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Microfluidic synthesis of stimuli-responsive hydrogel particles
Hydrogel microspheres are a class of hydrophilic polymeric particles in microscale, which has been developed as a new type of functional biomaterials for wide-range biomedical applications in recent years. This review provides a comprehensive overview of the preparation methods for hydrogel microspheres, including droplet microfluidics, electrospray and emulsion was first summarized. At the same time, we analyze the impacts of these methods on the properties of hydrogel microspheres and explore various functionalization strategies for enhancing their bioactivity and expanding their biomedical applications. In addition, we discuss the recent advances and the further prospect of hydrogel microspheres in life science applications, particularly in cell biology research, bioanalysis and detection, as well as tissue repair and regeneration. By synthesizing the latest developments, this review aims to offer valuable insights and strategies for optimizing hydrogel microspheres in diverse application scenarios and inspire future research and practical innovations.
Continuous and in situ detection of biomarkers in biofluids (for example, sweat) can provide critical health data but is limited by biofluid accessibility. Here we report a sensor design that enables in situ detection of solid-state biomarkers ubiquitously present on human skin. We deploy an ionic–electronic bilayer hydrogel to facilitate the sequential dissolution, diffusion and electrochemical reaction of solid-state analytes. We demonstrate continuous monitoring of water-soluble analytes (for example, solid lactate) and water-insoluble analytes (for example, solid cholesterol) with ultralow detection limits of 0.51 and 0.26 nmol cm⁻², respectively. Additionally, the bilayer hydrogel electrochemical interface reduces motion artefacts by a factor of three compared with conventional liquid-sensing electrochemical interfaces. In a clinical study, solid-state epidermal biomarkers measured by our stretchable wearable sensors showed a high correlation with biomarkers in human blood and dynamically correlated with physiological activities. These results present routes to universal platforms for biomarker monitoring without the need for biofluid acquisition.
Ideal hydrogel fibers with high toughness and environmental tolerance are indispensable for their long-term application in flexible electronics as actuating and sensing elements. However, current hydrogel fibers exhibit poor mechanical properties and environmental instability due to their intrinsically weak molecular (chain) interactions. Inspired by the multilevel adjustment of spider silk network structure by ions, bionic hydrogel fibers with elaborated ionic crosslinking and crystalline domains are constructed. Bionic hydrogel fibers show a toughness of 162.25 ± 21.99 megajoules per cubic meter, comparable to that of spider silks. The demonstrated bionic structural engineering strategy can be generalized to other polymers and inorganic salts for fabricating hydrogel fibers with broadly tunable mechanical properties. In addition, the introduction of inorganic salt/glycerol/water ternary solvent during constructing bionic structures endows hydrogel fibers with anti-freezing, water retention, and self-regeneration properties. This work provides ideas to fabricate hydrogel fibers with high mechanical properties and stability for flexible electronics.
Hydrogels are an attractive category of biointerfacing materials with adjustable mechanical properties, diverse biochemical functions, and good ionic conductivity. Despite these advantages, their application in electronics has been restricted because of their lack of semiconducting properties, and they have traditionally only served as insulators or conductors. We developed single- and multiple-network hydrogels based on a water-soluble n-type semiconducting polymer, endowing conventional hydrogels with semiconducting capabilities. These hydrogels show good electron mobilities and high on/off ratios, enabling the fabrication of complementary logic circuits and signal amplifiers with low power consumption and high gains. We demonstrate that hydrogel electronics with good bioadhesive and biocompatible interface can sense and amplify electrophysiological signals with enhanced signal-to-noise ratios.
In this study, a capillary microfluidic device was constructed, and sodium alginate solution and a pH-sensitive hydrophobic polymer (p(BMA-co-DAMA-co-MMA)) solution were introduced into the device for the preparation of hydrogel fibers loaded with polymer microspheres. The structure of the microsphere fiber, including the size and spacing of the microspheres, could be controlled by flow rate, and the microspheres were able to degrade and release cargo responding to acidic pH conditions. By modification with carboxymethylcellulose (CMC), alginate hydrogel exhibited enhanced pH sensitivity (shrunk in acidic while swollen in basic condition). This led to an impact on the diffusion rate of the molecules released from the inner microspheres. The microsphere fiber showed dramatic and negligible degradation and drug release in tumor cell (i.e., A431 and A549 cells) and normal cell environments, respectively. These results indicated that the microsphere fiber prepared in this study showed selective drug release in acidic environments, such as tumor and inflammation sites, which could be applied as a smart surgical dressing with normal tissue protective properties.
Penetrating corneal wounds can cause severe vision impairment and require prompt intervention to restore globe integrity and minimize the risk of infection. Tissue adhesives have emerged as a promising alternative to suturing for mitigating postoperative complications. However, conventional water‐soluble adhesives suffer formidable challenges in sealing penetrating corneal wounds due to dilution or loss in a moist environment. Inspired by the robust adhesion of mussels in aquatic conditions, an injectable photocurable bioadhesive hydrogel (referred to as F20HD5) composed of polyether F127 diacrylate and dopamine‐modified hyaluronic acid methacrylate is developed for sutureless closure of corneal full‐thickness wounds. F20HD5 exhibits high transparency, wound‐sealing ability, proper viscosity, biodegradability, and excellent biocompatibility. It allows in situ cross‐linking via visible light, thereby providing sufficient mechanical strength and adhesiveness. In vivo, the adhesive hydrogel effectively closed penetrating linear corneal incisions and corneal injuries with minimal tissue loss in rabbits. During the 56‐day follow‐up, the hydrogel facilitates the repair of the injured corneas, resulting in more symmetrical curvatures and less scarring in distinction to the untreated control. Thus, bioinspired hydrogel holds promise as an effective adhesive for sealing full‐thickness corneal wounds.
Stimuli-responsive shape-changing hydrogels are attractive candidates for use as underwater soft robots. The bottleneck lies in the low actuation speed inherently limited by the water diffusion between hydrogels and their surrounding environment. In addition, accessing complex motions is restricted by the material fabrication methods. Here we report a hitherto unknown mechanism to achieve high-speed and programmable actuations for a disulfide crosslinked thermally responsive hydrogel. The dynamic photo-activated disulfide bond exchange allows photo-mechanical programming to introduce spatio-selective network anisotropy. This gives rise to an actuation behavior dominated by thermally driven conformation change of the locally oriented polymer chains instead of the common mass-diffusion-based mechanism. With the incorporation of photothermal fillers, light-powered oscillation at frequencies as high as 1.7 Hz is realized. This, coupled with the versatility of the programming, allows access to robots with diverse high-speed motions including continuous swimming, step-wise walking, and rotating.
Bulk hydrogel scaffolds are common in reconstructive surgery. They allow for the staged repair of soft tissue loss by providing a base for revascularization. Unfortunately, they are limited by both slow and random vascularization, which may manifest as treatment failure or suboptimal repair. Rapidly inducing patterned vascularization within biomaterials has profound translational implications for current clinical treatment paradigms and the scaleup of regenerative engineering platforms. To address this long‐standing challenge, a novel microsurgical approach and granular hydrogel scaffold (GHS) technology are co‐developed to hasten and pattern microvascular network formation. In surgical micropuncture (MP), targeted recipient blood vessels are perforated using a microneedle to accelerate cell extravasation and angiogenic outgrowth. By combining MP with an adjacent GHS with precisely tailored void space architecture, microvascular pattern formation as assessed by density, diameter, length, and intercapillary distance is rapidly guided. This work opens new translational opportunities for microvascular engineering, advancing reconstructive surgery, and regenerative medicine.
Postsurgical pericardial adhesions pose increased risks of sequelae, prolonged reoperation time, and reduced visibility in the surgical field. Here, we introduce an injectable Janus hydrogel, which exhibits asymmetric adhesiveness properties after photocrosslinking, sustained delivering induced pluripotent stem cell-derived cardiomyocyte exosomes (iCM-EXOs) for post-heart surgery adhesion reduction. Our findings reveal that iCM-EXOs effectively attenuate oxidative stress in hydrogen peroxide-treated primary cardiomyocytes by inhibiting the activation of the transcription factor nuclear factor erythroid 2-related factor 2. Notably, in rat cardiac postsurgery models, the Janus hydrogels loaded with iCM-EXOs demonstrate dual functionality, acting as antioxidants and antipericardial adhesion agents. These hydrogels effectively protect iCM-EXOs from GATA6+ cavity macrophage clearance by inhibiting the recruitment of macrophages from the thoracic cavity. These results highlight the promising potential of iCM-EXO-laden Janus hydrogels for clinical safety and efficacy validation in trials involving heart surgery patients, with the ultimate goal of routine administration during open-heart surgeries.
Cellulose nanocrystal (CNC) based optical devices with adjustable schemochrome have attracted immense interest. However, most of the previously reported structural colored CNC‐based materials can only achieve simple stress‐induced color change, which have difficulty achieving multimode control of complex patterning that can be accurately identified. Here, inspired by the nanostructure‐based color‐changing mechanism of neon tetra, this study presents a pressure/temperature dual‐responsive CNC‐based schemochrome hydrogel with adjustable dynamic chiral nematic structure. By incorporating abundant interfacial noncovalent interactions, dynamic correlations between adjustable helical pitch of the vertically stacked cholesteric liquid crystalline (LC) phase and responsiveness of flexible thermosensitive substrate are established, which further enable wide‐range optical characteristic (12°–213° in HSV color model and 421–734 nm in the UV–Vis spectra) and identifiable visualized patterning. The resultant hydrogels are applied in proof‐of‐concept demonstrations of on‐demand schemochrome patterning, including customizable patterned dual‐encryption label, smart digital display, temperature monitor, and intelligent recognition/control system. This study envisages that the bioinspired construction of structural colored nanomaterials will have promising applications in smart responsive photonic equipment including smart display, anticounterfeiting, and intelligent control systems.
The response rate of pancreatic cancer to chemotherapy or immunotherapy pancreatic cancer is low. Although minimally invasive irreversible electroporation (IRE) ablation is a promising option for irresectable pancreatic cancers, the immunosuppressive tumour microenvironment that characterizes this tumour type enables tumour recurrence. Thus, strengthening endogenous adaptive antitumour immunity is critical for improving the outcome of ablation therapy and post-ablation immune therapy. Here we present a hydrogel microsphere vaccine that amplifies post-ablation anti-cancer immune response via releasing its cargo of FLT3L and CD40L at the relatively lower pH of the tumour bed. The vaccine facilitates migration of the tumour-resident type 1 conventional dendritic cells (cDC1) to the tumour-draining lymph nodes (TdLN), thus initiating the cDC1-mediated antigen cross-presentation cascade, resulting in enhanced endogenous CD8⁺ T cell response. We show in an orthotopic pancreatic cancer model in male mice that the hydrogel microsphere vaccine transforms the immunologically cold tumour microenvironment into hot in a safe and efficient manner, thus significantly increasing survival and inhibiting the growth of distant metastases.
The formation of microparticles (MPs) of biocompatible and biodegradable hydrogels such as polyethylene glycol diacrylate (PEGDA) utilizing microfluidic devices is an attractive option for entrapment and encapsulation of active principles and microorganisms. Our research group has presented in previous studies a formulation to produce these hydrogels with adequate physical and mechanical characteristics for their use in the formation of MPs. In this work, hydrogel MPs are formed based on PEGDA using a microfluidic device with a T-junction design, and the MPs become hydrogel through a system of photopolymerization. The diameters of the MPs are evaluated as a function of the hydrodynamic condition flow rates of the continuous (Qc) and disperse (Qd) phases, measured by optical microscopy, and characterized through scanning electron microscopy. As a result, the following behavior is found: the diameter is inversely proportional to the increase in flow in the continuous phase (Qc), and it has a significant statistical effect that is greater than that in the flow of the disperse phase (Qd). While the diameter of the MPs is proportional to Qd, it does not have a significant statistical effect on the intervals of flow studied. Additionally, the MPs’ polydispersity index (PDI) was measured for each experimental hydrodynamic condition, and all values were smaller than 0.05, indicating high homogeneity in the MPs. The microparticles have the potential to entrap pharmaceuticals and microorganisms, with possible pharmacological and bioremediation applications.
Selective delivery of anticancer drug molecules to the tumor site enhances local drug dosages, which leads to the death of cancer cells while simultaneously minimizing the negative effects of chemotherapy on other tissues, thereby improving the patient’s quality of life. To address this need, we developed reduction-responsive chitosan-based injectable hydrogels via the inverse electron demand Diels–Alder reaction between tetrazine groups of disulfide-based cross-linkers and norbornene groups of chitosan derivatives, which were applied to the controlled delivery of doxorubicin (DOX). The swelling ratio, gelation time (90–500 s), mechanical strength (G’~350–850 Pa), network morphology, and drug-loading efficiency (≥92%) of developed hydrogels were investigated. The in vitro release studies of the DOX-loaded hydrogels were performed at pH 7.4 and 5.0 with and without DTT (10 mM). The biocompatibility of pure hydrogel and the in vitro anticancer activity of DOX-loaded hydrogels were demonstrated via MTT assay on HEK-293 and HT-29 cancer cell lines, respectively.
Exosomal microRNAs have been studied as a good source of noninvasive biomarkers due to their functions in genetic exchange between cells and have been already well documented in many biological activities; however, inaccuracy remains a key challenge for liver cancer surveillance. Herein, a versatile duplex photothermal digital polymerase chain reaction (PCR) strategy combined with a lipid nanoparticle‐based exosome capture approach is proposed to profile microRNAs expression through a 3‐h easy‐to‐operate process. The microfluidically‐generated molybdenum disulfide‐nanocomposite‐doped gelatin microcarriers display attractive properties as a 2–4 °C s⁻¹ ramping‐up rate triggered by near‐infrared and reversible sol–gel transforming in step with PCR activation. To achieve PCR thermocycling, the corresponding irradiation coordinating with fan cooling are automatically performed via a homemade control module with programs. Thus, taking the multiplexing capability of dual‐color labeling, 19–31 folds higher in exosomal microRNA‐200b‐3p and microRNA‐21‐5p, and tenfold lower in microRNA‐22‐3p expressions relative to the control microRNA‐26a‐5p are quantified in two liver cancer cells (Huh7 and HepG2) than in those from the healthy cells. It is believed that this exosomal microRNA genotyping method would be highly applicable for liver cancer diagnostics.
Chemotherapy is an essential postoperative treatment for pancreatic cancer, while due to the lack of effective drug evaluation platforms, the therapeutic outcomes are hampered by tumor heterogeneity among individuals. Here, a novel microfluidic encapsulated and integrated primary pancreatic cancer cells platform is proposed for biomimetic tumor 3D cultivation and clinical drug evaluation. These primary cells are encapsulated into hydrogel microcapsules of carboxymethyl cellulose cores and alginate shells based on a microfluidic electrospray technique. Benefiting from the good monodispersity, stability, and precise dimensional controllability of the technology, the encapsulated cells can proliferate rapidly and spontaneously form 3D tumor spheroids with highly uniform size and good cell viability. By integrating these encapsulated tumor spheroids into a microfluidic chip with concentration gradient channels and culture chambers, dynamic and high‐throughput drug evaluation of different chemotherapy regimens could be realized. It is demonstrated that different patient‐derived tumor spheroids show different drug sensitivity on‐chip, which is significantly consistent with the clinical follow‐up study after the operation. The results demonstrate that the microfluidic encapsulated and integrated tumor spheroids platform has great application potential in clinical drug evaluation. Primary human pancreatic cancer cells encapsulated in hydrogel microcapsules with good biocompatibility and accessibility can proliferate rapidly and spontaneously form 3D tumor spheroids with highly uniform size and good cell viability. Integrating these encapsulated tumor spheroids into a microfluidic chip with a gradient generator and culture chambers enables dynamic and high‐throughput drug evaluation of different chemotherapy regimens.
Paracrine is an important mechanism in mesenchymal stem cells (MSCs) that promotes tissue regeneration. However, anoikis is attributed to unsuitable adhesion microenvironment hindered this paracrine effect. In this study, a living and injectable porous hydrogel microsphere with long‐term paracrine activity is constructed via the freeze‐drying microfluidic technology and the incorporation of platelet‐derived growth factor‐BB (PDGF‐BB) and exogenous MSCs. Benefiting from the porous structure and superior mechanical property of methacrylate gelatin (GelMA) hydrogel microspheres (GMs), exogenous stem cells are able to adhere and proliferate on GMs, thereby facilitating cell‐to‐extracellular matrix (ECM) and cell‐to‐cell interactions and enhancing paracrine effect. Furthermore, the sustained release of PDGF‐BB can recruit endogenous MSCs to prolong the paracrine activity of the living GMs. In vitro and in vivo experiments validated that the living GMs exhibit superior secretion properties and anti‐inflammatory efficacy and can attenuate osteoarthritis (OA) progression by favoring the adherent microenvironment and utilizing the synergistic effect of exogenous and endogenous MSCs. Overall, a living injectable porous hydrogel microsphere that can enhance the paracrine activity of stem cells is fabricated and anticipated to hold the potential of future clinical translation in OA and other diseases.
At present, surgery is one of the main treatments for bone tumor. However, the risk of recurrence and the large area of bone defects after surgery pose a great challenge. Therefore, a Janus-inspired core-shell structure bone scaffold was designed to achieve the self-programmed release of melatonin at different concentrations, clearing the residual tumor cells at early stage after resection and promoting bone repair later. The layered differential load designs inspired by Janus laid the foundation for the differential release of melatonin, where sufficient melatonin inhibited tumor growth as low dose promoted osteogenesis. Then, the automatically programmed delivery of melatonin is achieved by the gradient degradation of the core-shell structure. In the material characterization, scanning electron microscopy revealed the core-shell structure. The drug release experiment and in vivo degradation experiment reflected the programmed differential release of melatonin. In the biological experiment part, in vivo and in vitro experiments not only confirmed the significant inhibitory effect of the core-shell hydrogel scaffold on tumor but also confirmed its positive effect on osteogenesis. Our Janus-inspired core-shell hydrogel scaffold provides a safe and efficient means to inhibit tumor recurrence and bone repair after bone tumor, and it also develops a new and efficient tool for differential and programmed release of other drugs.
Granular hydrogels have been increasingly exploited in biomedical applications, including wound healing and cardiac repair. Despite their utility, design guidelines for engineering their macroscale properties remain limited, as we do not understand how the properties of granular hydrogels emerge from collective interactions of their microgel building blocks. In this work, we related building block features (stiffness and size) to the macroscale properties of granular hydrogels using contact mechanics. We investigated the mechanics of the microgel packings through dynamic oscillatory rheology. In addition, we modeled the system as a collection of two-body interactions and applied the Zwanzig and Mountain formula to calculate the plateau modulus and viscosity of the granular hydrogels. The calculations agreed with the dynamic mechanical measurements and described how microgel properties and contact deformations define the rheology of granular hydrogels. These results support a rational design framework for improved engineering of this fascinating class of materials.
Intelligent biomaterials can modify their properties in response to physical, chemical, and biological stimuli. These smart characteristics drive the innovation of biomaterials in therapy and diagnostics for detecting diseases and providing treatment at early stages. Mainly, hydrogels have gained significant interest in developing smart materials due to their excellent biocompatibility and ability to interact with body fluids that host condition-specific stimuli. Temperature, pressure, pH, light, ROS, cell metabolites, and other physicochemical factors specific to specific disease conditions were studied as major stimuli for designing intelligent biomaterials. The stimuli-responsive characteristic mainly depends on the sensitivity of the biomaterial to the stimuli factor and the tunable macromolecular structure of the materials. The method of biomaterial fabrication is critical in determining the physical and chemical properties of the biomaterial. Surface functionalisation, material blending, and crosslinking are commonly used to synthesise intelligent hydrogels to change the macromolecular structure. The impact and mechanism of these fabrication methods on the macromolecular structure and stimuli responsiveness of intelligent materials remain unidentified. This review focuses on strategies for transforming conventional hydrogels into intelligent hydrogels, their concerning mechanisms of stimuli-responsiveness and their biomedical applications.
The localization and accumulation of drugs in the body determine their therapeutic effects; however, the specific microstructure of damaged tissues hinders drug delivery. Currently, there is a shortage of effective drug carriers to breach these barriers and achieve efficient tissue and cellular delivery of drugs. In this study, an injectable double positively charged functional hydrogel microsphere with “targeting cartilage extracellular matrix”, “cartilage penetration”, and “cellular phagocytosis” is designed for matching the structural characteristics of joints, addressing the difficulties of drug delivery in joints. The microspheres could be adsorbed on the negatively charged cartilage surface because of their positively charged poly‐lysine surface. Furthermore, the internally loaded positively charged polyamidoamine contained kartogenin, which helped further the penetration of the cartilage under the guidance of electrical charge. The microspheres could release kartogenin for more than 21 days. In in vivo experiments, the microspheres effectively improve the efficiency of drug delivery, inhibit the degradation of cartilage matrix and subchondral bone, and delay the development of osteoarthritis. As a double positively charged drug delivery system, the versatile microsphere has great potential for treating osteoarthritis and other diseases.
Artemisinin and its derivatives (artemisinins) are first‐line chemotherapeutic agents of lethal malaria, which also showed tremendous value in many other diseases including chronic inflammation. Unfortunately, almost all artemisinins are rapid‐acting medicines with an extremely short half‐life in vivo, which significantly limits their clinical application for these new adaptation diseases. In this study, a locally injectable long‐acting gene/artemisinin co‐delivery nano‐microplex consisting of a biodegradable hyaluronic acid (HA) microsphere and releasable gene/artemisinin co‐delivery nano‐lipoplex is developed first, to obtain an improved efficacy for rheumatoid arthritis (RA). Briefly, a cationic multicomponent drug‐embedded liposome with pharmacological activity is first reported based on two novel artemisinin derivatives (dAPC and dACC), which possess mimic phospholipids and cationic lipids, respectively. A cationic artemisinin‐embedded lipoplex is first reported as a medicative gene carrier here. An in situ injectable TNF‐α siRNA/artemisinin co‐delivery nano‐microplex (MTAsi@MG) is further prepared by immobilization of TNF‐α siRNA/lipoplex on porous microfluidic HA microspheres. Using this nano‐microplex for intra‐articular injection, the sustaining activity of gene therapy and artemisinin efficacy for RA long‐term treatment is first realized. Undoubtedly, this intra‐articular injectable TNF‐α siRNA/artemisinin co‐delivery nano‐microplex based on dAPC/dACC lipoplex and microfluidic microspheres would be one of the most potent gene/drug co‐delivery systems for RA therapy.
Flexible pressure sensors usually require functional materials with both mechanical compliance and appropriate electrical performance. Most sensors based on materials with limited compressibility can hardly balance between high sensitivity and broad pressure range. Here, we prepare a heterophasic ionogel with shape and stiffness memory for adaptive pressure sensors. By combining the microstructure alignment for stiffness changing and shape memory micro-inclusions for stiffness fixing, the heterophasic ionogels reveal tunable compressibility. This controllable pressure-deformation property of the ionogels results in the pressure sensors’ programmable pressure-resistance behavior with tunable pressure ranges, varied detection limits, and good resolution at high pressure. Broad pressure ranges to 220 and 380 kPa, and tunable detection limit from 120 to 330 and 950 Pa are realized by the stiffness memory ionogel sensors. Adaptive detection is also brought out to monitor tiny pressure changes at low stiffness and distinguish different human motions at high stiffness. Using shape and stiffness memory materials in pressure sensors is a general design to achieve programmable performance for more complex application scenarios.
The soft colloidal probe (SCP) assay is a highly versatile sensing principle employing micrometer-sized hydrogel particles as optomechanical transducer elements. We report the synthesis, optimization, and conjugation of SCPs with defined narrow size distribution and specifically tailored mechanical properties and functionalities for integration into a microinterferometric optomechanical biosensor platform. Droplet-based microfluidics was used to crosslink polyethylene glycol (PEG) macromonomers by photocrosslinking and thiol-Michael addition. The effect of several synthesis parameters, i.e. PEG and radical initiator solid content, molecular weight and architecture of macromonomers, as well as UV exposure time and energy, were examined. SCPs were characterized with regard to the conversion of contained functional groups, morphology and mechanical properties by bright-field, confocal laser scanning and reflection interference contrast microscopy, as well as force spectroscopy. Functional groups were introduced during SCP synthesis and by several post-synthesis procedures, based on photoradical grafting and thiol-Michael addition. Preparation of SCPs by thiol-Michael addition and subsequent coupling of maleimide derivatives to unreacted thiols proved to be the superior strategy, while other approaches were associated with changes in the properties of the SCP. The newly developed SCPs were tested for their sensing capabilities employing the biotin-streptavidin-system. Biotin detection in the range of 10-7 to 10-10 M verified the concept of the synthesis strategy and the advantage of using monodisperse SCPs for easier and faster sensing applications of the SCP assay.
Biofabrication technologies are of importance for the construction of organ models and functional tissue replacements. Microfluidic manipulation, a promising biofabrication technique with micro‐scale resolution, can not only help to realize the fabrication of specific microsized structures but also build biomimetic microenvironments for biofabricated tissues. Therefore, microfluidic manipulation has attracted attention from researchers in the manipulation of particles and cells, biochemical analysis, tissue engineering, disease diagnostics, and drug discovery. Herein, biofabrication based on microfluidic manipulation technology is reviewed. The application of microfluidic manipulation technology in the manufacturing of biomaterials and biostructures with different dimensions and the control of the microenvironment is summarized. Finally, current challenges are discussed and a prospect of microfluidic manipulation technology is given. The authors hope this review can provide an overview of microfluidic manipulation technologies used in biofabrication and thus steer the current efforts in this field.
Bone defects caused by trauma, tumor, or osteoarthritis remain challenging due to the lack of effective treatments in clinic. Stem cell transplantation has emerged as an alternative approach for bone repair and attracted widespread attention owing to its excellent biological activities and therapy effect. The attempts to develop this therapeutic approach focus on the generation of effective cell delivery vehicles, since the shortcomings of direct injection of stem cells into target tissues. Here, we developed a novel core-shell microcapsule with a stem cell-laden core and a biomass shell by using all-aqueous phase microfluidic electrospray technology. The designed core-shell microcapsules showed a high cell viability during the culture procedure. In addition, the animal experiments exhibited that stem cell-laden core-shell microcapsules have good biocompatibility and therapeutic effect for bone defects. This study indicated that the core-shell biomass microcapsules generated by microfluidic electrospray have promising potential in tissue engineering and regenerative medicine.
Purpose
Small extracellular vesicles (sEV) play an irreplaceable role in cell–cell communication. However, sEV in solution aggregate with each other during preservation, leading to impairment of the structures, contents, and functions of sEV. Therefore, there is a need to develop an optimal preservation method that combines high recovery rate, low cost, convenience, and easy-transportation in one. In this study, a new preservation strategy different from the cryopreservation or lyophilization was developed by reducing sEV particles aggregation.
Methods
The sEV were encapsulated in thermoresponsive gelatin methacryloyl (GelMA) hydrogels at 4°C to reduce particles aggregation during the reversible cross-linking process. The sEV movement was visualized in different mediums and particles’ number, size, structure and protein of 28 days preserved sEV were compared to fresh sEV. Human umbilical vein endothelial cells (HUVEC) and rat adipose-derived stromal stem cells (rASC) were isolated and cultured with fresh and preserved sEV to test the cellular response. A mice subcutaneous model was adopted to detect controlled release and angiogenesis ability of preserved sEV.
Results
Through particles tracks visualization, GelMA hydrogels significantly decreased the sEV movement. After 28 days preservation in GelMA at 4°C, the particles number, size, structure and protein of sEV were similar to fresh sEV. In vitro, preserved sEV had the same ability to promote cell proliferation, migration and angiogenesis as fresh sEV. In vivo, preserved sEV-GelMA could artificially regulate the absorptivity of GelMA hydrogels and controlled released sEV for therapeutic application, and preserved sEV encapsulated in GelMA significantly promoted angiogenesis in mice.
Conclusion
Our results demonstrated that sEV encapsulated in GelMA could be a novel strategy for long-term preservation of sEV for therapeutic application.
Lymphocytes play a vital role in immunosurveillance through sensing biomolecules and eliminating targeted invaders. Compared with conventional therapies that depend on drug loading, lymphocytes are advantageous as they are able to ensure self-regulated therapeutics. Here, novel multi-compartmental DNA hydrogel particles were synthesized using a microfluidic assembly for intelligent cancer treatment via the logic-based control of siRNA release without external stimulation. The sensing sequence (D1) was compartmentalized from the treatment sequence (D2) with the use of core-shell DNA hydrogel particles. When D1 detects a cancer-associated biomarker, miRNA-21, a sequence cascade is triggered to release siRNA from D2, effectively eliminating the targeted cancer cells via lymphocyte-inspired precision medicine.
The wet and highly dynamic environment of the mouth makes local treatment of oral mucosal diseases challenging. To overcome this, a photo-crosslinking hydrogel adhesive is developed inspired by the success of light-curing techniques in dentistry. The adhesive operates on a fast (within 5 s) phototriggered S-nitrosylation coupling reaction and employs imine anchoring to connect to host tissues. Unlike other often-used clinical agents that adhere weakly and for short durations, this thin, elastic, adhesive, and degradable cyclic o-nitrobenzyl-modified hyaluronic acid gel protects mucosal wounds from disturbance by liquid rinsing, oral movement, and friction for more than 24 h. The results from both rat and pig oral mucosa repair models demonstrate that this new gel adhesive creates a favorable microenvironment for tissue repair and can shorten tissue healing time. This study thus illustrates a therapeutic strategy with the potential to advance the treatment of oral mucosal defects in the clinic.
To optimize the anti-tumor efficacy of combination therapy with paclitaxel (PTX) and imatinib (IMN), we used coaxial electrospray to prepare sequential-release core–shell microparticles composed of a PTX-loaded sodium hyaluronate outer layer and an IMN-loaded PLGA core. The morphology, size distribution, drug loading, differential scanning calorimetry (DSC), Fourier transform infrared spectra (FTIR), in vitro release, PLGA degradation, cellular growth inhibition, in vivo vaginal retention, anti-tumor efficacy, and local irritation in a murine orthotopic cervicovaginal tumor model after vaginal administration were characterized. The results show that such core–shell microparticles were of spherical appearance, with an average size of 14.65 μm and a significant drug-loading ratio (2.36% for PTX, 19.5% for IMN, w/w), which might benefit cytotoxicity against cervical-cancer-related TC-1 cells. The DSC curves indicate changes in the phase state of PTX and IMN after encapsulation in microparticles. The FTIR spectra show that drug and excipients are compatible with each other. The release profiles show sequential characteristics in that PTX was almost completely released in 1 h and IMN was continuously released for 7 days. These core–shell microparticles showed synergistic inhibition in the growth of TC-1 cells. Such microparticles exhibited prolonged intravaginal residence, a >90% tumor inhibitory rate, and minimal mucosal irritation after intravaginal administration. All results suggest that such microparticles potentially provide a non-invasive local chemotherapeutic delivery system for the treatment of cervical cancer by the sequential release of PTX and IMN.
Chemical reaction networks (CRN) embedded in hydrogels can transform responsive materials into complex self‐regulating materials that generate feedback to counter the effect of external stimuli. This study presents hydrogels containing the β‐cyclodextrin (CD) and ferrocene (Fc) host–guest pair as supramolecular crosslinks where redox‐responsive behavior is driven by the enzyme–fuel couples horse radish peroxidase (HRP)–H2O2 and glucose oxidase (GOx)–d‐glucose. The hydrogel can be tuned from a responsive to a self‐regulating supramolecular system by varying the concentration of added reduction fuel d‐glucose. The onset of self‐regulating behavior is due to formation of oxidation fuel in the hydrogel by a cofactor intermediate GOx[FADH2]. UV/Vis spectroscopy, rheology, and kinetic modeling were employed to understand the emergence of out‐of‐equilibrium behavior and reveal the programmable negative feedback response of the hydrogel, including the adaptation of its elastic modulus and its potential as a glucose sensor.
This article provides a systematic review of the crosslinking strategies used to produce microgel particles in microfluidic chips. Various ionic crosslinking methods for the gelation of charged polymers are discussed, including external gelation via crosslinkers dissolved or dispersed in the oil phase; internal gelation methods using crosslinkers added to the dispersed phase in their non-active forms, such as chelating agents, photo-acid generators, sparingly soluble or slowly hydrolyzing compounds, and methods involving competitive ligand exchange; rapid mixing of polymer and crosslinking streams; and merging polymer and crosslinker droplets. Covalent crosslinking methods using enzymatic oxidation of modified biopolymers, photo-polymerization of crosslinkable monomers or polymers, and thiol-ene “click” reactions are also discussed, as well as methods based on the sol−gel transitions of stimuli responsive polymers triggered by pH or temperature change. In addition to homogeneous microgel particles, the production of structurally heterogeneous particles such as composite hydrogel particles entrapping droplet interface bilayers, core−shell particles, organoids, and Janus particles are also discussed. Microfluidics offers the ability to precisely tune the chemical composition, size, shape, surface morphology, and internal structure of microgels by bringing multiple fluid streams in contact in a highly controlled fashion using versatile channel geometries and flow configurations, and allowing for controlled crosslinking.
The generation of hydrogel droplets using droplet microfluidics has emerged as a powerful tool with many applications in biology and medicine. Here, a microfluidic system to control the position of particles (beads or astrocyte cells) in hydrogel droplets using bulk acoustic standing waves is presented. The chip consisted of a droplet generator and a 380 µm wide acoustic focusing channel. Droplets comprising hydrogel precursor solution (polyethylene glycol tetraacrylate or a combination of polyethylene glycol tetraacrylate and gelatine methacrylate), photoinitiator and particles were generated. The droplets passed along the acoustic focusing channel where a half wavelength acoustic standing wave field was generated, and the particles were focused to the centre line of the droplets (i.e. the pressure nodal line) by the acoustic force. The droplets were cross-linked by exposure to UV-light, freezing the particles in their positions. With the acoustics applied, 89 ± 19% of the particles (polystyrene beads, 10 µm diameter) were positioned in an area ± 10% from the centre line. As proof-of-principle for biological particles, astrocytes were focused in hydrogel droplets using the same principle. The viability of the astrocytes after 7 days in culture was 72 ± 22% when exposed to the acoustic focusing compared with 70 ± 19% for samples not exposed to the acoustic focusing. This technology provides a platform to control the spatial position of bioparticles in hydrogel droplets, and opens up for the generation of more complex biological hydrogel structures.
Objective
Self-healing of bone from damage caused by infection, trauma, or surgical removal of cysts is limited. Generally, external intervention is needed to increase bone repair and regeneration. In this study, biocompatible light-cured hyaluronic acid hydrogels loaded with nano-hydroxyapa tite and chitosan were prepared using a new photoinitiating system based on riboflavin for bone regeneration applications.
Method
Four light-cured hydrogel groups were prepared as follows: Group I, a control group with no additions; Group II, loaded with nano-hydroxyapatite; Group III, loaded with chitosan; and Group IV, loaded with both nano-hydroxyapatite and chitosan. The new photoinitiating system consisted of riboflavin as a photoinitiator, dimethylaminoethyl methacrylate (DMAEMA) as a coinitiator (being used with riboflavin for the first time), and diphenyliodonium chloride as an accelerator. For each group, X-ray-diffraction, surface morphology by scanning electron microscope, mechanical properties, water uptake (%), and cell viability (%) were tested. The osteogenic potential was then tested in a rabbit model, and histomorphometric assessment was conducted.
Results
In the four groups, the light-cured hydrogels were obtained after a short irradiation time of 10 s using a dental light-curing unit. The prepared hydrogels were biocompatible. Simultaneous addition of nano-hydroxyapatite and chitosan increased the mechanical properties threefold and the osteogenic potential, twofold, with a statistically significant difference compared with the control group.
Conclusions
Light-cured hyaluronic acid composite hydrogels loaded with nano-hydroxyapatite and chitosan—prepared by using the new photoinitiating system—are promising materials that can be used in bone regeneration applications.
Filamentous fungi are competitive hosts for the production of drugs, proteins, and chemicals. However, their utility is limited by screening methods and low throughput. In this work, a universal high-throughput system for optimizing protein production in filamentous fungi was described. Droplet microfluidics was used to encapsulate large mutant strain pools in biocompatible core-shell microdroplets designed to avoid mycelial punctures and thus sustain prolonged culture. The self-assembled split GFP was then used to characterize the secretory capacity of the strains and isolate strains with superior production titers according to the fluorescence signals. The platform was applied to optimize the α-amylase secretion of Aspergillus niger, resulting in the isolation of a strain with 2.02-fold higher secretion capacity. The system allows the analysis of >105 single cells per h and will facilitate ultrahigh-throughput screening experiments of filamentous fungi. This method could help identify improved hosts for the large-scale production of biotechnology-relevant proteins. This is a broadly applicable system that can be equally used in other hosts.
A hydrogel is a three-dimensional (3D) network structure formed through polymer crosslinking, and these have emerged as a popular research topic in recent years. Hydrogel crosslinking can be classified as physical, chemical, or enzymatic, and photocrosslinking is a branch of chemical crosslinking. Compared with other methods, photocrosslinking can control the hydrogel crosslinking initiation, crosslinking time, and crosslinking strength using light. Owing to these properties, photocrosslinked hydrogels have important research prospects in tissue engineering, in situ gel formation, 3D bioprinting, and drug delivery. Methacrylic anhydride modification is a common method for imparting photocrosslinking properties to polymers, and graft-substituted polymers can be photocrosslinked under UV irradiation. In this review, we first introduce the characteristics of common natural polysaccharide- and protein-based hydrogels and the processes used for methacrylate group modification. Next, we discuss the applications of methacrylated natural hydrogels in tissue engineering. Finally, we summarize and discuss existing methacrylated natural hydrogels in terms of limitations and future developments. We expect that this review will help researchers in this field to better understand the synthesis of methacrylate-modified natural hydrogels and their applications in tissue engineering.
Unlabelled:
Gelatin-methacryloyl hydrogels (GelMA) have demonstrated their utility as scaffolds in a variety of tissue engineering applications.
Objectives:
In this study, a highly functionalized GelMA hydrogel was synthesized and assessed for degree of functionalization. As the proposed GelMA hydrogel was coupled to a visible-light photoinitiator, we hypothesized it might serve as base to formulate a model dentin primer for application in restorative dentistry.
Methods:
GelMA was mixed with photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), photopolymerized for 0-40 s using a dental light-curing device and tested for extrudability, degree of photo-crosslinking (DPxlink), water sorption/solubility/swelling (WS/SL/SW) and apparent modulus of elasticity (AE). Model dentin primer was prepared by mixing GelMA+LAP with a primer of a commercial three-step etch-and-rinse adhesive. After application of GelMA-based primer to acid-etched dentin, samples were bonded with correspondent adhesive agent, photopolymerized and had their immediate bond strength compared to control samples primed and bonded with the same commercial material.
Results:
Extrudability of hydrogel was confirmed using a microsyringe to write the acronym "CDMI". DPxlink of GelMA+LAP changed significantly as a function of photopolymerization time (20 s < 30 s ≤ 40 s). WS, SL and SW were significantly reduced in hydrogels polymerized for 30 and 40 s. AE of hydrogels varied significantly as a function of photopolymerization time (20 s < 30 s ≤ 40 s; 20 s ‡ 40 s). Bond strength of dentin primed with GelMA-based primer was lower (∼29.3 MPa) but not significantly of that of control (∼34.6 MPa).
Conclusions:
Optimization of a GelMA-based dentin primers can lead to the development of promising biomimetic adhesives for dentin rehabilitation.
The global demand for healthier food products has led to the introduction of functional foods fortified with bioactive agents, such as vitamins and nutraceuticals. Many of these bioactive agents have poor water-solubility, chemical stability, and bioavailability, which means delivery vehicles are needed to overcome these problems. The pharmaceutical industry also requires delivery vehicles to increase the administration and efficacy of drugs. In this study, we focus on the utilization of hydrogel-based delivery vehicles to overcome this problem. Hydrogels are formed by cross-linking biopolymer molecules to form a three-dimensional network that retains relatively high amounts of water. They can be used to encapsulate, protect, and control the release of bioactive agents in foods and pharmaceuticals. The functional attributes of hydrogels can be tuned for specific applications by manipulating their composition, dimensions, and internal structure. For instance, they can be designed to control the rate and extent of bioactive release under different conditions (such as pH, ionic strength, temperature, or enzyme activities). In this review, we discuss the structure, properties, and release mechanisms of hydrogels. In addition, we review previous applications of hydrogel-based delivery system for the targeted release of bioactive agents.
As our research on the physiopathology of intervertebral disc degeneration (IVD degeneration, IVDD) has advanced and tissue engineering has rapidly evolved, cell-, biomolecule- and nucleic acid-based hydrogel grafting strategies have been widely investigated for their ability to overcome the harsh microenvironment of IVDD. However, such single delivery systems suffer from excessive external dimensions, difficult performance control, the need for surgical implantation, and difficulty in eliminating degradation products. Stimulus-responsive composite hydrogels have good biocompatibility and controllable mechanical properties and can undergo solution-gel phase transition under certain conditions. Their combination with ready-to-use particles to form a multiscale delivery system may be a breakthrough for regenerative IVD strategies. In this paper, we focus on summarizing the progress of research on the stimulus response mechanisms of regenerative IVD-related biomaterials and their design as macro-, micro- and nanoparticles. Finally, we discuss multi-scale delivery systems as bioinks for bio-3D printing technology for customizing personalized artificial IVDs, which promises to take IVD regenerative strategies to new heights.
Although nano-systems can promote the cellular internalization, polymeric nanoparticles (NPs) are easily to be destroyed or eliminated in the gastrointestinal tract, which leads to premature drug release or insufficient colonic accumulation. A novel oral nano-in-micro system for the efficient colonic delivery of curcumin (Cur) was developed in this study, in which the hyaluronic acid (HA)/zein complex NPs loading Cur were embedded in alginate/chitosan hydrogel microparticles ([email protected]) with the electrospray technology. The [email protected] showed uniform-sized sphere with an average size of 218.36 ± 10 μm, encapsulating [email protected]/zein NPs with the average size of 148.64 ± 3.21 nm. Cur in NMPs showed the sustained drug-release profiles in simulated gastric fluid, whereas it showed the rapid release in simulated colonic fluid. Mediated by the HA-CD44 receptor recognition, [email protected]/zein NPs had the increased cellular uptake efficiency in macrophages. Furthermore, NMPs had the significant colon-retention and bio-adhesiveness capacity in colon tissues. As expected, the oral administration of [email protected] significantly mitigated colitis symptoms in DSS-induced UC mice by inhibiting TLR4/NF-κB pathway. The above results can provide a useful drug delivery strategy for Cur in the treatment of UC by retaining the advantages of nano- and micro-scaled carriers.
Soft biomaterials hold great potential for a plethora of biomedical applications because of their deformability, biodegradability, biocompatibility, high bioactivity, and low antigenicity. Multicomponent soft biomaterials are particularly attractive as a way of accommodating components made of different materials and generating combinative functions. Microfluidic technology has emerged as an outstanding tool in generating multicomponent materials with elaborate structures and constituents, in that it can manipulate multiphasic flows precisely on the micron scale. In recent decades, much progress has been achieved in the microfluidic fabrication of multicomponent soft biomaterials with finely defined physicochemical properties capable of controllable therapeutics delivery, three-dimensional (3D) cell culture, flexible devices and wearable electronics, and biosensing for molecules. In the paper, we summarize current progress in multicomponent soft biomaterials derived from microfluidics and emphasize their applications in biomedical fields. We also provide an outlook of the remaining challenges and future trends in this field.
Core-shell microparticles, composed of solid, liquid, or gas bubbles surrounded by a protective shell, are gaining considerable attention as intelligent and versatile carriers that show great potential in biomedical fields. In this review, an overview is given of recent developments in design and applications of biodegradable core-shell systems. Several emerging methodologies including self-assembly, gas-shearing, and coaxial electrospray are discussed and microfluidics technology is emphasized in detail. Furthermore, the characteristics of core-shell microparticles in artificial cells, drug release and cell culture applications are discussed and the superiority of these advanced multi-core microparticles for the generation of artificial cells is highlighted. Finally, the respective developing orientations and limitations inherent to these systems are addressed. It is hoped that this review can inspire researchers to propel the development of this field with new ideas.
Droplet microfluidic technology provides a new platform for controllable generation of microdroplets and droplet-derived materials. In particular, because of the ability in high-throughput production and accurate control of the size, structure, and function of these materials, droplet microfluidics presents unique advantages in the preparation of functional microcarriers, i.e., microsized liquid containers or solid particles that serve as substrates of biomolecules or cells. These microcarriers could be extensively applied in the areas of cell culture, tissue engineering, and drug delivery. In this review, we focus on the fabrication of microcarriers from droplet microfluidics, and discuss their applications in the biomedical field. We start with the basic principle of droplet microfluidics, including droplet generation regimes and its control methods. We then introduce the fabrication of biomedical microcarriers based on single, double, and multiple emulsion droplets, and emphasize the various applications of microcarriers in biomedical field, especially in 3D cell culture, drug development and biomedical detection. Finally, we conclude this review by discussing the limitations and challenges of droplet microfluidics in preparing microcarriers.
Statement of Significance
: Because of its precise control and high throughput, droplet microfluidics has been employed to generate functional microcarriers, which have been widely used in the areas of drug development, tissue engineering, and regenerative medicine. This review is significant because it emphasizes recent progress in research on droplet microfluidics in the preparation and application of biomedical microcarriers. In addition, this review suggests research directions for the future development of biomedical microcarriers based on droplet microfluidics by presenting existing shortcomings and challenges.
Drug releasing particles are valued for their ability to deliver therapeutics to targeted locations and for their controllable release patterns. The development of microfluidic technologies, which are designed specifically to manipulate small amounts of fluids, to manufacture particles for drug delivery applications reflects a recent trend due to the advantages they confer in terms of control over particle size and material composition. This review takes a comprehensive look at the different types of microfluidic devices used to fabricate such particles from different types of biomaterials, and at how the on-chip features enable the production of particles with different types of properties. The review concludes by suggesting avenues for future work that will enable these technologies to fulfill their potential and be used in industrial settings for the manufacture of drug releasing particles with unique capabilities.
The purpose of this study was to construct a delivery system using a microfluidic chip to protect procyanidins (PCs) and to achieve their pH-controlled release in simulated gastrointestinal fluid. The microfluidic chip was designed and fabricated to generate water-in-water-in-oil (W/W/O) templates for the preparation of sodium alginate/chitosan microparticles with a uniform size and core-shell structure, using an internal-external gelation method. Compared with free PCs, the stability of PCs embedded in microparticles was improved and a pH stimulus-responsive release of PCs from microparticles was observed under neutral pH conditions. The delivery system of microparticles was nontoxic and showed an inhibitory effect on the decrease of mitochondrial membrane potential in Caco-2 cells caused by H2O2 and acrylamide. This work provided a method for fabricating compact microfluidic chips to prepare a pH stimulus-responsive PCs delivery system with improved stability, which may have potential applications in the delivery of other nutrients.