Well-Defined Nanostructured Biointerfaces: Strengthened Cellular Interaction for Circulating Tumor Cells Isolation

The topographic features at the cell–material biointerface are critical for cellular sensing of the extracellular environment (ECM) and have gradually been recognized as key factors that regulate cell adhesion behavior. Herein, a well-defined nanostructured biointerface is fabricated via a new generation of mussel-inspired polymer coating to mimic the native ECM structures. Upon the bioinert background presence and biospecific ligands conjugation, the affinity of cancer cells to the resulting biofunctional surfaces, which integrate topographic features and biochemical cues, is greatly strengthened. Both the conjugated bioligand density, filopodia formation, and focal adhesion expression are significantly enhanced by the surficial nano-features with an optimized size-scale. Thus, this nanostructured biointerface exhibits high capture efficiency for circulating tumor cells (CTCs) with high sensitivity, high biospecificity, and high purity. Benefiting from the unique bioligands conjugation chemistry herein, the captured cancer cells can be responsively detached from the biointerfaces without damage for downstream analysis. The present biofunctional nanostructured interfaces offer a good solution to address current challenges to efficiently isolate rare CTCs from blood samples for earlier cancer diagnosis

Surface Roughness Gradients Reveal Topography‐Specific Mechanosensitive Responses in Human Mesenchymal Stem Cells

The topographic features of an implant, which mechanically regulate cell behaviors and functions, are critical for the clinical success in tissue regeneration. How cells sense and respond to the topographical cues, e.g., interfacial roughness, is yet to be fully understood and even debatable. Here, the mechanotransduction and fate determination of human mesenchymal stem cells (MSCs) on surface roughness gradients are systematically studied. The broad range of topographical scales and high‐throughput imaging is achieved based on a catecholic polyglycerol coating fabricated by a one‐step‐tilted dip‐coating approach. It is revealed that the adhesion of MSCs is biphasically regulated by interfacial roughness. The cell mechanotransduction is investigated from focal adhesion to transcriptional activity, which explains that cellular response to interfacial roughness undergoes a direct force‐dependent mechanism. Moreover, the optimized roughness for promoting cell fate specification is explored

A fast open-source Fiji-macro to quantify virus infection and transfection on single-cell level by fluorescence microscopy

The ability to automatically analyze large quantities of image data is a valuable tool for many biochemical assays, as it rapidly provides reliable data. Here, we describe a fast and robust Fiji macro for the analysis of cellular fluorescence microscopy images with single-cell resolution. The macro presented here was validated by successful reconstruction of fluorescent and non-fluorescent cell mixing ratios (for fluorescence fractions ranging between 0 and 100%) and applied to quantify the efficiency of transfection and virus infection inhibition. It performed well compared with manually obtained image quantification data. Its use is not limited to the cases shown here but is applicable for most monolayered cellular assays with nuclei staining. We provide a detailed description of how the macro works and how it is applied to image data. It can be downloaded free of charge and may be used by and modified according to the needs of the user. • Rapid, simple, and reproducible segmentation of eukaryotic cells in confluent cellular assays • Open-source software for use without technical or computational expertise • Single-cell analysis allows identification and quantification of virus infected cell populations and infection inhibitio

Mussel-Inspired Polyglycerol Coatings for Surface Modification with Tunable Architecture

Mussel-inspired coatings, known for their outstanding substrate-independent adhesive capabilities, have numerous potential applications in materials science and biomedical fields. To improve the understanding of how these polymers’ molecular structure and chemical composition affect their coating mechanisms and resulting coating properties, herein three mussel-inspired polymers are developed: dendritic polyglycerol with 40% catechol groups and 60% amines (dPG40), linear polyglycerol with 80% catechols and 20% amines (lPG80), and finally lPG40 with 40% catechols and 60% amines. After a series of characterizations, it is found that chemical surface modification with a monolayer coating can be easily achieved with lPG40, and that robust and well-defined nano- to micro-structural surface coatings are possible with lPG80 and dPG40. Tunable properties are found to include not only coating speed, but coating thickness, roughness, and surficial topography. This diverse suite of controllable attributes enables mussel-inspired polyglycerol (MiPG) coatings to satisfy a wide-range of applications on multiple material

Scaffold Flexibility Controls Binding of Herpes Simplex Virus Type 1 with Sulfated Dendritic Polyglycerol Hydrogels Fabricated by Thiol-Maleimide Click Reaction

Herpes Simplex Virus-1 (HSV-1) with a diameter of 155–240 nm uses electrostatic interactions to bind with the heparan sulfate present on the cell surface to initiate infection. In this work, the initial contact using polysulfate-functionalized hydrogels is aimed to deter. The hydrogels provide a large contact surface area for viral interaction and sulfated hydrogels are good mimics for the native heparan sulfate. In this work, hydrogels of different flexibilities are synthesized, determined by rheology. Gels are prepared within an elastic modulus range of 10–1119 Pa with a mesh size of 80–15 nm, respectively. The virus binding studies carried out with the plaque assay show that the most flexible sulfated hydrogel performs the best in binding HSV viruses. These studies prove that polysulfated hydrogels are a viable option as HSV-1 antiviral compounds. Furthermore, such hydrogel networks are also physically similar to naturally occurring mucus gels and therefore may be used as mucus substitutes

Charge Matters: Mutations in Omicron Variant Favor Binding to Cells

Evidence is strengthening to suggest that the novel SARS-CoV-2 mutant Omicron, with its more than 60 mutations, will spread and dominate worldwide. Although the mutations in the spike protein are known, the molecular basis for why the additional mutations in the spike protein that have not previously occurred account for Omicron's higher infection potential, is not understood. We propose, based on chemical rational and molecular dynamics simulations, that the elevated occurrence of positively charged amino acids in certain domains of the spike protein (Delta: +4; Omicron: +5 vs. wild type) increases binding to cellular polyanionic receptors, such as heparan sulfate due to multivalent charge-charge interactions. This observation is a starting point for targeted drug development

Virus removal from aqueous environments with polyelectrolyte coatings on a polypropylene fleece

The adsorption of viruses from aqueous solution is frequently performed to detect viruses. Charged filtration materials capture viruses via electrostatic interactions, but lack the specificity of biological virus-binding substances like heparin. Herein, we present three methods to immobilize heparin-mimicking, virus-binding polymers to a filter material. Two mussel-inspired approaches are used, based on dopamine or mussel-inspired dendritic polyglycerol, and post-functionalized with a block-copolymer consisting of linear polyglycerol sulfate and amino groups as anchor (lPGS-b-NH2). As third method, a polymer coating based on lPGS with benzophenone anchor groups is tested (lPGS-b-BPh). All three methods yield dense and stable coatings. A positively charged dye serves as a tool to quantitatively analyze the sulfate content on coated fleece. Especially lPGS-b-BPh is shown to be a dense polymer brush coating with about 0.1 polymer chains per nm2. Proteins adsorb to the lPGS coated materials depending on their charge, as shown for lysozyme and human serum albumin. Finally, herpes simplex virus type 1 (HSV-1) and severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) can be removed from solution upon incubation with coated fleece materials by about 90% and 45%, respectively. In summary, the presented techniques may be a useful tool to collect viruses from aqueous environments

Topology-Matching Design of an Influenza-Neutralizing Spiky Nanoparticle-Based Inhibitor with a Dual Mode of Action

In this study, we demonstrate the concept of "topology-matching design" for virus inhibitors. With the current knowledge of influenzaA virus (IAV), we designed a nanoparticle-based inhibitor (nano-inhibitor) that has a matched nanotopology to IAV virions and shows heteromultivalent inhibitory effects on hemagglutinin and neuraminidase. The synthesized nano-inhibitor can neutralize the viral particle extracellularly and block its attachment and entry to the host cells. The virus replication was significantly reduced by 6 orders of magnitude in the presence of the reverse designed nano-inhibitors. Even when used 24hours after the infection, more than 99.999% inhibition is still achieved, which indicates such a nano-inhibitor might be a potent antiviral for the treatment of influenza infection

Surface-Initiated Grafting of Dendritic Polyglycerol from Mussel-Inspired Adhesion-Layers for the Creation of Cell-Repelling Coatings

Biofouling is a major challenge in the application of textiles, biosensors, and biomedical implants. In the current work, a straightforward method for the solvent-free polymerization of antifouling dendritic polyglycerol (dPG) from mussel-inspired dendritic polyglycerol (MI-dPG) coatings on hydrophilic titanium dioxide (TiO2) and hydrophobic polydimethylsiloxane (PDMS) is reported. Surface characterization is performed by static water contact angle (CA) measurements, X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy (SEM). Significant lower CA values are obtained after dPG grafting from MI-dPG-coated TiO2 and MI-dPG coated PDMS. Furthermore, XPS shows a time-dependent increase of the C-O bond content upon dPG grafting from MI-dPG-coated TiO2 and MI-dPG-coated PDMS. Analysis of the surface morphology by SEM shows a clear time-dependent increase in the surface roughness upon dPG grafting from MI-dPG-coated TiO2 and MI-dPG-coated PDMS. When the viability of two adhesive cell types is studied via LIVE/DEAD staining, a strong reduction in the cell density is observed after the dPG grafting from MI-dPG-coated TiO2 and MI-dPG-coated PDMS (a decrease of >95% in all cases). The combined results show that biocompatible but highly cell-repelling surfaces are efficiently constructed via the grafting of dPG from MI-dPG-coated TiO2 and MI-dPG-coated PDMS

Mucin-Inspired, High Molecular Weight Virus Binding Inhibitors Show Biphasic Binding Behavior to Influenza A Viruses

Multivalent binding inhibitors are a promising new class of antivirals that prevent virus infections by inhibiting virus binding to cell membranes. The design of these inhibitors is challenging as many properties, for example, inhibitor size and functionalization with virus attachment factors, strongly influence the inhibition efficiency. Here, virus binding inhibitors are synthesized, the size and functionalization of which are inspired by mucins, which are naturally occurring glycosylated proteins with high molecular weight (MDa range) and interact efficiently with various viruses. Hyperbranched polyglycerols (hPGs) with molecular weights ranging between 10 and 2600 kDa are synthesized, thereby hitting the size of mucins and allowing for determining the impact of inhibitor size on the inhibition efficiency. The hPGs are functionalized with sialic acids and sulfates, as suggested from the structure of mucins, and their inhibition efficiency is determined by probing the inhibition of influenza A virus (IAV) binding to membranes using various methods. The largest, mucin-sized inhibitor shows potent inhibition at pm concentrations, while the inhibition efficiency decreases with decreasing the molecular weight. Interestingly, the concentration-dependent IAV inhibition shows a biphasic behavior, which is attributed to differences in the binding affinity of the inhibitors to the two IAV envelope proteins, neuraminidase, and hemagglutinin