3D Microenvironment Research Articles

Tunneling Nanotubes between Cells Migrating in ECM Mimicking Fibrous Environments.

Simple SummaryTumor cells grow, spread, and invade in a three-dimensional manner, but most experimental approaches in cancer cell biology focus on the behavior of cells in two-dimensional spaces. Patterns of cell invasion and spread in 2D may not accurately depict cell behavior in 3D tumors. We cultivated and investigated malignant mesothelioma cells on a 3D scaffold composed of nanofibers positioned in parallel and crosshatched network configurations to overcome this barrier. We focused on studying long extensions called tunneling nanotubes (TNTs) protruding from cells and connecting with other cells, which have been shown to transmit signals from cell to cell and extend into the tumor microenvironments in a 3D manner. This manuscript describes the biophysics of the formation and function of cancer cell TNTs. These findings will impact the research community by accurately assessing how TNTs affect cancer cell invasion and migration in their natural 3D microenvironment using a novel bioengineered platform.Tunneling nanotubes (TNTs) comprise a unique class of actin-rich nanoscale membranous protrusions. They enable long-distance intercellular communication and may play an integral role in tumor formation, progression, and drug resistance. TNTs are three-dimensional, but nearly all studies have investigated them using two-dimensional cell culture models. Here, we applied a unique 3D culture platform consisting of crosshatched and aligned fibers to fabricate synthetic suspended scaffolds that mimic the native fibrillar architecture of tumoral extracellular matrix (ECM) to characterize TNT formation and function in its native state. TNTs are upregulated in malignant mesothelioma; we used this model to analyze the biophysical properties of TNTs in this 3D setting, including cell migration in relation to TNT dynamics, rate of TNT-mediated intercellular transport of cargo, and conformation of TNT-forming cells. We found that highly migratory elongated cells on aligned fibers formed significantly longer but fewer TNTs than uniformly spread cells on crossing fibers. We developed new quantitative metrics for the classification of TNT morphologies based on shape and cytoskeletal content using confocal microscopy. In sum, our strategy for culturing cells in ECM-mimicking bioengineered scaffolds provides a new approach for accurate biophysical and biologic assessment of TNT formation and structure in native fibrous microenvironments.

Particulate leaching improves spheroid formation in PEG and gelatin-based matrices for 3D tumor model

Spheroids have been developed and used as in vitro tumor models for various applications. However, the most commonly used hanging drop and liquid overlay methods to culture spheroids are challenging due to lack of sturdiness and reproducibility of spheroids and poor microenvironment mimicking features such as matrix stiffness and 3D architecture. Consequently, scaffold-based platforms to culture spheroids have gained momentum. It is already well established that scaffold features such as architecture, surface characteristics and mechanical properties influence the formation of spheroids. In present study, polyethylene glycol diacrylate (PEGDA) and gelatin methacryloyl (GelMA) based scaffolds were fabricated to provide a 3D microenvironment for melanoma spheroids, and pore size of scaffolds was found to be an important parameter influencing the spheroid size. In this study, we have demonstrated that fabricating matrices using a combination of particulate leaching using sodium chloride (NaCl) and freeze-drying enhances the pore size of scaffolds for subsequent formation of larger spheroids (~300 µm) required for various applications. PEGDA-GelMA matrix fabricated in this study provides a sturdy platform for scaffold-based spheroid formation and can be used for long-term culture of multicellular spheroids for development of in vitro tumor models.

Self-organization of human stem cells into spheroids in a multinode acoustic levitation

We present an approach based on acoustic levitation to grow the cells without wall interactions in order to promote 3D cell architecture (spheroïd, organoïd). The 3D microenvironment is indeed closer to In Vivo physiological behavior. The suspended cells are moved toward the acoustic pressure nodes where they are trapped and maintained in acoustic levitation in perfectly straight monolayers reaching very quickly the classical confluency of the cells in 2D culture. Interestingly, by maintaining the monolayers of MSCs in culture over a 24-h period, the MSCs spontaneously self-organized from cell sheets to cell spheroids with a characteristic time of about 10 h. This approach of 3D cell culture is based on the use of the acoustic wave coupled with microfluidics. We designed a standing wave cavity to generate a large acoustic radiation force (ARF) with an optical access, which allows the characterization of the self-organization dynamics. This 3D cell culture method has been validated on MSCs over 24h experiments. The MSCs viability has been checked. Moreover, they show a higher differentiation capacity compared to standard 2D culture conditions. These results open the path to long-time cell culture in acoustic levitation of cell sheets or spheroids for any type of cells.

3D hybrid scaffold with aligned nanofiber yarns embedded in injectable hydrogels for monitoring and repairing chronic wounds

Pressure ulcers (PU) as a major clinical concern are disproportionately affecting the older adult population and resulting in a lot of medical expenses. The issues of preventing the incidence of PU and treating chronic wounds are urgent to be solved at the present stage. Here, a 3D hybrid scaffold with conductive and antibacterial properties is designed to monitor and repair chronic wounds. The 3D hybrid scaffold is prepared by encapsulating nanofiber yarns (NFY) network into the injectable hydrogel which is cross-linked by Schiff base . The aligned NFY networks assembled by weaving nanofiber yarns consisting of polyacrylonitrile (PAN) and reduced graphene oxide (rGO) can induce cellular elongation and alignment on their 3D peripheral surface. The injectable hydrogels outside not only provide the protection for the cells on the surfaces of the NFYs, but also mimic a native extracellular matrix (ECM) to supply a much more suitable 3D microenvironment for the cells proliferation and arrangement. The addition of ions within the hydrogel endows the hydrogels with excellent conductivity , making the hydrogels the potential flexible strain sensor to exactly monitor the wound pressure. The 3D hybrid scaffold with aligned nanofiber yarns embedded in injectable hydrogels displays favorable biocompatibility, self-healing and antibacterial properties, showing promising medical applications especially in tissue engineering and wearable electronics .

Tuning the viscoelastic response of hydrogel scaffolds with covalent and dynamic bonds

The processes of growth, proliferation and differentiation of stem cells encapsulated in 3D hydrogel microenvironments are strongly affected by the viscoelastic properties of the platforms. As the viscoelastic response of a hydrogel is determined by the rates of thermally induced dissociation of reversible cross-links, its modulation by introduction of several types of supramolecular and/or dynamic covalent bonds with different characteristic lifetimes has recently become a hot topic.To reduce the number of experiments needed for design of hydrogel microenvironments with required mechanical properties, a model is developed for the viscoelastic and viscoplastic responses of hydrogels with multiple networks bridged by covalent and physical bonds. An advantage of the model is that it (i) involves a small number of material parameters, (ii) describes observations in rheological and mechanical tests in a unified manner, and (iii) predicts conventional measures of viscoelasticity used in the analysis of viability of cells.

Multimodal microwheel swarms for targeting in three-dimensional networks

Microscale bots intended for targeted drug delivery must move through three-dimensional (3D) environments that include bifurcations, inclined surfaces, and curvature. In previous studies, we have shown that magnetically actuated colloidal microwheels (µwheels) reversibly assembled from superparamagnetic beads can translate rapidly and be readily directed. Here we show that, at high concentrations, µwheels assemble into swarms that, depending on applied magnetic field actuation patterns, can be designed to transport cargo, climb steep inclines, spread over large areas, or provide mechanical action. We test the ability of these multimodal swarms to navigate through complex, inclined microenvironments by characterizing the translation and dispersion of individual µwheels and swarms of µwheels on steeply inclined and flat surfaces. Swarms are then studied within branching 3D vascular models with multiple turns where good targeting efficiencies are achieved over centimeter length scales. With this approach, we present a readily reconfigurable swarm platform capable of navigating through 3D microenvironments.

Polymeric Hydrogels for In Vitro 3D Ovarian Cancer Modeling.

Ovarian cancer (OC) grows and interacts constantly with a complex microenvironment, in which immune cells, fibroblasts, blood vessels, signal molecules and the extracellular matrix (ECM) coexist. This heterogeneous environment provides structural and biochemical support to the surrounding cells and undergoes constant and dynamic remodeling that actively promotes tumor initiation, progression, and metastasis. Despite the fact that traditional 2D cell culture systems have led to relevant medical advances in cancer research, 3D cell culture models could open new possibilities for the development of an in vitro tumor microenvironment more closely reproducing that observed in vivo. The implementation of materials science and technology into cancer research has enabled significant progress in the study of cancer progression and drug screening, through the development of polymeric scaffold-based 3D models closely recapitulating the physiopathological features of native tumor tissue. This article provides an overview of state-of-the-art in vitro tumor models with a particular focus on 3D OC cell culture in pre-clinical studies. The most representative OC models described in the literature are presented with a focus on hydrogel-based scaffolds, which guarantee soft tissue-like physical properties as well as a suitable 3D microenvironment for cell growth. Hydrogel-forming polymers of either natural or synthetic origin investigated in this context are described by highlighting their source of extraction, physical-chemical properties, and application for 3D ovarian cancer cell culture.

Nanostructure Formation and Cell Spheroid Morphogenesis of a Peptide Supramolecular Hydrogel.

Peptide-based hydrogels have attracted much attention due to their extraordinary applications in biomedicine and offer an excellent mimic for the 3D microenvironment of the extracellular matrix. These hydrated matrices comprise fibrous networks held together by a delicate balance of intermolecular forces. Here, we investigate the hydrogelation behavior of a designed decapeptide containing a tetraleucine self-assembling backbone and fibronectin-related tripeptides near both ends of the strand. We have observed that this synthetic peptide can produce hydrogel matrices entrapping >99% wt/vol % water. Ultrastructural analyses combining atomic force microscopy, small-angle neutron scattering, and X-ray diffraction revealed that amyloid-like fibrils form cross-linked networks endowed with remarkable thermal stability, the structure of which is not disrupted up to temperatures >80 °C. We also examined the interaction of peptide hydrogels with either NIH3T3 mouse fibroblasts or HeLa cells and discovered that the matrices sustain cell viability and induce morphogenesis into grape-like cell spheroids. The results presented here show that this decapeptide is a remarkable building block to prepare highly stable scaffolds simultaneously endowed with high water retention capacity and the ability to instruct cell growth into tumor-like spheroids even in noncarcinoma lineages.

Three-Dimensional Silk Fibroin/Chitosan Based Microscaffold for Anticancer Drug Screening.

Traditional monolayer cell cultures often fail to accurately predict the anticancer activity of drug candidates, as they do not recapitulate the natural microenvironment. Recently, three-dimensional (3D) culture systems have been increasingly applied to cancer research and drug screening. Materials with good biocompatibility are crucial to create a 3D tumor microenvironment involved in such systems. In this study, natural silk fibroin (SF) and chitosan (CS) were selected as the raw materials to fabricate 3D microscaffolds; Besides, sodium tripolyphosphate (TPP), and 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) were used as cross-linking agents. The physicochemical properties of obtained scaffolds were characterized with kinds of testing methods, including emission scanning electron microscopy, x-ray photoelectron spectroscopy, fourier transform infrared spectroscopy, water absorption, and swelling ratio analysis. Cancer cell lines (LoVo and MDA-MB-231) were then seeded on scaffolds for biocompatibility examination and drug sensitivity tests. SEM results showed that EDC cross-linked scaffolds had smaller and more uniform pores with great interconnection than the TPP cross-linked scaffolds, and the EDC cross-linked scaffold exhibited a water absorption ratio around 1000% and a swelling ratio of about 72%. These spatial structures and physical properties could provide more adhesion sites and sufficient nutrients for cell growth. Moreover, both LoVo and MDA-MB-231 cells cultured on the EDC cross-linked scaffold exhibited good adhesion and spreading. CCK8 results showed that increased chemotherapeutic drug sensitivity was observed in 3D culture compared with 2D culture, particularly in the condition of low drug dose (<1 M). The proposed SF/CS microscaffold can provide a promising in vitro platform for the efficacy prediction and sensitivity screening of anticancer drugs.

Decellularized Avian Cartilage, a Promising Alternative for Human Cartilage Tissue Regeneration.

Articular cartilage defects, and subsequent degeneration, are prevalent and account for the poor quality of life of most elderly persons; they are also one of the main predisposing factors to osteoarthritis. Articular cartilage is an avascular tissue and, thus, has limited capacity for healing and self-repair. Damage to the articular cartilage by trauma or pathological causes is irreversible. Many approaches to repair cartilage have been attempted with some potential; however, there is no consensus on any ideal therapy. Tissue engineering holds promise as an approach to regenerate damaged cartilage. Since cell adhesion is a critical step in tissue engineering, providing a 3D microenvironment that recapitulates the cartilage tissue is vital to inducing cartilage regeneration. Decellularized materials have emerged as promising scaffolds for tissue engineering, since this procedure produces scaffolds from native tissues that possess structural and chemical natures that are mimetic of the extracellular matrix (ECM) of the native tissue. In this work, we present, for the first time, a study of decellularized scaffolds, produced from avian articular cartilage (extracted from Gallus Gallus domesticus), reseeded with human chondrocytes, and we demonstrate for the first time that human chondrocytes survived, proliferated and interacted with the scaffolds. Morphological studies of the decellularized scaffolds revealed an interconnected, porous architecture, ideal for cell growth. Mechanical characterization showed that the decellularized scaffolds registered stiffness comparable to the native cartilage tissues. Cell growth inhibition and immunocytochemical analyses showed that the decellularized scaffolds are suitable for cartilage regeneration.

Think outside the box: 3D bioprinting concepts for biotechnological applications – recent developments and future perspectives

3D bioprinting – the fabrication of geometrically complex 3D structures from biocompatible materials containing living cells using additive manufacturing technologies – is a rapidly developing research field with a broad range of potential applications in fundamental research, regenerative medicine and industry. Currently, research into 3D bioprinting is mostly focused on new therapeutic concepts for the treatment of injured or degenerative tissue by fabrication of functional tissue equivalents or disease models, utilizing mammalian cells. However, 3D bioprinting also has an enormous potential in biotechnology. Due to the defined spatial arrangement of biologically active (non-mammalian) cells in a biomaterial matrix, reaction compartments can be designed according to specific needs, or co-cultures of different cell types can be realized in a highly organized manner to exploit cell-cell interactions. Thus, 3D bioprinting technology can enable new biotechnological concepts, for example, by implementing perfusion systems while protecting shear sensitive cells or performing cascaded bioreactions. Here, we review the use of 3D bioprinting to manufacture defined 3D microenvironments for biotechnological applications using bacteria, fungi, microalgae, plant cells and co-cultures of different cell types. We discuss recent approaches to apply 3D bioprinting in biotechnological applications and – as it is a particular challenge – concepts for the real-time monitoring of the physiological state, growth and metabolic activity of the embedded cells in 3D bioprinted constructs. With these insights, we outline new applications of 3D bioprinting in biotechnology, engineered living materials and space research.

Bioinks Enriched with ECM Components Obtained by Supercritical Extraction.

Extracellular matrix (ECM)-based bioinks have been steadily gaining interest in the field of bioprinting to develop biologically relevant and functional tissue constructs. Herein, we propose the use of supercritical carbon dioxide (scCO2) technology to extract the ECM components of cell-sheets that have shown promising results in creating accurate 3D microenvironments replicating the cell’s own ECM, to be used in the preparation of bioinks. The ECM extraction protocol best fitted for cell sheets was defined by considering efficient DNA removal with a minor effect on the ECM. Cell sheets of human dermal fibroblasts (hDFbs) and adipose stem cells (hASCs) were processed using a customised supercritical system by varying the pressure of the reactor, presence, exposure time, and type of co-solvent. A quantification of the amount of DNA, protein, and sulfated glycosaminoglycans (sGAGs) was carried out to determine the efficiency of the extraction in relation to standard decellularization methodologies. The bioinks containing the extracted ECM were fabricated by combining them with alginate as a support polymer. The influence of the alginate (1%, 2% w/vol) and ECM (0.5% and 1.5% w/vol) amounts on the printability of the blends was addressed by analysing the rheological behaviour of the suspensions. Finally, 3D printed constructs were fabricated using an in-house built extrusion-based bioprinter, and the impact of the extrusion process on cell viability was assessed. The optimised scCO2 protocol allowed efficient removal of DNA while preserving a higher number of proteins and sGAGs than the standard methodologies. The characterization of extract’s composition also revealed that the ECM produced by hDFbs (fECM) and hASCs (aECM) is distinctively affected by the extraction protocols. Furthermore, rheological analysis indicated an increase in viscosity with increasing ECM composition, an effect even more prominent in samples containing aECM. 3D printing of alginate/ECM constructs demonstrated that cell viability was only marginally affected by the extrusion process, and this effect was also dependent on the ECM source. Overall, this work highlights the benefits of supercritical fluid-based methods for ECM extraction and strengthens the relevance of ECM-derived bioinks in the development of printed tissue-like constructs.

Mast Cells Tryptase Promotes Intestinal Fibrosis in Natural Decellularized Intestinal Scaffolds.

Standard two-dimensional (2D) culture has confirmed the mechanism of mast cells (MCs) in the pathogenesis of inflammatory bowel disease(IBD), but the regulation of signaling responses of MCs may well differ in three-dimensional (3D) microenvironments. The aim of the study was to develop a 3D culture model based on decellularized intestinal scaffolds (DIS) and verify how MCs influenced fibroblasts phenotype in the 3D model. DIS were achieved using the detergent technique and extracellular matrix (ECM) components were verified by histologic analysis, quantification and scanning electron microscope. After human colon fibroblasts recellularized into the scaffolds and activated by MCs tryptase and TGFβ1, the changes in genes and signaling pathways during fibroblasts activation in 3D were studied and compared with the changes in 2D cell culture on plastic plates. Decellularization process effectively removed native cell debris while retaining natural ECM components and structure. The engrafted fibroblasts could penetrate into the scaffolds and maintain its phenotype. No matter whether fibroblasts were cultured in 2D or 3D, MCs tryptase and transforming growth factor β1 (TGF-β1) could promote the differentiation of fibroblasts into fibrotic-phenotype myofibroblasts through Akt and Smad2/3 signaling pathways. Furthermore, the pro-collagen1α1 and fibronectin synthesis of myofibroblasts in 3D was higher than in 2D culture. Our results demonstrated that the DIS can be used as a bioactive microenvironment for the study of intestinal fibrosis, providing an innovative platform for future intestinal disease modeling and screening of genes and signaling pathways.

Cardiac Tissue-like 3D Microenvironment Enhances Route towards Human Fibroblast Direct Reprogramming into Induced Cardiomyocytes by microRNAs.

The restoration of cardiac functionality after myocardial infarction represents a major clinical challenge. Recently, we found that transient transfection with microRNA combination (miRcombo: miR-1, miR-133, miR-208 and 499) is able to trigger direct reprogramming of adult human cardiac fibroblasts (AHCFs) into induced cardiomyocytes (iCMs) in vitro. However, achieving efficient direct reprogramming still remains a challenge. The aim of this study was to investigate the influence of cardiac tissue-like biochemical and biophysical stimuli on direct reprogramming efficiency. Biomatrix (BM), a cardiac-like extracellular matrix (ECM), was produced by in vitro culture of AHCFs for 21 days, followed by decellularization. In a set of experiments, AHCFs were transfected with miRcombo and then cultured for 2 weeks on the surface of uncoated and BM-coated polystyrene (PS) dishes and fibrin hydrogels (2D hydrogel) or embedded into 3D fibrin hydrogels (3D hydrogel). Cell culturing on BM-coated PS dishes and in 3D hydrogels significantly improved direct reprogramming outcomes. Biochemical and biophysical cues were then combined in 3D fibrin hydrogels containing BM (3D BM hydrogel), resulting in a synergistic effect, triggering increased CM gene and cardiac troponin T expression in miRcombo-transfected AHCFs. Hence, biomimetic 3D culture environments may improve direct reprogramming of miRcombo-transfected AHCFs into iCMs, deserving further study.

Clinical Application Perspectives of Lung Cancers 3D Tumor Microenvironment Models for In Vitro Cultures.

Despite the enormous progress and development of modern therapies, lung cancer remains one of the most common causes of death among men and women. The key element in the development of new anti-cancer drugs is proper planning of the preclinical research phase. The most adequate basic research exemplary for cancer study are 3D tumor microenvironment in vitro models, which allow us to avoid the use of animal models and ensure replicable culture condition. However, the question tormenting the scientist is how to choose the best tool for tumor microenvironment research, especially for extremely heterogenous lung cancer cases. In the presented review we are focused to explain the key factors of lung cancer biology, its microenvironment, and clinical gaps related to different therapies. The review summarized the most important strategies for in vitro culture models mimicking the tumor–tumor microenvironmental interaction, as well as all advantages and disadvantages were depicted. This knowledge could facilitate the right decision to designate proper pre-clinical in vitro study, based on available analytical tools and technical capabilities, to obtain more reliable and personalized results for faster introduction them into the future clinical trials.

Modeling analysis of microenvironment of 3D cell mechanics based on machine vision

Abstract Aiming at the problem of poor construction accuracy of the cellular three-dimensional (3D) mechanical microenvironment, this article studies the cellular 3D mechanical microenvironment based on machine vision. The gelatin methacrylate microgel column was prepared by NIH/3T3 mouse fibroblast and precursor solution of gelatin methacrylate microgel. The gelatin methacrylate microgel array with magnetic end was adopted. The external magnetic field was used to load microgel array and build 3D mechanics microenvironment model. The deformed pictures of hydrogel under magnetic field were obtained by fluorescence microscope. The scanning electron microscope was used to characterize the pore structure of gelatin methacrylate hydrogel. The pictures obtained by machine vision method were used to calculate the deformed parameters of sample. The machine vision method adopted the discrete cosine transform for autofocus, and then used the image analysis and processing technology to identify and estimate the cell motion parameters. After getting the cell motion parameters, Comsol multiphysics (COMSOL) multiphysics multifield coupling finite element analysis software was adopted. The correlative numerical simulation method and gel deformed simulation method were used to obtain the mechanical changes of cells in the 3D mechanical microenvironment. Experimental results show that the modulus of gelatin methacrylate microgel is changed significantly during the tensile loading. The tensile strain and the cell spreading area are nonlinearly related. The increase in stiffness of the hydrogel substrate helps to promote cell proliferation to a certain extent.

Quantitative mapping of keratin networks in 3D.

Mechanobiology requires precise quantitative information on processes taking place in specific 3D microenvironments. Connecting the abundance of microscopical, molecular, biochemical, and cell mechanical data with defined topologies has turned out to be extremely difficult. Establishing such structural and functional 3D maps needed for biophysical modeling is a particular challenge for the cytoskeleton, which consists of long and interwoven filamentous polymers coordinating subcellular processes and interactions of cells with their environment. To date, useful tools are available for the segmentation and modeling of actin filaments and microtubules but comprehensive tools for the mapping of intermediate filament organization are still lacking. In this work, we describe a workflow to model and examine the complete 3D arrangement of the keratin intermediate filament cytoskeleton in canine, murine, and human epithelial cells both, in vitro and in vivo. Numerical models are derived from confocal airyscan high-resolution 3D imaging of fluorescence-tagged keratin filaments. They are interrogated and annotated at different length scales using different modes of visualization including immersive virtual reality. In this way, information is provided on network organization at the subcellular level including mesh arrangement, density and isotropic configuration as well as details on filament morphology such as bundling, curvature, and orientation. We show that the comparison of these parameters helps to identify, in quantitative terms, similarities and differences of keratin network organization in epithelial cell types defining subcellular domains, notably basal, apical, lateral, and perinuclear systems. The described approach and the presented data are pivotal for generating mechanobiological models that can be experimentally tested.

Illuminating the Regenerative Microenvironment: Emerging Quantitative Imaging Technologies for Craniofacial Bone Tissue Engineering.

Tissue engineering has the potential to revolutionize treatments for patients suffering from critical-sized craniofacial bone defects, but it has yet to make a substantial impact in clinical practice. One of the barriers to improving the design of tissue-engineered bone grafts (TEBGs) is the lack of adequate techniques to study how transplanted cells, host cells, and biomaterials interact to facilitate the dynamic healing process. In this perspective, we discuss recent advances in quantitative imaging that may be adapted to provide high spatiotemporal resolution of the 3D tissue microenvironment during cranial bone regeneration. The adoption and application of these imaging technologies will provide a more rigorous framework for evaluating TEBG performance and enable the development of next-generation TEBGs for craniofacial repair.

3D Bioprinting of an In Vitro Model of a Biomimetic Urinary Bladder with a Contract-Release System.

The development of curative therapy for bladder dysfunction is usually hampered owing to the lack of reliable ex vivo human models that can mimic the complexity of the human bladder. To overcome this issue, 3D in vitro model systems offering unique opportunities to engineer realistic human tissues/organs have been developed. However, existing in vitro models still cannot entirely reflect the key structural and physiological characteristics of the native human bladder. In this study, we propose an in vitro model of the urinary bladder that can create 3D biomimetic tissue structures and dynamic microenvironments to replicate the smooth muscle functions of an actual human urinary bladder. In other words, the proposed biomimetic model system, developed using a 3D bioprinting approach, can recreate the physiological motion of the urinary bladder by incorporating decellularized extracellular matrix from the bladder tissue and introducing cyclic mechanical stimuli. The results showed that the developed bladder tissue models exhibited high cell viability and proliferation rate and promoted myogenic differentiation potential given dynamic mechanical cues. We envision the developed in vitro bladder mimicry model can serve as a research platform for fundamental studies on human disease modeling and pharmaceutical testing.

Regenerating the Injured Spinal Cord at the Chronic Phase by Engineered iPSCs-Derived 3D Neuronal Networks.

Cell therapy using induced pluripotent stem cell‐derived neurons is considered a promising approach to regenerate the injured spinal cord (SC). However, the scar formed at the chronic phase is not a permissive microenvironment for cell or biomaterial engraftment or for tissue assembly. Engineering of a functional human neuronal network is now reported by mimicking the embryonic development of the SC in a 3D dynamic biomaterial‐based microenvironment. Throughout the in vitro cultivation stage, the system's components have a synergistic effect, providing appropriate cues for SC neurogenesis. While the initial biomaterial supported efficient cell differentiation in 3D, the cells remodeled it to provide an inductive microenvironment for the assembly of functional SC implants. The engineered tissues are characterized for morphology and function, and their therapeutic potential is investigated, revealing improved structural and functional outcomes after acute and chronic SC injuries. Such technology is envisioned to be translated to the clinic to rewire human injured SC.