Biofabrication最新文献

The curvature of cell adhesion substrates has emerged as a critical geometric parameter influencing cellular fate determination. While its regulatory role is increasingly recognized, the osteogenic effects of complex three-dimensional (3D) curved surfaces remain insufficiently explored. In this study, high-precision two-photonic polymerization 3D printing was utilized to fabricate scaffolds with controlled curvature distributions, achieving unprecedented fidelity between manufactured surfaces and their digital models. Comparative analysis of triply periodic minimal surface (TPMS) scaffolds and conventional truss scaffolds revealed distinct osteogenic mechanisms: zero mean curvature enhanced osteogenic differentiation through improved scaffold permeability, while negative Gaussian curvature promoted bone formation through combined effects of permeability controlling and guided cellular organization. Notably, scaffolds exhibiting broader ranges of negative Gaussian curvature demonstrated superior osteogenesis inductive capacity, as evidenced by enhanced new bone formation in bothin vitroandin vivomodels. These findings provide mechanistic insights into curvature-dependent osteogenesis, quantitative design principles for TPMS-based bone scaffolds, and experimental validation of curvature optimization strategies. The study establishes a geometric framework for rational scaffold design, advancing the development of high-performance regenerative implants.Keyworks.TPMS, Gaussian curvature, two-photonic polymerization, osteogenesis, bone regeneration.

Congenital heart diseases, including single ventricle heart defects such as hypoplastic left and right heart syndromes, remain a leading cause of neonatal death and long-term morbidity. Regenerative medicine approaches hold great therapeutic promise for treating single ventricle disease, specifically through the use of human pluripotent stem cell-derived cardiomyocytes (iPSC-CM) to generate pulsatile conduits capable of growing and developing over time within the recipient. However, current strategies for rapidly fabricating large-scale engineered heart muscle to create such conduits face limitations, including the shear stress generated during most bioprinting processes along with harsh enzymatic treatments required for initial singularization of cells prior to bioprinting, which together can compromise cell viability and downstream tissue function. Here, we explored the use of derived cardiovascular progenitors (iPSC-CVP) as an alternative to fully differentiated cardiomyocytes as a potential cell source for future biomanufacturing efforts. We demonstrate that iPSC-CVP can be used to form functional engineered heart tissues with similar electrophysiological properties to tissues formed from fully differentiated iPSC-CM, while also being more amenable to enzymatic dissociation and mechanical manipulation. Our results suggest that iPSC-CVP may be an ideal cell population for future efforts in biofabrication of contractile structures such as engineered heart muscle and pulsatile conduits.

The use of bioprinters as depositional tools for bioinks and cells has expanded greatly over the past two decades. Bioprinting combines hydrogels with cells to produce customized constructs for personalized medicine. However, several challenges hinder the clinical use of these constructs. Quality control metrics for bioprinting involve the assessment of critical quality attributes at every stage of production. Currently, bioprinted constructs are manually assessed using destructive methods that occur post-production, requiring the creation of multiple products per patient. Reproducing printed constructs is difficult due to time-sensitive bioink properties that require lengthy optimization processes to print with accuracy. In addition, the collection, processing, and testing of cell bioactivity for each printed construct greatly increases production costs. To address these challenges, non-destructive, real-time monitoring can be integrated into the bioprinting process. The goal of this review paper is to reimagine the function of a bioprinter from a simple tool of production to one capable of evaluating constructs in real-time. This review features recent advances in the field for real-time monitoring with a focus on time-sensitive bioink properties, print accuracy, and cell health. Automated assessment and quantification of time-sensitive bioink qualities such as mixing, pH, temperature, and viscosity will enhance construct quality by enabling the rapid optimization of printing parameters. Meanwhile, real-time monitoring of cell health through concentration, viability, and type serves as an indicator for bioactivity. Construct accuracy and reproducibility are also improved through the identification, prediction, and correction of defects during printing. Incorporating real-time monitoring into the bioprinting process using closed-loop feedback would improve the reproducibility, quality, and translation of constructs into the clinic.

Drug-induced liver injury (DILI) remains a major cause of acute liver failure, clinical trial attrition, and post-marketing drug withdrawal, yet predictivein vitromodels are limited in accuracy, scalability, and human relevance. Here, we present a Liver-on-Micropillar (LoM) platform a fully animal-free, high-throughput, miniaturized human liver model designed for early-stage hepatotoxicity screening. The system combines a xeno-free medium with a xeno-free bioink to support co-culture of four human liver-relevant cell types: differentiated HepaRG, LX-2, HMEC-1, and differentiated THP-1 cells. Microlivers are bioprinted onto micropillar arrays compatible with standard 96-well plate formats. Functional characterization confirmed stable cell viability, albumin and urea production, as well as inducible CYP expression. To evaluate DILI predictivity, ten reference drugs were tested using assays to measure ATP content, XTT metabolic activity, and albumin secretion. Half-maximal inhibitory concentrations (IC₅₀) were experimentally determined, and margins of safety (MOS) were calculated by dividing IC₅₀by clinical maximum plasma concentration (C_max). The LoM platform correctly classified 90% of the tested compounds using a MOS threshold of 100. This scalable and reproducible model provides a human-relevant, regulatory-aligned alternative to animal testing and supports broader efforts to implement non-animal methodologies in drug safety evaluation.

Organs in the human body exist within a highly integrated and dynamically interacting environment, and their interactions are critical for maintaining normal physiological processes. Traditional cell culture models and animal models fail to meet the needs of preclinical research, as they struggle to fully recapitulatein vivophysiology and pathology. Thus, innovativein vivoplatforms are urgently needed to bridge the gaps between preclinical research and clinical translation. Multiorgan-on-a-chip (multi-OoC), an emerging field in bioengineering, offers precise control over cellular microenvironments and recapitulates organ-level functions and interorgan crosstalk. By mimicking complex human physiology and pathophysiology, multi-OoC systems provide novel opportunities for disease modeling, drug discovery, and personalized medicine. This paper will systematically elaborate on the necessity of developing multi-OoC systems, delve into their structural design and biomanufacturing strategies, and highlight their recent applications in biomedical research. Additionally, it will analyze key challenges such as the establishment of standardized operating procedures and the validation of model outputs, and envision their application prospects in the field of personalized medicine. The aim is to provide a reference for promoting the standardization and clinical translation of this technology.

Recently, significant advancements have been witnessed in variousin vitrotreatment evaluation models, especially organoids and organs-on-chips.In vitroculture of cancer cells and drug screening are key technical components in functional oncology precision medicine. However, most studies primarily focus on constructing models using established cell lines, with limited integration with clinical diagnosis or patient treatment. This review provides a brief overview of precision medicine models, followed by discussions on the broad spectrum of applications involving two-dimensional tumor cell culture, patient-derived tumor xenograft models, tumor organoids, and tumors-on-chips. It highlights the success rate of patient-derived tumor organoids construction and their application in clinical trials. Recent advancements in tumors-on-chips and organoids-on-chips are elaborated on, alongside with integration of other new generation technologies. Additionally, this review summarizes the advantages and constraints associated with tumor organoids and tumors-on-chips, underscoring their crucial role in the advancement of personalized medicine.

Suspension bath bioprinting, whereby bioinks are extruded into a yield stress bath with rapid recovery from shearing, has enabled the printing of low viscosity bioinks into constructs with high geometric complexity. Previous studies have often relied upon external stabilisation of the suspension bath (e.g. collagen) in order to culture soft materials without loss of printed structure. Here, we report a systematic investigation of suspension bath properties that support the printing, fusion, and culture of spheroid-based bioinks without added stabilisation. Specifically, agarose fluid gels of varied polymer concentrations and dilutions were produced and characterised morphologically and rheologically. Juvenile bovine chondrocytes or mesenchymal stromal cells (MSCs) were formed into spheroids of ∼150µm in diameter and investigated within agarose suspension baths either for their fusion in hanging drop cultures or as jammed bioinks. MSC spheroids were also printed when mixed with hydrogel microparticles to demonstrate additional versatility to the approach. Suspension baths of lower polymer concentrations and increased dilution enabled faster spheroid fusion; however, the most heavily diluted suspension bath was unable to maintain print fidelity. Other formulations supported the printing, fusion, and culture of spheroid-based inks, either as simple lines or more complex patterns. These findings help to inform the design of suspension baths for bioprinting and culture.

The intervertebral disc (IVD) primarily comprises an outer ring of collagen fibers (annulus fibrosus, AF), which encases a soft, gelatinous core (nucleus pulposus, NP). Existingin vitromodels have failed to integrate these two tissues effectively or accurately replicate their intricate organization. By combining two biofabrication techniques, we developed a novel 3Din vitromodel that closely mimics the organization of an ovine IVD. Our approach employs a polycaprolactone (PCL) frame produced via melt electrowriting to recreate the multilamellar architecture of the AF. Ovine primary cells, encapsulated in a photocrosslinkable alginate hydrogel, were precisely extruded within the multilamellar structure, thereby mimicking the native shape and size of an ovine disc. The bioink containing the NP cells was deposited at the center of the construct, while the bioink with the AF cells was strategically layered in between the lamellae of the PCL frame. Photocrosslinking was optimized to match the native stiffness of the disc. The constructs were maintained in culture for 28 d, during which we thoroughly assessed reproducibility, stability, and cell viability and phenotype. The results unequivocally demonstrated that the PCL frame effectively guided the alignment and proliferation of AF cells, while the alginate hydrogel preserved NP cell phenotype. This model successfully replicates the organization of the IVD, providing a promising platform for advancing our understanding of disc biology and driving the development of novel therapeutic strategies.

Extensive peripheral nerve injuries often lead to the loss of neurological function due to slow regeneration and limited recovery over large gaps. Current clinical interventions, such as nerve guidance conduits (NGCs), face challenges in creating biomimetic microenvironments that effectively support nerve repair. The developedGrooveNeuroTubeis composed of hyaluronic acid methacrylate and gelatin methacrylate hydrogel, incorporating active agents (growth factors and antibacterial agents) encapsulated within an NGC conduit made of 3D-printed PCL grid fibers.In vitrostudies showed thatGrooveNeuroTubesignificantly promoted migration of dorsal root ganglion (DRG) neuronal cells, 3D bioprinted at the far ends of the conduit to imitate a proximal nerve injury as a novelex vivomodel. A long-term culture of up to 60 d was employed to better mimicin vivoconditions. This model tested the effects of pulsed electromagnetic field stimulation on neural tissue development. After 60 d,GrooveNeuroTubeshowed a 32% cell migration increase compared to the growth-factor-group and 105% compared to the no-growth-factor condition. These results confirm that theGrooveNeuroTubesystem can effectively support sustained neural cell migration and maturation over extended periods, proving a new technology for testing peripheral nerve injuryex vivo.