Three-dimensional Microenvironment Research Articles

Three-dimensional hydrogel with human Wharton jelly-derived mesenchymal stem cells towards nucleus pulposus niche.

Introduction: A regenerative strategy employing extracellular matrix (ECM)-based biomaterials and stem cells provide a better approach to mimicking the three-dimensional (3D) microenvironment of intervertebral disc for endogenous tissue regeneration. However, there is currently limited understanding regarding the human Wharton Jelly derived-mesenchymal stem cells (hWJ-MSCs) towards nucleus pulposus (NP)-like cells. Our study focused on the development of 3D bioengineered hydrogel based on the predominant ECM of native NP, including type II collagen (COLII) and hyaluronic acid (HA), which aims to tailor the needs of the microenvironment in NP. Methods: We have fabricated a 3D hydrogel using from COLII enriched with HA by varying the biomacromolecule concentration and characterised it for degradation, stability and swelling properties. The WJ-MSC was then encapsulated in the hydrogel system to guide the cell differentiation into NP-like cells. Results: We successfully fabricated COLII hydrogel (2mg/ml) and HA 10mg/ml at a weight ratio of HA and COLII at 1:9 and 4.5:9, and both hydrogels physically maintained their 3D sphere-shaped structure after complete gelation. The higher composition of HA in the hydrogel system indicated a higher water intake capacity in the hydrogel with a higher amount of HA. All hydrogels showed over 60% hydrolytic stability over a month. The hydrogel showed an increase in degradation on day 14. The hWJ-MSCs encapsulated in hydrogel showed a round morphology shape that was homogenously distributed within the hydrogel of both groups. The viability study indicated a higher cell growth of hWJ-MSCs encapsulated in all hydrogel groups until day 14. Discussion: Overall, our findings demonstrate that HA/COLII hydrogel provides an optimal swelling capacity, stability, degradability, and non-cytotoxic, thus mimics the NP microenvironment in guiding hWJ-MSCs towards NP phenotype, which is potentially used as an advanced cell delivery system for intervertebral disc regeneration.

Harnessing decellularised extracellular matrix microgels into modular bioinks for extrusion-based bioprinting with good printability and high post-printing cell viability.

The printability of bioink and post-printing cell viability is crucial for extrusion-based bioprinting. A proper bioink not only provides mechanical support for structural fidelity, but also serves as suitable three-dimensional (3D) microenvironment for cell encapsulation and protection. In this study, a hydrogel-based composite bioink was developed consisting of gelatin methacryloyl (GelMA) as the continuous phase and decellularised extracellular matrix microgels (DMs) as the discrete phase. A flow-focusing microfluidic system was employed for the fabrication of cell-laden DMs in a high-throughput manner. After gentle mixing of the DMs and GelMA, both rheological characterisations and 3D printing tests showed that the resulting DM-GelMA hydrogel preserved the shear-thinning nature, mechanical properties, and good printability from GelMA. The integration of DMs not only provided an extracellular matrix-like microenvironment for cell encapsulation, but also considerable shear-resistance for high post-printing cell viability. The DM sizes and inner diameters of the 3D printer needles were correlated and optimised for nozzle-based extrusion. Furthermore, a proof-of-concept bioink composedg of RSC96 Schwann cells encapsulated DMs and human umbilical vein endothelial cell-laden GelMA was successfully bioprinted into 3D constructs, resulting in a modular co-culture system with distinct cells/materials distribution. Overall, the modular DM-GelMA bioink provides a springboard for future precision biofabrication and will serve in numerous biomedical applications such as tissue engineering and drug screening.

Mechanics of the cellular microenvironment as probed by cells in vivo during zebrafish presomitic mesoderm differentiation

Tissue morphogenesis, homoeostasis and repair require cells to constantly monitor their three-dimensional microenvironment and adapt their behaviours in response to local biochemical and mechanical cues. Yet the mechanical parameters of the cellular microenvironment probed by cells in vivo remain unclear. Here, we report the mechanics of the cellular microenvironment that cells probe in vivo and in situ during zebrafish presomitic mesoderm differentiation. By quantifying both endogenous cell-generated strains and tissue mechanics, we show that individual cells probe the stiffness associated with deformations of the supracellular, foam-like tissue architecture. Stress relaxation leads to a perceived microenvironment stiffness that decreases over time, with cells probing the softest regime. We find that most mechanical parameters, including those probed by cells, vary along the anteroposterior axis as mesodermal progenitors differentiate. These findings expand our understanding of in vivo mechanosensation and might aid the design of advanced scaffolds for tissue engineering applications.

Mice Placental ECM Components May Provide A Three-Dimensional Placental Microenvironment.

Bioethical limitations impair deeper studies in human placental physiology, then most studies use human term placentas or murine models. To overcome these challenges, new models have been proposed to mimetize the placental three-dimensional microenvironment. The placental extracellular matrix plays an essential role in several processes, being a part of the establishment of materno-fetal interaction. Regarding these aspects, this study aimed to investigate term mice placental ECM components, highlighting its collagenous and non-collagenous content, and proposing a potential three-dimensional model to mimetize the placental microenvironment. For that, 18.5-day-old mice placenta, both control and decellularized (n = 3 per group) were analyzed on Orbitrap Fusion Lumos spectrometer (ThermoScientific) and LFQ intensity generated on MaxQuant software. Proteomic analysis identified 2317 proteins. Using ECM and cell junction-related ontologies, 118 (5.1%) proteins were filtered. Control and decellularized conditions had no significant differential expression on 76 (64.4%) ECM and cell junction-related proteins. Enriched ontologies in the cellular component domain were related to cell junction, collagen and lipoprotein particles, biological process domain, cell adhesion, vasculature, proteolysis, ECM organization, and molecular function. Enriched pathways were clustered in cell adhesion and invasion, and labyrinthine vasculature regulation. These preserved ECM proteins are responsible for tissue stiffness and could support cell anchoring, modeling a three-dimensional structure that may allow placental microenvironment reconstruction.

Three-dimensional heart extracellular matrix enhances chemically induced direct cardiac reprogramming

Direct cardiac reprogramming has emerged as a promising therapeutic approach for cardiac regeneration. Full chemical reprogramming with small molecules to generate cardiomyocytes may be more amenable than genetic reprogramming for clinical applications as it avoids safety concerns associated with genetic manipulations. However, challenges remain regarding low conversion efficiency and incomplete cardiomyocyte maturation. Furthermore, the therapeutic potential of chemically induced cardiomyocytes (CiCMs) has not been investigated. Here, we report that a three-dimensional microenvironment reconstituted with decellularized heart extracellular matrix can enhance chemical reprogramming and cardiac maturation of fibroblasts to cardiomyocytes. The resultant CiCMs exhibit elevated cardiac marker expression, sarcomeric organization, and improved electrophysiological features and drug responses. We investigated the therapeutic potential of CiCMs reprogrammed in three-dimensional heart extracellular matrix in a rat model of myocardial infarction. Our platform can facilitate the use of CiCMs for regenerative medicine, disease modeling, and drug screening.

Multiplexed Detection of Molecular Interactions with DNA Origami Engineered Cells in 3D Collagen Matrices.

The interactions of cells with signaling molecules present in their local microenvironment maintain cell proliferation, differentiation, and spatial organization and mediate progression of diseases such as metabolic disorders and cancer. Real-time monitoring of the interactions between cells and their extracellular ligands in a three-dimensional (3D) microenvironment can inform detection and understanding of cell processes and the development of effective therapeutic agents. DNA origami technology allows for the design and fabrication of biocompatible and 3D functional nanodevices via molecular self-assembly for various applications including molecular sensing. Here, we report a robust method to monitor live cell interactions with molecules in their surrounding environment in a 3D tissue model using a microfluidic device. We used a DNA origami cell sensing platform (CSP) to detect two specific nucleic acid sequences on the membrane of B cells and dendritic cells. We further demonstrated real-time detection of biomolecules with the DNA sensing platform on the surface of dendritic cells in a 3D microfluidic tissue model. Our results establish the integration of live cells with membranes engineered with DNA nanodevices into microfluidic chips as a highly capable biosensor approach to investigate subcellular interactions in physiologically relevant 3D environments under controlled biomolecular transport.

Carboxymethyl Chitosan and Gelatin Hydrogel Scaffolds Incorporated with Conductive PEDOT Nanoparticles for Improved Neural Stem Cell Proliferation and Neuronal Differentiation

Tissue engineering scaffolds provide biological and physiochemical cures to guide tissue recovery, and electrical signals through the electroactive materials possess tremendous potential to modulate the cell fate. In this study, a novel electroactive hydrogel scaffold was fabricated by assembling poly(3,4-ethylenedioxythiophene) (PEDOT) nanoparticles on a carboxymethyl chitosan/gelatin (CMCS/Gel) composite hydrogel surface via in situ chemical polymerization. The chemical structure, morphology, conductivity, porosity, swelling rate, in vitro biodegradation, and mechanical properties of the prepared hydrogel samples were characterized. The adhesion, proliferation, and differentiation of neural stem cells (NSCs) on conductive hydrogels were investigated. The CMCS/Gel-PEDOT hydrogels exhibited high porosity, excellent water absorption, improved thermal stability, and adequate biodegradability. Importantly, the mechanical properties of the prepared hydrogels were similar to those of brain tissue, with electrical conductivity up to (1.52 ± 0.15) × 10-3 S/cm. Compared to the CMCS/Gel hydrogel, the incorporation of PEDOT nanoparticles significantly improved the adhesion of NSCs, and supported long-term cell growth and proliferation in a three-dimensional (3D) microenvironment. In addition, under the differentiation condition, the conductive hydrogel also significantly enhanced neuronal differentiation with the up-regulation of β-tubulin III expression. These results suggest that CMCS/Gel-PEDOT hydrogels may be an attractive conductive substrate for further studies on neural tissue repair and regeneration.

Vascularization Strategies in 3D Cell Culture Models: From Scaffold-Free Models to 3D Bioprinting

The discrepancies between the findings in preclinical studies, and in vivo testing and clinical trials have resulted in the gradual decline in drug approval rates over the past decades. Conventional in vitro drug screening platforms employ two-dimensional (2D) cell culture models, which demonstrate inaccurate drug responses by failing to capture the three-dimensional (3D) tissue microenvironment in vivo. Recent advancements in the field of tissue engineering have made possible the creation of 3D cell culture systems that can accurately recapitulate the cell-cell and cell-extracellular matrix interactions, as well as replicate the intricate microarchitectures observed in native tissues. However, the lack of a perfusion system in 3D cell cultures hinders the establishment of the models as potential drug screening platforms. Over the years, multiple techniques have successfully demonstrated vascularization in 3D cell cultures, simulating in vivo-like drug interactions, proposing the use of 3D systems as drug screening platforms to eliminate the deviations between preclinical and in vivo testing. In this review, the basic principles of 3D cell culture systems are briefly introduced, and current research demonstrating the development of vascularization in 3D cell cultures is discussed, with a particular focus on the potential of these models as the future of drug screening platforms.

Investigations of an organic–inorganic nanotheranostic hybrid for pancreatic cancer therapy using cancer-in-a-dish and in vivo models

The incidence of highly aggressive pancreatic cancer is increasing across the globe and is projected to increase to 18.6% by 2050. The mortality rate for this form of cancer is very high and the 5 y relative survival rate is only about 9%–10%. The 3D pancreatic cancer microenvironment exerts a major influence on the poor survival rate. A key factor is the prevention of the penetration of the chemotherapeutic drugs in the three-dimensional (3D) microenvironment leading to the development of chemoresistance which is a major contributor to the survival rates. Hence, in vitro studies using 3D cultures represent a better approach to understand the effect of therapeutic formulations on the cancer cells when compared to conventional 2D cultures. In the present study, we have explored three different conditions for the development of a 3D pancreatic tumour spheroid model from MiaPaCa-2 and PanC1 cells cultured for 10 days using Matrigel matrix. This optimized spheroid model was employed to evaluate a multi-functional nanotheranostic system fabricated using chitosan nanoparticles co-encapsulated with the chemotherapeutic agent gemcitabine and gold-capped iron oxide nanoparticles for multimodal imaging. The effect of the single and multiple-dose regimens of the theranostic system on the viability of 3D spheroids formed from the two pancreatic cancer cell lines was studied. It was observed that the 3D tumour spheroids cultured for 10 days exhibited resistance towards free gemcitabine drug, unlike the 2D culture. The administration of the multifunctional nanotheranostic system on alternate days effectively reduced the cancer cell viability after five doses to about 20% when compared with other groups. The repeated doses of the nanotheranostic system were found to be more effective than the single dose. Cell line-based differences in internalization of the carrier was also reflected in their response to the nanocarrier with PanC1 showing better sensitivity to the treatment. In vivo studies revealed that the combination of gemcitabine and magnetic field induced hypothermia produced superior regression in cancer when compared with the chemotherapeutic agent alone by a combination of activating the pro-apoptotic pathway and heat-induced necrosis. Our results reveal that this multi-functional system holds promise to overcome the current challenges to treat pancreatic cancers.

Three-dimensional microenvironment regulates gene expression, function, and tight junction dynamics of iPSC-derived blood–brain barrier microvessels

The blood–brain barrier (BBB) plays a pivotal role in brain health and disease. In the BBB, brain microvascular endothelial cells (BMECs) are connected by tight junctions which regulate paracellular transport, and express specialized transporter systems which regulate transcellular transport. However, existing in vitro models of the BBB display variable accuracy across a wide range of characteristics including gene/protein expression and barrier function. Here, we use an isogenic family of fluorescently-labeled iPSC-derived BMEC-like cells (iBMECs) and brain pericyte-like cells (iPCs) within two-dimensional confluent monolayers (2D) and three-dimensional (3D) tissue-engineered microvessels to explore how 3D microenvironment regulates gene expression and function of the in vitro BBB. We show that 3D microenvironment (shear stress, cell-ECM interactions, and cylindrical geometry) increases BBB phenotype and endothelial identity, and alters angiogenic and cytokine responses in synergy with pericyte co-culture. Tissue-engineered microvessels incorporating junction-labeled iBMECs enable study of the real-time dynamics of tight junctions during homeostasis and in response to physical and chemical perturbations.

Breast Cancer Cells Reprogram the Oncogenic lncRNAs/mRNAs Coexpression Networks in Three-Dimensional Microenvironment.

Organotypic three-dimensional (3D) cell cultures more accurately mimic the characteristics of solid tumors in vivo in comparison with traditional two-dimensional (2D) monolayer cell models. Currently, studies on the regulation of long non-coding RNAs (lncRNAs) have not been explored in breast cancer cells cultured in 3D microenvironments. In the present research, we studied the expression and potential roles of lncRNAs in estrogen receptor-positive luminal B subtype BT-474 breast cancer cells grown over extracellular matrix proteins-enriched 3D cultures. Global expression profiling using DNA microarrays identifies 290 upregulated and 183 downregulated lncRNAs in 3D cultures relative to 2D condition. Using a co-expression analysis approach of lncRNAs and mRNAs pairs expressed in the same experimental conditions, we identify hundreds of regulatory axes modulating genes involved in cancer hallmarks, such as responses to estrogens, cell proliferation, hypoxia, apical junctions, and resistance to endocrine therapy. In addition, we identified 102 lncRNAs/mRNA correlations in 3D cultures, which were similar to those reported in TCGA datasets obtained from luminal B breast cancer patients. Interestingly, we also found a set of mRNAs transcripts co-expressed with LINC00847 and CTD-2566J3.1 lncRNAs, which were predictors of pathologic complete response and overall survival. Finally, both LINC00847 and CTD -2566J3.1 were co-expressed with essential genes for cancer genetic dependencies, such as FOXA1 y GINS2. Our experimental and predictive findings show that co-expressed lncRNAs/mRNAs pairs exhibit a high degree of similarity with those found in luminal B breast cancer patients, suggesting that they could be adequate pre-clinical tools to identify not only biomarkers related to endocrine therapy response and PCR, but to understand the biological behavior of cancer cells in 3D microenvironments.

Reprogramming of three-dimensional microenvironments for in vitro hair follicle induction.

During embryonic development, reciprocal interactions between epidermal and mesenchymal layers trigger hair follicle morphogenesis. This study revealed that microenvironmental reprogramming via control over these interactions enabled hair follicle induction in vitro. A key approach is to modulate spatial distributions of epithelial and mesenchymal cells in their spontaneous organization. The de novo hair follicles with typical morphological features emerged in aggregates of the two cell types, termed hair follicloids, and hair shafts sprouted with near 100% efficiency in vitro. The hair shaft length reached ~3 mm in culture. Typical trichogenic signaling pathways were up-regulated in hair follicloids. Owing to replication of hair follicle morphogenesis in vitro, melanosome production and transportation were also monitored in the hair bulb region. This in vitro hair follicle model might be valuable for better understanding hair follicle induction, evaluating hair growth and inhibition of hair growth by drugs, and modeling gray hairs in a well-defined environment.

Bioinspired sandwich-like hybrid surface functionalized scaffold capable of regulating osteogenesis, angiogenesis, and osteoclastogenesis for robust bone regeneration.

Recently, strategies that focus on biofunctionalized implant surfaces to enhance bone defect healing through the synergistic regulation of osteogenesis, angiogenesis, and osteoclastogenesis have attracted increasing attention in the bone tissue engineering field. Studies have shown that the Wnt/β-catenin signaling pathway has an imperative effect of promoting osteogenesis and angiogenesis while reducing osteoclastogenesis. However, how to prepare biofunctionalized bone implants with balanced osteogenesis, angiogenesis, and osteoclastogenesis by activating the Wnt/β-catenin pathway has seldom been investigated. Herein, through a bioinspired dopamine chemistry and self-assembly method, BML-284 (BML), a potent and highly selective Wnt signaling activator, was loaded on a mussel-inspired polydopamine (PDA) layer that had been immobilized on the porous beta-tricalcium calcium phosphate (β-TCP) scaffold surface and subsequently modified by a biocompatible carboxymethyl chitosan hydrogel to form a sandwich-like hybrid surface. β-TCP provides a biomimetic three-dimensional porous microenvironment similar to that of natural cancellous bone, and the BML-loaded sandwich-like hybrid surface endows the scaffold with multifunctional properties for potential application in bone regeneration. The results show that the sustained release of BML from the sandwich-like hybrid surface significantly facilitates the adhesion, migration, proliferation, spreading, and osteogenic differentiation of MC3T3-E1 cells as well as the angiogenic activity of human umbilical vein endothelial cells. In addition to osteogenesis and angiogenesis, the hybrid surface also exerts critical roles in suppressing osteoclastic activity. Remarkably, in a critical-sized cranial defect model, the biofunctionalized β-TCP scaffold could potentially trigger a chain of biological events: stimulating the polarization of M2 macrophages, recruiting endogenous stem cells and endothelial cells at the injury site to enable a favorable microenvironment for greatly accelerating bone ingrowth and angiogenesis while compromising osteoclastogenesis, thereby promoting bone healing. Therefore, these surface-biofunctionalized β-TCP implants, which regulate the synergies of osteogenesis, angiogenesis, and anti-osteoclastogenesis, indicate strong potential for clinical application as advanced orthopedic implants.

Perfusable Vessel-on-a-Chip for Antiangiogenic Drug Screening with Coaxial Bioprinting.

Vessel-on-a-chips, which can be used to study microscale fluid dynamics, tissue-level biological molecules delivery and intercellular communication under favorable three-dimensional (3D) extracellular matrix microenvironment, are increasingly gaining traction. However, not many of them can allow for long-term perfusion and easy observation of angiogenesis process. Since angiogenesis is necessary for the expansion of tumor, antiangiogenic drugs play a significant role in cancer treatment. In this study, we established an innovative and reliable antiangiogenic drug screening chip that was highly modularly integrated for long-term perfusion (up to 10 days depending on the hydrogel formula) and real-time monitoring. To maintain an unobstructed flow of cell-laden tubes for subsequent perfusion culture on the premise of excellent bioactivities, a polycaprolactone stent inspired by coronary artery stents was introduced to hold up the tubular lumen from the inside, while the perfusion chip was also elaborately designed to allow for convenient observation. After 3 days of perfusion screening, distinct differences in human umbilical vein endothelial cell sprouting were observed for a gradient of concentrations of bevacizumab, which pointed to the effectiveness and reliability of the drug screening perfusion system. Overall, a perfusion system for antiangiogenic drug screening was developed, which can not only conduct drug evaluation, but also be potentially useful in other vessel-mimicking scenarios in the area of tissue engineering, drug screening, pharmacokinetics, and regenerative medicine.

Enhanced Proliferation of Human Mesenchymal Stem Cells by Self-Assembled Peptide Hydrogel Modified with Heparin Mimetic Peptide

The development of three-dimensional (3D) microenvironments that mimic the role of native extracellular matrix (ECM) components is very crucial. Self-assembling peptide (SAP) hydrogels have been utilized as scaffolds for encapsulation, 3D culture, proliferation, and differentiation of cells and also for repairing defects in cartilage tissue. SAPs can be biofunctionalized with short peptide epitopes to form biomimetic scaffolds. Herein, KLD (KLDLKLDLKLDL) SAP was functionalized with a short bioactive motif, EGDK, to fabricate KLD-HM (KLDLKLDLKLDL-EGDK) SAP hydrogel and self-assembled. In this study, it was aimed to investigate the effect of developed KLD-HM SAP hydrogels on the viability and proliferation of human mesenchymal stem cells (hMSCs). For ensuring the stability of SAPs, the rheological properties and degradation profile of produced SAP hydrogels were assessed. After the encapsulation of hMSCs in SAP hydrogels, MTT assay and Live and Dead staining assay were conducted. We showed that these biomimetic peptide hydrogel scaffolds provided a proper microenvironment for encapsulated hMSCs and the developed SAP hydrogels promoted the adhesion, viability, and proliferation of hMSCs. Our results suggest that designed bioactive SAP hydrogel scaffolds might be useful for promoting the regeneration of cartilage tissue.

Engineering a dynamic three-dimensional cell culturing microenvironment using a ‘sandwich’ structure-liked microfluidic device with 3D printing scaffold

Most of in vivo tissue cells reside in 3D extracellular matrix (ECM) with fluid flow. To better study cell physiology and pathophysiology, there has been an increasing need in the development of methods for culturing cells in in vivo like microenvironments with a number of strategies currently being investigated including hydrogels, spheroids, tissue scaffolds and very promising microfluidic systems. In this paper, a ‘sandwich’ structure-liked microfluidic device integrated with a 3D printing scaffold is proposed for three-dimensional and dynamic cell culture. The device consists of three layers, i.e. upper layer, scaffold layer and bottom layer. The upper layer is used for introducing cells and fixing scaffold, the scaffold layer mimicking ECM is used for providing 3D attachment areas, and the bottom layer mimicking blood vessels is used for supplying dynamic medium for cells. Thermally assisted electrohydrodynamic jet (TAEJ) printing technology and microfabrication technology are combined to fabricate the device. The flow field in the chamber of device is evaluated by numerical simulation and particle tracking technology to investigate the effects of scaffold on fluid microenvironment. The cell culturing processes are presented by the flow behaviors of inks with different colors. The densities and viabilities of HeLa cells are evaluated and compared after 72 h of culturing in the microfluidic devices and 48-well plate. The dose-dependent cell responses to doxorubicin hydrochloride (DOX) are observed after 24 h treatment at different concentrations. These experimental results, including the evaluation of cell proliferation and in vitro cytotoxicity assessment of DOX in the devices and plate, demonstrate that the presented microfluidic device has good biocompatibility and feasibility, which have great potential in providing native microenvironments for in vitro cell studies, tissue engineering and drug screening for tumor therapy.

Strategies for Generating Human Pluripotent Stem Cell-Derived-Organoid Culture for Disease Modeling, Drug Screening, and Regenerative Therapy

Human pluripotent stem cells (hPSCs) have become a powerful tool to generate the various kinds of cell types comprising the human body. Recently, organoid technology has emerged as a platform to generate a physiologically relevant tissue-like structure from PSCs. Compared to an actual human organ, this structure more closely represents a three-dimensional microenvironment than the conventional monolayer culture system for transplantation, disease modeling, and drug development. Despite its advantages, however, the organoid culture system still has various problems related to culture methods, which have become a challenge for attempts to obtain similar physiological properties to their original tissue counterparts. Here, we discuss the current development of organoid culture methods, including the problems that may arise from the currently available culture systems, as well as a possible approach for overcoming their current limitations and improving their optimum utilization for translational application purposes.

Biomimetic Hydrogels in the Study of Cancer Mechanobiology: Overview, Biomedical Applications, and Future Perspectives

Hydrogels are biocompatible polymers that are tunable to the system under study, allowing them to be widely used in medicine, bioprinting, tissue engineering, and biomechanics. Hydrogels are used to mimic the three-dimensional microenvironment of tissues, which is essential to understanding cell–cell interactions and intracellular signaling pathways (e.g., proliferation, apoptosis, growth, and survival). Emerging evidence suggests that the malignant properties of cancer cells depend on mechanical cues that arise from changes in their microenvironment. These mechanobiological cues include stiffness, shear stress, and pressure, and have an impact on cancer proliferation and invasion. The hydrogels can be tuned to simulate these mechanobiological tissue properties. Although interest in and research on the biomedical applications of hydrogels has increased in the past 25 years, there is still much to learn about the development of biomimetic hydrogels and their potential applications in biomedical and clinical settings. This review highlights the application of hydrogels in developing pre-clinical cancer models and their potential for translation to human disease with a focus on reviewing the utility of such models in studying glioblastoma progression.

Exploring the function of stromal cells in cholangiocarcinoma by three-dimensional bioprinting immune microenvironment model.

The tumor immune microenvironment significantly affects tumor progression, metastasis, and clinical therapy. Its basic cell components include tumor-associated endothelial cells, fibroblasts, and macrophages, all of which constitute the tumor stroma and microvascular network. However, the functions of tumor stromal cells have not yet been fully elucidated. The three-dimensional (3D) model created by 3D bioprinting is an efficient way to illustrate cellular interactions in vitro. However, 3D bioprinted model has not been used to explore the effects of stromal cells on cholangiocarcinoma cells. In this study, we fabricated 3D bioprinted models with tumor cells and stromal cells. Compared with cells cultured in two-dimensional (2D) environment, cells in 3D bioprinted models exhibited better proliferation, higher expression of tumor-related genes, and drug resistance. The existence of stromal cells promoted tumor cell activity in 3D models. Our study shows that 3D bioprinting of an immune microenvironment is an effective way to study the effects of stromal cells on cholangiocarcinoma cells.

Influence of circadian rhythm on drug metabolism in 3D hepatic spheroids.

Circadian rhythms are characterized as oscillations that fluctuate based on a 24 h cycle and are responsible for regulation of physiological functions. While the internal clock synchronizes gene expression using external cues like light, a similar synchronization can be induced in vitro by incubating the cells with an increased percentage of serum followed by its rapid removal. Previous studies have suggested that synchronization of HepG2 cell line induced the rhythmic expression of drug-metabolizing enzymes (DME) most specifically the cytochrome P450 enzymes. However, there is a lack of evidence demonstrating the influence of three-dimensional microenvironment on the rhythmicity of these genes. To understand this interplay, gene expression of the circadian machinery and CYP450s were compared using the model human hepatocarcinoma cell line, HepG2. Upon serum shock synchronization, gene and protein expression of core clock regulators was assessed and rhythmic expression of these genes was demonstrated. Further insight into the interrelations between various gene pairs was obtained using statistical analysis. Using RNA sequencing, an in-depth understanding of the widespread effects of circadian regulation on genes involved in metabolic processes in the liver was obtained. This study aids in the better understanding of chronopharmacokinetic events in humans using physiologically relevant 3D culture systems.