Hydrogel microparticles (microgels) have significant potential for use as building blocks in tissue engineering, as bioinks for 3D bioprinting, and as drug and cell carriers for cell-based therapies targeting damaged and diseased tissues. Various fabrication techniques have been developed for producing microgels with predefined shapes and sizes. However, for practical applications in biological laboratories and clinics, it is necessary to reduce time costs and simplify instrumentation and synthesis protocols, improving their reproducibility and reliability. Here we demonstrate a three-step experimental approach to develop microfluidic flow-focusing droplet generators that enable the introduction of all liquids by creating negative pressure in the outlet reservoir for the generation of spherical, core-shell, and Janus alginate microgels with living cells. This approach allows the use of a simple experimental setup that is easy to operate and robust and provides highly reproducible results, achieving a synthesis performance of up to 200 μL of microgels per hour. The size and the structure of the microgels were determined by the chip design and remained stable under pressure variations within the operating range of -7 to -15 kPa. This enabled the reliable and reproducible encapsulation of CT26 and HepG2 cells into core-shell and Janus alginate microgels with diameters ranging from 80 to 120 μm, maintaining over 80% cell viability during long-term incubation. Our findings offer a new perspective for the automation and scaling of multicomponent alginate microgel fabrication, paving the way for their implementation in tissue engineering and 3D bioprinting.

Self-healing and stimuli-responsive hydrogels are highly desirable in biomedicine. A promising strategy to achieve these properties is dynamic covalent cross-linking via boronic ester bonds between boronic acids and diols. However, most hydrogels using this mechanism are unstable under physiological conditions, which limits their biomedical applicability. Here, we present the synthesis and application of a star-shaped poly(2-methyloxazoline) functionalized with boronic acid derivatives, designed to form stable hydrogels in the presence of poly(vinyl alcohol) under physiological conditions. By tuning the overall polymer content, boronic acid/diol molar ratio, and molecular weight, hydrogel stiffness could be precisely adjusted. The resulting hydrogels exhibited rapid self-healing, high stretchability, injectability, and responsiveness to the pH value and glucose levels, as demonstrated by insulin release studies. Cytotoxicity and cell encapsulation experiments confirmed the cytocompatibility of the polymer and hydrogels.

Hydrogel microfibers provide a versatile platform for cell encapsulation and tissue engineering, but their fabrication typically involves harsh or clog-prone processes that limit cytocompatibility or scalability. Here, we report a clog-free, all-aqueous water-in-water-in-water microfluidic approach for continuous and cytocompatible fabrication of hydrogel microfibers. This approach uses immiscible aqueous polymer solutions to enable fiber formation and controlled crosslinking without organic solvents and rapid solidification, preventing channel clogging during continuous operation. Using alginate as a model biomaterial, we generate meter-long microfibers that can be spun into macroscopic films. Systematic experiments and laminar flow modeling reveal a universal scaling law, , showing that the fiber diameter Df depends solely on the collection capillary diameter Dc and the ratio of inner flow rate Qi to the total flow rate of the middle and outer phases Qsum. Finally, we demonstrate encapsulation of pancreatic β-cells, which retain glucose-responsive insulin secretion comparable to that of unencapsulated cells. This work establishes the physical basis of all-aqueous microfiber fabrication and provides a robust, scalable, and cytocompatible approach for cell encapsulation.

Enhancing microbial induced calcium carbonate precipitation (MICCP) in self-healing concrete by addressing cell encapsulation: Investigating the impact of exogenous carbonate ions

Leak-free encapsulation of EGF-expressing yeast in a hyaluronic acid-based hydrogel for diabetes wound healing.

Temperature-responsive hydrogels are sophisticated stimuli-responsive biomaterials that undergo rapid, reversible sol–gel phase transitions in response to subtle thermal stimuli, most notably around physiological temperature. This inherent thermosensitivity enables non-invasive, precise spatiotemporal control of material properties and bioactive payload release, rendering them highly promising for advanced biomedical applications. This review critically surveys recent advances in the design, synthesis, and translational potential of thermo-responsive hydrogels, emphasizing nanoscale and hybrid architectures optimized for superior tunability and biological performance. Foundational systems remain dominated by poly(N-isopropylacrylamide) (PNIPAAm), which exhibits a sharp lower critical solution temperature near 32 °C, alongside Pluronic/Poloxamer triblock copolymers and thermosensitive cellulose derivatives. Contemporary developments increasingly exploit biohybrid and nanocomposite strategies that incorporate natural polymers such as chitosan, gelatin, or hyaluronic acid with synthetic thermo-responsive segments, yielding materials with markedly enhanced mechanical robustness, biocompatibility, and physiologically relevant transition behavior. Cross-linking methodologies—encompassing covalent chemical approaches, dynamic physical interactions, and radiation-induced polymerization are rigorously assessed for their effects on network topology, swelling/deswelling kinetics, pore structure, and degradation characteristics. Prominent applications include on-demand drug and gene delivery, injectable in situ gelling systems, three-dimensional matrices for cell encapsulation and organoid culture, tissue engineering scaffolds, self-healing wound dressings, and responsive biosensing platforms. The integration of multi-stimuli orthogonality, nanotechnology, and artificial intelligence-guided materials discovery is anticipated to deliver fully programmable, patient-specific hydrogels, establishing them as pivotal enabling technologies in precision and regenerative medicine.

Hydrogels have found utility as both tissue scaffolds and cell delivery vehicles. As tissue engineering scaffolds, they provide physical and biochemical cues that guidetissue remodeling processes. Degradation of the gel over time is a desirable material property that allows for the complete integration of imbued cells with the native tissue. Here, novel alginate-based hydrogels with tunable degradability were formulated by synthesizing diblock polymers with a degradable domain. Alginate and poly(D, L-lactic acid) (PLA) of varying molecular weights were end-modified using bio-orthogonal chemistry and linked covalently to form alginate-b-PLA diblock polymers. These polymers retained both the ionic-cross-linking properties of alginate and the hydrolytic degradation properties of PLA. The hydrogel degradation rate was determined by the size of the constituent domains as well as the diblock content in blended gels and enabled the tunable, temporal release of encapsulated cells. This material platform is useful where tunable degradation is advantageous, such as in regenerative medicine or drug delivery.

Alginate microgels are attractive platforms for cell encapsulation, yet conventional gelation strategies often lead to heterogeneous crosslinking, unstable droplets, and reduced cell viability. Here, we present a paraffin oil-based flow-focusing microfluidic system that integratesin situandex situgelation to generate structurally homogeneous and monodisperse Ca-ALG microgels. Unlike conventional approaches that often suffer from unstable droplet formation or incomplete gelation, our method reliably produced uniform microgels with coefficients of variation consistently below 5% and maintained spherical morphology across a wide range of flow conditions. Scanning electron microscopy revealed a hierarchical porous architecture that supported nutrient and metabolite transport while providing structural stability. Encapsulated HEK-293 cells remained highly viable for more than two weeks, and spontaneous spheroid formation occurred within 24 h-an outcome rarely achieved in comparable systems and underscoring the functional relevance of this platform. Compared with existing microfluidic methods, this paraffin oil-driven dual gelation strategy offered superior reproducibility, droplet stability, and encapsulation efficiency. This study integrates and optimizes previously reported dual gelation strategies by employing paraffin oil in a flow-focusing device, establishing a simple, practical, and scalable solution to long-standing challenges in microgel-based encapsulation with strong potential to advance 3D culture, tissue engineering, and regenerative medicine.

Objectives: To evaluate the biocompatibility of blue light-activated methacrylated hyaluronic acid (BL-MeHA) hydrogel using the L929 cell line. Methods: Biocompatibility was assessed using three different assays. In the indirect cytotoxicity assay, L929 cells were cultured in conditioned media that had been exposed to BL-MeHA for 24 hours, followed by an MTT assay to evaluate cell viability. In the 2D culture assay, L929 cells were seeded on top of the BL-MeHA hydrogel, and cell viability was measured on days 1, 3, 5, and 12 using the resazurin assay. For the encapsulation culture assay, L929 cells were embedded within the BL-MeHA hydrogel, and viability was similarly assessed on days 1, 3, 5, and 12 using the resazurin assay. Additionally, L929 cell morphology was examined using scanning electron microscopy (SEM). Results: The indirect cytotoxicity assay demonstrated that L929 cells remained viable when cultured with the BL-MeHA extract. In both the 2D and encapsulation culture assays, L929 cells initially exhibited slower growth compared to the control group but reached comparable levels by day 12. Notably, there was no significant difference in cell viability between BL-MeHA samples cured for 60 and 90 seconds. Conclusions: The BL-MeHA hydrogel exhibited no cytotoxic effects on L929 cells, indicating good biocompatibility. These findings support its potential use as a scaffold for future applications in cell encapsulation or drug delivery for soft tissue engineering.

This study presents a novel approach for the synthesis of biodegradable cell-laden microgels using stop-flow lithography (SFL), addressing critical challenges in the field of tissue engineering. Traditional methods for creating 3D cell cultures often rely on non-biodegradable materials, which limit their application and raise concerns about cell viability. In this work, we successfully replace poly(ethylene glycol) diacrylate (PEGDA) with dextran-2-hydroxyethyl methacrylate (dex-HEMA), a biocompatible and biodegradable alternative. Furthermore, we introduce a technical solution for sterile cell encapsulation, validated through assessments of cell growth and viability alongside the biodegradation rate of the microgel matrix. Our results demonstrate the potential of the self-assembly technique to form organized structures with high spatial resolution. By encapsulating relevant cell lines, Caco-2 and HT-29, within distinct microgel types, we pave the way for the development of sophisticated 3D co-culture models. These advancements hold significant promise for replicating the structural and functional complexities found in native tissues, thereby enhancing the relevance of in vitro studies in biomedical research.

Over the past two decades, microfluidic approaches have been extensively explored for the production of monodisperse alginate hydrogel microparticles, particularly for applications such as 3D in vitro cell culture in tissue engineering. However, conventional methods often suffer from satellite droplet contamination and inefficient oil-to-water phase transfer, limiting purity and scalability. Here, we present an integrated microfluidic platform for the continuous, on-chip production, gelation, extraction, and purification of monodisperse alginate hydrogel microparticles. The device combines a flow-focusing droplet generator with a deterministic lateral displacement (DLD) array, which guides alginate precursor droplets (mean diameter ∼73 µm; CV ∼2.7%) through a water-in-oil emulsion containing calcium ions to induce in-flow ionic crosslinking. The DLD array effectively separates gelled particles from satellite droplets (<37 µm) and transfers them across the oil-water interface into an aqueous stream, eliminating the need for off-chip centrifugation. We demonstrate the platform's applicability to cell encapsulation using MCF-7 cells, achieving a post-processing viability of ∼80%. This work highlights the potential of integrating DLD-based trajectory control with in-flow chemical reactions to enable scalable, high-purity microparticle fabrication for biomedical applications.

A versatile and efficient method to quantify purity and degree of substitution in alginate derivatives.

This article examines the stability of alginate-chitosan capsules containing live Lactobacillus plantarum L1 cultures over a three-month storage period. Culture fluid and biomass precipitated from it were used. The relevance of this study lies in the critical importance of maintaining high viability of probiotic microorganisms at all stages—from production to delivery to the target gastrointestinal tract. Encapsulation of live cells is considered a key technological approach for solving two fundamental problems in probiotic therapy: ensuring targeted delivery to the intestine, bypassing the aggressive acidic environment of the stomach, and significantly increasing the shelf life of the drug by creating a protective shell. A simple and easily scalable syringe-based droplet method was used as the primary microencapsulation method. Formation of the capsule's main polymer matrix is based on the ionic cross-linking reaction of sodium alginate with calcium chloride. A double-coating strategy using chitosan was implemented to increase mechanical strength and reduce permeability of the final capsule shell. The key control parameters for assessing encapsulation efficiency and system stability were the colony-forming unit (CFU) titer, reflecting the viability of the microflora, and the capsule moisture level, indicating their permeability to the surrounding liquid environment. The previous phase of the study included technology refinement and optimization to achieve optimal conditions. An initial stability assessment was performed visually by observing the color development of the capsules in distilled water. The variable parameters included the cross-linking time of sodium alginate with calcium chloride, the reaction time with chitosan, and the concentration of the initial sodium alginate solution. The degree of chitosan deacetylation was also tested as an incoming control parameter.

Engineering mammalian protein secretion: Toward the convergence of high-throughput biology and computational methods.

Abstract Injectable and in situ‐forming hydrogels are attractive for tissue engineering and drug delivery, yet challenges remain in balancing gelation speed, mechanical strength, and cytocompatibility. Here, we report a thermally activatable SpyCatcher/SpyTag (TASpy) system that undergoes rapid chemical crosslinking at physiological temperature. Inspired by the CnaB2 domain, thermally activatable SpyCatcher (TASpyC) fuses SpyCatcher with a non‐reactive SpyTag mutant, creating a single polypeptide that remains folded and minimally reactive at lower temperatures yet partially unfolds at 37 °C to expose a covalent binding site. This design enables controlled isopeptide‐bond formation, facilitating the formation of robust injectable protein hydrogels. We demonstrate improved mechanical properties, enhanced cell encapsulation, and accelerated gelation upon heating—supporting high cell viability and in vivo retention. Notably, subcutaneous injections of TASpy‐based precursors solidify within minutes, degrade over five weeks, and exhibit excellent biocompatibility, underscoring the potential of TASpy hydrogels for advanced biomedical applications.

Injectable and in situ-forming hydrogels are attractive for tissue engineering and drug delivery, yet challenges remain in balancing gelation speed, mechanical strength, and cytocompatibility. Here, we report a thermally activatable SpyCatcher/SpyTag (TASpy) system that undergoes rapid chemical crosslinking at physiological temperature. Inspired by the CnaB2 domain, thermally activatable SpyCatcher (TASpyC) fuses SpyCatcher with a non-reactive SpyTag mutant, creating a single polypeptide that remains folded and minimally reactive at lower temperatures yet partially unfolds at 37°C to expose a covalent binding site. This design enables controlled isopeptide-bond formation, facilitating the formation of robust injectable protein hydrogels. We demonstrate improved mechanical properties, enhanced cell encapsulation, and accelerated gelation upon heating-supporting high cell viability and in vivo retention. Notably, subcutaneous injections of TASpy-based precursors solidify within minutes, degrade over five weeks, and exhibit excellent biocompatibility, underscoring the potential of TASpy hydrogels for advanced biomedical applications.

Peritoneal fibrosis is a significant complication arising from long-term peritoneal dialysis (PD), primarily due to the loss of peritoneal mesothelial cells (PMCs). Recent studies have implicated periostin (POSTN) in the progression of various fibrotic diseases; however, its specific role in PD-induced peritoneal fibrosis remains unclear. Sodium alginate (SA) microgels have emerged as promising carriers for cell encapsulation in tissue engineering and regenerative medicine. This study investigated the therapeutic potential of PMCs encapsulated in SA microgels (SA/PMC) for reducing PD-induced peritoneal fibrosis, with a focus on the modulation by the periostin/nuclear factor kappa-B (NF-κB)/CXCL8 signaling pathway. Primary human peritoneal mesothelial cells (PHPMCs) were isolated from the PD effluent of patients. The effect of SA encapsulation on PMCs proliferation was evaluated using a Cell Counting Kit 8 (CCK-8) assay. The expression levels of POSTN, NF-κB p65, and CXCL8, as well as fibrosis markers, including α-smooth muscle actin (α-SMA), collagen I, transforming growth factor-β (TGF-β), and fibronectin, were evaluated in patients undergoing PD and a PD mouse model. Patients undergoing PD for 1 year exhibited elevated levels of POSTN, NF-κB p65, CXCL8, and fibrosis markers compared with those undergoing PD for 1 week. Consistent results from in vivo and in vitro models demonstrated that PD and hyperglycemic conditions upregulated the expression of POSTN, NF-κB p65, CXCL8, and profibrotic markers, leading to peritoneal thickening and fibrotic progression. Treatment with SA/PMC microgels ameliorated these effects. By modulating the POSTN/NF-κB/CXCL8 pathway and enhancing PMCs survival, SA/PMC microgels may have therapeutic potential in mitigating peritoneal fibrosis in PD patients.

Despite the rapid progress of perovskite solar cells (PSCs) toward commercialization, perovskite layers are unstable and contain toxic lead. Encapsulation represents an efficient strategy for improving the stability of PSCs as well as suppressing lead leakage. However, the damage to encapsulation materials during outdoor applications inevitably deteriorates the packaging effect and increases lead leakage. Here we report a damage perception and rapid self-healing encapsulant consisting of alkoxy polyvinylimidazole bis(trifluoromethanesulphonyl)imide (EP). The dynamic ion aggregates in the encapsulant can easily drive the molecular chain movement of EP, thereby achieving rapid damage repair to maintain device stability and inhibit lead leakage. The damaged cracks of EP completely self-heal within 6 minutes at 50°C. The EP encapsulated devices exhibit a lead sequestration efficiency of more than 99% under poor weather. After 1500 hours of damp heat test and 300 thermal cycles, the EP encapsulated devices retain 95.17 and 93.53% of their initial efficiency, respectively.

Engineering mechanically resilient hydrogels from naturally derived proteins, such as collagen and gelatin, remains a key challenge in tissue regeneration, particularly when cell compatibility and structural integrity are simultaneously required. Here, a bioorthogonal crosslinking strategy using rhodamine and polyethylene glycol (PEG) is reported to fabricate dense, mechanically reinforced collagen hydrogels. PEG-mediated dehydration induces spontaneous peptide bond formation between rhodamine and collagen without the need for additional catalysts, yielding fibrous protein networks with enhanced stiffness. To enable anisotropic tissue engineering, this crosslinking method is integrated with wet-spinning to produce uniaxially aligned collagen filaments. These constructs exhibit high mechanical strength and support human adipose-derived stem cell (hASC) encapsulation. Mechanotransductive signaling, including cytoskeletal organization and myogenic gene expression, is effectively activated within the aligned filaments. The applicability of cell-laden filaments in a murine volumetric muscle loss model is demonstrated, which promoted in vitro differentiation and in vivo functional muscle regeneration. This strategy offers a scalable and cytocompatible platform for generating aligned protein-based scaffolds with tunable mechanical and biological properties, thereby expanding the toolkit for regenerative medicine.

Enhanced electron microscopy imaging for a detailed structural study of alginate hydrogel containing the encapsulated cells.