Topographic cues guide the attachment of diatom cells and algal zoospores.

Surface topography plays a key role in the colonization of substrata by the colonizing stages of marine fouling organisms. For the innovation of marine antifouling coatings, it is essential to understand how topographic cues affect the settlement of these organisms. In this study, tapered, spiked microstructures and discrete honeycombs of varying feature dimensions were designed and fabricated in order to examine the influence of topography on the attachment of zoospores of the green macroalga Ulva linza and cells of the diatom (microalga) Navicula incerta. Contrasting results were obtained with these two species of algae. Indeed, the preferred location of cells of N. incerta was dominated by attachment point theory, which suggested a positive correlation between the density of cells adhering and the amount of available attachment points, while the settlement of spores of U. linza was mainly regulated by both Wenzel roughness and local binding geometry.

Reactive superhydrophobic surface and its photoinduced disulfide-ene and thiol-ene (bio)functionalization.

Reactive superhydrophobic surfaces are highly promising for biotechnological, analytical, sensor, or diagnostic applications but are difficult to realize due to their chemical inertness. In this communication, we report on a photoactive, inscribable, nonwettable, and transparent surface (PAINTS), prepared by polycondensation of trichlorovinylsilane to form thin transparent reactive porous nanofilament on a solid substrate. The PAINTS shows superhydrophobicity and can be conveniently functionalized with the photoclick thiol-ene reaction. In addition, we show for the first time that the PAINTS bearing vinyl groups can be easily modified with disulfides under UV irradiation. The effect of superhydrophobicity of PAINTS on the formation of high-resolution surface patterns has been investigated. The developed reactive superhydrophobic coating can find applications for surface biofunctionalization using abundant thiol or disulfide bearing biomolecules, such as peptides, proteins, or antibodies.

Shear-Induced Detachment of Polystyrene Beads from SAM-Coated Surfaces.

In this work we experimentally and theoretically analyze the detachment of microscopic polystyrene beads from different self-assembled monolayer (SAM) surfaces in a shear flow in order to develop a mechanistic model for the removal of cells from surfaces. The detachment of the beads from the surface is treated as a thermally activated process applying an Arrhenius Ansatz to determine the activation barrier and attempt frequency of the rate determing step in bead removal. The statistical analysis of the experimental shear detachment data obtained in phosphate-buffered saline buffer results in an activation energy around 20 kJ/mol, which is orders of magnitude lower than the adhesion energy measured by atomic force microscopy (AFM). The same order of magnitude for the adhesion energy measured by AFM is derived from ab initio calculations of the van der Waals interaction energy between the polystyrene beads and the SAM-covered gold surface. We conclude that the rate determing step for detachment of the beads is the initiation of rolling on the surface (overcoming static friction) and not physical detachment, i.e., lifting the particle off the surface.

Holographic microscopy provides new insights into the settlement of zoospores of the green alga Ulva linza on cationic oligopeptide surfaces.

Interaction of zoospores of Ulva linza with cationic, arginine-rich oligopeptide self-assembled monolayers (SAMs) is characterized by rapid settlement. Some spores settle (ie permanently attach) in a 'normal' manner involving the secretion of a permanent adhesive, retraction of the flagella and cell wall formation, whilst others undergo 'pseudosettlement' whereby motile spores are trapped (attached) on the SAM surface without undergoing the normal metamorphosis into a settled spore. Holographic microscopy was used to record videos of swimming zoospores in the vicinity of surfaces with different cationic oligopeptide concentrations to provide time-resolved insights into processes associated with attachment of spores. The data reveal that spore attachment rate increases with increasing cationic peptide content. Accordingly, the decrease in swimming activity in the volume of seawater above the surface accelerated with increasing surface charge. Three-dimensional trajectories of individual swimming spores showed a 'hit and stick' motion pattern, exclusively observed for the arginine-rich peptide SAMs, whereby spores were immediately trapped upon contact with the surface.

Plerixafor induces the rapid and transient release of stromal cell-derived factor-1 alpha from human mesenchymal stromal cells and influences the migration behavior of human hematopoietic progenitor cells.

The interaction between the stromal cell-derived factor-1 alpha (SDF-1α, CXCL12) and its chemokine receptor CXCR4 has been reported to regulate stem cell migration, mobilization and homing. The CXCR4 antagonist plerixafor is highly efficient in mobilizing hematopoietic progenitor cells (HPCs). However, the precise regulatory mechanisms governing the CXCR4/SDF-1α axis between the bone marrow niche and HPCs remain unclear. In this study, we quantify the impact of plerixafor on the interaction between human bone marrow derived mesenchymal stromal cells (MSCs) and human CD34+ HPCs. An assessment of SDF-1α levels in the supernatant of MSC cultures revealed that exposure to plerixafor led to a transient increase but had no long-term effect. In Transwell experiments, we observed that the addition of SDF-1α significantly stimulated HPC migration; this stimulation was almost completely antagonized by the addition of plerixafor, confirming the direct impact of the CXCR4/SDF-1α interaction on the migration capacity of HPCs. We also developed a new microstructural niche model to determine the chemotactic sensitivity of HPCs. Time-lapse microscopy demonstrated that HPCs migrated actively along an SDF-1α gradient within the microchannels and the quantitative assessment of the required minimum gradient initiating this chemotaxis revealed a surprisingly high sensitivity of HPCs. These data demonstrate the fine-tuned balance of the CXCR4/SDF-1α axis and the synergistic effects of plerixafor on HPCs and MSCs, which most likely represent the key mechanisms for the consecutive mobilization of HPCs from the bone marrow niche into the circulating blood.

Direct three-dimensional imaging of polymer-water interfaces by nanoscale hard X-ray phase tomography.

We report three-dimensional (3D) direct imaging of complex surface-liquid interfaces by hard X-ray phase contrast tomography as a non-destructive approach for the morphological characterization of surfaces at the micro- and nanoscale in contact with water. Specifically, we apply this method to study the solid-air-water interface in hydrophobic macroporous polymethacrylate surfaces, and the solid-oil-water interface in slippery liquid-infused porous surfaces (SLIPS). Varying the isotropic spatial resolution allows the 3D quantitative characterization of individual polymer globules, globular clusters (porosity) as well as the infused lubricant layer on SLIPS. Surface defects were resolved at the globular level. We show the first application of X-ray nanotomography to hydrated surface characterizations and we anticipate that X-ray nanoscale imaging will open new ways for various surface/interface studies.

Possible mechanism of adhesion in a mica supported phospholipid bilayer.

Phospholipid bilayers supported on hydrophilic solids like silica and mica play a substantial role in fundamental studies and technological applications of phospholipid membranes. In both cases the molecular mechanism of adhesion between the bilayer and the support is of primary interest. Since the possibilities of experimental methods in this specific area are rather limited, the methods of computer simulation acquire great importance. In this paper we use the grand canonical Monte Carlo technique and an atomistic force field to simulate the behavior of a mica supported phospholipid bilayer in pure water as a function of the distance between the bilayer and the support. The simulation reveals a possible adhesion mechanism, where the adhesion is due to individual lipid molecules that protrude from the bilayer and form widely spaced links with the support. Simultaneously, the bilayer remains separated from the bilayer by a thin water interlayer which maintains the bilayer fluidity.

Direct UV-induced functionalization of surface hydroxy groups by thiol-ol chemistry.

A novel UV-initiated surface modification method for the direct functionalization of surface hydroxy groups with thiol-containing molecules (termed "thiol-ol" modification) is described. This method is based on the oxidative conjugation of thiols to hydroxy groups. We demonstrate that different thiol-containing molecules, such as fluorophores, thiol-terminated poly(ethylene glycol) (PEG-SH), and a cysteine-containing peptide, can be attached onto the surface of porous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate). Direct functionalization of other hydroxy-group-bearing surfaces, fabrication of micropatterns, and double patterning have been also demonstrated using the thiol-ol method.

Hot embossed microtopographic gradients reveal morphological cues that guide the settlement of zoospores.

Among different surface cues, the settlement of cells and larvae of marine macrofouling organisms has been found to be strongly influenced by surface microtopographies. In this article, the settlement of zoospores of the green alga Ulva linza on a surface topographic gradient has been investigated. "Honeycomb" gradient structures with feature sizes ranging from 1 to 10 µm were prepared by hot embossing, and the effect on the density of spores that attached in settlement assays was quantified. The highest density of spores was found when the size of the microstructures was similar to or larger than the size of the spores. With decreasing size of the structures, spore settlement density decreased. Interestingly, spore settlement density correlated with the Wenzel roughness of the surfaces. "Kink sites" on the surface played an important role and resembled preferred attachment positions. Furthermore, the gradients allowed the minimum pit size that the spores were able to squeeze into to be determined.

Computer simulation of adhesion between hydrophilic and hydrophobic self-assembled monolayers in water.

The grand canonical Monte Carlo technique and atomistic force fields are used to calculate the force-distance relations and free energies of adhesion between carboxyl and methyl terminated alkanethiolate self-assembled monolayers (SAMs) in water. Both symmetric and asymmetric confinements are considered, as formed by like and unlike SAMs, respectively. As the confinement is increased, water confined by the hydrophobic methyl terminated SAMs experiences capillary evaporation. As a consequence, the adhesion energy is determined by the direct interaction between bare SAMs. In the asymmetric system, an incomplete capillary evaporation is observed, with the number of water molecules dropped by more than an order of magnitude. The remaining water molecules are all adsorbed on the hydrophilic SAM, while the hydrophobic SAM is separated from the rest of the system by a thin vapor layer. The calculated free energies of adhesion are in acceptable agreement with experiment.

Conditioning of surfaces by macromolecules and its implication for the settlement of zoospores of the green alga Ulva linza.

Conditioning, ie the adsorption of proteins and other macromolecules, is the first process that occurs in the natural environment once a surface is immersed in seawater, but no information is available either regarding the conditioning of surfaces by artificial seawater or whether conditioning affects data obtained from laboratory assays. A range of self-assembled monolayers (SAMs) with different chemical terminations was used to investigate the time-dependent formation of conditioning layers in commercial and self-prepared artificial seawaters. Subsequently, these results were compared with conditioning by solutions in which zoospores of the green alga Ulva linza had been swimming. Spectral ellipsometry and contact angle measurements as well as infrared reflection absorption spectroscopy (IRRAS) were used to reveal the thickness and chemical composition of the conditioning layers. The extent that surface preconditioning affected the settlement of zoospores of U. linza was also investigated. The results showed that in standard spore settlement bioassays (45-60 min), the influence of a molecular conditioning layer is likely to be small, although more substantial effects are possible at longer settlement times.

Shear stress regulates adhesion and rolling of CD44+ leukemic and hematopoietic progenitor cells on hyaluronan.

Leukemic cells and human hematopoietic progenitor cells expressing CD44 receptors have the ability to attach and roll on hyaluronan. We investigated quantitatively the adhesion behavior of leukemic cell lines and hematopoietic progenitor cells on thin films of the polysaccharides hyaluronan and alginate in a microfluidic system. An applied flow enhances the interaction between CD44-positive cells and hyaluronan if a threshold shear stress of 0.2 dyn/cm(2) is exceeded. At shear stress ∼1 dyn/cm(2), the cell rolling speed reaches a maximum of 15 µm/s. Leukemic Jurkat and Kasumi-1 cells lacking CD44-expression showed no adhesion or rolling on the polysaccharides whereas the CD44-expressing leukemic cells KG-1a, HL-60, K-562, and hematopoietic progenitor cells attached and rolled on hyaluronan. Interestingly, the observations of flow-induced cell rolling are related to those found in the recruitment of leukocytes to inflammatory sites and the mechanisms of stem-cell homing into the bone marrow.

Reduced hydrophobicity of the colonic mucosal surface in ulcerative colitis as a hint at a physicochemical barrier defect.

PURPOSE: There is increasing evidence that a defect of the gastrointestinal mucosal barrier is important for the development of inflammatory bowel diseases (IBD). The hydrophobicity of the colonic mucosal surface is a measure of its resistance to luminal antigens, e.g. of bacterial origin. Therefore, the purpose of this study was to determine this parameter in patients suffering from IBD. METHODS: Nineteen patients with ulcerative colitis (UC), ten patients with Crohn's disease (CD) and 20 controls were examined. All underwent colonic surgery at the University Hospital Heidelberg. Clinical disease activity was determined. From every subject, colonic tissue specimens were obtained, and hydrophobicity of the mucosal surface was determined with a goniometer by multiple plateau contact angle measurements. Histological evaluation of disease activity was performed in directly adjacent tissue specimens. RESULTS: Hydrophobicity of the colonic mucosal surface, expressed as plateau contact angles, was significantly reduced in patients with UC (mean ± SEM, 47.8° ± 3.4°) compared to those with CD (72.0° ± 5.2°) and controls (72.5° ± 5.6°; over-all P = 0.0004; UC versus controls, P < 0.001; UC versus CD, P < 0.05; CD versus controls, P > 0.05). Between mucosal hydrophobicity and clinical disease activity, as well as mucosal hydrophobicity and histological disease activity, no significant correlation was found. CONCLUSIONS: The results suggest a defective physicochemical barrier as an essential factor in the pathogenesis of UC, but not CD. The fact that no correlation was found between mucosal hydrophobicity and disease activity may indicate that the loss of mucosal hydrophobicity in UC is not exclusively a secondary effect due to inflammation.

Flow conditions in the vicinity of microstructured interfaces studied by holography and implications for the assembly of artificial actin networks.

Microstructured fluidic devices have successfully been used for the assembly of free standing actin networks as mechanical model systems on the top of micropillars. The assembly occurs spontaneously at the pillar heads when preformed filaments are injected into the channel. In order to reveal the driving mechanism of this localization, we studied the properties of the flow profile by holographic tracking. Despite the strong optical disturbances originating from the pillar field, 2 µm particles were traced with digital in-line holographic microscopy (DIHM). Trajectories in the pillar free region and local alterations of the flow profile induced by the channel structure in the pillar decorated region can be distinguished. Velocity histograms at different z-positions reveal that the laminar flow profile across the channel shows a difference between the minimum in the z-component of the velocity field and the maximum of the overall velocity. This minimum drag in vertical direction is present at the top of the pillars and explains why biopolymer networks readily assemble in this region instead of forming a homogeneous three-dimensional network in between the pillars. On the basis of the observations we propose a new mechanism for actin network formation on top of the microstructures.

Quasistatic computer simulations of shear behavior of water nanoconfined between mica surfaces.

We combine the grand canonical Monte Carlo and molecular dynamics techniques to simulate the shear response of water under a 9.2 Å confinement between two parallel sheets of muscovite mica. The shear deformation is modeled in the quasistatic regime corresponding to an infinitely small shear rate. It is found that the confined water film is capable of sustaining shear stress, as is characteristic of solids, while remaining fluid-like in respect of molecular mobility and lateral order. An important information is obtained by splitting the stress tensor components into contributions arising from the interaction of the opposing mica sheets between themselves and their interaction with water. The mica-mica contributions to shear stress show a strong anisotropy dictated by the alignment of the surface K(+) ions in chains along the x axis. On shearing in this direction, the mica-mica contribution to shear stress is negligible, so that the shear resistance is determined by the water interlayer. By contrast, in the y direction, the mica-mica contribution to shear resistance is dominant. The water-mica contribution is slightly less in magnitude but opposite in sign. As a consequence, the mica-mica contribution is largely canceled out. The physics behind this cancellation is the screening of the electrostatic interactions of the opposing surface K(+) ions by water molecules.

Building a community to engineer synthetic cells and organelles from the bottom-up.

Employing concepts from physics, chemistry and bioengineering, 'learning-by-building' approaches are becoming increasingly popular in the life sciences, especially with researchers who are attempting to engineer cellular life from scratch. The SynCell2020/21 conference brought together researchers from different disciplines to highlight progress in this field, including areas where synthetic cells are having socioeconomic and technological impact. Conference participants also identified the challenges involved in designing, manipulating and creating synthetic cells with hierarchical organization and function. A key conclusion is the need to build an international and interdisciplinary research community through enhanced communication, resource-sharing, and educational initiatives.

In situ characterization of thermo-responsive poly(N-isopropylacrylamide) films with sum-frequency generation spectroscopy.

The thermo-responsive behaviour of thiol modified poly(N-isopropylacrylamide) (pNIPAM) films immobilized on gold are probed by in situ broadband sum-frequency generation (SFG) spectroscopy. The pNIPAM films were prepared by atom transfer radical polymerization (ATRP) using a nitro-biphenyl-thiol (NBT)-SAM on a polycrystalline gold surface as a substrate. Additionally, Raman and infrared reflection absorption spectroscopy (IRRAS) are applied to spin-coated pNIPAM films. Molecular groups involved in the reorientation and disordering of the polymer chains during the LCST (lower critical solution temperature) transition of pNIPAM are identified. The characteristic vibrations of the CH(3) groups show a gradual reorientation of the isopropyl groups within the pNIPAM film and instantaneous reorientation of the outermost CH(3) groups around 32 degrees C.

Quantification of the adhesion strength of fibroblast cells on ethylene glycol terminated self-assembled monolayers by a microfluidic shear force assay.

The adhesion strength of cells depends on the properties of the surface they attach to. Varying the surface properties can trigger different cellular responses such as differentiation. In order to study cell adhesion quantitatively, we developed a microfluidic shear force assay which allows the variation of applied shear stress by five orders of magnitude. With this device we can determine the critical shear stress which is necessary to remove 50% of the adherent cells. As an application we investigated the adhesion strength of cells on a series of oligo(ethylene glycol) (OEG) containing self-assembled monolayers (SAMs). By varying the number of ethylene oxide units, the hydration properties of the monolayers are changed. We found that cell adhesion strength for mammalian fibroblasts decreases if the hydration of the surface is increased. As the cell spreading area changes with the substrate properties, the adhesion strength per unit area was additionally determined.

The role of "inert" surface chemistry in marine biofouling prevention.

The settlement and colonization of marine organisms on submerged man-made surfaces is a major economic problem for many marine industries. The most apparent detrimental effects of biofouling are increased fuel consumption of ships, clogging of membranes and heat exchangers, disabled underwater sensors, and growth of biofoulers in aquaculture systems. The presently common-but environmentally very problematic-way to deal with marine biofouling is to incorporate biocides, which use biocidal products in the surface coatings to kill the colonizing organisms, into the surface coatings. Since the implementation of the International Maritime Organization Treaty on biocides in 2008, the use of tributyltin (TBT) is restricted and thus environmentally benign but effective surface coatings are required. In this short review, we summarize the different strategies which are pursued in academia and industry to better understand the mechanisms of biofouling and to develop strategies which can be used for industrial products. Our focus will be on chemically "inert" model surface coatings, in particular oligo- and poly(ethylene glycol) (OEG and PEG) functionalized surface films. The reasons for choosing this class of chemistry as an example are three-fold: Firstly, experiments on spore settlement on OEG and PEG coatings help to understand the mechanism of non-fouling of highly hydrated interfaces; secondly, these studies defy the common assumption that surface hydrophilicity-as measured by water contact angles-is an unambiguous and predictive tool to determine the fouling behavior on the surface; and thirdly, choosing this system is a good example for "interfacial systems chemistry": it connects the behavior of unicellular marine organisms with the antifouling properties of a hydrated surface coating with structural and electronic properties as derived from ab initio quantum mechanical calculations using the electronic wave functions of oxygen, hydrogen, and carbon. This short review is written to outline for non-experts the hierarchical structure in length- and timescale of marine biofouling and the role of surface chemistry in fouling prevention. Experts in the field are referred to more specialized recent reviews.

Droplet-Microarray: Miniaturized Platform for High-Throughput Screening of Antimicrobial Compounds.

Currently, there are no time-saving and cost-effective high-throughput screening methods for the evaluation of bacterial drug-resistance. In this study, a droplet microarray (DMA) system is established as a miniaturized platform for high-throughput screening of antibacterial compounds using the emerging, opportunistic human pathogen Pseudomonas aeruginosa (P. aeruginosa) as a target. Based on the differences in wettability of DMA slides, a rapid method for generating microarrays of nanoliter-sized droplets containing bacteria is developed. The bacterial growth in droplets is evaluated using fluorescence. The new method enables immediate screening with libraries of antibiotics. A novel simple colorimetric readout method compatible with the nanoliter size of the droplets is established. Furthermore, the drug-resistance of P. aeruginosa 49, a multi-resistant strain from an environmental isolate, is investigated. This study demonstrates the potential of the DMA platform for the rapid formation of microarrays of bacteria for high-throughput drug screening.