Nanocolloidal hydrogel mimics the structure and nonlinear mechanical properties of biological fibrous networks.

Fibrous networks formed by biological polymers such as collagen or fibrin exhibit nonlinear mechanical behavior. They undergo strong stiffening in response to weak shear and elongational strains, but soften under compressional strain, in striking difference with the response to the deformation of flexible-strand networks formed by molecules. The nonlinear properties of fibrous networks are attributed to the mechanical asymmetry of the constituent filaments, for which a stretching modulus is significantly larger than the bending modulus. Studies of the nonlinear mechanical behavior are generally performed on hydrogels formed by biological polymers, which offers limited control over network architecture. Here, we report an engineered covalently cross-linked nanofibrillar hydrogel derived from cellulose nanocrystals and gelatin. The variation in hydrogel composition provided a broad-range change in its shear modulus. The hydrogel exhibited both shear-stiffening and compression-induced softening, in agreement with the predictions of the affine model. The threshold nonlinear stress and strain were universal for the hydrogels with different compositions, which suggested that nonlinear mechanical properties are general for networks formed by rigid filaments. The experimental results were in agreement with an affine model describing deformation of the network formed by rigid filaments. Our results lend insight into the structural features that govern the nonlinear biomechanics of fibrous networks and provide a platform for future studies of the biological impact of nonlinear mechanical properties.

Design, characterization and applications of nanocolloidal hydrogels.

Nanocolloidal gels (NCGs) are an emerging class of soft matter, in which nanoparticles act as building blocks of the colloidal network. Chemical or physical crosslinking enables NCG synthesis and assembly from a broad range of nanoparticles, polymers, and low-molecular weight molecules. The synergistic properties of NCGs are governed by nanoparticle composition, dimensions and shape, the mechanism of nanoparticle bonding, and the NCG architecture, as well as the nature of molecular crosslinkers. Nanocolloidal gels find applications in soft robotics, bioengineering, optically active coatings and sensors, optoelectronic devices, and absorbents. This review summarizes currently scattered aspects of NCG formation, properties, characterization, and applications. We describe the diversity of NCG building blocks, discuss the mechanisms of NCG formation, review characterization techniques, outline NCG fabrication and processing methods, and highlight most common NCG applications. The review is concluded with the discussion of perspectives in the design and development of NCGs.

Polymer-Tethered Nanoparticles: From Surface Engineering to Directional Self-Assembly.

Current interest in nanoparticle ensembles is motivated by their collective synergetic properties that are distinct from or better than those of individual nanoparticles and their bulk counterparts. These new advanced optical, electronic, magnetic, and catalytic properties can find applications in advanced nanomaterials and functional devices, if control is achieved over nanoparticle organization. Self-assembly offers a cost-efficient approach to produce ensembles of nanoparticles with well-defined and predictable structures. Nanoparticles functionalized with polymer molecules are promising building blocks for self-assembled nanostructures, due to the comparable dimensions of macromolecules and nanoparticles, the ability to synthesize polymers with various compositions, degrees of polymerization, and structures, and the ability of polymers to self-assemble in their own right. Moreover, polymer ligands can endow additional functionalities to nanoparticle assemblies, thus broadening the range of their applications.In this Account, we describe recent progress of our research groups in the development of new strategies for the self-assembly of nanoparticles tethered to macromolecules. At the beginning of our journey, we developed a new approach to patchy nanoparticles and their self-assembly. In a thermodynamically driven strategy, we used poor solvency conditions to induce homopolymer surface segregation in pinned micelles (patches). Patchy nanoparticles underwent self-assembly in a well-defined and controlled manner. Following this work, we overcame the limitation of low yield of the generation of patchy nanoparticles, by using block copolymer ligands. For block copolymer-capped nanoparticles, patch formation and self-assembly were "staged" by using distinct stimuli for each process. We expanded this work to the generation of patchy nanoparticles via dynamic exchange of block copolymer molecules between the nanoparticle surface and micelles in the solution. The scope of our work was further extended to a series of strategies that utilized the change in the configuration of block copolymer ligands during nanoparticle interactions. To this end, we explored the amphiphilicity of block copolymer-tethered nanoparticles and complementary interactions between reactive block copolymer ligands. Both approaches enabled exquisite control over directional and self-limiting self-assembly of complex hierarchical nanostructures. Next, we focused on the self-assembly of chiral nanostructures. To enable this goal, we attached chiral molecules to the surface of nanoparticles and organized these hybrid building blocks in ensembles with excellent chiroptical properties. In summary, our work enables surface engineering of polymer-capped nanoparticles and their controllable and predictable self-assembly. Future research in the field of nanoparticle self-assembly will include the development of effective characterization techniques, the synthesis of new functional polymers, and the development of environmentally responsive self-assembly of polymer-capped nanoparticles for the fabrication of nanomaterials with tailored functionalities.

Stimulus-Responsive Transport Properties of Nanocolloidal Hydrogels.

Applications of polymer hydrogels in separation technologies, environmental remediation, and drug delivery require control of hydrogel transport properties that are largely governed by the pore dimensions. Stimulus-responsive change in pore size offers the capability to change gel's transport properties "on demand". Here, we report a nanocolloidal hydrogel that exhibits temperature-controlled increase in pore size and, as a result, enhanced transport of encapsulated species from the gel. The hydrogel was formed by the covalent cross-linking of aldehyde-modified cellulose nanocrystals and chitosan carrying end-grafted poly(N-isopropylacrylamide) (pNIPAm) molecules. Owing to the temperature-mediated coil-to-globule transition of pNIPAm grafts, they acted as a temperature-responsive "gate" in the hydrogel. At elevated temperature, the size of the pores showed up to a 4-fold increase, with no significant changes in volume, in contrast with conventional pNIPAm-derived gels exhibiting a reduction in both pore size and volume in similar conditions. Temperature-mediated transport properties of the gel were explored by studying diffusion of nanoparticles with different dimensions from the gel, leading to the established correlation between the kinetics of diffusion-governed nanoparticle release and the ratio nanoparticle dimensions-to-pore size. The proposed approach to stimulus-responsive control of hydrogel transport properties has many applications, including their use in nanomedicine and tissue engineering.

Trends in Droplet Microfluidics: From Droplet Generation to Biomedical Applications.

Over the past decade, droplet microfluidics has attracted growing interest in biology, medicine, and engineering. In this feature article, we review the advances in droplet microfluidics, primarily focusing on the research conducted by our group. Starting from the introduction to the mechanisms of microfluidic droplet formation and the strategies for cell encapsulation in droplets, we then focus on droplet transformation into microgels. Furthermore, we review three biomedical applications of droplet microfluidics, that is, 3D cell culture, single-cell analysis, and in vitro organ and disease modeling. We conclude with our perspective on future directions in the development of droplet microfluidics for biomedical applications.

Stimulus-Responsive Nanoconjugates Derived from Phytoglycogen Nanoparticles.

Plant-derived phytoglycogen nanoparticles (PhG NPs) have the advantages of size uniformity, dispersibility in water, excellent lubrication properties, and lack of cytotoxicity; however, their chemical functionalization may lead to loss of NP structural integrity. Here, we report a straightforward approach to the generation of PhG NP conjugates with biologically active molecules. Hydrogen bonding of bovine serum albumin with electroneutral PhG NPs endows them with additional ligand binding affinity and enables the electrostatically governed attachment of methotrexate (MTX), a therapeutic agent commonly used in the treatment of cancer and arthritis diseases, to the protein-capped NPs. We showed stimuli-responsive release of MTX from the PhG-based nanoconjugates under physiological cues such as temperature and ionic strength. The results of this study stimulate future exploration of biomedical applications of nanoconjugates of PhG NPs.

Surface patterning of nanoparticles with polymer patches.

Patterning of colloidal particles with chemically or topographically distinct surface domains (patches) has attracted intense research interest. Surface-patterned particles act as colloidal analogues of atoms and molecules, serve as model systems in studies of phase transitions in liquid systems, behave as 'colloidal surfactants' and function as templates for the synthesis of hybrid particles. The generation of micrometre- and submicrometre-sized patchy colloids is now efficient, but surface patterning of inorganic colloidal nanoparticles with dimensions of the order of tens of nanometres is uncommon. Such nanoparticles exhibit size- and shape-dependent optical, electronic and magnetic properties, and their assemblies show new collective properties. At present, nanoparticle patterning is limited to the generation of two-patch nanoparticles, and nanoparticles with surface ripples or a 'raspberry' surface morphology. Here we demonstrate nanoparticle surface patterning, which utilizes thermodynamically driven segregation of polymer ligands from a uniform polymer brush into surface-pinned micelles following a change in solvent quality. Patch formation is reversible but can be permanently preserved using a photocrosslinking step. The methodology offers the ability to control the dimensions of patches, their spatial distribution and the number of patches per nanoparticle, in agreement with a theoretical model. The versatility of the strategy is demonstrated by patterning nanoparticles with different dimensions, shapes and compositions, tethered with various types of polymers and subjected to different external stimuli. These patchy nanocolloids have potential applications in fundamental research, the self-assembly of nanomaterials, diagnostics, sensing and colloidal stabilization.

Enhanced electrocatalytic CO2 reduction via field-induced reagent concentration.

Electrochemical reduction of carbon dioxide (CO2) to carbon monoxide (CO) is the first step in the synthesis of more complex carbon-based fuels and feedstocks using renewable electricity. Unfortunately, the reaction suffers from slow kinetics owing to the low local concentration of CO2 surrounding typical CO2 reduction reaction catalysts. Alkali metal cations are known to overcome this limitation through non-covalent interactions with adsorbed reagent species, but the effect is restricted by the solubility of relevant salts. Large applied electrode potentials can also enhance CO2 adsorption, but this comes at the cost of increased hydrogen (H2) evolution. Here we report that nanostructured electrodes produce, at low applied overpotentials, local high electric fields that concentrate electrolyte cations, which in turn leads to a high local concentration of CO2 close to the active CO2 reduction reaction surface. Simulations reveal tenfold higher electric fields associated with metallic nanometre-sized tips compared to quasi-planar electrode regions, and measurements using gold nanoneedles confirm a field-induced reagent concentration that enables the CO2 reduction reaction to proceed with a geometric current density for CO of 22 milliamperes per square centimetre at -0.35 volts (overpotential of 0.24 volts). This performance surpasses by an order of magnitude the performance of the best gold nanorods, nanoparticles and oxide-derived noble metal catalysts. Similarly designed palladium nanoneedle electrocatalysts produce formate with a Faradaic efficiency of more than 90 per cent and an unprecedented geometric current density for formate of 10 milliamperes per square centimetre at -0.2 volts, demonstrating the wider applicability of the field-induced reagent concentration concept.

Oxidative Elimination and Reductive Addition of Thiol-Terminated Polymer Ligands to Metal Nanoparticles.

Metal nanoparticles (NPs) stabilized with thiol- (HS-) terminated polymers have applications in medicine, optoelectronics, and catalysis. It is assumed that upon exposure to oxidants or even air, these NPs lose colloidal stability, due to the oxidation of the HS-end-group and elimination of polymer ligands from the NP surface, however, this mechanism does not explain the unsuccessful recovery of the NP stability by adding fresh HS-terminated polymers. Here we propose the oxidation of the surface metal atoms as a mechanism for the oxidative elimination of polymer from the NP surface. Based on this mechanism, we reversed NP aggregation by reducing the oxidized metal surface and re-attaching HS-terminated polymer ligands. This mechanism is general for various metal NPs and different HS-terminated polymers. We show that oxidative elimination and reductive addition reactions can improve the colloidal stability of polymer-capped metal NPs and control their redox stimuli-responsive self-assembly.

Nanofibrillar Hydrogel Recapitulates Changes Occurring in the Fibrotic Extracellular Matrix.

Fibrosis is a pathological condition that leads to excessive deposition of collagen and increased tissue stiffness. Understanding the mechanobiology of fibrotic tissue necessitates the development of effective in vitro models that recapitulate its properties and structure; however, hydrogels that are currently used for this purpose fail to mimic the filamentous structure and mechanical properties of the fibrotic extracellular matrix (ECM). Here, we report a nanofibrillar hydrogel composed of cellulose nanocrystals and gelatin, which addresses this challenge. By altering the composition of the hydrogel, we mimicked the changes in structure, mechanical properties, and chemistry of fibrotic ECM. Furthermore, we decoupled the variations in hydrogel structure, properties, and ligand concentration. We demonstrate that this biocompatible hydrogel supports the three-dimensional culture of cells relevant to fibrotic diseases. This versatile hydrogel can be used for in vitro studies of fibrosis of different tissues, thus enabling the development of novel treatments for fibrotic diseases.

Composite Microgels for Imaging-Monitored Tracking of the Delivery of Vascular Endothelial Growth Factor to Ischemic Muscles.

Monitoring the supply of vascular endothelial growth factor (VEGF) to ischemic tissues provides information on its biodistribution and delivery to meet the requirements of therapeutic angiogenesis and tissue engineering applications. We herein report the use of microfluidically generated microgels containing VEGF-conjugated fluorescent carbon dots (CDs) (VEGF-CDs), a gelatin-phenol conjugate, and silk fibroin for imaging-monitored tracking of VEGF delivery to ischemic muscles. An in vitro release study and a bioactivity assay indicated that the VEGF-CDs were released in a sustained manner with high bioactivity. The microgels showed a high angiogenesis potential, along with a strong fluorescent signal, for the chicken chorioallantoic membrane and chick embryo. Imaging and studies of therapeutic modalities of the composite microgels indicated their effective localization in ischemic tissues and sustained VEGF release, which resulted in enhanced therapeutic angiogenesis of ischemic muscles. This work reveals the success of using VEGF-loaded composite polymer microgels for efficient and monitored VEGF delivery by intramuscular administration for ischemic disease treatment.

Matrix Stiffness-Regulated Growth of Breast Tumor Spheroids and Their Response to Chemotherapy.

Interactions between tumor cells and the extracellular matrix (ECM) are an important factor contributing to therapy failure in cancer patients. Current in vitro breast cancer spheroid models examining the role of mechanical properties on spheroid response to chemotherapy are limited by the use of two-dimensional cell culture, as well as simultaneous variation in hydrogel matrix stiffness and other properties, e.g., hydrogel composition, pore size, and cell adhesion ligand density. In addition, currently used hydrogel matrices do not replicate the filamentous ECM architecture in a breast tumor microenvironment. Here, we report a collagen-alginate hydrogel with a filamentous architecture and a 20-fold variation in stiffness, achieved independently of other properties, used for the evaluation of estrogen receptor-positive breast cancer spheroid response to doxorubicin. The variation in hydrogel mechanical properties was achieved by altering the degree of cross-linking of alginate molecules. We show that soft hydrogels promote the growth of larger MCF-7 tumor spheroids with a lower fraction of proliferating cells and enhance spheroid resistance to doxorubicin. Notably, the stiffness-dependent chemotherapeutic response of the spheroids was temporally mediated: it became apparent at sufficiently long cell culture times, when the matrix stiffness has influenced the spheroid growth. These findings highlight the significance of decoupling matrix stiffness from other characteristics in studies of chemotherapeutic resistance of tumor spheroids and in development of drug screening platforms.

Carbon Dots Conjugated with Vascular Endothelial Growth Factor for Protein Tracking in Angiogenic Therapy.

One of the challenges of using growth factors for tissue regeneration is to monitor their biodistributions and delivery to injured tissues for minimally invasive detection. In the present study, tracking of human vascular endothelial growth factor (VEGF) was achieved by chemically linking it to photoluminescent carbon dots (CDs). Carbon dots were synthesized by the hydrothermal method and, subsequently, conjugated with VEGF using carbodiimide coupling. ELISA and western blot analysis revealed that VEGF-conjugated CDs preserve the binding affinity of VEGF to its antibodies. We also show that VEGF-conjugated CDs maintain the functionality of VEGF for tube formation and cell migration. The VEGF-conjugated CDs were also used for in vitro imaging of human umbilical vein endothelial cells. The results of this work suggest that cell-penetrating VEGF-conjugated CDs can be used for growth factor protein tracking in therapeutic and tissue engineering applications.

Periodic assembly of nanoparticle arrays in disclinations of cholesteric liquid crystals.

An important goal of the modern soft matter science is to discover new self-assembly modalities to precisely control the placement of small particles in space. Spatial inhomogeneity of liquid crystals offers the capability to organize colloids in certain regions such as the cores of the topological defects. Here we report two self-assembly modes of nanoparticles in linear defects-disclinations in a lyotropic colloidal cholesteric liquid crystal: a continuous helicoidal thread and a periodic array of discrete beads. The beads form one-dimensional arrays with a periodicity that matches half a pitch of the cholesteric phase. The periodic assembly is governed by the anisotropic surface tension and elasticity at the interface of beads with the liquid crystal. This mode of self-assembly of nanoparticles in disclinations expands our ability to use topological defects in liquid crystals as templates for the organization of nanocolloids.

Helicoidal Patterning of Gold Nanorods by Phase Separation in Mixed Polymer Brushes.

The spatial distribution of polymer ligands on the surface of nanoparticles (NPs) is of great importance because it determines their interactions with each other and with the surrounding environment. Phase separation in mixtures of polymer brushes has been studied for spherical NPs; however, the role of local surface curvature of nonspherical NPs in the surface phase separation of end-grafted polymer ligands remains an open question. Here, we examined phase separation in mixed monolayers of incompatible polystyrene and poly(ethylene glycol) brushes end-capping the surface of gold nanorods in a good solvent. By varying the molar ratio between these polymers, we generated a range of surface patterns, including uniform and nonuniform polystyrene shells, randomly distributed polystyrene surface patches, and, most interestingly, a helicoidal pattern of polystyrene patches wrapping around the nanorods. The helicoidally patterned nanorods exhibited long-term colloidal stability in a good solvent. The helicoidal wrapping of the nanorods was achieved for the mixtures of polymers with different molecular weights and preserved when the quality of the solvent for the polymers was reduced. The helicoidal organization of polymer patches on the surface of nanorods can be used for templating the synthesis or self-assembly of helicoidal multicomponent nanomaterials.

Staged Surface Patterning and Self-Assembly of Nanoparticles Functionalized with End-Grafted Block Copolymer Ligands.

Using two orthogonal external stimuli, programmable staged surface patterning and self-assembly of inorganic nanoparticles (NPs) was achieved. For gold NPs capped with end-grafted poly(styrene-block-(4-vinylbenzoic acid)), P(St-block-4VBA), block copolymer ligands, surface-pinned micelles (patches) formed from NP-adjacent PSt blocks under reduced solvency conditions (Stimulus 1); solvated NP-remote P(4VBA) blocks stabilized the NPs against aggregation. Subsequent self-assembly of patchy NPs was triggered by crosslinking the P(4VBA) blocks with copper(II) ions (Stimulus 2). Block copolymer ligand design has a strong effect on NP self-assembly. Small, well-defined clusters assembled from NPs functionalized with ligands with a short P(4VBA) block, while NPs tethered with ligands with a long P(4VBA) block formed large irregularly shaped assemblies. This approach is promising for high-yield fabrication of colloidal molecules and their assemblies with structural and functional complexity.

Helicoidal Patterning of Nanorods with Polymer Ligands.

Chiral packing of ligands on the surface of nanoparticles (NPs) is of fundamental and practical importance, as it determines how NPs interact with each other and with the molecular world. Herein, for gold nanorods (NRs) capped with end-grafted nonchiral polymer ligands, we show a new mechanism of chiral surface patterning. Under poor solvency conditions, a smooth polymer layer segregates into helicoidally organized surface-pinned micelles (patches). The helicoidal morphology is dictated by the polymer grafting density and the ratio of the polymer ligand length to nanorod radius. Outside this specific parameter space, a range of polymer surface structures was observed, including random, shish-kebab, and hybrid patches, as well as a smooth polymer layer. We characterize polymer surface morphology by theoretical and experimental state diagrams. The helicoidally organized polymer patches on the NR surface can be used as a template for the helicoidal organization of other NPs, masked synthesis on the NR surface, as well as the exploration of new NP self-assembly modes.

Shear-Induced Alignment of Anisotropic Nanoparticles in a Single-Droplet Oscillatory Microfluidic Platform.

Flow-induced alignment of shape-anisotropic colloidal particles is of great importance in fundamental research and in the fabrication of structurally anisotropic materials; however, rheo-optical studies of shear-induced particle orientation are time- and labor-intensive and require complicated experimental setups. We report a single-droplet oscillatory microfluidic strategy integrated with in-line polarized light imaging as a strategy for studies of shear-induced alignment of rod-shape nanoparticles. Using an oscillating droplet of an aqueous isotropic suspension of cellulose nanocrystals (CNCs), we explore the effect of the shear rate and suspension viscosity on the flow-induced CNC alignment and subsequent relaxation to the isotropic state. The proposed microfluidic strategy enables high-throughput studies of shear-induced orientations in structured liquid under precisely controlled experimental conditions. The results of such studies can be used in the development of structure-anisotropic materials.

Patterning of Structurally Anisotropic Composite Hydrogel Sheets.

Compositional and structural patterns play a crucial role in the function of many biological tissues. In the present work, for nanofibrillar hydrogels formed by chemically cross-linked cellulose nanocrystals (CNC) and gelatin, we report a microextrusion-based 3D printing method to generate structurally anisotropic hydrogel sheets with CNCs aligned in the direction of extrusion. We prepared hydrogels with a uniform composition, as well as hydrogels with two different types of compositional gradients. In the first type of gradient hydrogel, the composition of the sheet varied parallel to the direction of CNC alignment. In the second hydrogel type, the composition of the sheet changed orthogonally to the direction of CNC alignment. The hydrogels exhibited gradients in structure, mechanical properties, and permeability, all governed by the compositional patterns, as well as cytocompatibility. These hydrogels have promising applications for both fundamental research and for tissue engineering and regenerative medicine.

Compound droplets derived from a cholesteric suspension of cellulose nanocrystals.

This study reports microfluidic generation of Janus droplets that consist of a liquid crystal component (a cholesteric aqueous suspension of cellulose nanocrystals, Ch-CNC) and a mineral oil (MO) component. The composition of the droplets was controlled by varying the relative flow rates of MO and Ch-CNC suspension. The shape of the Ch-CNC component of the droplets was changed from a truncated sphere to a hemisphere to a crescent moon. For these three Ch-CNC phase shapes, the Ch packing of the CNC pseudolayers was preserved, however the Ch pitch reduced, which was ascribed to the change in CNC orientation at the Ch-CNC/MO interface from perpendicular to parallel. The shape of the compound droplets was tuned from a dumbbell to a sphere by reducing interfacial tension between the Ch-CNC suspension and MO phases. Photopolymerization of the monomer mixture introduced in the Ch-CNC phase of the droplets and the removal of the sacrificial MO phase enabled the generation of Ch microgels. The results of this work can be used for exploring new applications of Janus colloids and the fabrication of programmable active soft matter.