Cell Surface Engineering Research Articles

Growth and site-specific organization of micron-scale biomolecular devices on living mammalian cells

Mesoscale molecular assemblies on the cell surface, such as cilia and filopodia, integrate information, control transport and amplify signals. Designer cell-surface assemblies could control these cellular functions. Such assemblies could be constructed from synthetic components ex vivo, making it possible to form such structures using modern nanoscale self-assembly and fabrication techniques, and then oriented on the cell surface. Here we integrate synthetic devices, micron-scale DNA nanotubes, with mammalian cells by anchoring them by their ends to specific cell surface receptors. These filaments can measure shear stresses between 0-2 dyn/cm2, a regime important for cell signaling. Nanotubes can also grow while anchored to cells, thus acting as dynamic cell components. This approach to cell surface engineering, in which synthetic biomolecular assemblies are organized with existing cellular architecture, could make it possible to build new types of sensors, machines and scaffolds that can interface with, control and measure properties of cells.

Poly(γ-glutamic acid) Nanocoating To Enhance the Viability of Pseudomonas stutzeri NRCB010 through Cell Surface Engineering.

Microbial inoculants can enhance soil quality, promote plant nutrient acquisition, and alleviate problems caused by the excessive use of chemical fertilizers. However, susceptibility to harsh conditions during transport and storage, as well as the short shelf-life of plant growth-promoting rhizobacteria (PGPR), limit industrial application. Herein, a novel strategy to form nanocoating on bacterial surfaces to enhance viability was proposed. The nanocoating was composed of N-hydroxysuccinimide (NHS)-modified poly (γ-glutamic acid) (γ-PGA) and calcium ions, which could adhere to the surface of bacteria by forming covalent bonds and ionic bonds with the bacteria. The bacteria encapsulated in the coating had better resistance against harsh conditions than bare bacteria. The viability of coated bacteria was also increased by 2.38 times compared with bare bacteria after 4 weeks of storage. The pot experiment showed that coated Pseudomonas stutzeri NRCB010 had better growth-promoting properties compared with free P. stutzeri NRCB010. These results indicate that cell surface engineering is an effective method to enhance the resistance of bacteria against harsh conditions and is expected to promote the widespread use of PGPR.

Revisiting the Cytotoxicity of Cationic Polyelectrolytes as a Principal Component in Layer-by-Layer Assembly Fabrication.

Polycations are an essential part of layer-by-layer (LbL)-assembled drug delivery systems, especially for gene delivery. In addition, they are used for other related applications, such as cell surface engineering. As a result, an assessment of the cytotoxicity of polycations and elucidation of the mechanisms of polycation toxicity is of paramount importance. In this study, we examined in detail the effects of a variety of water-soluble, positively charged synthetic polyelectrolytes on in vitro cytotoxicity, cell and nucleus morphology, and monolayer expansion changes. We have ranked the most popular cationic polyelectrolytes from the safest to the most toxic in relation to cell cultures. 3D cellular cluster formation was disturbed by addition of polyelectrolytes in most cases in a dose-dependent manner. Atomic force microscopy allowed us to visualize in detail the structures of the polyelectrolyte–DNA complexes formed due to electrostatic interactions. Our results indicate a relationship between the structure of the polyelectrolytes and their toxicity, which is necessary for optimization of drug and gene delivery systems.

界面微生物工学

界面微生物工学

Biomaterials for Cell-Surface Engineering and Their Efficacy.

Literature in the field of stem cell therapy indicates that, when stem cells in a state of single-cell suspension are injected systemically, they show poor in vivo survival, while such cells show robust cell survival and regeneration activity when transplanted in the state of being attached on a biomaterial surface. Although an attachment-deprived state induces anoikis, when cell-surface engineering technology was adopted for stem cells in a single-cell suspension state, cell survival and regenerative activity dramatically improved. The biochemical signal coming from ECM (extracellular matrix) molecules activates the cell survival signal transduction pathway and prevents anoikis. According to the target disease, various therapeutic cells can be engineered to improve their survival and regenerative activity, and there are several types of biomaterials available for cell-surface engineering. In this review, biomaterial types and application strategies for cell-surface engineering are presented along with their expected efficacy.

Cell surface engineering of Saccharomyces cerevisiae for simultaneous valorization of corn cob and cheese whey via ethanol production

The viability of 2nd generation bioethanol processes is dependent on achieving high ethanol titers, which requires the use of high solid loadings that will negatively affect the fermentative microorganism besides increasing enzyme-associated costs. To solve this, and also problems of feedstock availability, lignocellulosic biomass can be mixed with dairy by-products to increase carbon content. In this study, industrial strains of Saccharomyces cerevisiae, with improved thermotolerance and stress resistance, were engineered for the cell surface display of cellulolytic enzymes and were evaluated in consolidated bioprocessing of cellulose. Additionally, β-galactosidase was also displayed to enable lactose consumption, resulting in high ethanol titers (>50 g/L) from the simultaneous use of cheese whey and pretreated corn cob as substrate. The multi-feedstock valorization approach together with this lactose-consuming cellulolytic yeast allowed the reduction on materials costs by 60% with a 2.5-fold increase in the annual ethanol production, therefore contributing to the establishment of economic viable ethanol processes.

Natural Photosensitizers in Antimicrobial Photodynamic Therapy.

Health problems and reduced treatment effectiveness due to antimicrobial resistance have become important global problems and are important factors that negatively affect life expectancy. Antimicrobial photodynamic therapy (APDT) is constantly evolving and can minimize this antimicrobial resistance problem. Reactive oxygen species produced when nontoxic photosensitizers are exposed to light are the main functional components of APDT responsible for microbial destruction; therefore, APDT has a broad spectrum of target pathogens, such as bacteria, fungi, and viruses. Various photosensitizers, including natural extracts, compounds, and their synthetic derivatives, are being investigated. The main limitations, such as weak antimicrobial activity against Gram-negative bacteria, solubility, specificity, and cost, encourage the exploration of new photosensitizer candidates. Many additional methods, such as cell surface engineering, cotreatment with membrane-damaging agents, nanotechnology, computational simulation, and sonodynamic therapy, are also being investigated to develop novel APDT methods with improved properties. In this review, we summarize APDT research, focusing on natural photosensitizers used in in vitro and in vivo experimental models. In addition, we describe the limitations observed for natural photosensitizers and the methods developed to counter those limitations with emerging technologies.

Functional bionanomaterials for cell surface engineering in cancer immunotherapy.

The cell surface is the forward position in cancer immunotherapy, with surface ligand and receptor interactions between various cells for determining immune privilege or recognition. Therefore, cell surface engineering (CSE) that manipulates the surface interactions between the immune effector cells (IECs) and tumor cells represents a promising means for eliciting effective anticancer immunity. Specifically, taking advantage of the development in biomaterials and nanotechnology, the use of functional bionanomaterials for CSE is attracting more and more attention in recent years. Rationally designed functional biomaterials have been applied to construct artificial functional modules on the surface of cells through genetic engineering, metabolic labeling, chemical conjugation, hydrophobic insertion, and many other means, and the CSE process can be performed both ex vivo and in vivo, on either IECs or tumor cells, and results in enhanced anticancer immunity and various new cancer immunity paradigms. In this review, we will summarize the recent exciting progresses made in the application of functional bionanomaterials for CSE especially in establishing effective recognition and interaction between IECs and tumor cells.

Functional bionanomaterials help optimize surface engineering to improve cancer immunotherapy

A review summarizes using bionanomaterials for cell surface engineering – both ex vivo and in vivo – in cancer immunotherapy.

Controlling Cell Dynamics by Cell-surface Modification

Although the concept of a drug delivery system (DDS) is usually applied to conventional drug therapy, it is also important for cell-based therapy. The surface manipulation of living cells represents a powerful tool for controlling cell behaviors in the body, such as enhancement of cell-cell interactions, targeted delivery of cells, and protection from immunological rejection. Functional groups, including amines, thiols, and carbonyls, offer excellent opportunities for chemical modification through the formation of covalent bonds with exogenous molecules. Non-natural reactive groups introduced by metabolic labeling were recently utilized for targeted chemical modification. On the other hand, noncovalent strategies are also available; two major examples are electrostatic interaction with a negative charge on the cell surface and hydrophobic insertion or interaction with the cell membrane. In this study, we analyzed factors affecting cell surface modifications using PEG-lipid and succeeded in enhancing the efficacy of modification by cyclodextrin. Then, mesenchymal stem cells (MSCs), whose therapeutic effect has been demonstrated at the clinical stage and which have been clinically used as a drug, were decorated with PEG-lipid conjugates having a targeted ligand such as peptide or scFv, which are recognized by ICAM1. The peptide or scFv decoration enhanced the cell adhesion of MSCs on cytokine treated-endothelial cells. This technique will prompt the targeted delivery of MSCs to intended therapy sites, and underscores the promise of cell surface engineering as a tool for improving cell-based therapy.

Cell Surface Engineering Using DNAzymes: Metal Ion Mediated Control of Cell-Cell Interactions.

Regulating cell-cell interactions and cell behaviors via cell surface engineering is of significance for biological research such as cell fate control and cell therapy. While extensive efforts have been made to induce cell-cell assembly via various cell surface modifications triggered by macromolecules or organic metabolites, controllable cell-cell interactions that include both assembly and disassembly triggered by metal ions remain a challenge. Herein, we report a strategy based on DNAzymes to realize controllable cell-cell interactions, triggered by metal ions. The metal-dependent DNAzyme-based cleavage can effectively manipulate cell behaviors, including cell-cell conjunctions and disaggregation. Using a Zn2+-specific DNAzyme, a Mg2+-specific DNAzyme, and their respective substrate strands as the building blocks, the corresponding DNA double-chain switches enabling two-factor disassembly are demonstrated. Moreover, the method has been applied to control the assembly and disassembly between two cell spheroids. Since a wide variety of metal-specific DNAzymes are available, this method can be readily applied to construct cell dynamic systems controlled by other metal ions, providing a smart and versatile platform to regulate dynamic cell behavior.

Assembly Pathway Selection with DNA Reaction Circuits for Programming Multiple Cell-Cell Interactions.

The manipulation of cell-cell interactions promotes the study of multicellular behavior, but it remains a great challenge for programming multicellular assembly in complex reaction pathways with multiple cell types. Here we report a DNA reaction circuit-based approach to cell-surface engineering for the programmable regulation of multiple cell-cell interactions. The DNA circuits are designed on the basis of a stem-loop-integrated DNA hairpin motif, which has the capability of programming diverse molecular self-assembly and disassembly pathways by sequential allosteric activation. Modifying the cell surface with such DNA reaction circuits allows for performing programmable chemical functions on cell membranes and the control of multicellular self-assembly with selectivity. We demonstrate the selective control of targeting the capability of natural killer (NK) cells to two types of tumor cells, which show selectively enhanced cell-specific adaptive immunotherapy efficacy. We hope that our method provides new ideas for the programmable control of multiple cell-cell interactions in complex reaction pathways and potentially promotes the development of cell immunotherapy.

Engineering organoids

Organoids are in vitro miniaturized and simplified model systems of organs that have gained enormous interest for modelling tissue development and disease, and for personalized medicine, drug screening and cell therapy. Despite considerable success in culturing physiologically relevant organoids, challenges remain to achieve real-life applications. In particular, the high variability of self-organizing growth and restricted experimental and analytical access hamper the translatability of organoid systems. In this Review, we argue that many limitations of traditional organoid culture can be addressed by engineering approaches at all levels of organoid systems. We investigate cell surface and genetic engineering approaches, and discuss stem cell niche engineering based on the design of matrices that allow spatiotemporal control of organoid growth and shape-guided morphogenesis. We examine how microfluidic approaches and lessons learnt from organs-on-a-chip enable the integration of mechano-physiological parameters and increase accessibility of organoids to improve functional readouts. Applying engineering principles to organoids increases reproducibility and provides experimental control, which will, ultimately, be required to enable clinical translation.

In Situ Synthesis of Photoactive Polymers on a Living Cell Surface via Bio-Palladium Catalysis for Modulating Biological Functions.

Cell surface engineering with functional polymers is an effective strategy to modulate cell activity. Here, a bio-palladium catalyzed polymerization strategy was developed for in situ synthesis of conjugated polymers on living cell surfaces. Through Sonagashira polymerization, photoactive polyphenyleneethynylene (PPE) is synthesized on the cell surface via cell-generated bio-Pd catalyst. The in situ formed PPE is identified by excellent light-harvest capacity and blue fluorescence on the surfaces of E. coli and C. pyrenoidosa. Besides imaging microbes for tracing the polymerization process, PPE also exhibits enhanced antibacterial activity against E. coli. It can also augment the ATP synthesis of C. pyrenoidosa through enlarging the light absorption and accelerating the cyclic electron transport of the algae. With this bio-metal catalyzed polymerization method, functional polymers can be synthesized in situ on the living cell surface.

A Novel Strategy for the Microbial Removal of Heavy Metals: Cell-surface Display of Peptides

Cell surface engineering is a rapidly developing technology of microorganism that achieves modification of cell surface function by joining external functional peptides with surface anchoring proteins, for example, Outer Member Protein (OMP). On account of these proteins possessing metal responsive motifs, they can be specifically used for metal adsorption and dissociation. To elucidate the problems caused by heavy metals and develop various technologies for their removel or recovery, various metal-binding proteins/peptides fused on microorganism cell surface have been applied as novel methods. During the past few years, bacterial cell surface display strategy has received growing attention for their availability to eliminate heavy metals. In this paper, the existing problems, progress, and suggestions for furture of the peptide displaying system are summarized.

Synthetic DNA for Cell-Surface Engineering.

The cell membrane is not only a physical barrier, but also a functional organelle that regulates the communication between a cell and its environment. The ability to functionalize the cell membrane with synthetic molecules or nanostructures would advance cellular functions beyond what evolution has provided. The aim of this Minireview is to introduce recent progress in using synthetic DNA and DNA-based nanostructures for cell-surface engineering. We first introduce chemical conjugation and physical binding methods for monovalent and polyvalent surface engineering. We then introduce the application of these methods for either the promotion or inhibition of cell-environment communication in numerous applications, including the promotion of cell-cell recognition, regulation of intracellular pathways, protection of therapeutic cells, and sensing of the intracellular and extracellular microenvironments. Lastly, we summarize current challenges existing in this area and potential solutions to solve these challenges.

Surface-Anchored Nanogel Coating Endows Stem Cells with Stress Resistance and Reparative Potency via Turning Down the Cytokine-Receptor Binding Pathways.

Stem cell‐based therapy has great potential in regenerative medicine. However, the survival and engraftment rates of transplanted stem cells in disease regions are poor and limit the effectiveness of cell therapy due to the fragility of stem cells. Here, an approach involving a single‐cell coating of surface‐anchored nanogel to regulate stem cell fate with anti‐apoptosis capacity in the hypoxic and ischemic environment of infarcted hearts is developed for the first time. A polysialic acid‐based system is used to anchor microbial transglutaminase to the external surface of the cell membrane, where it catalyzes the crosslinking of gelatin. The single‐cell coating with surface‐anchored nanogel endows mesenchymal stem cells (MSCs) with stress resistance by blocking the activity of apoptotic cytokines including the binding of tumor necrosis factor α (TNFα) to tumor necrosis factor receptor, which in turn maintains mitochondrial integrity, function and protects MSCs from TNFα‐induces apoptosis. The administration of surface engineered MSCs to hearts results in significant improvements in engraftment, cardiac function, infarct size, and vascularity compared with using uncoated MSCs in treating myocardial infarction. The surface‐anchored, biocompatible cell surface engineering with nanogel armor provides a new way to produce robust therapeutic stem cells and may explore immense potentials in cell‐based therapy.

Cell-surface engineering of yeasts for whole-cell biocatalysts.

Due to the unique advantages comparing with traditional free enzymes and chemical catalysis, whole-cell biocatalysts have been widely used to catalyze reactions effectively, simply and environment friendly. Cell-surface display technology provides a novel and effective approach for improved whole-cell biocatalysts expressing heterologous enzymes on the cell surface. They can overcome the substrate transport limitation of the intracellular expression and provide the enzymes with enhanced properties. Among all the host surface-displaying microorganisms, yeast is ideally suitable for constructing whole cell-surface-displaying biocatalyst, because of the large cell size, the generally regarded as safe (GRAS) status, and the perfect post-translational processing of secreted proteins. Yeast cell-surface display system has been a promising and powerful method for development of novel and improved engineered biocatalysts. In this review, the characterization and principles of yeast cell-surface display and the applications of yeast cell-surface display in engineered whole-cell biocatalysts as well as the improvement of the enzyme efficiency are summarized and discussed.

Cell surface modulation of endothelial cells by cell surface engineering for improvement of graft survival in kidney transplantation

Cell surface modulation of endothelial cells by cell surface engineering for improvement of graft survival in kidney transplantation

Improved ammonia production from soybean residues by cell surface-displayed l-amino acid oxidase on yeast.

Ammonia is critical for agricultural and chemical industries. The extracellular production of ammonia by yeast (Saccharomyces cerevisiae) using cell surface engineering can be efficient approach because yeast can avoid growth deficiencies caused by knockout of genes for ammonia assimilation. In this study, we produced ammonia outside the yeast cells by displaying an l-amino acid oxidase with a wide substrate specificity derived from Hebeloma cylindrosporum (HcLAAO) on yeast cell surfaces. The HcLAAO-displaying yeast successfully produced 12.6 m m ammonia from a mixture of 20 proteinogenic amino acids (the theoretical conversion efficiency was 63%). We also succeeded in producing ammonia from a food processing waste, soybean residues (okara) derived from tofu production. The conversion efficiency was 88.1%, a higher yield than reported in previous studies. Our study demonstrates that ammonia production outside of yeast cells is a promising strategy to utilize food processing wastes.