John P. Fisher

The rapid growth of additive manufacturing technologies continues to drive the commercial development of standardized materials to streamline the process. Despite the growing range of available materials, few still achieve a balance between flexibility and biocompatibility. Here, we evaluated the cytotoxicity of Formlabs Biomed Elastic 50A, Biomed Flexible 80A, and Biomed Durable resin using normal human dermal fibroblasts and human mesenchymal stromal cells. The sol fraction of the crosslinked and uncrosslinked materials was measured to determine the amount of leachable components in the polymer network. Following ISO 10993‐5 guidelines, cytotoxicity was evaluated through direct, indirect, or extract testing for 24 h. All three materials did not elicit a cytotoxic response when assessed through fluorescence imaging and LDH assay. In each test, cells showed similar viability as cells cultured with a known noncytotoxic control, polycaprolactone, and showed significantly higher viability compared to a cytotoxic control, a polyurethane film containing 0.1% zinc diethyldithiocarbamate (ZDEC). Post‐processing parameters were shown to minimize the effects of cytotoxicity when post‐treated with an isopropyl alcohol wash, UV post‐curing, or ethanol sterilization. Overall, the Formlabs Biomed Elastic 50A, Biomed Flexible 80A, and Biomed Durable resins were shown to exhibit minimal cytotoxicity after printing and post‐processing. Further, the presented findings can be adapted to standardize in vitro testing of a wide range of commercial photocurable resins for tissue‐interfacing parts.

As improvements in material compatibility are achieved, stereolithography (SLA) 3D printing allows for the prototyping of soft tissue constructs with high reproducibility and precision. To streamline the Food and Drug Administration approval process, clinical-grade manufacturing must be validated. Here, we present a manufacturing process and post-manufacturing analysis for soft-tissue implants used to reconstruct the nipple-areolar complex (NAC). The NAC is a specialized soft tissue structure that influences aesthetic outcomes in breast reconstruction. Physiologically-sized NAC implants were designed and printed in a good manufacturing practice facility using Formlabs Biomed Elastic 50 A (E 50 A) and Biomed Flexible 80 A (F 80 A) resins. Documentation and procedures were established to standardize and control macroporous soft-tissue implant manufacturing. Post-manufacturing classification based on architectural and mechanical features defined the minimum acceptable criteria for future production. E 50 A macroporous implants resembled the modulus of soft tissue, while F 80 A geometries showed higher reproducibility and preservation of fine architectural features. SLA printing enabled the patterning of implants with porosities of 74.65% and 72.74% for 0.4- and 0.5- mm geometries, respectively. 3D printed implants supported the infusion of gelatin methacryloyl for the delivery of biological cues, with a filling efficiency greater than 97%. This manufacturing platform achieves reproducible fabrication of soft-tissue implants that meet both structural and mechanical requirements. Further, the presented methodology can be adapted for a wide range of complex soft tissues to advance regenerative medicine solutions toward regulatory approval and eventual clinical integration.

Acellular methods are needed to vascularize 3D‐printed tissue‐engineered constructs for faster clinical translation. One method is to create bioactive materials that encourage migration and network formation of a patient's own cells. This study utilizes QK peptide, which replicates a binding sequence of vascular endothelial growth factor. This peptide has previously shown to maintain bioactivity when modified with an acrylate group and covalently bound to gelatin methacrylate (GelMA). However, this binding interferes with the crosslinking of the hydrogel matrix, requiring characterization of the modified material. We investigated how binding QK peptide impacts GelMA crosslinking and material properties. A factorial design was employed to investigate the relationships between GelMA concentration, GelMA degree of substitution, and QK peptide concentration. These experiments were used to inform a rheology study, where the impact of QK peptide on crosslinking was investigated over a range of photocrosslinking times. We then investigated the printability of QK‐GelMA bioink using rheology and a filament collapse test. The bioactivity induced by the peptide on the hydrogel surface was evaluated using endothelial cell network formation. QK peptide impacted key GelMA characteristics, including swelling behavior, mesh size, and storage modulus, potentially through inhibition of temperature‐induced chain entanglement. While bound QK peptide impacts GelMA bioink at the nanoscale, QK‐GelMA bioinks maintain high print fidelity and increase endothelial network formation on the surface of the hydrogel. The addition of QK peptide increases the vascularization potential of 3D‐printed tissue‐engineered constructs.

This perspective, marking the 30th anniversary of the Tissue Engineering journal, discusses the exciting trends in the global commercialization of tissue engineering technology. Within a historical context, we present an evolution of challenges and a discussion of the last 5 years of global commercial successes and emerging market trends, highlighting the continued expansion of the field in the northeastern United States. This leads to an overview of the last 5 years’ progress in clinical trials for tissue-engineered therapeutics, including an analysis of trends in success and failure. Finally, we provide a broad overview of preclinical research and a perspective on where the state-of-the-art lies on the horizon.

Bone defects due to age, trauma, and surgery, which are exacerbated by medication side effects and common diseases like osteoporosis, diabetes, and rheumatoid arthritis, are a problem of epidemic scale. The present clinical standard for treating these defects includes autografts and allografts. While both treatments can promote robust regenerative outcomes, they fail to strike a desirable balance of availability, side effect profile, consistent regenerative efficacy, and affordability. This difficulty has contributed to the rise of bone tissue engineering (BTE) as a potential avenue through which enhanced bone regeneration could be delivered. BTE is founded upon a paradigm of using biomaterials, bioactive factors, osteoblast lineage cells (ObLCs), and vascularization to cue deficient bone tissue into a state of regeneration. Despite promising preclinical results, BTE has had modest success in being translated into the clinical setting. One barrier has been the simplicity of its paradigm relative to the complexity of biological bone. Therefore, this paradigm must be critically examined and expanded to better account for this complexity. One potential avenue for this is a more detailed consideration of osteoclast lineage cells (OcLCs). While these cells ostensibly oppose ObLCs and bone regeneration through their resorptive functions, myriad investigations have shed light on their potential to influence bone equilibrium in more complex ways through their interactions with both ObLCs and bone matrix. Most BTE research has not systematically evaluated their influence. Yet contrary to expectations associated with the paradigm, a selection of BTE investigations have demonstrated that this influence can enhance bone regeneration in certain contexts. Additionally, much work has elucidated the role of many controllable scaffold parameters in both inhibiting and stimulating the activity of OcLCs in parallel to bone regeneration. Therefore, this review aims to detail and explore the implications of OcLCs in BTE, and how they can be leveraged to improve upon the existing BTE paradigm.

Fabrication of engineered thin membranous tissues (TMTs) presents a significant challenge to researchers, as these structures are small in scale, but present complex anatomies containing multiple stratified cell layers. While numerous methodologies exist to fabricate such tissues, many are limited by poor mechanical properties, need for post‐fabrication, or lack of cytocompatibility. Extrusion bioprinting can address these issues, but lacks the resolution necessary to generate biomimetic, microscale TMT structures. Therefore, our goal was to develop a strategy that enhances bioprinting resolution below its traditional limit of 150 μm and delivers a viable cell population. We have generated a system to effectively shrink printed gels via electrostatic interactions between anionic and cationic polymers. Base hydrogels are composed of gelatin methacrylate type A (cationic), or B (anionic) treated with anionic alginate, and cationic poly‐L‐lysine, respectively. Through a complex coacervation‐like mechanism, the charges attract, causing compaction of the base GelMA network, leading to reduced sample dimensions. In this work, we evaluate the role of both base hydrogel and shrinking polymer charge on effective print resolution and cell viability. The alginate anion‐mediated system demonstrated the ability to reach bioprinting resolutions of 70 μm, while maintaining a viable cell population. To our knowledge, this is the first study that has produced such significant enhancement in extrusion bioprinting capabilities, while also remaining cytocompatible.

Osteochondral defects, characterized by structural compromises to articular cartilage and subchondral bone, can cause pain and lead to progressive cartilage damage and eventual osteoarthritis. Unfortunately, repairing these defects remains difficult because of the poor regenerative properties of cartilage and complex mechanical demands of the joint. As such, the field of tissue engineering aims to develop multiphasic implants that replace pathological cartilage and bone tissue and restore mechanical functionality to the joint. Recent bone physiology investigations have demonstrated that osteoclast (OC) lineage cells are inextricably involved in osteoblastic bone formation through an extensive network of anabolic signaling pathways, and so the codelivery OC and osteoblast (OB) lineage cells within scaffolds is being actively explored for bone tissue engineering purposes. However, it remains unclear how these cells can be incorporated into the design of multiphasic osteochondral scaffolds to potentially enhance subchondral bone formation and subsequent implant osseointegration. To explore this question, we examined direct surface seeding and hydrogel encapsulation as potential scaffold cellularization strategies. First, we examined how OC precursor cells and peripheral blood monocytes (PBMCs) influence early-stage bone matrix development and osteogenesis in 2D coculture. Then, we evaluated the osteogenic potential of mesenchymal stem cells (MSCs) and PBMCs cocultures encapsulated within a gelatin methacrylate (GelMA) hydrogel system. Our findings demonstrate that coculturing PBMCs with MSCs in 2D cultures significantly enhanced cell proliferation, early bone matrix deposition, and the formation of cell clusters by Day 28. However, we observed no significant difference in type I collagen deposition between GelMA hydrogel scaffolds cultured in basal and OC conditions during the same period. In addition, we found that the GelMA hydrogel system with MSC/PBMC cocultures in OC conditions exhibited decreased osteogenic activity by Day 28. Collectively, our findings support the osteogenic potential of OC-lineage cells in 2D culture conditions, and the potential benefits of surface-seeding for the codelivery of OC-lineage cells and MSCs in osteo-scaffolds for enhanced osteochondral regeneration and broader bone tissue engineering purposes.

Osteoarthritis is a debilitating chronic joint disorder that affects millions of people worldwide. Since palliative and surgical treatments cannot completely regenerate hyaline cartilage within the articulating joint, osteochondral (OC) tissue engineering has been explored to heal these challenging defects. Using computational simulations and 3D printing, we aimed to build a strategy to fabricate OC scaffolds with enhanced biomechanics. First, computational simulations revealed that interfacial fibrils within a bilayer alter OC scaffold deformation patterns by redirecting load-induced stresses toward the top of the cartilage layer. Principal component analysis (PCA) revealed that scaffolds with 800 m long fibrils (scaffolds 8A-8H) possessed optimal biomechanical properties to withstand compression and shear forces. While compression testing indicated that OC scaffolds with 800 m fibrils did not have greater compressive moduli than other scaffolds, interfacial shear tests indicated that scaffold 8H possessed the greatest shear strength. Lastly, failure analysis demonstrated that yielding or buckling models describe interfacial fibril failure depending on fibril slenderness, S. Specifically for scaffolds with packing density n=6 and n=8, the yielding failure model fits experimental loads with S10 slenderness. The research presented here provides critical insights into designing 3D printed interfacial scaffolds with refined biomechanics toward improving OC tissue engineering outcomes.