalent Binding of an Antibacterial Polymer to Hydroxyapatite for Protection against Escherichia coli Colonization.

Orthopedic-device-related infections are notorious for causing physical and psychological trauma to patients suffering from them. Traditional methods of treating these infections have relied heavily on antibiotics and are becoming ineffectual due to the rise of antibiotic-resistant bacteria. Mimics of antimicrobial peptides have emerged as exciting alternatives due to their favorable antibacterial properties and lack of propensity for generating resistant bacteria. In this study, the efficacy of an antibacterial polymer as a coating material for hydroxyapatite and glass surfaces, two materials with wide ranging application in orthopedics and the biomedical sciences, is demonstrated. Both physical and covalent modes of attachment of the polymer to these materials were explored. Polymer attachment to the material surfaces was confirmed via X-ray photoelectron spectroscopy and contact angle measurements. The modified surfaces exhibited significant antibacterial activity against the Gram-negative bacterium E. coli, and the activity was retained for a prolonged period on the surfaces of the covalently modified materials.

Discovering an unknown territory using atom probe tomography: Elemental exchange at the bioceramic scaffold/bone tissue interface.

Here we report the first atom probe study to reveal the atomic-scale composition of in vivo bone formed in a bioceramic scaffold (strontium-hardystonite-gahnite) after 12-month implantation in a large bone defect in sheep tibia. The composition of the newly formed bone tissue differs to that of mature cortical bone tissue, and elements from the degrading bioceramic implant, particularly aluminium (Al), are present in both the newly formed bone and in the original mature cortical bone tissue at the perimeter of the bioceramic implant. Atom probe tomography confirmed that the trace elements are released from the bioceramic and are actively transported into the newly formed bone. NanoSIMS mapping, as a complementary technique, confirmed the distribution of the released ions from the bioceramic into the newly formed bone tissue within the scaffold. This study demonstrated the combined benefits of atom probe and nanoSIMS in assessing nanoscopic chemical composition changes at precise locations within the tissue/biomaterial interface. Such information can assist in understanding the interaction of scaffolds with surrounding tissue, hence permitting further iterative improvements to the design and performance of biomedical implants, and ultimately reducing the risk of complications or failure while increasing the rate of tissue formation. STATEMENT OF SIGNIFICANCE: The repair of critical-sized load-bearing bone defects is a challenge, and precisely engineered bioceramic scaffold implants is an emerging potential treatment strategy. However, we still do not understand the effect of the bioceramic scaffold implants on the composition of newly formed bone in vivo and surrounding existing mature bone. This article reports an innovative route to solve this problem, the combined power of atom probe tomography and nanoSIMS is used to spatially define elemental distributions across bioceramic implant sites. We determine the nanoscopic chemical composition changes at the Sr-HT Gahnite bioceramic/bone tissue interface, and importantly, provide the first report of in vivo bone tissue chemical composition formed in a bioceramic scaffold.

Ceramic Omnidirectional Bioprinting in Cell-laden Suspensions for the Generation of Bone Analogs.

Structurally, bone tissue is an inorganic-organic composite containing metabolically active cells embedded within a hierarchical, highly mineralized matrix. This organization is challenging to replicate due to the heterogeneous environment of bone. Ceramic omnidirectional bioprinting in cell-suspensions (COBICS) is a microgel-based bioprinting technique that uniquely replicates the mineral and cellular structure of bone. COBICS prints complex, biologically relevant constructs without the need for sacrificial support materials or harsh postprocessing steps (e.g., radiation and high-temperature sintering), which are two of the biggest challenges in the additive manufacturing of bone mimetic constructs. This technique is enabled via the freeform extrusion of a novel calcium phosphate-based ink within a gelatin-based microgel suspension. The yield-stress properties of the suspension allow deposition and support the printed bone structure. UV crosslinking and nanoprecipitation then "lock" it in place. The ability to print nanostructured bone-mimetic ceramics within cell-laden biomaterials provides spatiotemporal control over macro- and micro-architecture and facilitates the real-time fabrication of complex bone constructs in clinical settings.

Low-Temperature Synthesis of Hollow ß-Tricalcium Phosphate Particles for Bone Tissue Engineering Applications.

ß-Tricalcium phosphate (ß-TCP) has been extensively used in bone tissue engineering in the form of scaffolds, granules, or as reinforcing phase in organic matrices. Solid-state reaction route at high temperatures (>1000 °C) is the most widely used method for the preparation of ß-TCP. The high-temperature synthesis, however, results in the formation of hard agglomerates and fused particles which necessitates postprocessing steps such as milling and sieving operations. This, inadvertently, could lead to introducing unwanted trace elements, promoting particle shape irregularity as well as compromising the biodegradability and bioactivity of ß-TCP because of the solid microstructure of particles. In this study, we introduce a one-pot wet-chemical method at low temperatures (between 160 and 170 °C) to synthesize hollow ß-TCP (hß-TCP) submicron particles of an average size of 300 nm with a uniform rhombohedral shape. We assessed the cytocompatibility of the hß-TCP using primary human osteoblasts (HOB), adipose-derived stem cells (ADSC), and antigen-presenting cells (APCs). We demonstrate the bioactivity of the hß-TCP when cultured with HOB, ADSC, and APCs at a range of particle concentrations (up to 1000 µg/mL) for up to 7 days. hß-TCP significantly enhances osteogenic differentiation of ADSC without the addition of osteogenic supplements. These findings offer a new type of ß-TCP particles prepared at low temperatures, which present various opportunities for developing ß-TCP based biomaterials.

Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

Surface topography is one of the key factors in regulating interactions between materials and cells. While topographies presented to cells in vivo are non-symmetrical and in complex shapes, current fabrication techniques are limited to replicate these complex geometries. In this study, we developed a microcasting technique and successfully produced imprinted hydroxyapatite (HAp) surfaces with nature-inspired (honeycomb, pillars, and isolated islands) topographies. The in vitro biological performance of the developed non-symmetrical topographies was evaluated using adipose-derived stem cells (ADSCs). We demonstrated that ADSCs cultured on all HAp surfaces, except honeycomb patterns, presented well-defined stress fibers and expressed focal adhesion protein (paxillin) molecules. Isolated islands topographies significantly promoted osteogenic differentiation of ADSCs with increased alkaline phosphatase activity and upregulation of key osteogenic markers, compared to the other topographies and the control unmodified (flat) HAp surface. In contrast, honeycomb topographies hampered the ability of the ADSCs to proliferate and differentiate to the osteogenic lineage. This work presents a facile technique to imprint nature-derived topographies on the surface of bioceramics which opens up opportunities for the development of bioresponsive interfaces in tissue engineering and regenerative medicine.

A modular design strategy to integrate mechanotransduction concepts in scaffold-based bone tissue engineering.

Repair or regeneration of load-bearing bones has long been an incentive for the tissue engineering community to develop a plethora of synthetic bone scaffolds. Despite the key role of physical forces and the mechanical environment in bone regeneration, the mechanotransduction concept has rarely been incorporated in structural design of bone tissue scaffolds, particularly those made of bioactive materials such as hydrogels and bioceramics. Herein, we introduce a modular design strategy to fabricate a load bearing device that can support a wide range of hydrogel- and ceramic-based scaffolds against complex in-vivo loading conditions to induce desirable mechanical strains for bone regeneration within the scaffolds. The device is comprised of a fenestrated polymeric shell and ceramic structural pillars arranged in a sophisticated configuration to provide ample internal space for the scaffold, also enabling it to purposely regulate the levels of strains and stresses within the scaffolds. Utilizing this top-down design approach, we demonstrate that the failure load of alginate hydrogels increases 3200-fold in compression, 300-fold in shear and 75-fold in impact, achieving the values that enable them to withstand physiological loads in weight-bearing sites, while allowing generation of osteoinductive strains (i.e., 0.2-0.4%) in the hydrogel. This modular design approach opens a broad range of opportunities to utilize various bioactive but mechanically weak scaffolds for the treatment of load-bearing defects and exploiting mechanobiology strategies to improve bone regeneration.

iPSC Bioprinting: Where are We at?

Here, we present a concise review of current 3D bioprinting technologies applied to induced pluripotent stem cells (iPSC). iPSC have recently received a great deal of attention from the scientific and clinical communities for their unique properties, which include abundant adult cell sources, ability to indefinitely self-renew and differentiate into any tissue of the body. Bioprinting of iPSC and iPSC derived cells combined with natural or synthetic biomaterials to fabricate tissue mimicked constructs, has emerged as a technology that might revolutionize regenerative medicine and patient-specific treatment. This review covers the advantages and disadvantages of bioprinting techniques, influence of bioprinting parameters and printing condition on cell viability, and commonly used iPSC sources, and bioinks. A clear distinction is made for bioprinting techniques used for iPSC at their undifferentiated stage or when used as adult stem cells or terminally differentiated cells. This review presents state of the art data obtained from major searching engines, including Pubmed/MEDLINE, Google Scholar, and Scopus, concerning iPSC generation, undifferentiated iPSC, iPSC bioprinting, bioprinting techniques, cartilage, bone, heart, neural tissue, skin, and hepatic tissue cells derived from iPSC.

3D Printing of Bioceramic Scaffolds-Barriers to the Clinical Translation: From Promise to Reality, and Future Perspectives.

In this review, we summarize the challenges of the three-dimensional (3D) printing of porous bioceramics and their translational hurdles to clinical applications. The state-of-the-art of the major 3D printing techniques (powder-based and slurry-based), their limitations and key processing parameters are discussed in detail. The significant roadblocks that prevent implementation of 3D printed bioceramics in tissue engineering strategies, and medical applications are outlined, and the future directions where new research may overcome the limitations are proposed. In recent years, there has been an increasing demand for a nanoscale control in 3D fabrication of bioceramic scaffolds via emerging techniques such as digital light processing, two-photon polymerization, or large area maskless photopolymerization. However, these techniques are still in a developmental stage and not capable of fabrication of large-sized bioceramic scaffolds; thus, there is a lack of sufficient data to evaluate their contribution. This review will also not cover polymer matrix composites reinforced with particulate bioceramics, hydrogels reinforced with particulate bioceramics, polymers coated with bioceramics and non-porous bioceramics.

Architectural Design of 3D Printed Scaffolds Controls the Volume and Functionality of Newly Formed Bone.

The successful regeneration of functional bone tissue in critical-size defects remains a significant clinical challenge. To address this challenge, synthetic bone scaffolds are widely developed, but remarkably few are translated to the clinic due to poor performance in vivo. Here, it is demonstrated how architectural design of 3D printed scaffolds can improve in vivo outcomes. Ceramic scaffolds with different pore sizes and permeabilities, but with similar porosity and interconnectivity, are implanted in rabbit calvaria for 12 weeks, and then the explants are harvested for microcomputed tomography evaluation of the volume and functionality of newly formed bone. The results indicate that scaffold pores should be larger than 390 µm with an upper limit of 590 µm to enhance bone formation. It is also demonstrated that a bimodal pore topology-alternating large and small pores-enhances the volume and functionality of new bone substantially. Moreover, bone formation results indicate that stiffness of new bone is highly influenced by the scaffold's permeability in the direction concerned. This study demonstrates that manipulating pore size and permeability in a 3D printed scaffold architecture provides a useful strategy for enhancing bone regeneration outcomes.