James F. Schumacher

Engineered antifouling microtopographies – effect of feature size, geometry, and roughness on settlement of zoospores of the green alga Ulva

Abstract The effect of feature size, geometry, and roughness on the settlement of zoospores of the ship fouling alga Ulva was evaluated using engineered microtopographies in polydimethylsiloxane elastomer. The topographies studied were designed at a feature spacing of 2 μm and all significantly reduced spore settlement compared to a smooth surface. An indirect correlation between spore settlement and a newly described engineered roughness index (ERI) was identified. ERI is a dimensionless ratio based on Wenzels roughness factor, depressed surface fraction, and the degree of freedom of spore movement. Uniform surfaces of either 2 μm diameter circular pillars (ERI = 5.0) or 2 μm wide ridges (ERI = 6.1) reduced settlement by 36% and 31%, respectively. A novel multi-feature topography consisting of 2 μm diameter circular pillars and 10 μm equilateral triangles (ERI = 8.7) reduced spore settlement by 58%. The largest reduction in spore settlement, 77%, was obtained with the Sharklet AF™ topography (ERI = 9.5).

Engineered antifouling microtopographies - : correlating wettability with cell attachment

Abstract Bioadhesion and surface wettability are influenced by microscale topography. In the present study, engineered pillars, ridges and biomimetic topography inspired by the skin of fast moving sharks (Sharklet AF™) were replicated in polydimethylsiloxane elastomer. Sessile drop contact angle changes on the surfaces correlated well (R2 = 0.89) with Wenzel and Cassie and Baxters relationships for wettability. Two separate biological responses, i.e. settlement of Ulva linza zoospores and alignment of porcine cardiovascular endothelial cells, were inversely proportional to the width (between 5 and 20 μm) of the engineered channels. Zoospore settlement was reduced by ∼85% on the finer (ca 2 μm) and more complex Sharklet AF™ topographies. The response of both cell types suggests their responses are governed by the same underlying thermodynamic principles as wettability.

Impact of engineered surface microtopography on biofilm formation of Staphylococcus aureus

The surface of an indwelling medical device can be colonized by human pathogens that can form biofilms and cause infections. In most cases, these biofilms are resistant to antimicrobial therapy and eventually necessitate removal or replacement of the device. An engineered surface microtopography based on the skin of sharks, Sharklet AFTM, has been designed on a poly(dimethyl siloxane) elastomer (PDMSe) to disrupt the formation of bacterial biofilms without the use of bactericidal agents. The Sharklet AFTM PDMSe was tested against smooth PDMSe for biofilm formation of Staphylococcus aureus over the course of 21 days. The smooth surface exhibited early-stage biofilm colonies at 7 days and mature biofilms at 14 days, while the topographical surface did not show evidence of early biofilm colonization until day 21. At 14 days, the mean value of percent area coverage of S. aureus on the smooth surface was 54% compared to 7% for the Sharklet AFTM surface (p<0.01). These results suggest that surface modification of indwelling medical devices and exposed sterile surfaces with the Sharklet AFTM engineered topography may be an effective solution in disrupting biofilm formation of S. aureus.

Species-specific engineered antifouling topographies: correlations between the settlement of algal zoospores and barnacle cyprids

Abstract Novel, non-toxic antifouling technologies are focused on the manipulation of surface topography to deter settlement of the dispersal stages of fouling organisms. This study investigated the effect of the aspect ratio (feature height/feature width) of topographical features engineered in polydimethylsiloxane, on the settlement of cyprids of Balanus amphitrite and zoospores of Ulva linza. The correlation of relative aspect ratios to antifouling efficacy was proven to be significant. An increase in aspect ratio resulted in an increase of fouling deterrence for both zoospores and cyprids. The spore density of Ulva was reduced 42% with each unit increase in aspect ratio of the Ulva-specific Sharklet AF™ topography. Similarly, the number of settled cyprids was reduced 45% with each unit increase in aspect ratio. The newly described barnacle-specific Sharklet AF™ topography (40 μm feature height, aspect ratio of 2) reduced cyprid settled by 97%. Techniques have been developed to superimpose the smaller Ulva-specific topographies onto the barnacle-specific surfaces into a hierarchical structure to repel both organisms simultaneously. The results for spore settlement on first-generation hierarchical surfaces provide insight for the efficacious design of such structures when targeting multiple settling species.

Antifouling Potential of Lubricious, Micro-engineered, PDMS Elastomers against Zoospores of the Green Fouling Alga Ulva (Enteromorpha)

The settlement and release of Ulva spores from chemically modified, micro-engineered surface topographies have been investigated using poly(dimethyl siloxane) elastomers (PDMSe) with varying additions of non-network forming poly(dimethyl siloxane) based oils. The topographic features were based on 5u2009μm wide pillars or ridges separated by 5, 10, or 20u2009μm wide channels. Pattern depths were 5 or 1.5u2009μm. Swimming spores showed no marked difference in settlement on smooth surfaces covered with excess PDMS oils. However, incorporation of oils significantly reduced settlement density on many of the surfaces with topographic features, in particular, the 5u2009μm wide and deep channels. Previous results, confirmed here, demonstrate preferences by the spores to settle in channels and against pillars with spatial dimensions of 5u2009μm, 10u2009μm and 20u2009μm. The combination of lubricity and pillars significantly reduced the number of attached spores compared to the control, smooth, unmodified PDMSe surfaces when exposed to turbulent flow in a flow channel. The results are discussed in relation to the energy needs for spores to adhere to various surface features and the concepts of ultrahydrophobic surfaces. A factorial, multi-level experimental design was analyzed and a 2nd order polynomial model was regressed for statistically significant effects and interactions to determine the magnitude and direction of influence on the spore density measurements between factor levels.

Engineered nanoforce gradients for inhibition of settlement (attachment) of swimming algal spores.

Current antifouling strategies are focused on the development of environmentally friendly coatings that protect submerged surfaces from the accumulation of colonizing organisms (i.e., biofouling). One ecofriendly approach is the manipulation of the surface topography on nontoxic materials to deter settlement of the dispersal stages of fouling organisms. The identification of effective antifouling topographies typically occurs through trial-and-error rather than predictive models. We present a model and design methodology for the identification of nontoxic, antifouling surface topographies for use in the marine environment by the creation of engineered nanoforce gradients. The design and fabrication of these gradients incorporate discrete micrometer-sized features that are associated with the species-specific surface design technique of engineered topography and the concepts of mechanotransduction. The effectiveness of designed nanoforce gradients for antifouling applications was tested by evaluating the settlement behavior of zoospores of the alga Ulva linza. The surfaces with nanoforce gradients ranging from 125 to 374 nN all significantly reduced spore settlement relative to a smooth substrate, with the highest reduction, 53%, measured on the 374 nN gradient surface. These results confirm that the designed nanoforce gradients may be an effective tool and predictive model for the design of unique nontoxic, nonfouling surfaces for marine applications as well as biomedical surfaces in the physiological environment.

A model that predicts the attachment behavior of Ulva linza zoospores on surface topography

A predictive model for the attachment of spores of the green alga Ulva on patterned topographical surfaces was developed using a constant refinement approach. This ‘attachment model’ incorporated two historical data sets and a modified version of the previously-described Engineered Roughness Index. Two sets of newly-designed surfaces were used to evaluate the effect of two components of the model on spore settlement. Spores attached in fewer numbers when the area fraction of feature tops increased or when the number of distinct features in the design increased, as predicted by the model. The model correctly predicted the spore attachment density on three previously-untested surfaces relative to a smooth surface. The two historical data sets and two new data sets showed high correlation (R 2 = 0.88) with the model. This model may be useful for designing new antifouling topographies.

Potential for Tunable Static and Dynamic Contact Angle Anisotropy on Gradient Microscale Patterned Topographies

Translationally symmetric topographies can be designed to induce anisotropy of static and dynamic contact angles. The validity of ignoring directionality of topography in contact angle characterization was evaluated using microscale patterned topographies. Seven patterned topographies comprising elongated discontinuous microfeatures oriented along parallel paths and one topography comprising ridges were fabricated in a poly(dimethyl siloxane) elastomer (PDMSe). The static contact angle, advancing contact angle, receding contact angle, contact angle hysteresis, and slip angle were measured using water on each surface at three in-plane perspectives, with respect to the feature orientation. Static and dynamic contact angle anisotropies were investigated on the topographies to evaluate the effect of discontinuities along the feature lengths on the anisotropy that has been shown on channels or ridges in previous reports. Discontinuous feature topographies exhibited a statistically significant anisotropy of 2 degrees-6 degrees between the perpendicular and parallel directions, with respect to the static and dynamic contact angles. The ridges topography exhibited much larger 5 degrees-42 degrees anisotropy in the contact angles. The discontinuities along the feature lengths greatly reduced, but did not eliminate, the anisotropies compared to the ridges. This evidence of contact angle anisotropy indicates a need to identify the orientation of topography, in relation to contact angle measurements. It also implies a need to consider directionality in the design of microfluidic devices and self-cleaning surfaces.

Engineering high-density endothelial cell monolayers on soft substrates

This study demonstrates that a confluent monolayer of endothelial cells (ECs) can be tissue engineered on a soft substrate with a cell density and morphology that approximates in vivo conditions. We achieved formation of a confluent EC monolayer on polydimethylsiloxane (PDMS) elastomer by microcontact printing of fibronectin (FN) in a square lattice array of 3microm diameter circular islands at a 6microm pitch. Uniform coatings of FN or serum proteins on PDMS or on tissue-culture-treated polystyrene failed to support the equivalent EC density and/or confluence. The ECs on the FN micropatterned PDMS achieved a density of 1,536+/-247cellsmm(-2), close to the 3,215+/-336cellsmm(-2) observed in vivo from porcine pulmonary artery and significantly higher (2- to 5-fold) than EC density on other materials. The probable mechanism for enhanced EC adhesion, growth and density is increased focal adhesion (FA) formation between the ECs and the substrate. After 14days culture, the micropatterned FN surface increased the average number of FAs per cell to 35+/-10, compared to 7+/-6 for ECs on PDMS uniformly coated with FN. Thus, microscale patterning of FN into FA-sized, circular islands on PDMS elastomer promotes the formation of EC monolayers with in vivo-like cell density and morphology.