Switchable Enzyme/DNAzyme Cascades by the Reconfiguration of DNA Nanostructures

Mimicking cellular transformations and signal transduction pathways by means of biocatalytic cascades proceeding in organized media is a scientific challenge. We describe two DNA machines that enable the “ON/OFF” switchable activation and deactivation of three‐component biocatalytic cascades. One system consists of a reconfigurable DNA tweezers‐type structure, whereas in the second system the catalytic cascade proceeds on a switchable DNA clamp scaffold. The three‐component catalytic cascades consist of β‐galactosidase (β‐Gal), glucose oxidase (GOx), and the K⁺‐ion‐stabilized hemin‐G‐quadruplex horseradish peroxidase (HRP)‐mimicking DNAzyme. The hemin‐G‐quadruplex‐bridged closed structure of the tweezers or clamp allows the biocatalytic cascades to operate (switched “ON′′), whereas separation of the hemin‐G‐quadruplex by means of 18‐crown‐6‐ether opens the tweezers/clamp structures, thus blocking the catalytic cascade (switched ”OFF“). This study is complemented by two‐component, switchable biocatalytic cascades composed of GOx and hemin‐G‐quadruplex assembled on hairpin‐bridged DNA tweezers or clamp nanostructures.     

Redox-responsive supramolecular polymeric networks having double-threaded inclusion complexes

Stimuli-responsive hydrogels have attracted attention as soft actuators that act similarly to muscles. In this work, hydrogel actuators controlled by host–guest interactions have been developed. The introduction of a 1:1 inclusion complex into a hydrogel is a popular design for achieving a change in cross-linking density. To realize faster and larger deformation properties, the introduction of a 1:2 inclusion complex is effective because the alteration in cross-linking density in a hydrogel with 1:2 complexes is larger than that in a hydrogel with 1:1 complexes. A redox-responsive hydrogel actuator cross-linked with 1:2 inclusion complexes is designed, where γ-cyclodextrin (γCD) and viologens modified with an alkyl chain derivative (VC11) were employed as the host and guest units, respectively. γCD includes two VC11 molecules in its cavity. The obtained γCD–VC11 hydrogel cross-linked with the 1:2 complex showed faster and larger deformation behaviour than the αCD–VC11 and the βCD–VC11 hydrogels cross-linked with a 1:1 complex. The deformation ratio and response speed of the γCD–VC11 hydrogel, which forms a supramolecular cross-linking structure by stimuli, are 3 and 11 times larger, respectively, than those of our previous hydrogel consisting of a βCD/ferrocene 1:1 inclusion complex.

A hydrogel actuator with a 1:2 host–guest complex controlled by redox stimuli has been developed to realize faster and larger deformation.     

Directional molecular sliding movement in peptide hydrogels accelerates cell proliferation

Adjusting the mechanical cues generated in cellular microenvironments is important for manipulating cell behaviour. Here we report on mechanically dynamic hydrogels undergoing directional domain sliding motion and investigate the effect of the well-defined mechanical motion on accelerating cell proliferation. The mechanically dynamic hydrogels were prepared via self-assembly of an amphiphilic peptide consisting of two alternating polar and nonpolar domains cross-linked by disulfide bonds at a nonsymmetrical position. The cross-linked peptide assembled into entangled nanofibers driven by the hydrophobic collapse involving a partial-length sequence due to the covalent constraint. Reduction of the disulfide bonds led to formation of non-equilibrated peptide bilayers, which underwent directional domain sliding motion along each promoted by the thermodynamically favourable transition from the partial to full hydrophobic collapse. The mechanical cues resulting from the directional domain sliding motion within the mechanically dynamic hydrogels accelerated cell proliferation when incubating cells on the hydrogel, compared to the thermodynamically static counterparts, via a mechanotransduction mechanism as supported by the facilitated translocation of yes-associated proteins into the nucleus of the cells. Our finding demonstrates the great potential of mechanically dynamic hydrogels as new-generation biomimetic extracellular matrices in tissue engineering and regeneration.

Dynamic peptide hydrogels undergoing directional domain sliding movement upon release of covalent constraint accelerate cell proliferation through a mechanotransduction pathway.     

pH-responsive and switchable triplex-based DNA hydrogels

New methods for the preparation of reversible pH-responsive DNA hydrogels based on Hoogsteen triplex structures are described. One system consists of a hydrogel composed of duplex DNA units that bridge acrylamide chains at pH = 7.4 and undergoes dissolution at pH = 5.0 through the reconfiguration of one of the duplex bridging units into a protonated CG·C⁺ triplex structure. The second system consists of a hydrogel consisting of acrylamide chains crosslinked in the presence of an auxiliary strand by Hoogsteen TA·T triplex interaction at pH = 7.0. The hydrogel transforms into a liquid phase at pH = 10.0 due to the separation of the triplex bridging units. The two hydrogel systems undergo reversible and cyclic hydrogel/solution transitions by subjecting the systems to appropriate pH values. The anti-cancer drug, coralyne, binds specifically to the TA·T triplex-crosslinked hydrogel thereby increasing its stiffness. The pH-controlled release of the coralyne from the hydrogel is demonstrated.     

Dynamic,multimodal hydrogel actuators using porphyrin-based visible light photoredox catalysis in a thermoresponsive polymer network

Hydrogels that can respond to multiple external stimuli represent the next generation of advanced functional biomaterials. Here, a series of multimodal hydrogels were synthesized that can contract and expand reversibly over several cycles while changing their mechanical properties in response to blue and red light, as well as heat (∼50 °C). The light-responsive behavior was achieved through a photoredox-based mechanism consisting of photoinduced electron transfer from a zinc porphyrin photocatalyst in its excited state to oligoviologen-based macrocrosslinkers, both of which were integrated into the hydrogel polymer network during gel formation. Orthogonal thermoresponsive properties were also realized by introducing N-isopropyl acrylamide (NIPAM) monomer simultaneously with hydroxyethyl acrylate (HEA) in the pre-gel mixture to produce a statistical 60 : 40 HEA : NIPAM polymer network. The resultant hydrogel actuators – crosslinked with either a styrenated viologen dimer (2V4+-St) or hexamer (6V12+-St) – were exposed to red or blue light, or heat, for up to 5 h, and their rate of contraction, as well as the corresponding changes in their physical properties (i.e., stiffness, tensile strength, Young''s modulus, etc.), were measured. The combined application of blue light and heat to the 6V12+-St-based hydrogels was also demonstrated, resulting in hydrogels with more than two-fold faster contraction kinetics and dramatically enhanced mechanical robustness when fully contracted. We envision that the reported materials and the corresponding methods of remotely manipulating the dynamic hydrogels may serve as a useful blueprint for future adaptive materials used in biomedical applications.

Orthogonal modes of activation in thermoresponsive hydrogel actuators using porphyrin-based visible light photoredox catalysis, viologen-based crosslinkers, and poly(N-isopropylacrylamide).     

双重响应性生物基自修复凝胶的制备及其性能

为拓宽多重响应性凝胶在生物医学领域中的应用,本文基于生物大分子构筑具有pH响应、糖响应性的可自修复性水凝胶。 本文选用3-氨基苯硼酸(APBA)和2,3-环氧丙基三甲基氯化铵(CHGTA)分别对聚谷氨酸(γ-PGA)和瓜尔胶(GG)进行改性制备了聚谷氨酸-g-氨基苯硼酸(γ-PGA-g-APBA)和阳离子瓜尔胶,在此基础上,对γ-PGA-g-APBA和阳离子瓜尔胶进行物理共混制备生物基凝胶。 通过傅里叶变换红外光谱仪(FTIR)、核磁共振波谱仪(¹H NMR)和流变仪对聚合物化学结构、接枝率、流变性能和力学性能进行表征,并考察了凝胶在不同pH值及糖浓度下刺激响应性。 结果表明,凝胶具有自修复性,修复效率可达100%;具有pH响应性,在环境pH值较高时更易形成凝胶,且凝胶强度随pH值升高而增大;同时所制凝胶具有糖响应性,在4 g/L的葡萄糖溶液中浸泡后即可导致凝胶解体。 这些结果说明功能基团APBA的引入可赋予凝胶多重响应性。 所制的双重响应性生物基凝胶具有良好的生物相容性,有望应用于生物医学、功能器件、传感等领域。     

Preparation of a novel pH-responsive silver nanoparticle/poly(HEMA-PEGMA-MAA) composite hydrogel

In this paper, series of novel pH-responsive silver (Ag) nanoparticle/poly(2-hydroxyethyl methacrylate (HEMA)-poly(ethylene glycol) methyl ether methacrylate (PEGMA)-methacrylic acid (MAA)) composite hydrogel were successfully prepared by in situ reducing Ag⁺ ions anchored in the hydrogel by the deprotonized carboxyl acid groups. X-ray diffraction (XRD), UV-vis spectrophotometry, transmission electron microscopy (TEM) and electric conductivity tests were used to characterize the composite system. It was found that the size and morphology of the reduced Ag nanoparticles in the composite hydrogels could be changed by loading the Ag⁺ ions at various swelling ratios of hydrogel. Moreover, compared to the pure poly(HEMA-PEGMA-MAA) hydrogel, not only did the Ag nanoparticle/poly(HEMA-PEGMA-MAA) composite hydrogels exhibit much higher swelling ratio and faster deswelling rate, but also higher pH switchable electrical properties upon controlling the interparticle distance under pH stimulus. The pH responsive nanocomposite hydrogel reported here might be a potentially smart material in the range of applications including electronics, biosensors and drug-delivery devices.     

Light-driven release of cucurbit[8]uril from a bivalent cage

Temporal control over supramolecular systems has great potential for the modulation of binding and assembly events, such as providing orthogonal control over protein activity. Especially light controlled triggering provides unique entries for supramolecular systems to interface in a controlled manner with enzymes. Here we report on the light-induced release of cucurbit[8]uril (CB[8]) from a bivalent cage molecule and its subsequent activation of a proteolytic enzyme, caspase-9, that itself is unresponsive to light. Central to the design is the bivalent binding of the cage with high affinity to CB[8], 100-fold stronger than the UV-inactivated products. The affinity switching occurs in the (sub-)micromolar concentration regime, matching the concentration characteristics required for dimerizing and activating caspase-9 by CB[8]. The light-responsive caged CB[8] concept presented offers a novel platform for tuning and application of switchable cucurbiturils and beyond.

Photo-switchable supramolecular systems offer unique entries to control biomolecular process, as illustrated via the light-induced release of cucurbit[8]uril from a bivalent cage molecule and its subsequent activation of the caspase-9 enzyme.     

“Smart” carboxymethylchitosan hydrogels crosslinked with poly(N-isopropylacrylamide) and poly(acrylic acid) for controlled drug release

Carboxymethylchitosan (CMC) hydrogels containing thermo-responsive poly(N-isopropylacrylamide) (poly(NIPAAm)) and pH-responsive poly(acrylic acid) (poly(AA)) were prepared via a free radical polymerization in the presence of hexamethylene-1,6-di-(aminocarboxysulfonate) crosslinking agents. A proper ratio of CMC to NIPAAm and AA used in the reaction was investigated such that the thermo- and pH-responsive properties of the hydrogels were obtained. Water swelling of the hydrogels was improved when the solution pH was in basic conditions (pH 10) or the temperature was below its lower critical solution temperature (LCST). Effects of the change in solution temperature and pH on water swelling properties of the hydrogel as well as the releasing rate of an entrapped drug were also investigated. The hydrogels were not toxic and showed antibacterial activity against Straphylococcus aureus (S. aureus). The pH- and thermo-responsive properties of this novel “smart” hydrogel might be efficiently used as dual triggering mechanisms in controlled drug release applications.     

Tunable biohybrid hydrogels from coacervation of hyaluronic acid and PEO-based block copolymers

Accurately tuning the macroscopic properties of biopolymer-based hydrogels remains challenging due to the ill-defined molecular architecture of the natural building blocks. Here, we report a biohybrid coacervate hydrogel, combining the biocompatibility and biodegradability of naturally occurring hyaluronic acid (HA) with the tunability of a synthetic polyethylene oxide (PEO) -based ABA-triblock copolymer. Coacervation of the cationic ammonium or guanidinium-functionalized copolymer A-blocks with the anionic HA leads to hydrogel formation. Both mechanical properties and water content of the self-healing hydrogels can be controlled independently by altering the copolymer structure. By controlling the strength of the interaction between the polymer network and small-molecule cargo, both release rate and maximum release are controlled. Finally, we show that coacervation of HA and the triblock copolymer leads to increased biostability upon exposure to hyaluronidase. We envision that noncovalent crosslinking of HA hydrogels through coacervation is an attractive strategy for the facile synthesis of tunable hydrogels for biomedical applications.     

基于动态共价键的自愈性水凝胶及其在医学领域的应用

自愈性水凝胶作为一种新型仿生智能材料受到了科研人员的广泛关注.近年来,人们利用动态共价键、超分子作用,发展了一系列自愈性水凝胶,并将其应用于药物控释、细胞三维培养、组织工程等生物医用领域.本文总结和评述了基于动态共价键的自愈性水凝胶及这些水凝胶作为药物载体的相关研究,并展望了基于动态化学的自愈性水凝胶的未来发展.     

Head group modulated pH-responsive hydrogel of amino acid-based amphiphiles: entrapment and release of cytochrome c and vitamin B12

The present study describes the rational design and synthesis of amino acid-based amphiphilic hydrogelators, which were systemically fine-tuned at the head group to develop pH-responsive hydrogels. To understand the basic structural requirements of a low molecular weight amphiphilic hydrogelator, 10 analogous amphiphiles based on L-phenylalanine and L-tyrosine with structurally related head group were synthesized. Among them, three with quaternary ammonium substitution at the head group formed transparent hydrogels at room temperature while others were unable to gelate water. To establish correlations between the head group architecture of the gelators and their supramolecular arrangements, a variety of spectroscopic and microscopic techniques were investigated that showed that a balance between hydrophilicity and hydrophobicity is required to achieve hydrogelation. Interestingly, the gelator with tyrosinate in its head group showed remarkable response toward external pH. All hydrogels including the pH-responsive one were used in the controlled and/or pH-triggered release of entrapped (with in hydrogels) vitamin B12 and cytochrome c at different pHs. Since the hydrogels were formed at room temperature without heating, this could be very important during the entrapment of biomolecules such as proteins because of their heat sensitivity. At biological pH (7.4), the release of entrapped biomolecules from all three hydrogels was caused by diffusion through the gel structure, but at endosomal pH (approximately 5.5) and further lower pH, the release rate of biomolecules from the pH-responsive hydrogel with tyrosinate head group (pKa approximately equal to 7.2) increased by 9-10-fold compared to that observed at physiological pH, because of gel dissolution. Retention of the structure and activity of released biomolecule has established the prospect of the hydrogel as an efficient drug delivery vehicle.     

Thermo- and pH-Responsive Gelatin/Polyphenolic Tannin/Graphene Oxide Hydrogels for Efficient Methylene Blue Delivery

Gelatin (GE), amino-functionalized polyphenolic tannin derivative (TN), and graphene oxide (GO) were associated to yield thermo- and pH-responsive hydrogels for the first time. Durable hydrogel assemblies for drug delivery purposes were developed using the photosensitizer methylene blue (MB) as a drug model. The cooling GE/TN blends provide brittle physical assemblies. To overcome this disadvantage, different GO contents (between 0.31% and 1.02% wt/wt) were added to the GE/TN blend at 89.7/10.3 wt/wt. FTIR and RAMAN spectroscopy analyses characterized the materials, indicating GO presence in the hydrogels. Incorporation studies revealed a total MB (0.50 mg/mL) incorporation into the GE/TN-GO hydrogel matrices. Additionally, the proposed systems present a mechanical behavior similar to gel. The GO presence in the hydrogel matrices increased the elastic modulus from 516 to 1650 Pa. SEM revealed that hydrogels containing MB present higher porosity with interconnected pores. Dissolution and swelling degree studies revealed less stability of the GE/TN-GO-MB hydrogels in SGF medium (pH 1.2) than SIF (pH 6.8). The degradation increased in SIF with the GO content, making the polymeric matrices more hydrophilic. MB release studies revealed a process controlled by Fickian diffusion. Our results point out the pH-responsible behavior of mechanically reinforced GE/TN-GO-MB hydrogels for drug delivery systems purposes.     

Spatial and temporal diffusion-control of dynamic multi-domain self-assembled gels

The dynamic assembly of a pH-responsive low-molecular-weight gelator (LMWG) within the pre-formed matrix of a second LMWG has been achieved via diffusion of an acid from a reservoir cut into the gel. Self-assembly of the acid-triggered LMWG as it converts from micellar aggregates at basic pH into gel nanofibers at lower pH values can be both spatially and temporally controlled. The pH-responsive LMWG has an impact on the stiffness of the pre-formed gel in the domains in which it assembles. When low acid concentrations are used, LMWG assembly is transient – after the initial proton diffusion phase, the pH rises and disassembly occurs as the system equilibrates. Re-application of additional acid as ‘fuel’ can then re-assemble the LMWG network. Using glucono-δ-lactone (which slowly hydrolyses to gluconic acid) instead of HCl gives slower, more spatially-restricted assembly, and creates longer-lasting pH gradients within the gel. The presence of an agarose polymer gel network improves the mechanical strength of the gels and appears to slightly enhance the rate of proton diffusion. More sophisticated reservoir shapes can be cut into these more mechanically robust gels, enabling the creation of diffusion waves with different geometries, and hence different patterns of LMWG activation. Multiple reservoirs can be used to create overlapping proton diffusion waves, hence achieving differentiated pH patterns in the gel. Using acid diffusion in this way within gels is an intriguing and powerful way of dynamic patterning. The ability to temporally-evolve spatially-resolved patterns using biocompatible weak acids, and the change in rheological performance of the triggered domains, suggest potential future applications of this strategy in tissue engineering.

The assembly of a pH-sensitive LMWG within a pre-formed network of a second LMWG can be achieved by diffusing acids from pre-cut reservoirs, giving rise to patterned gels in which the rheological properties evolve with spatial and temporal control.     

In vivo oral insulin delivery via covalent organic frameworks

With diabetes being the 7th leading cause of death worldwide, overcoming issues limiting the oral administration of insulin is of global significance. The development of imine-linked-covalent organic framework (nCOF) nanoparticles for oral insulin delivery to overcome these delivery barriers is herein reported. A gastro-resistant nCOF was prepared from layered nanosheets with insulin loaded between the nanosheet layers. The insulin-loaded nCOF exhibited insulin protection in digestive fluids in vitro as well as glucose-responsive release, and this hyperglycemia-induced release was confirmed in vivo in diabetic rats without noticeable toxic effects. This is strong evidence that nCOF-based oral insulin delivery systems could replace traditional subcutaneous injections easing insulin therapy.

We report the successful use of a gastro-resistant covalent organic framework for in vivo oral delivery of insulin.     

Long-lived lanthanide emission via a pH-sensitive and switchable LRET complex

Lanthanide-based luminescence resonance energy transfer (LRET) can be used as a tool to enhance lanthanide emission for time-resolved cellular imaging applications. By shortening lanthanide emission lifetimes whilst providing an alternative radiative pathway to the formally forbidden, weak lanthanide-only emission, the photon flux of such systems is increased. With this aim in mind, we investigated energy transfer in differently spaced donor–acceptor terbium–rhodamine pairs with the LRET “on” (low pH) and LRET “off” (high pH). Results informed the design, preparation and characterisation of a compound containing terbium, a spectrally-matched pH-responsive fluorophore and a receptor-targeting group. By combining these elements, we observed switchable LRET, where the targeting group sensitises lanthanide emission, resulting in an energy transfer to the rhodamine dye with an efficiency of E = 0.53. This strategy can be used to increase lanthanide emission rates for brighter optical probes.

A pH-sensitive luminescence resonance energy transfer (LRET) was explored as a method to increase photon flux in a terbium-rhodamine-receptor targeting group construct. At low pH, long-lived dye emission and shorter terbium lifetimes were observed.     

Stimuli‐Responsive Biomolecule‐Based Hydrogels and Their Applications

This Review presents polysaccharides, oligosaccharides, nucleic acids, peptides, and proteins as functional stimuli‐responsive polymer scaffolds that yield hydrogels with controlled stiffness. Different physical or chemical triggers can be used to structurally reconfigure the crosslinking units and control the stiffness of the hydrogels. The integration of stimuli‐responsive supramolecular complexes and stimuli‐responsive biomolecular units as crosslinkers leads to hybrid hydrogels undergoing reversible triggered transitions across different stiffness states. Different applications of stimuli‐responsive biomolecule‐based hydrogels are discussed. The assembly of stimuli‐responsive biomolecule‐based hydrogel films on surfaces and their applications are discussed. The coating of drug‐loaded nanoparticles with stimuli‐responsive hydrogels for controlled drug release is also presented.     

Sodium caseinate as a particulate emulsifier for making indefinitely recycled pH-responsive emulsions

pH-responsive emulsions are one of the simplest and most readily implementable stimuli-responsive systems. However, their practical uses have been greatly hindered by cyclability. Here, we report a robust pH-responsive emulsion prepared by utilizing pure sodium caseinate (NaCas) as the sole emulsifier. We demonstrate that the emulsification/demulsification of the obtained NaCas-stabilized emulsion can be triggered by simply changing the pH value over 100 cycles, which has never been observed in any protein-stabilized emulsion system. The NaCas-stabilized emulsion maintains its pH-responsive properties even in a saturated salt solution (NaCl ∼ 6.1 M) or seawater. We illustrate how NaCas functions in pH-responsive emulsions and show that when conventional nanoparticles such as zein protein or bare SiO₂ particles were coated with a layer of NaCas, the resulting formulated emulsions could be switched on and off over 10 cycles. The unique properties of NaCas thus enable the engineering of conventional Pickering emulsions to pH-responsive Pickering emulsions. Finally, we have integrated catalytically active gold (Au) nanoclusters (NCs) into the NaCas protein and then utilized them to produce emulsions. Remarkably, these NaCas–Au NCs assembled at the oil–water interface exhibited excellent catalytic activity and cyclability, not only in aqueous solution, but also in complicated seawater environments.

An unprecedented pH-responsive emulsion is shaped by utilizing pure sodium caseinate (NaCas) as the sole emulsifier for recyclable interfacial catalysis. This emulsion could be reversibly switched on and off over 100 cycles.     

Controlling the charge of pH-responsive redox hydrogels by means of redox-silent biocatalytic processes. A biocatalytic off/on switch

Coupling of redox-silent biocatalytic processes for analyte detection with enzyme-catalyzed redox reactions for signal generation is proposed by the modulation of electrostatic interactions between a pH-responsive polymer and a redox enzyme to control the off–on transition for electrochemical signal generation. Glassy carbon electrodes are modified with a poly(vinyl)imidazole Os(bipyridine)₂Cl redox hydrogel film entrapping urease and PQQ-dependent glucose dehydrogenase, while glucose is present in the solution. The off–on transition is based on the detection of urea as model analyte which is hydrolyzed to ammonia by urease within the hydrogel film concomitantly increasing the local pH value thus invoking deprotonation of the imidazole groups at the polymer backbone. The decrease of positive charges at the polymer decreases electrostatic repulsion between the polymer and the positively charged PQQ-dependent glucose dehydrogenase. Hence, electron transfer rates between polymer-bound Os complexes and PQQ inside the enzyme are enhanced activating electrocatalytic oxidation of glucose. This process generates the electrochemical signal for urea detection.     

Self-healing and shape memory functions exhibited by supramolecular liquid-crystalline networks formed by combination of hydrogen bonding interactions and coordination bonding

We here report a new approach to develop self-healing shape memory supramolecular liquid-crystalline (LC) networks through self-assembly of molecular building blocks via combination of hydrogen bonding and coordination bonding. We have designed and synthesized supramolecular LC polymers and networks based on the complexation of a forklike mesogenic ligand with Ag⁺ ions and carboxylic acids. Unidirectionally aligned fibers and free-standing films forming layered LC nanostructures have been obtained for the supramolecular LC networks. We have found that hybrid supramolecular LC networks formed through metal–ligand interactions and hydrogen bonding exhibit both self-healing properties and shape memory functions, while hydrogen-bonded LC networks only show self-healing properties. The combination of hydrogen bonds and metal–ligand interactions allows the tuning of intermolecular interactions and self-assembled structures, leading to the formation of the dynamic supramolecular LC materials. The new material design presented here has potential for the development of smart LC materials and functional LC membranes with tunable responsiveness.

New supramolecular hybrid liquid-crystalline networks exhibiting self-healing and shape memory properties are developed by self-assembly of small components through hydrogen bonding interactions and coordination bonding.