VMT/ACP/Dextran composite nanosheets against dental caries through promoting mineralization of dentin tubules, pH buffering, and antibacterial.

Dental caries is a worldwide public healthcare concern, and is closely related to the acidic environment that caused by bacterial decomposition of food. In this study, a two-step ion exchange liquid-phase stripping method was applied to strip out vermiculite (VMT) nanosheets, then amorphous calcium phosphate (ACP) and dextran were inserted between the VMT nanosheets interlayer to obtain a composite two-dimension nanosheets (VMT/ACP/Dextran). VMT/ACP/Dextran composite nanosheets exhibited excellent biocompatibility and could provide exogenous Ca2+and PO43- from ACP, provide SiO44-, Mg2+, Fe2+ and obtain buffering pH and antibacterial properties from VMT, as well as improve suspension stability and targeting Streptococcus mutans through glucan. The in vitro study showed that the composite materials could promote the mineralization and sealing of dentin tubules by releasing active ions, buffer pH 4.5 (a value close to the pH in the dental plaque environment) to pH 6.6-7.1 (values close to the pH in human saliva) through ion exchange, and exert antibacterial effects by targeting Streptococcus mutans and exerting oxidase like and peroxidase like activities to produce reactive oxygen species (ROS). The in vivo animal study showed that daily cleaning teeth using VMT/ACP/Dextran composite nanosheets could effectively reduce the incidence rate and severity of dental caries in rats. Taking together, the developed VMT/ACP/Dextran composite nanosheets, which integrated the excellent properties of VMT, ACP and dextran, can effectively prevent dental caries through a combination of factors such as buffering acids, antibacterial properties, and promoting calcification, and may be used as an active ingredient for daily oral hygiene or filling materials to prevent and treat dental caries.

Support Enzyme Loading Influences the Effect of Aldehyde Dextran Modification on the Specificity of Immobilized Ficin for Large Proteins.

It has been reported that the modification of immobilized glyoxyl-ficin with aldehyde dextran can promote steric hindrances that greatly reduce the activity of the immobilized protease against hemoglobin, while the protease still maintained a reasonable level of activity against casein. In this paper, we studied if this effect may be different depending on the amount of ficin loaded on the support. For this purpose, both the moderately loaded and the overloaded glyoxyl-ficin biocatalysts were prepared and modified with aldehyde dextran. While the moderately loaded biocatalyst had a significantly reduced activity, mainly against hemoglobin, the activity of the overloaded biocatalyst was almost maintained. This suggests that aldehyde dextran was able to modify areas of the moderately loaded enzyme that were not available when the enzyme was overloaded. This modification promoted a significant increase in biocatalyst stability for both biocatalysts, but the stability was higher for the overloaded biocatalyst (perhaps due to a combination of inter- and intramolecular crosslinking).

Thermo-responsive aqueous two-phase system for two-level compartmentalization.

Hierarchical compartmentalization responding to changes in intracellular and extracellular environments is ubiquitous in living eukaryotic cells but remains a formidable task in synthetic systems. Here we report a two-level compartmentalization approach based on a thermo-responsive aqueous two-phase system (TR-ATPS) comprising poly(N-isopropylacrylamide) (PNIPAM) and dextran (DEX). Liquid membraneless compartments enriched in PNIPAM are phase-separated from the continuous DEX solution via liquid-liquid phase separation at 25 °C and shrink dramatically with small second-level compartments generated at the interface, resembling the structure of colloidosome, by increasing the temperature to 35 °C. The TR-ATPS can store biomolecules, program the spatial distribution of enzymes, and accelerate the overall biochemical reaction efficiency by nearly 7-fold. The TR-ATPS inspires on-demand, stimulus-triggered spatiotemporal enrichment of biomolecules via two-level compartmentalization, creating opportunities in synthetic biology and biochemical engineering.

Marangoni Droplets of Dextran in PEG Solution and Its Motile Change Due to Coil-Globule Transition of Coexisting DNA.

Motile droplets using Marangoni convection are attracting attention for their potential as cell-mimicking small robots. However, the motion of droplets relative to the internal and external environments that generate Marangoni convection has not been quantitatively described. In this study, we used an aqueous two-phase system [poly(ethylene glycol) (PEG) and dextran] in an elongated chamber to generate motile dextran droplets in a constant PEG concentration gradient. We demonstrated that dextran droplets move by Marangoni convection, resulting from the PEG concentration gradient and the active transport of PEG and dextran into and out of the motile dextran droplet. Furthermore, by spontaneously incorporating long DNA into the dextran droplets, we achieved cell-like motility changes controlled by coexisting environment-sensing molecules. The DNA changes its position within the droplet and motile speed in response to external conditions. In the presence of Mg2+, the coil-globule transition of DNA inside the droplet accelerates the motile speed due to the decrease in the droplet's dynamic viscosity. Globule DNA condenses at the rear part of the droplet along the convection, while coil DNA moves away from the droplet's central axis, separating the dipole convections. These results provide a blueprint for designing autonomous small robots using phase-separated droplets, which change the mobility and molecular distribution within the droplet in reaction with the environment. It will also open unexplored areas of self-assembly mechanisms through phase separation under convections, such as intracellular phase separation.

Ultrahigh-Resolution Visualization of Vascular Heterogeneity in Brain Tumors via Magnetic Nanoparticles-Enhanced Susceptibility-Weighted Imaging.

The precise assessment of vascular heterogeneity in brain tumors is vital for diagnosing, grading, predicting progression, and guiding treatment decisions. However, currently, there is a significant shortage of high-resolution imaging approaches. Herein, we propose a contrast-enhanced susceptibility-weighted imaging (CE-SWI) utilizing the minimalist dextran-modified Fe3O4 nanoparticles (Dextran@Fe3O4 NPs) for ultrahigh-resolution mapping of vasculature in brain tumors. The Dextran@Fe3O4 NPs are prepared via a facile coprecipitation method under room temperature, and exhibit small hydrodynamic size (28 nm), good solubility, excellent biocompatibility, and high transverse relaxivity (r2*, 159.7 mM-1 s-1) under 9.4 T magnetic field. The Dextran@Fe3O4 NPs-enhanced SWI can increase the contrast-to-noise ratio (CNR) of cerebral vessels to 2.5 times that before injection and achieves ultrahigh-spatial-resolution visualization of microvessels as small as 0.1 mm in diameter. This advanced imaging capability not only allows for the detailed mapping of both enlarged peritumoral drainage vessels and the intratumoral microvessels, but also facilitates the sensitive imaging detection of vascular permeability deterioration in a C6 cells-bearing rat glioblastoma model. Our proposed Dextran@Fe3O4 NPs-enhanced SWI provides a powerful imaging technique with great clinical translation potential for the precise theranostics of brain tumors.

Slowly hydrolyzable property of microbial dextrans at the small intestinal α-glucosidase levels leads to the modulated glycemic responses in the mouse model.

Dextran-type α-glucans have been known as non-digestible ingredients that can be considered prebiotics to promote colon health. However, recent studies have revealed that various α-linked glucosyl units are hydrolyzed to glucose by small intestinal α-glucosidases. This study analyzed the structural characteristics of exopolysaccharides (EPSs) from Weissella species, and the hydrolysis properties at both in vitro/in vivo levels were investigated. Compared with a previous in vitro digestion model using fungal α-hydrolytic enzymes, dextrans, which mainly consist of α-1,6 linkages with small amounts of α-1,3 linked glucose units, were slowly hydrolyzed to glucose by mammalian mucosal α-glucosidases, resulting in attenuation of the initial glycemic response following administration of EPS samples to mice via oral gavage. The results of this study demonstrate the concept of dextran-type α-glucans as glycemic carbohydrates rather than dietary fibers or prebiotics. Slowly digestible dextrans can be applied as a functional ingredient to regulate postprandial glucose delivery throughout the gastrointestinal tract.

Facile roll-to-roll production of nanoporous fiber coatings for advanced wound care sutures.

Theranostic sutures are derived from innovative ideas to enhance wound healing results by adding wound diagnostics and therapeutics to typical sutures by functionalizing them with additional materials. Here, we present a new direct electrospinning method for the fast, continuous, inexpensive, and high-throughput production of versatile nanofibrous-coated suture threads, with precise control over various essential microstructural and physical characteristics. The thickness of the coating layer and the alignment of nanofibers with the thread's direction can be adjusted by the user by varying the spooling speed and the displacement between the spinneret needle and thread. To show the flexibility of our method for a range of different materials selected, gelatin, polycaprolactone, silk fibroin, and PEDOT:PSS (poly(3,4-ethylene dioxythiophene):poly(styrene sulfonate)) were the resultant nanofibers characterized by scanning electron microscopy (SEM) imaging and conductivity tests. In a series of in vitro and ex vivo tests (pig skin), sutures were successfully tested for their flexibility and mechanical properties when used as weaving and knotting sutures, and their biocompatibility with a keratinocyte cell line. For temperature-based drug-releasing tests, two fluorescent molecules as drug models with high and low molecular weight, namely fluorescein isothiocyanate-dextran (20 kDa) and rhodamine B (470 Da), were used, and their steady release with incremental increase of temperature to 37 °C over 120 min was seen, which is appropriate for bacterial treatment drugs. Given the advantages of the presented technique, it seems to have promising potential to be used in future medical applications for wound closure and bacterial infection treatment via a temperature-triggered drug release strategy.

Effects of enzymatic hydrolysis combined with glycation on the emulsification characteristics and emulsion stability of peanut protein isolate.

Peanut protein isolate (PPI) has high nutritional value, but its poor function limits its application in the food industry. In this study, peanut protein isolate was modified by enzymatic hydrolysis combined with glycation. The structure, emulsification and interface properties of peanut protein isolate hydrolysate (HPPI) and dextran (Dex) conjugate (HPPI-Dex) were studied. In addition, the physicochemical properties, rheological properties, and stability of the emulsion were also investigated. The results showed that the graft degree increased with the increase of Dex ratio. Fourier transform infrared spectroscopy (FTIR) confirmed that the glycation of HPPI and Dex occurred. The microstructure showed that the structure of HPPI-Dex was expanded, and the molecular flexibility was enhanced. When the ratio of HPPI to Dex was 1:3, the emulsifying activity and the interface pressure of glycated HPPI reached the highest value, and the emulsifying activity (61.08 m2/g) of HPPI-Dex was 5.28 times that of PPI. The HPPI-Dex stabilized emulsions had good physicochemical properties and rheological properties. In addition, HPPI-Dex stabilized emulsions had high stability under heat treatment, salt ion treatment and freeze-thaw cycle. According to confocal laser scanning microscopy (CLSM), the dispersion of HPPI-Dex stabilized emulsions was better after 28 days of storage. This study provides a theoretical basis for developing peanut protein emulsifier and further expanding the application of peanut protein in food industry.

A biguanide chitosan-based hydrogel adhesive accelerates the healing of bacterial-infected wounds.

The development of tissue adhesives with good biocompatibility and potent antimicrobial properties is crucial for addressing the high incidence of surgical site infections in emergency and clinical settings. Herein, an injectable hydrogel adhesive composed of chitosan biguanidine (CSG), oxidized dextran (ODex) and tannin (TA) was synthesized primarily through Schiff-base reactions, hydrogen bonding, and electrostatic interactions. TA was introduced into the CSG/ODex hydrogel to prepare a physicochemically double cross-linked hydrogel. The hydrogel formulation incorporating 2 wt% TA (CSG/ODex-TA2) exhibited rapid gelation, moderate mechanical properties, good tissue adhesion, and sustained release behavior of TA. Both in vitro and in vivo studies demonstrated that CSG/ODex-TA2 showed significantly enhanced adhesion and antibacterial effectiveness compared to the CSG/ODex hydrogel and commercial fibrin glue. Leveraging the positive charge of CSG, the CSG/ODex-TA2 hydrogel demonstrated a strong contact antibacterial effect, while the sustained release of TA provided diffusion antibacterial capabilities. By integrating contact and diffusion antibacterial mechanisms into the hydrogel, a promising approach was developed to boost antibacterial efficiency and accelerate the healing of wounds infected with methicillin-resistant Staphylococcus aureus (MRSA). The CSG/ODex-TA2 hydrogel has excellent biocompatibility, hemostatic properties, and antibacterial capabilities, making it a promising candidate for improving in vivo wound care and combating bacterial infections.

A pilot study on endoscopic delivery of injectable bioadhesive for esophageal repair in a porcine model.

Endoscopic submucosal dissection (ESD) is the gold-standard surgical procedure for superficial esophageal cancer. A significant and challenging complication of this technique is post-ESD esophageal stricture. In this study, the feasibility of endoscopic catheter delivery of bioadhesive to esophageal lesions in a porcine model was tested. Injectable bioadhesive was composed of oxidized dextran (ODA) and chitosan hydrochloride (CS), its physicochemical properties, injectability, antibacterial activity, and cytocompatibility were investigated beforein vivotest. ODA-CS bioadhesive was delivered to the wound bed of the esophageal tissue using a custom-made catheter device after ESD in a porcine model. Our results show that the ODA-CS bioadhesive is of good injectability, tissue adhesive strength, antibacterial capacity, and blood compatibility.In vivodelivery was achieved by endoscopic spraying of ODA and CS in separate catheters fixed on the endoscopic probe. ODA and CS can be mixed well to allow in situ bioadhesive formation and firmly adhere to the esophageal wound surface. After two weeks, the bioadhesive maintained structural integrity and adhered to the surface of esophageal wounds. However, histological analysis reveals that the ODA-CS bioadhesive did not show improvement in attenuating inflammatory response after ESD. This pilot study demonstrates the feasibility of ODA-CS bioadhesive for shielding esophageal wounds after ESD, whereas efforts need to improve its anti-inflammatory activity to reduce fibrosis for stricture prevention.

Multimodal evaluation of blood-brain barrier opening in mice in response to low-intensity ultrasound and a claudin-5 binder.

Background: The blood-brain barrier (BBB) is a major bottleneck in delivering therapeutics to the brain. Treatment strategies to transiently open this barrier include focused ultrasound combined with intravenously injected microbubbles (FUS+MB) and targeting of molecules that regulate BBB permeability. Methods: Here, we investigated BBB opening mediated by the claudin-5 binder cCPEm (a microorganismal toxin in a truncated form) and FUS+MB at a centre frequency of 1 MHz, assessing dextran uptake, broadband emission, and endogenous immunoglobulin G (IgG) extravasation. Results: FUS+MB-induced BBB opening was detectable at a pressure ≥0.35 MPa when assessed for leakage of 10 and 70 kDa dextran, and at ≥0.2 MPa for uptake of endogenous IgG. Treating mice with 20 mg/kg cCPEm failed to open the BBB, and pre-treatment with cCPEm followed by FUS+MB at 0.2 and 0.3 MPa did not overtly increase BBB opening compared to FUS+MB alone. Using passive cavitation detection (PCD), we found that broadband emission correlated with the peak negative pressure (PNP) and dextran leakage, indicating the possibility of using broadband emission for developing a feedback controller to monitor BBB opening. Conclusions: Together, our study highlights the challenges in developing combinatorial approaches to open the BBB and presents an additional IgG-based histological detection method for BBB opening.

Cofunctionalization of Macroporous Dextran Hydrogels with Adhesive Peptides and Growth Factors Enables Vascular Spheroid Sprouting.

Ensuring good definition of scaffolds used for 3D cell culture is a prominent challenge that hampers the development of tissue engineering platforms. Since dextran repels cell adhesion, using dextran-based materials biofunctionalized through a bottom-up approach allows for precise control over material definition. Here, we report the design of dextran hydrogels displaying a fully interconnected macropore network for the culture of vascular spheroids in vitro. We biofunctionalized the hydrogels with the RGD peptide sequence to promote cell adhesion. We used an affinity peptide pair, the E/K coiled coil, to load the gels with epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF). Dual functionalization with adhesive and proliferative cues allows vascular spheroids to colonize naturally cell-repellant dextran. In supplement-depleted medium, we report improved colonization of the macropores compared to that of unmodified dextran. Altogether, we propose a well-defined and highly versatile platform for tissue engineering and tissue vascularization applications.

The Discovery, Molecular Cloning, and Characterization of Dextransucrase LmDexA and Its Active Truncated Mutant from Leuconostoc mesenteroides NN710.

Dextransucrases play a crucial role in the production of dextran from economical sucrose; therefore, there is a pressing demand to explore novel dextransucrases with better performance. This study characterized a dextransucrase enzyme, LmDexA, which was identified from the Leuconostoc mesenteroides NN710. This bacterium was isolated from the soil of growing dragon fruit in Guangxi province, China. We successfully constructed six different N-terminal truncated variants through sequential analysis. Additionally, a truncated variant, ΔN190LmDexA, was constructed by removing the 190 amino acids fragment from the N-terminal. This truncated variant was then successfully expressed heterologously in Escherichia coli and purified. The purified ΔN190LmDexA demonstrated optimal hydrolysis activity at a pH of 5.6 and a temperature of 30 °C. Its maximum specific activity was measured to be 126.13 U/mg, with a Km of 13.7 mM. Results demonstrated a significant improvement in the heterologous expression level and total enzyme activity of ΔN190LmDexA. ΔN190LmDexA exhibited both hydrolytic and transsaccharolytic enzymatic activities. When sucrose was used as the substrate, it primarily produced high-molecular-weight dextran (>400 kDa). However, upon the addition of maltose as a receptor, it resulted in the production of a significant amount of oligosaccharides. Our results can provide valuable information for enhancing the characteristics of recombinant dextransucrase and potentially converting sucrose into high-value-added dextran and oligosaccharides.

Towards Self-regeneration: Exploring the Limits of Protein Synthesis in the Protein Synthesis Using Recombinant Elements (PURE) Cell-free Transcription-Translation System.

Self-regeneration is a key function of living systems that needs to be recapitulated in vitro to create a living synthetic cell. A major limiting factor for protein self-regeneration in the PURE cell-free transcription-translation system is its high protein concentration, which far exceeds the system's protein synthesis rate. Here, we were able to drastically reduce the nonribosomal PURE protein concentration up to 97.3% while increasing protein synthesis efficiency. Although crowding agents were not effective in the original PURE formulation, we found that in highly dilute PURE formulations, addition of 6% dextran considerably increased protein synthesis rate and total protein yield. These new PURE formulations will be useful for many cell-free synthetic biology applications, and we estimate that PURE can now support the complete self-regeneration of all 36 nonribosomal proteins, which is a critical step toward the development of a universal biochemical constructor and living synthetic cell.

A Tween-80 modified hypoxia/esterase dual stimulus-activated nanomicelle as a delivery platform for carmustine - Design, synthesis, and biological evaluation.

As a clinical anti-glioma agent, the therapeutic effect of carmustine (BCNU) was largely decreased because of the drug resistance mediated by O6-alkylguanine-DNA alkyltransferase (AGT) and the blood-brain barrier (BBB). To overcome these obstacles, we synthesized a BCNU-loaded hypoxia/esterase dual stimulus-activated nanomicelle, abbreviated as T80-HACB/BCNU NPs. In this nano-system, Tween 80 acts as the functional coating on the surface of the micelle to facilitate transport across the BBB. Hyaluronic acid (HA) with active tumor-targeting capability was linked with the hypoxia-sensitive AGT inhibitors (O6-azobenzyloxycarbonyl group) via an esterase-activated ester bond. The obtained T80-HACB/BCNU NPs had an average particle size of 232.10 ± 10.66 nm, the zeta potential of -18.13 ± 0.91 mV, and it showed high drug loading capacity, eximious biocompatibility and dual activation of hypoxia/esterase drug release behavior. The obtained T80-HACB/BCNU NPs showed enhanced cytotoxicity against hypoxic T98G and SF763 cells with IC50 at 132.2 µM and 133.1 µM, respectively. T80 modification improved the transportation of the micelle across an in vitro BBB model. The transport rate of the T80-HACB/Cou6 NPs group was 12.37 %, which was 7.6-fold (p<0.001) higher than the micelle without T80 modification. T80-HACB/BCNU NPs will contribute to the development of novel CENUs chemotherapies with high efficacy.

Dextran-based nanoparticles with 5-FU-conjugated polymethacrylate segments for drug delivery: Synthesis of amphiphilic graft copolymers by mechanochemical solid-state polymerization and characterization.

Dextran (Dx) is a biodegradable and biocompatible polysaccharide, thus promising as a drug delivery carrier for tumor therapy. Herein, we applied mechanical energy to a high molecular weight Dx to control its molecular weight and simultaneously generate mechanoradicals. The solid-state polymerization of methacrylate- or methacrylamide derivatives initiated with Dx mechanoradicals showed polymer conversion of >95%, yielding Dx-based graft copolymers with molecular weights of approximately 30,000 g mol-1. The Dx-based graft copolymers with hydrophobic segments formed nanoparticles with a particle size of 25-35 nm in an aqueous solution. The anti-pancreatic tumor drug 5-fluorouracil (5-FU) was covalently conjugated onto the hydrophobic segments of the amphiphilic Dx, and the nanoparticles were also prepared. The drug release profile from 5-FU-conjugated nanoparticles corresponded well to the Korsmeyer-Peppas model applied to drug release from matrix substrates, and was also immensely predicted by the Logistic and Gompertz curves. The 5-FU-conjugated nanoparticles showed cytotoxicity against the pancreatic adenocarcinoma cell lines (BxPC-3) that were not significantly inferior to the 5-FU positive group. Furthermore, the fluorescein-labeled nanoparticles internalized into BxPC-3 within 6 h and actively migrated into the cytosol. These results suggest that Dx-based graft copolymers with hydrophobic segments might be used to enhance therapeutic activity.

Copper peroxide loaded gelatin/oxide dextran hydrogel with temperature and pH responsiveness for antibacterial and wound healing activity.

Bacterial infection and tissue hypoxia always prevent wound healing, so multifunctional platforms with antimicrobial and oxygen-supplying functions were developed. However, they face many difficulties such as complex preparation and low oxygen release. To address this challenge, a copper peroxide loaded gelatin/oxide dextran hydrogel (CGO) was prepared. Surprisingly, CGO hydrogel as a wound dressing not only had good biocompatibility, injectivity, and mechanical properties, but also exhibited mild photothermal properties, temperature responsiveness, and pH responsiveness. After being applied to wounds infected with bacteria, CGO hydrogel released copper peroxide under near-infrared laser irradiation, which produced copper ions and hydrogen peroxide, combined with PTT to kill bacteria. After the bacteria were cleared from the wound and the pH of the wound was changed to be acidic, CGO hydrogel released copper peroxide via pH response. Copper ions and oxygen produced from copper peroxide accelerated wound healing by promoting angiogenesis. The multi-responsive and multi-mode treatment platform provided a potential strategy for treating bacteria-infected wounds.

Membrane nanotubes transform into double-membrane sheets at condensate droplets.

Cellular membranes exhibit a multitude of highly curved morphologies such as buds, nanotubes, cisterna-like sheets defining the outlines of organelles. Here, we mimic cell compartmentation using an aqueous two-phase system of dextran and poly(ethylene glycol) encapsulated in giant vesicles. Upon osmotic deflation, the vesicle membrane forms nanotubes, which undergo surprising morphological transformations at the liquid-liquid interfaces inside the vesicles. At these interfaces, the nanotubes transform into cisterna-like double-membrane sheets (DMS) connected to the mother vesicle via short membrane necks. Using super-resolution (stimulated emission depletion) microscopy and theoretical considerations, we construct a morphology diagram predicting the tube-to-sheet transformation, which is driven by a decrease in the free energy. Nanotube knots can prohibit the tube-to-sheet transformation by blocking water influx into the tubes. Because both nanotubes and DMSs are frequently formed by cellular membranes, understanding the formation and transformation between these membrane morphologies provides insight into the origin and evolution of cellular organelles.

Synthetic biodegradable microporous hydrogels for in vitro 3D culture of functional human bone cell networks.

Generating 3D bone cell networks in vitro that mimic the dynamic process during early bone formation remains challenging. Here, we report a synthetic biodegradable microporous hydrogel for efficient formation of 3D networks from human primary cells, analysis of cell-secreted extracellular matrix (ECM) and microfluidic integration. Using polymerization-induced phase separation, we demonstrate dynamic in situ formation of microporosity (5-20 µm) within matrix metalloproteinase-degradable polyethylene glycol hydrogels in the presence of living cells. Pore formation is triggered by thiol-Michael-addition crosslinking of a viscous precursor solution supplemented with hyaluronic acid and dextran. The resulting microporous architecture can be fine-tuned by adjusting the concentration and molecular weight of dextran. After encapsulation in microporous hydrogels, human mesenchymal stromal cells and osteoblasts spread rapidly and form 3D networks within 24 hours. We demonstrate that matrix degradability controls cell-matrix remodeling, osteogenic differentiation, and deposition of ECM proteins such as collagen. Finally, we report microfluidic integration and proof-of-concept osteogenic differentiation of 3D cell networks under perfusion on chip. Altogether, this work introduces a synthetic microporous hydrogel to efficiently differentiate 3D human bone cell networks, facilitating future in vitro studies on early bone development.

A 3D-printed tumor-on-chip: user-friendly platform for the culture of breast cancer spheroids and the evaluation of anti-cancer drugs.

Tumor-on-chips (ToCs) are useful platforms for studying the physiology of tumors and evaluating the efficacy and toxicity of anti-cancer drugs. However, the design and fabrication of a ToC system is not a trivial venture. We introduce a user-friendly, flexible, 3D-printed microfluidic device that can be used to culture cancer cells or cancer-derived spheroids embedded in hydrogels under well-controlled environments. The system consists of two lateral flow compartments (left and right sides), each with two inlets and two outlets to deliver cell culture media as continuous liquid streams. The central compartment was designed to host a hydrogel in which cells and microtissues can be confined and cultured. We performed tracer experiments with colored inks and 40 kDa fluorescein isothiocyanate dextran to characterize the transport/mixing performances of the system. We also cultured homotypic (MCF7) and heterotypic (MCF7-BJ) spheroids embedded in gelatin methacryloyl hydrogels to illustrate the use of this microfluidic device in sustaining long-term micro-tissue culture experiments. We further demonstrated the use of this platform in anticancer drug testing by continuous perfusion of doxorubicin, a commonly used anti-cancer drug for breast cancer. In these experiments, we evaluated drug transport, viability, glucose consumption, cell death (apoptosis), and cytotoxicity. In summary, we introduce a robust and friendly ToC system capable of recapitulating relevant aspects of the tumor microenvironment for the study of cancer physiology, anti-cancer drug transport, efficacy, and safety. We anticipate that this flexible 3D-printed microfluidic device may facilitate cancer research and the development and screening of strategies for personalized medicine.