Photodegradable Hydrogels for Capture,Detection, and Release of Live Cells

Cells may be captured and released using a photodegradable hydrogel (photogel) functionalized with antibodies. Photogel substrates were used to first isolate human CD4 or CD8 T‐cells from a heterogeneous cell suspension and then to release desired cells or groups of cells by UV‐induced photodegradation. Flow cytometry analysis of the retrieved cells revealed approximately 95 % purity of CD4 and CD8 T‐cells, suggesting that this substrate had excellent specificity. To demonstrate the possibility of sorting cells according to their function, photogel substrates that were functionalized with anti‐CD4 and anti‐TNF‐α antibodies were prepared. Single cells captured and stimulated on such substrates were identified by the fluorescence “halo” after immunofluorescent staining and could be retrieved by site‐specific exposure to UV light through a microscope objective. Overall, it was demonstrated that functional photodegradable hydrogels enable the capture, analysis, and sorting of live cells.     

Surface Molding of Microscale Hydrogels with Microactuation Functionality

This work describes the fabrication of numerous hydrogel microstructures (μ‐gels) via a process called “surface molding.” Chemically patterned elastomeric‐assembly substrates were used to organize and manipulate the geometry of liquid prepolymer microdroplets, which, following photo‐initiated crosslinking, maintained the desired morphology. By adjusting the state of strain during the crosslinking process, a continua of structures could be created using one pattern. These arrays of μ‐gels have stimuli‐responsive properties that are directly applicable to actuation where the basis shape and array geometry of the μ‐gels can be used to rationally generate microactuators with programmed motions. As a method, “surface molding,” represents a powerful addition to the soft‐lithographic toolset that can be readily applied to the simultaneous synthesis of large numbers of geometrically and functionally distinct polymeric microstructures.     

Photodegradable Hydrogels for Capture,Detection, and Release of Live Cells

Cells may be captured and released using a photodegradable hydrogel (photogel) functionalized with antibodies. Photogel substrates were used to first isolate human CD4 or CD8 T‐cells from a heterogeneous cell suspension and then to release desired cells or groups of cells by UV‐induced photodegradation. Flow cytometry analysis of the retrieved cells revealed approximately 95 % purity of CD4 and CD8 T‐cells, suggesting that this substrate had excellent specificity. To demonstrate the possibility of sorting cells according to their function, photogel substrates that were functionalized with anti‐CD4 and anti‐TNF‐α antibodies were prepared. Single cells captured and stimulated on such substrates were identified by the fluorescence “halo” after immunofluorescent staining and could be retrieved by site‐specific exposure to UV light through a microscope objective. Overall, it was demonstrated that functional photodegradable hydrogels enable the capture, analysis, and sorting of live cells.     

软光刻法制备具有表面微结构的角蛋白膜

近年来,使用微纳米制造工艺将蛋白质或多肽进行高精度空间图案化,推动了细胞生物学、组织工程学、药物科学等领域的发展.同时,羊毛角蛋白作为一种储量大的天然生物蛋白质,具有优异的水溶性、良好的生物相容性和可控的降解性,但羊毛角蛋白通常不能自组装形成凝胶网络或其他不溶形式,因此,使用羊毛角蛋白制备如纤维、薄膜、凝胶等的成型结构存在很大困难.本工作通过使用化学修饰的方法,在角蛋白上接枝功能基团,使角蛋白获得光敏感性,探究了共价交联法制备具有表面微结构角蛋白膜的可行性.并用3D激光扫描显微镜、紫外可见近红外光谱仪和傅里叶变换显微红外光谱仪对薄膜结构进行了表征.结果表明,使用软光刻法可以得到表面微结构完整度很高的角蛋白膜.本工作对羊毛角蛋白共价交联法进行了实验探索,实验结果不仅为人们提供了一种软光刻技术制备具有表面微结构的角蛋白膜的方法,而且为羊毛角蛋白制备成型结构提供了新的途径.     

Large Polyacrylamide Hydrogels for Large-Batch Cell Culture and Mechanobiological Studies

The rigidity of a cell's substrate or extracellular matrix plays a vital role in regulating cell and tissue functions. Polyacrylamide (PAAm) hydrogels are one of the most widely used cell culture substrates that provide a physiologically relevant range of stiffness. However, it is still arduous and time-consuming to prepare PAAm substrates in large batches for high-yield or multiscale cell cultures. In this communication, a simple method to prepare PAAm hydrogels with less time cost and easily accessible materials is presented. The hydrogel is mechanically uniform and supports cell culture in a large batch. It is further shown that the stiffness of the hydrogel covers a large range of Young's modulus and is sensed by cells, regulating various cell features including changes in cell morphology, proliferation, and contractility. This method improves the reproducibility of mechanobiology studies and can be easily applied for mechanobiology research requiring large numbers of cells or experimental groups.     

Degradable Poly(amidoamine) Hydrogels as Scaffolds for In Vitro Culturing of Peripheral Nervous System Cells

This paper reports on the synthesis and physico‐chemical, mechanical, and biological characterization of two sets of poly(amidoamine) (PAA) hydrogels with potential as scaffolds for in vivo peripheral nerve regeneration. They are obtained by polyaddition of piperazine with N,N′‐methylenebis(acrylamide) or 1,4‐bis(acryloyl)piperazine with 1,2‐diaminoethane as cross‐linking agent and exhibit a combination of relevant properties, such as mechanical strength, biocompatibility, biodegradability, ability to induce adhesion and proliferation of Schwann cells (SCs) preserving their viability. Moreover, the most promising hydrogels, that is those deriving from 1,4‐bis(acryloyl)piperazine, allow the in vitro growth of the sensitive neurons of the dorsal root ganglia, thus getting around a critical point in the design of conduits for nerve regeneration.

     

二维平面细胞微图案化技术及其生物学应用

可以控制细胞粘附形状、大小的方法统称为细胞图案化技术.这些方法结合微纳米制备、表面化学、电化学和光化学等手段可以动态控制细胞的粘附、迁移、分化及其相互作用,为细胞生物学研究提供了一个新平台.本文介绍了二维平面细胞图案化的各种方法,并对其优缺点进行了总结,评述了细胞图案化技术在细胞生物学基础研究、组织工程以及基于细胞的生物传感器领域的应用.     

Molecular Encryption and Reconfiguration for Remodeling of Dynamic Hydrogels

Dynamic materials have been widely studied for regulation of cell adhesion that is important to a variety of biological and biomedical applications. These materials can undergo changes mainly through one of the two mechanisms: ligand release in response to chemical, physical, or biological stimuli, and ligand burial in response to mechanical stretching or the change of electrical potential. This study demonstrates an encrypted ligand and a new hydrogel that are capable of inducing and inhibiting cell adhesion, which is controlled by molecular reconfiguration. The ligand initially exhibits an inert state; it can be reconfigured into active and inert states by using unblocking and recovering molecules in physiological conditions. Since molecular reconfiguration does not require the release of the ligand from the hydrogels, inhibiting and inducing cell adhesion on the hydrogels can be repeated for multiple cycles.     

Modified Hyaluronan Hydrogels Support the Maintenance of Mouse Embryonic Stem Cells and Human Induced Pluripotent Stem Cells

These studies provide evidence for the ability of a commercially available, defined, hyaluronan‐gelatin hydrogel, HyStem‐C?, to maintain both mouse embryonic stem cells (mESCs) and human induced pluripotent stem cells (hiPSCs) in culture while retaining their growth and pluripotent characteristics. Growth curve and doubling time analysis show that mESCs and hiPSCs grow at similar rates on HyStem‐C? hydrogels and mouse embryonic fibroblasts and Matrigel?, respectively. Immunocytochemistry, flow cytometry, gene expression and karyotyping reveal that both human and murine pluripotent cells retain a high level of pluripotency on the hydrogels after multiple passages. The addition of fibronectin to HyStem‐C? enabled the attachment of hiPSCs in a xeno‐free, fully defined medium.

     

Stiff,Multistimuli‐Responsive Supramolecular Hydrogels as Unique Molds for 2D/3D Microarchitectures of Live Cells

Supramolecular hydrogels constructed through molecular self‐assembly of small molecules have unique stimuli‐responsive properties; however, they are mechanically weak in general, relative to conventional polymer gels. Very recently, we developed a zwitterionic amino acid tethered amphiphilic molecule 1 , which gave rise to a remarkably stiff hydrogel comparable with polymer‐based agarose gel, retaining reversible thermal‐responsive properties. In this study, we describe that rational accumulation of multiple and orthogonal noncovalent interactions in the supramolecular nanofibers of 1 played crucial roles not only in the mechanical reinforcement but also in the multistimuli responsiveness. That is, the zwitterionic amino acid moiety and the C C double bond unit of the hydrogelator 1 can function as a pH‐responsive unit and a light‐responsive unit, respectively. We also demonstrated that this stiff and multistimuli‐responsive supramolecular hydrogel 1 is applied as a unique mold for 2D and 3D‐patterning of various substances. More significantly, we succeeded in the fabrication of a collagen gel for spatial patterning, culturing, and differentiation of live cells by using hydrogel 1 molds equipped with 2D/3D microspace channels (100–200 μm in diameter).     

Polyurethane Molecular Stamps for the in situ Synthesis of DNA Microarray

Fabrication of polyurethane molecular stamps (PU stamps) based on polypropylene glycol (PPG) and toluene dissocyanate (TDI), using 3,3′-dichloro-4,4′-methylenedianiline (MOCA) as the crosslinker ,is reported. It was shown from the contact angle measurement that PU stamps surface has good affinity with acetonitrile,guaranteeing the well distribution of DNA monomers on patterned stamps. Laser confocal fluorescence microscopy images of oligonucleotide arrays after hybridization confirmed polyurethane is an excellent material for molecular stamps when ransferring polar chemicals and conducting rections on interfaces by stamping.     

Emerging Technology of Nanofiber-Composite Hydrogels for Biomedical Applications

Hydrogels and nanofibers have been firmly established as go-to materials for various biomedical applications. They have been mostly utilized separately, rarely together, because of their distinctive attributes and shortcomings. However, the potential benefits of integrating nanofibers with hydrogels to synergistically combine their functionalities while attenuating their drawbacks are increasingly recognized. Compared to other nanocomposite materials, incorporating nanofibers into hydrogel has the distinct advantage of emulating the hierarchical structure of natural extracellular environment needed for cell and tissue culture. The most important technological aspect of developing “nanofiber-composite hydrogel” is generating nanofibers made of various polymers that are cross-linked and short enough to maintain stable dispersion in hydrated environment. In this review, recent research efforts to develop nanofiber-composite hydrogels are presented, with added emphasis on nanofiber processing techniques. Several notable examples of implementing nanofiber-composite hydrogels for biomedical applications are also introduced.     

Nitrile Oxide‐Norbornene Cycloaddition as a Bioorthogonal Crosslinking Reaction for the Preparation of Hydrogels

This communication describes the first application of cycloaddition between an in situ generated nitrile oxide with norbornene leading to a polymer crosslinking reaction for the preparation of poly(ethylene glycol) hydrogels under physiological conditions. Hydrogels with high water content and robust physical strength are readily formed within 2–5 min by a simple two‐solution mixing method which allows 3D encapsulation of neuronal cells. This bioorthogonal crosslinking reaction provides a simple yet highly effective method for preparation of hydrogels to be used in bioengineering.

     

Preparation of the Cationic Dendrimer-Based Hydrogels for Controlled Heparin Release

We introduce a cationic polyamidoamine (PAMAM) dendrimers and tetronic (Te) based hydrogels in which precursor copolymers were prepared with simple methods. In the synthetic process, tyramine-conjugated tetronic (TTe) was prepared via activation of its four terminal hydroxyl groups by nitrophenyl chloroformate (NPC) and then substitution of tyramine (TA) into the activated product to obtain TTe. Cationic PAMAM dendrimers G3.0 functionalized with p-hydroxyphenyl acetic acid (HPA) by use of carbodiimide coupling agent (EDC) to obtain Den-HPA. ¹H-NMR confirmed the amount of HPA and TA conjugations. The aqueous TTe and Den-HPA copolymer solution rapidly formed the cationic hydrogels in the presence of horseradish peroxidase enzyme (HRP) and hydrogen peroxide (H₂O₂) at physiological conditions. The gelation time of the hydrogels could be modulated ranging from 7 to 73 secs, when the concentrations of HRP and H₂O₂ varied. The hydrogels exhibited minimal swelling degree and low degradation under physical condition. In vitro cytotoxicity study indicated that the hydrogels were highly cytocompatible as prepared at 0.15 mg/mL HRP and 0.063 wt% of H₂O₂ concentration. Heparin release profiles show that the cationic hydrogels can sustainably release the anionic anticoagulant drug. The obtained results demonstrated a great potential of the cationic hydrogels for coating medical devices or delivering anionic drugs.     

Hydrogels for the Detection and Management of Protease Levels

The design of hydrogels that simultaneously report protease activity and remove excess protease from solution is elucidated. The hydrogels, based on amino‐PEGA, combine enzyme‐specific peptides flanked with FRET complimented by charged amino acid residues that facilitate protease uptake via short range electrostatic interactions. Enzymatic response was analysed using a combination of fluorescence spectroscopy, two‐photon microscopy and UV/Vis spectroscopy. An optimised elastase‐responsive hydrogel resulted in lowering of elastase levels below those typical of chronic wounds. The versatility of the modular‐design approach was demonstrated by development of matrix metalloprotease and chymotrypsin sensitive systems.

     

Injectable Polysaccharide Hydrogels as Biocompatible Platforms for Localized and Sustained Delivery of Antibiotics for Preventing Local Infections

Biocompatible and antibacterial hydrogels have received increasing attention for preventing local bacterial infections. In this study, a type of polysaccharide hydrogels is prepared via the Schiff‐based reaction at physiological conditions. The gelation time and mechanical property of the hydrogels are found to be dependent on the polysaccharide concentration and the polysaccharide weight ratio. 3‐(4,5‐Dimethyl‐thiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide assay and live/dead assay indicate that the hydrogels display nontoxicity in vitro. After subcutaneous injection into rats, the hydrogels exhibit an acceptable biocompatibility in vivo. Furthermore, the bacterial inhibition tests by shaking flask method and agar disc‐diffusion method demonstrate that the ceftriaxone‐sodium‐loaded hydrogels have remarkable antibacterial properties in vitro. The in vivo anti‐infective tests further display that the antibiotic‐loaded hydrogels display excellent anti‐infective efficacies in both superficial and deep tissue infection. Consequently, the injectable and biocompatible polysaccharide hydrogels may serve as promising platforms for localized, sustained delivery of antibiotics for preventing local infections.