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.

Printing of 3D biomimetic structures for the study of bone metastasis: A review.

Bone metastasis primarily occurs when breast, prostate, or lung cancers disseminate tumoral cells into bone tissue, leading to a range of complications in skeletal tissues and, in severe cases, paralysis resulting from spinal cord compression. Unfortunately, our understanding of pathophysiological mechanisms is incomplete and the translation of bone metastasis research into the clinic has been slow, mainly due to the lack of credible ex vivo and in vivo models to study the disease progression. Development of reliable and rational models to study how tumor cells become circulating cells and then invade and sequentially colonize the bone are in great need. Advances in tissue engineering technologies offers reliable 3D tissue alternatives which answer relevant research questions towards the understanding of cancer evolution and key functional properties of metastasis progression as well as prognosis of therapeutic approach. Here we performed an overview of cellular mechanisms involved in bone metastasis including a short summary of normal bone physiology and metastasis initiation and progression. Also, we comprehensively summarized current advances and methodologies in fabrication of reliable bone tumor models based on state-of-the-art printing technologies which recapitulate structural and biological features of native tissue. STATEMENT OF SIGNIFICANCE: This review provides a comprehensive summary of the collective findings in relation to various printed bone metastasis models utilized for investigating specific bone metastasis diseases, related characteristic functions and chemotherapeutic drug screening. These tumoral models are comprehensively evaluated and compared, in terms of their ability to recapitulate physiological metastasis microenvironment. Various biomaterials (natural and synthetic polymers and ceramic based substrates) and printing strategies and design architecture of models used for printing of 3D bone metastasis models are discussed here. This review clearly out-lines current challenges and prospects for 3D printing technologies in bone metastasis research by focusing on the required perspective models for clinical application of these technologies in chemotherapeutic drug screening.

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.

Tumor Extracellular Matrix Stiffness Promptly Modulates the Phenotype and Gene Expression of Infiltrating T Lymphocytes.

The immune system is a fine modulator of the tumor biology supporting or inhibiting its progression, growth, invasion and conveys the pharmacological treatment effect. Tumors, on their side, have developed escaping mechanisms from the immune system action ranging from the direct secretion of biochemical signals to an indirect reaction, in which the cellular actors of the tumor microenvironment (TME) collaborate to mechanically condition the extracellular matrix (ECM) making it inhospitable to immune cells. TME is composed of several cell lines besides cancer cells, including tumor-associated macrophages, cancer-associated fibroblasts, CD4+ and CD8+ lymphocytes, and innate immunity cells. These populations interface with each other to prepare a conservative response, capable of evading the defense mechanisms implemented by the host's immune system. The presence or absence, in particular, of cytotoxic CD8+ cells in the vicinity of the main tumor mass, is able to predict, respectively, the success or failure of drug therapy. Among various mechanisms of immunescaping, in this study, we characterized the modulation of the phenotypic profile of CD4+ and CD8+ cells in resting and activated states, in response to the mechanical pressure exerted by a three-dimensional in vitro system, able to recapitulate the rheological and stiffness properties of the tumor ECM.