Novel grooved substrata stimulate macrophage fusion, CCL2 and MMP-9 secretion.

Rough surface topographies on implants attract macrophages but the influence of topography on macrophage fusion to produce multinucleated giant cells (MGC) and foreign body giant cells (FBGC) is unclear. Two rough novel grooved substrata, G1 and G2, fabricated by anisotropic etching of Silicon <110> crystals without the use of photolithographic patterning, and a control smooth surface (Pol) were produced and replicated in epoxy. The surfaces were compared for their effects on RAW264.7 macrophage morphology, gene expression, cyto/chemokine secretion, and fusion for one and five days. Macrophages on grooved surfaces exhibited an elongated morphology similar to M2 macrophages and increased cell alignment with surface directionality, roughness and cell culture time. Up-regulated expression of macrophage chemoattractants at gene and protein level was observed on both grooved surfaces relative to Pol. Grooved surfaces showed time-dependent increase in soluble mediators involved in cell fusion, CCL2 and MMP-9, and an increased proportion of multinucleated cells at Day 5. Collectively, this study demonstrated that a rough surface with surface directionality produced changes in macrophage shape and macrophage attractant chemokines and soluble mediators involved in cell fusion. These in vitro results suggest a possible explanation for the observed accumulation of macrophages and MGCs on rough surfaced implants in vivo. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2243-2254, 2016.

Orthogonal Morphological Feature Size and Density Gradients for Exploring Synergistic Effects in Biology.

Gradient surfaces enable rapid screening and high-throughput investigations in various fields, such as biology and tribology. A new method is described for the preparation of material-independent morphological gradients, in which the density and height of roughness features are varied along two orthogonal axes. A polystyrene-particle-density gradient was produced by a dip-coating process on titanium-oxide-coated silicon wafers. A controlled exposure to ultraviolet light enabled the generation of a particle-height gradient in the orthogonal direction. These gradients were replicated to generate material-independent morphology gradients. MC3T3 cell proliferation studies were performed on titanium-coated replicas and showed a higher cell density on the high-feature-density region of the gradient. The cell area coverage was found to increase with decreasing particle height.

Ultrathin, freestanding, stimuli-responsive, porous membranes from polymer hydrogel-brushes.

The fabrication of freestanding, sub-100 nm-thick, pH-responsive hydrogel membranes with controlled nano-morphology, based on modified poly(hydroxyethyl methacrylate) (PHEMA) is presented. Polymer hydrogel-brush films were first synthesized by surface-initiated atom transfer radical polymerization (SI-ATRP) and subsequently detached from silicon substrates by UV-induced photo-cleavage of a specially designed linker within the initiator groups. The detachment was also assisted by pH-induced osmotic forces generated within the films in the swollen state. The mechanical properties and morphology of the freestanding films were studied by atomic force microscopy (AFM). Inclusion of nanopores of controlled diameter was accomplished by performing SI-ATRP from initiator-coated surfaces that had previously been patterned with polystyrene nanoparticles. Assembly parameters and particle sizes could be varied, in order to fabricate nanoporous hydrogel-brush membranes with tunable pore coverage and characteristics. Additionally, due to the presence of weak polyacid functions within the hydrogel, the membranes exhibited pH-dependent thickness in water and reversible opening/closing of the pores.

Patterning gradients.

Surface-chemical and -morphological gradients can be extremely useful in cell-biological research as high-throughput screening tools-for example, exposing a given set of cells to many different surface conditions at once, under identical ambient conditions, in order to monitor cell behavior such as proliferation or specific gene expression. They can also be used to investigate the effects of gradients themselves on cell behavior, such as migration. A number of simple, reliable techniques for both chemical- and morphological-gradient fabrication have been developed in our laboratories and are described in detail in the following.

Template-stripped, ultraflat gold surfaces with coplanar, embedded titanium micropatterns.

Ultraflat gold surfaces with coplanar, embedded titanium micropatterns, exhibiting extremely low roughness over the entire surface, have been obtained by a modified template-stripping procedure. Titanium is deposited onto photolithographically predefined regions of a silicon template. Following photoresist lift-off, the entire surface is backfilled with gold, template stripping is conducted, and an ultraflat micropatterned surface is revealed. Atomic force microscopy confirms a roughness of <0.5 nm RMS on both Ti and Au regions, with a topographically indistinguishable gold-titanium interface. Detailed surface-chemical maps of the patterned surfaces have been obtained by means of imaging X-ray photoelectron spectroscopy (i-XPS) as well as time-of-flight secondary-ion mass spectrometry (ToF-SIMS). They confirm the presence of well-separated Ti and Au regions, with a chemical contrast that is sharp (as determined by ToF-SIMS) and complete (as determined by i-XPS) across the Ti-Au interface. Thus, a surface has been fabricated that is physically homogeneous down to the nanoscale incorporating chemically distinct micropatterns consisting of two different metals, with totally contrasting surface chemistries.