Removal of Free Liquid Layer from Liquid-Infused Catheters Reduces Silicone Loss into the Environment while Maintaining Adhesion Resistance.

Silicone urinary catheters infused with silicone liquid offer an effective alternative to antibiotic coatings, reducing microbial adhesion while decreasing bladder colonization and systemic dissemination. However, loss of free silicone liquid from the surface into the host system is undesirable. To reduce the potential for liquid loss, free silicone liquid was removed from the surface of liquid-infused catheters by either removing excess liquid from fully infused samples or by partial infusion. The effect on bacterial and host protein adhesion was then assessed. Removing the free liquid from fully infused samples resulted in a ~64% decrease in liquid loss into the environment compared to controls, with no significant increase in deposition of the host protein fibrinogen or the adhesion of the common uropathogen Enterococcus faecalis. Partially infusing samples decreased liquid loss as total liquid content decreased, with samples infused to 70-80% of their maximum capacity showing a ~85% reduction in liquid loss compared to fully infused controls. Furthermore, samples above 70% infusion showed no significant increase in fibrinogen or E. faecalis adhesion. Together, the results suggest that eliminating free liquid layer, mechanically or through partial infusion, can reduce liquid loss from liquid-infused catheters while preserving functionality.

DESIGNER fraction concept unmasks minor bioactive constituents in red clover (Trifolium pratense L.).

In botanical extracts, highly abundant constituents can mask or dilute the effects of other, and often, more relevant biologically active compounds. To facilitate the rational chemical and biological assessment of these natural products with wide usage in human health, we introduced the DESIGNER approach of Depleting and Enriching Selective Ingredients to Generate Normalized Extract Resources. The present study applied this concept to clinical Red Clover Extract (RCE) and combined phytochemical and biological methodology to help rationalize the utility of RCE supplements for symptom management in postmenopausal women. Previous work has demonstrated that RCE reduces estrogen detoxification pathways in breast cancer cells (MCF-7) and, thus, may serve to negatively affect estrogen metabolism-induced chemical carcinogenesis. Clinical RCE contains ca. 30% of biochanin A and formononetin, which potentially mask activities of less abundant compounds. These two isoflavonoids are aryl hydrocarbon receptor (AhR) agonists that activate P450 1A1, responsible for estrogen detoxification, and P450 1B1, producing genotoxic estrogen metabolites in female breast cells. Clinical RCE also contains the potent phytoestrogen, genistein, that downregulates P450 1A1, thereby reducing estrogen detoxification. To identify less abundant bioactive constituents, countercurrent separation (CCS) of a clinical RCE yielded selective lipophilic to hydrophilic metabolites in six enriched DESIGNER fractions (DFs 01-06). Unlike solid-phase chromatography, CCS prevented any potential loss of minor constituents or residual complexity (RC) and enabled the polarity-based enrichment of certain constituents. Systematic analysis of estrogen detoxification pathways (ERα-degradation, AhR activation, CYP1A1/CYP1B1 induction and activity) of the DFs uncovered masked bioactivity of minor/less abundant constituents including irilone. These data will allow the optimization of RCE with respect to estrogen detoxification properties. The DFs revealed distinct biological activities between less abundant bioactives. The present results can inspire future carefully designed extracts with phytochemical profiles that are optimized to increase in estrogen detoxification pathways and, thereby, promote resilience in women with high-risk for breast cancer. The DESIGNER approach helps to establish links between complex chemical makeup, botanical safety and possible efficacy parameters, yields candidate DFs for (pre)clinical studies, and reveals the contribution of minor phytoconstituents to the overall safety and bioactivity of botanicals, such as resilience promoting activities relevant to women's health.

Breast cancer prevention with liquiritigenin from licorice through the inhibition of aromatase and protein biosynthesis in high-risk women's breast tissue.

Breast cancer risk continues to increase post menopause. Anti-estrogen therapies are available to prevent postmenopausal breast cancer in high-risk women. However, their adverse effects have reduced acceptability and overall success in cancer prevention. Natural products such as hops (Humulus lupulus) and three pharmacopeial licorice (Glycyrrhiza) species have demonstrated estrogenic and chemopreventive properties, but little is known regarding their effects on aromatase expression and activity as well as pro-proliferation pathways in human breast tissue. We show that Gycyrrhiza inflata (GI) has the highest aromatase inhibition potency among these plant extracts. Moreover, phytoestrogens such as liquiritigenin which is common in all licorice species have potent aromatase inhibitory activity, which is further supported by computational docking of their structures in the binding pocket of aromatase. In addition, GI extract and liquiritigenin suppress aromatase expression in the breast tissue of high-risk postmenopausal women. Although liquiritigenin has estrogenic effects in vitro, with preferential activity through estrogen receptor (ER)-ß, it reduces estradiol-induced uterine growth in vivo. It downregulates RNA translation, protein biosynthesis, and metabolism in high-risk women's breast tissue. Finally, it reduces the rate of MCF-7 cell proliferation, with repeated dosing. Collectively, these data suggest that liquiritigenin has breast cancer prevention potential for high-risk postmenopausal women.

Continuous, Nondestructive Detection of Microorganism Growth at Buried Interfaces with Vascularized Polymers.

Evaluating surface bacterial growth at buried interfaces can be problematic due to the difficulties associated with obtaining samples. In this work, we present a new method to detect signals from microorganisms at buried interfaces that is nondestructive and can be conducted continuously. Inspired by vascular systems in nature that permit chemical communication between the surface and underlying tissues of an organism, we created a system in which an inert carrier fluid could be introduced into an empty vascular network embedded in a polymer matrix. When a microorganism layer was grown on top, small molecules produced by the growth process would diffuse down into the carrier fluid, which could then be collected and analyzed. We used this system to nondestructively detect signals from a surface layer of Escherichia coli using conductivity, ultraviolet-visible (UV-vis) absorbance spectroscopy, and high-performance liquid chromatography (HPLC) for organic acids, methods that ranged in sensitivity, time-to-result, and cost. Carrier fluid from sample vascularized polymers with surface bacterial growth recorded significantly higher values in both conductivity and absorbance at 350 nm compared to controls with no bacteria after 24 h. HPLC analysis showed three clear peaks that varied between the samples with bacteria and the controls without. Tests tracking the change in signals over 48 h showed clear trends that matched the bacterial growth curves, demonstrating the system's ability to monitor changes over time. A 2D finite element model of the system closely matched the experimental results, confirming the predictability of the system. Finally, tests using clinically relevant Staphylococcus aureus and Pseudomonas aeruginosa yielded differences in conductivity, absorbance, and HPLC peak areas unique to each species. This work lays the foundation for the use of vascularized polymers as an adaptive system for the continuous, nondestructive detection of surface microorganisms at buried interfaces in both industry and medicine.

Improved Recovery of Captured Airborne Bacteria and Viruses with Liquid-Coated Air Filters.

The COVID-19 pandemic has revealed the importance of the detection of airborne pathogens. Here, we present composite air filters featuring a bioinspired liquid coating that facilitates the removal of captured aerosolized bacteria and viruses for further analysis. We tested three types of air filters: commercial polytetrafluoroethylene (PTFE), which is well known for creating stable liquid coatings, commercial high-efficiency particulate air (HEPA) filters, which are widely used, and in-house-manufactured cellulose nanofiber mats (CNFMs), which are made from sustainable materials. All filters were coated with omniphobic fluorinated liquid to maximize the release of pathogens. We found that coating both the PTFE and HEPA filters with liquid improved the rate at which Escherichia coli was recovered using a physical removal process compared to uncoated controls. Notably, the coated HEPA filters also increased the total number of recovered cells by 57%. Coating the CNFM filters did not improve either the rate of release or the total number of captured cells. The most promising materials, the liquid-coated HEPA, filters were then evaluated for their ability to facilitate the removal of pathogenic viruses via a chemical removal process. Recovery of infectious JC polyomavirus, a nonenveloped virus that attacks the central nervous system, was increased by 92% over uncoated controls; however, there was no significant difference in the total amount of genomic material recovered compared to that of controls. In contrast, significantly more genomic material was recovered for SARS-CoV-2, the airborne, enveloped virus, which causes COVID-19, from liquid-coated filters. Although the amount of infectious SARS-CoV-2 recovered was 58% higher, these results were not significantly different from uncoated filters due to high variability. These results suggest that the efficient recovery of airborne pathogens from liquid-coated filters could improve air sampling efforts, enhancing biosurveillance and global pathogen early warning.

Inhibiting host-protein deposition on urinary catheters reduces associated urinary tract infections.

Microbial adhesion to medical devices is common for hospital-acquired infections, particularly for urinary catheters. If not properly treated these infections cause complications and exacerbate antimicrobial resistance. Catheter use elicits bladder inflammation, releasing host serum proteins, including fibrinogen (Fg), into the bladder, which deposit on the urinary catheter. Enterococcus faecalis uses Fg as a scaffold to bind and persist in the bladder despite antibiotic treatments. Inhibition of Fg-pathogen interaction significantly reduces infection. Here, we show deposited Fg is advantageous for uropathogens E. faecalis, Escherichia coli, Pseudomonas aeruginosa, K. pneumoniae, A. baumannii, and C. albicans, suggesting that targeting catheter protein deposition may reduce colonization creating an effective intervention for catheter-associated urinary tract infections (CAUTIs). In a mouse model of CAUTI, host-protein deposition was reduced, using liquid-infused silicone catheters, resulting in decreased colonization on catheters, in bladders, and dissemination in vivo. Furthermore, proteomics revealed a significant decrease in deposition of host-secreted proteins on liquid-infused catheter surfaces. Our findings suggest targeting microbial-binding scaffolds may be an effective antibiotic-sparing intervention for use against CAUTIs and other medical device infections.

Liquid-Infused Membranes Exhibit Stable Flux and Fouling Resistance.

Antifouling membranes that offer excellent operational lifetimes are critical technologies needed to meet the growing demand for clean water. In this study, we demonstrate antifouling membranes featuring an ultrathin oil layer that stayed immobilized on the surface and in the pore walls of poly(vinylidene fluoride) membranes for multiple cycles of operation at industrially relevant transmembrane pressures. An optimized quantity of a commercial Krytox oil with either a low (K103) or a high viscosity (K107) was infused onto the active surface and into the pores of membranes with a 0.45 µm pore size. The presence of the oil layer was qualitatively confirmed using crystal violet staining and variable pressure scanning electron microscopy. Using a dead-end stirred cell, a consistent pure water permeance value of 3000 L m-2 h-1 bar-1 was achieved for the K103 liquid-infused membranes for at least 10 operation cycles, which was expectedly lower than the permeance of bare control membranes (∼16 000 L m-2 h-1 bar-1), suggesting that a stable oil layer was formed on all membrane-active sites. To quantify if oil was lost during membrane operation, extensive thermogravimetric analysis was conducted on both the as-prepared and used membranes. When challenged with the microorganism, Escherichia coli K12, the liquid-infused membranes statistically reduced microbial attachment by ∼50% versus the control membranes. For the first time, we have demonstrated that by forming an immobilized, robust, and stable oil-coated membrane, we can generate high-performance membranes with stable permeance values that can be operated at relevant transmembrane pressures and provide long-lasting antifouling properties.

Characterization of novel cellulose nanofibril and phenolic acid-based active and hydrophobic packaging films.

Cellulose nanofibril (CNF) is a natural biodegradable biopolymer with excellent mechanical and barrier properties. However, it is susceptible to moisture-induced deterioration of its properties. Attachment of phenolic acids can improve its hydrophobicity and provide additional active functionalities such as antioxidant properties. In this study, CNF films were esterified to vanillic and syringic acid through two different reaction mechanisms. The films were investigated for evidence of modification, hydrophobicity, mechanical properties, crystallinity, thermal stability, and antioxidant properties. Results indicate that esterification with vanillic and syringic acids imparted antioxidant activity to CNF films, with a significantly higher ABTS+· scavenging activity (76 ± 18%) when compared to control CNF films (30 ± 6%). Similarly, esterification of phenolic acids significantly improved the hydrophobicity of the films with a water contact angle of 94 ± 3° when compared to control CNF films (46 ± 5°). Covalent attachment of phenolic acids can improve hydrophobicity while providing additional functionality to CNF important for food packaging applications.

3D printing direct to industrial roll-to-roll casting for fast prototyping of scalable microfluidic systems.

Microfluidic technologies have enormous potential to offer breakthrough solutions across a wide range of applications. However, the rate of scale-up and commercialization of these technologies has lagged significantly behind promising breakthrough developments in the lab, due at least in part to the problems presented by transitioning from benchtop fabrication methods to mass-manufacturing. In this work, we develop and validate a method to create functional microfluidic prototype devices using 3D printed masters in an industrial-scale roll-to-roll continuous casting process. There were no significant difference in mixing performance between the roll-to-roll cast devices and the PDMS controls in fluidic mixing tests. Furthermore, the casting process provided information on the suitability of the prototype microfluidic patterns for scale-up. This work represents an important step in the realization of high-volume prototyping and manufacturing of microfluidic patterns for use across a broad range of applications.

Functionality of Surface Mycelium Interfaces in Wood Bonding.

Filamentous fungi have been considered as candidates to replace petroleum-based adhesives and plastics in novel composite material production, particularly those containing lignocellulosic materials. However, the nature of the role of surface mycelium in the adhesion between lignocellulosic composite components is not well-known. The current study investigated the functionality of surface mycelium for wood bonding by incubating Trametes versicolor on yellow birch veneers and compared the lap-shear strengths after hot-pressing to evaluate if the presence of surface mycelium can improve the interface between two wood layers and consequently improve bonding. We found that the lap-shear strength of the samples was enhanced by the increase of surface mycelium coverage up to 8 days of incubation (up to 1.74 MPa) without a significant wood weight loss. We provide evidence that the bottom surface of the mycelium layer is more hydrophilic, contains more small-scale filamentous structure and contains more functional groups, resulting in better bonding with wood than the top surface. These observations confirm and highlight the functionality of the surface mycelium layer for wood bonding and provide useful information for future developments in fully biobased composites manufacturing.

6-Prenylnaringenin from Hops Disrupts ERα-Mediated Downregulation of CYP1A1 to Facilitate Estrogen Detoxification.

Botanical dietary supplements (BDS) containing hops are sold as women's health supplements due to the potent hop phytoestrogen, 8-prenylnaringenin (8-PN), and the cytoprotective chalcone, xanthohumol. Previous studies have shown a standardized hop extract to beneficially influence chemical estrogen carcinogenesis in vitro by fostering detoxified 2-hydroxylation over genotoxic 4-hydroxylation estrogen metabolism. In this study, hop extract and its bioactive compounds were investigated for its mechanism of action within the chemical estrogen carcinogenesis pathway, which is mainly mediated through the 4-hydroxylation pathway catalyzed by CYP1B1 that can form gentoxic quinones. Aryl hydrocarbon receptor (AhR) agonists induce CYP1A1 and CYP1B1, while estrogen receptor alpha (ERα) inhibits transcription of CYP1A1, the enzyme responsible for 2-hydroxylated estrogens and the estrogen detoxification pathway. An In-Cell Western MCF-7 cell assay revealed hop extract and 6-prenylnaringenin (6-PN) degraded ERα via an AhR-dependent mechanism. Reverse transcription PCR and xenobiotic response element luciferase assays showed hop extract and 6-PN-mediated activation of AhR and induction of CYP1A1. A reduction in estrogen-mediated DNA (cytosine-5)-methyltransferase 1 (DNMT1) downregulation of CYP1A1 accompanied this activity in a chromatin immunoprecipitation assay. Ultimately, hop extract and 6-PN induced preferential metabolism of estrogens to their detoxified form in vitro. These results suggest that the standardized hop extract and 6-PN activate AhR to attenuate epigenetic inhibition of CYP1A1 through degradation of ERα, ultimately increasing 2-hydroxylated estrogens. A new mechanism of action rationalizes the positive influence of hop BDS and 6-PN on oxidative estrogen metabolism in vitro and, thus, potentially on chemical estrogen carcinogenesis. The findings underscore the importance of elucidating various biological mechanisms of action and standardizing BDS to multiple phytoconstituents for optimal resilience promoting properties.

SAR Study on Estrogen Receptor α/ß Activity of (Iso)flavonoids: Importance of Prenylation, C-Ring (Un)Saturation, and Hydroxyl Substituents.

Many botanicals used for women's health contain estrogenic (iso)flavonoids. The literature suggests that estrogen receptor beta (ERß) activity can counterbalance estrogen receptor alpha (ERα)-mediated proliferation, thus providing a better safety profile. A structure-activity relationship study of (iso)flavonoids was conducted to identify ERß-preferential structures, overall estrogenic activity, and ER subtype estrogenic activity of botanicals containing these (iso)flavonoids. Results showed that flavonoids with prenylation on C8 position increased estrogenic activity. C8-prenylated flavonoids with C2-C3 unsaturation resulted in increased ERß potency and selectivity [e.g., 8-prenylapigenin (8-PA), EC50 (ERß): 0.0035 ± 0.00040 µM], whereas 4'-methoxy or C3 hydroxy groups reduced activity [e.g., icaritin, EC50 (ERß): 1.7 ± 0.70 µM]. However, nonprenylated and C2-C3 unsaturated isoflavonoids showed increased ERß estrogenic activity [e.g., genistein, EC50 (ERß): 0.0022 ± 0.0004 µM]. Licorice (Glycyrrhiza inflata, [EC50 (ERα): 1.1 ± 0.20; (ERß): 0.60 ± 0.20 µg/mL], containing 8-PA, and red clover [EC50 (ERα): 1.8 ± 0.20; (ERß): 0.45 ± 0.10 µg/mL], with genistein, showed ERß-preferential activity as opposed to hops [EC50 (ERα): 0.030 ± 0.010; (ERß): 0.50 ± 0.050 µg/mL] and Epimedium sagittatum [EC50 (ERα): 3.2 ± 0.20; (ERß): 2.5 ± 0.090 µg/mL], containing 8-prenylnaringenin and icaritin, respectively. Botanicals with ERß-preferential flavonoids could plausibly contribute to ERß-protective benefits in menopausal women.

Lateral field excited quartz crystal microbalances for biosensing applications.

The most common bulk acoustic wave device used in biosensing applications is the quartz crystal microbalance (QCM), in which a resonant pure shear acoustic wave is excited via electrodes on both major faces of a thin AT-cut quartz plate. For biosensing, the QCM is used to detect the capture of a target by a target-capture film. The sensitivity of the QCM is typically based solely on the detection of mechanical property changes, as electrical property change detection is limited by the electrode on its sensing surface. A modification of the QCM called the lateral field excited (LFE) QCM (LFE-QCM) has been developed with a bare sensing surface as both electrodes are now on a single face of the quartz plate. Compared to the QCM, the LFE-QCM exhibits significantly higher sensitivity to both electrical and mechanical property changes. This paper presents theoretical and experimental aspects of LFE-QCMs. In particular, the presence and strength of the usual and newfound LFE-QCM modes depend on the electrical properties of the film and/or sensing environment. This work also presents examples of experimental setups for measuring the response of an LFE-QCM, followed by results of LFE-QCMs used to detect liquid electrical and mechanical properties, chemical targets, and biological targets. Finally, details are given about the attachment of various target-capture films to the LFE-QCM surface to capture biomarkers associated with diseases such as cancer.

Vascularized Polymers Spatially Control Bacterial Cells on Surfaces.

Nature uses vascular systems to permit large-area control over the functionality of surfaces that lie above them. In this work, the application of this concept to the control of a hybrid living-nonliving system is demonstrated. Defined arrangements of vascular channels are created in agar using a fugitive ink printing method. The antibiotic gentamicin is then introduced into the vascular network where it diffuses to the surface and interacts with a model system of Escherichia coli cells. The cells either live or die depending on their distance from the underlying channels, permitting spatial control over the biological system. Using single-channel systems to define critical parameters, a theoretical model is developed to define the final surface pattern based solely on the arrangement of the underlying vascular channels. The model is then successfully used to create more complex arrangements of cells at the surface. Finally, by introducing different types of active compounds into separate vascular channels, a mixture of bacterial species is separated and localized at defined points. This work demonstrates the ability of bioinspired embedded vascular systems to predictably control a biological system at a surface, laying the groundwork for future spatially and temporally controlled biointerfaces in both industry and medicine.

Combining the geometry of folded paper with liquid-infused polymer surfaces to concentrate and localize bacterial solutions.

Point-of-care (POC) detection and diagnostic platforms provide critical information about health and safety conditions in austere and resource-limited settings in which medical, military, and disaster relief operations are conducted. In this work, low-cost paper materials commonly used in POC devices are coated with liquid-infused polymer surfaces and folded to produce geometries that precisely localize complex liquid samples undergoing concentration by evaporation. Liquid-infused polymer surfaces were fabricated by infusing silicone-coated paper with a chemically compatible polydimethylsiloxane oil to create a liquid overlayer. Tests on these surfaces showed no remaining bacterial cells after exposure to a sliding droplet containing a concentrated solution of Escherichia coli or Staphylococcus aureus, while samples without a liquid layer showed adhesion of both microdroplets and individual bacterial cells. Folding of the paper substrates with liquid-infused polymer surfaces into several functional 3D geometries enabled a clean separation and simultaneous concentration of a liquid containing rhodamine dye into discrete, predefined locations. When used with bacteria, which are known for their ability to adhere to nearly any surface type, functional geometries with liquid-infused polymer surfaces concentrated the cells at levels significantly higher than geometries with dry control surfaces. These results show the potential of synergistically combining paper-based materials with liquid-infused polymer surfaces for the manipulation and handling of complex samples, which may help the future engineering of POC devices.

Evidence for Chemopreventive and Resilience Activity of Licorice: Glycyrrhiza Glabra and G. Inflata Extracts Modulate Estrogen Metabolism in ACI Rats.

Women are increasingly using botanical dietary supplements (BDS) to reduce menopausal hot flashes. Although licorice (Glycyrrhiza sp.) is one of the frequently used ingredients in BDS, the exact plant species is often not identified. We previously showed that in breast epithelial cells (MCF-10A), Glycyrrhiza glabra (GG) and G. inflata (GI), and their compounds differentially modulated P450 1A1 and P450 1B1 gene expression, which are responsible for estrogen detoxification and genotoxicity, respectively. GG and isoliquiritigenin (LigC) increased CYP1A1, whereas GI and its marker compound, licochalcone A (LicA), decreased CYP1A1 and CYP1B1 The objective of this study was to determine the distribution of the bioactive licorice compounds, the metabolism of LicA, and whether GG, GI, and/or pure LicA modulate NAD(P)H quinone oxidoreductase (NQO1) in an ACI rat model. In addition, the effect of licorice extracts and compounds on biomarkers of estrogen chemoprevention (CYP1A1) as well as carcinogenesis (CYP1B1) was studied. LicA was extensively glucuronidated and formed GSH adducts; however, free LicA as well as LigC were bioavailable in target tissues after oral intake of licorice extracts. GG, GI, and LicA caused induction of NQO1 activity in the liver. In mammary tissue, GI increased CYP1A1 and decreased CYP1B1, whereas GG only increased CYP1A1 LigC may have contributed to the upregulation of CYP1A1 after GG and GI administration. In contrast, LicA was responsible for GI-mediated downregulation of CYP1B1 These studies highlight the polypharmacologic nature of botanicals and the importance of standardization of licorice BDS to specific Glycyrrhiza species and to multiple constituents.

Designing Liquid-Infused Surfaces for Medical Applications: A Review.

The development of new technologies is key to the continued improvement of medicine, relying on comprehensive materials design strategies that can integrate advanced therapeutic and diagnostic functions with a variety of surface properties such as selective adhesion, dynamic responsiveness, and optical/mechanical tunability. Liquid-infused surfaces have recently come to the forefront as a unique approach to surface coatings that can resist adhesion of a wide range of contaminants on medical devices. Furthermore, these surfaces are proving highly versatile in enabling the integration of established medical surface treatments alongside the antifouling capabilities, such as drug release or biomolecule organization. Here, the range of research being conducted on liquid-infused surfaces for medical applications is presented, from an understanding of the basics behind the interactions of physiological fluids, microbes, and mammalian cells with liquid layers to current applications of these materials in point-of-care diagnostics, medical tubing, instruments, implants, and tissue engineering. Throughout this exploration, the design parameters of liquid-infused surfaces and how they can be adapted and tuned to particular applications are discussed, while identifying how the range of controllable factors offered by liquid-infused surfaces can be used to enable completely new and dynamic approaches to materials and devices for human health.

Tunability of liquid-infused silicone materials for biointerfaces.

The ability to control the properties of bio-inspired liquid-infused surfaces is of interest in a wide range of applications. Liquid layers created using oil-infused polydimethylsiloxane elastomers offer a potentially simple way of accomplishing this goal through the adjustment of parameters such as curing agent ratio and oil viscosity. In this work, the effect of tuning these compositional parameters on the properties of the infused polymer are investigated, including infusion dynamics, stiffness, longevity in the face of continuous liquid overlayer removal, and resistance to bacterial adhesion. It is found that that curing agent concentration appears to have the greatest impact on the functionality of the system, with a lower base-to-curing agent ratio resulting in both increased longevity and improved resistance to adhesion by Escherichia coli. A demonstration of how these findings may be implemented to introduce patterned wettability to the surface of the infused polymers is presented by controlling the spatial arrangement of bacteria. These results demonstrate a new degree of control over immobilized liquid layers and will facilitate their use in future applications.

Red Clover Aryl Hydrocarbon Receptor (AhR) and Estrogen Receptor (ER) Agonists Enhance Genotoxic Estrogen Metabolism.

Many women consider botanical dietary supplements (BDSs) as safe alternatives to hormone therapy for menopausal symptoms. However, the effect of BDSs on breast cancer risk is largely unknown. In the estrogen chemical carcinogenesis pathway, P450 1B1 metabolizes estrogens to 4-hydroxylated catechols, which are oxidized to genotoxic quinones that initiate and promote breast cancer. In contrast, P450 1A1 catalyzed 2-hydroxylation represents a detoxification pathway. The current study evaluated the effects of red clover, a popular BDS used for women's health, and its isoflavones, biochanin A (BA), formononetin (FN), genistein (GN), and daidzein (DZ), on estrogen metabolism. The methoxy estrogen metabolites (2-MeOE1, 4-MeOE1) were measured by LC-MS/MS, and CYP1A1 and CYP1B1 gene expression was analyzed by qPCR. Nonmalignant ER-negative breast epithelial cells (MCF-10A) and ER-positive breast cancer cells (MCF-7) were derived from normal breast epithelial tissue and ER+ breast cancer tissue. Red clover extract (RCE, 10 µg/mL) and isoflavones had no effect on estrogen metabolism in MCF-10A cells. However, in MCF-7 cells, RCE treatments downregulated CYP1A1 expression and enhanced genotoxic metabolism (4-MeOE1/CYP1B1 > 2-MeOE1/CYP1A1). Experiments with the isoflavones showed that the AhR agonists (BA, FN) preferentially induced CYP1B1 expression as well as 4-MeOE1. In contrast, the ER agonists (GN, DZ) downregulated CYP1A1 expression likely through an epigenetic mechanism. Finally, the ER antagonist ICI 182,780 potentiated isoflavone-induced XRE-luciferase reporter activity and reversed GN and DZ induced downregulation of CYP1A1 expression. Overall, these studies show that red clover and its isoflavones have differential effects on estrogen metabolism in "normal" vs breast cancer cells. In breast cancer cells, the AhR agonists stimulate genotoxic metabolism, and the ER agonists downregulate the detoxification pathway. These data may suggest that especially breast cancer patients should avoid red clover and isoflavone based BDSs when making choices for menopausal symptom relief.