Automated Microfluidics-Assisted Hydrogel-Based Wet-Spinning for the Biofabrication of Biomimetic Engineered Myotendinous Junction.

The muscle-tendon junction (MTJ) plays a pivotal role in efficiently converting the muscular contraction into a controlled skeletal movement through the tendon. Given its complex biomechanical intricacy, the biofabrication of such tissue interface represents a significant challenge in the field of musculoskeletal tissue engineering. Herein, a novel method to produce MTJ-like hydrogel yarns using a microfluidics-assisted 3D rotary wet-spinning strategy is developed. Optimization of flow rates, rotational speed, and delivery time of bioinks enables the production of highly compartmentalized scaffolds that recapitulate the muscle, tendon, and the transient MTJ-like region. Additionally, such biofabrication parameters are validated in terms of cellular response by promoting an optimal uniaxial alignment for both muscle and tendon precursor cells. By sequentially wet-spinning C2C12 myoblasts and NIH 3T3 fibroblasts, a gradient-patterned cellular arrangement mirroring the intrinsic biological heterogeneity of the MTJ is successfully obtained. The immunofluorescence assessment further reveals the localized expression of tissue-specific markers, including myosin heavy chain and collagen type I/III, which demonstrate muscle and tenogenic tissue maturation, respectively. Remarkably, the muscle-tendon transition zone exhibits finger-like projection of the multinucleated myotubes in the tenogenic compartment, epitomizing the MTJ signature architecture.

Exploring swine oviduct anatomy through micro-computed tomography: a 3D modeling perspective.

The oviduct plays a crucial role in the reproductive process, serving as the stage for fertilization and the early stages of embryonic development. When the environment of this organ has been mimicked, it has been shown to enhance in vitro embryo epigenetic reprogramming and to improve the yield of the system. This study explores the anatomical intricacies of two oviduct regions, the uterotubal junction (UTJ) and the ampullary-isthmic junction (AIJ) by using micro-computed tomography (MicroCT). In this study, we have characterized and 3D-reconstructed the oviduct structure, by measuring height and width of the oviduct's folds, along with the assessments of fractal dimension, lacunarity and shape factor. Results indicate distinct structural features in UTJ and AIJ, with UTJ displaying small, uniformly distributed folds and high lacunarity, while AIJ shows larger folds with lower lacunarity. Fractal dimension analysis reveals values for UTJ within 1.189-1.1779, while AIJ values range from 1.559-1.770, indicating differences in structural complexity between these regions. Additionally, blind sacs or crypts are observed, akin to those found in various species, suggesting potential roles in sperm sequestration or reservoir formation. These morphological differences align with functional variations and are essential for developing an accurate 3D model. In conclusion, this research provides information about the oviduct anatomy, leveraging MicroCT technology for detailed 3D reconstructions, which can significantly contribute to the understanding of geometric-morphological characteristics influencing functional traits, providing a foundation for a biomimetic oviduct-on-a-chip.

Tuning Physical Properties of GelMA Hydrogels through Microarchitecture for Engineering Osteoid Tissue.

Gelatin methacryloyl (GelMA) hydrogels have gained significant attention due to their biocompatibility and tunable properties. Here, a new approach to engineer GelMA-based matrices to mimic the osteoid matrix is provided. Two cross-linking methods were employed to mimic the tissue stiffness: standard cross-linking (SC) based on visible light exposure (VL) and dual cross-linking (DC) involving physical gelation, followed by VL. It was demonstrated that by reducing the GelMA concentration from 10% (G10) to 5% (G5), the dual-cross-linked G5 achieved a compressive modulus of ∼17 kPa and showed the ability to support bone formation, as evidenced by alkaline phosphatase detection over 3 weeks of incubation in osteogenic medium. Moreover, incorporating poly(ethylene) oxide (PEO) into the G5 and G10 samples was found to hinder the fabrication of highly porous hydrogels, leading to compromised cell survival and reduced osteogenic differentiation, as a consequence of incomplete PEO removal.

Combining rotary wet-spinning biofabrication and electro-mechanical stimulation for thein vitroproduction of functional myo-substitutes.

In this work, we present an innovative, high-throughput rotary wet-spinning biofabrication method for manufacturing cellularized constructs composed of highly-aligned hydrogel fibers. The platform is supported by an innovative microfluidic printing head (MPH) bearing a crosslinking bath microtank with a co-axial nozzle placed at the bottom of it for the immediate gelation of extruded core/shell fibers. After a thorough characterization and optimization of the new MPH and the fiber deposition parameters, we demonstrate the suitability of the proposed system for thein vitroengineering of functional myo-substitutes. The samples produced through the described approach were first characterizedin vitroand then used as a substrate to ascertain the effects of electro-mechanical stimulation on myogenic maturation. Of note, we found a characteristic gene expression modulation of fast (MyH1), intermediate (MyH2), and slow (MyH7) twitching myosin heavy chain isoforms, depending on the applied stimulation protocol. This feature should be further investigated in the future to biofabricate engineered myo-substitutes with specific functionalities.

In vitro functional models for human liver diseases and drug screening: beyond animal testing.

Liver is one of the most important and complex organs in the human body, being characterized by a sophisticated microarchitecture and responsible for key physiological functions. Despite its remarkable ability to regenerate, acute liver failure and chronic liver diseases are major causes of morbidity and mortality worldwide. Therefore, understanding the molecular mechanisms underlying such liver disorders is critical for the successful development of novel therapeutics. In this frame, preclinical animal models have been portrayed as the most commonly used tool to address such issues. However, due to significant species differences in liver architecture, regenerative capacity, disease progression, inflammatory markers, metabolism rates, and drug response, animal models cannot fully recapitulate the complexity of human liver metabolism. As a result, translational research to model human liver diseases and drug screening platforms may yield limited results, leading to failure scenarios. To overcome this impasse, over the last decade, 3D human liver in vitro models have been proposed as an alternative to pre-clinical animal models. These systems have been successfully employed for the investigation of the etiology and dynamics of liver diseases, for drug screening, and - more recently - to design patient-tailored therapies, resulting in potentially higher efficacy and reduced costs compared to other methods. Here, we review the most recent advances in this rapidly evolving field with particular attention to organoid cultures, liver-on-a-chip platforms, and engineered scaffold-based approaches.

Hydrogel-Based Fiber Biofabrication Techniques for Skeletal Muscle Tissue Engineering.

The functional capabilities of skeletal muscle are strongly correlated with its well-arranged microstructure, consisting of parallelly aligned myotubes. In case of extensive muscle loss, the endogenous regenerative capacity is hindered by scar tissue formation, which compromises the native muscle structure, ultimately leading to severe functional impairment. To address such an issue, skeletal muscle tissue engineering (SMTE) attempts to fabricate in vitro bioartificial muscle tissue constructs to assist and accelerate the regeneration process. Due to its dynamic nature, SMTE strategies must employ suitable biomaterials (combined with muscle progenitors) and proper 3D architectures. In light of this, 3D fiber-based strategies are gaining increasing interest for the generation of hydrogel microfibers as advanced skeletal muscle constructs. Indeed, hydrogels possess exceptional biomimetic properties, while the fiber-shaped morphology allows for the creation of geometrical cues to guarantee proper myoblast alignment. In this review, we summarize commonly used hydrogels in SMTE and their main properties, and we discuss the first efforts to engineer hydrogels to guide myoblast anisotropic orientation. Then, we focus on presenting the main hydrogel fiber-based techniques for SMTE, including molding, electrospinning, 3D bioprinting, extrusion, and microfluidic spinning. Furthermore, we describe the effect of external stimulation (i.e., mechanical and electrical) on such constructs and the application of hydrogel fiber-based methods on recapitulating complex skeletal muscle tissue interfaces. Finally, we discuss the future developments in the application of hydrogel microfibers for SMTE.

Tackling Current Biomedical Challenges With Frontier Biofabrication and Organ-On-A-Chip Technologies.

In the last decades, biomedical research has significantly boomed in the academia and industrial sectors, and it is expected to continue to grow at a rapid pace in the future. An in-depth analysis of such growth is not trivial, given the intrinsic multidisciplinary nature of biomedical research. Nevertheless, technological advances are among the main factors which have enabled such progress. In this review, we discuss the contribution of two state-of-the-art technologies-namely biofabrication and organ-on-a-chip-in a selection of biomedical research areas. We start by providing an overview of these technologies and their capacities in fabricating advanced in vitro tissue/organ models. We then analyze their impact on addressing a range of current biomedical challenges. Ultimately, we speculate about their future developments by integrating these technologies with other cutting-edge research fields such as artificial intelligence and big data analysis.