Metabolic stability of 3-epi-1α,25-dihydroxyvitamin D3 over 1 α 25-dihydroxyvitamin D3: metabolism and molecular docking studies using rat CYP24A1.

3-epi-1α,25-dihydroxyvitamin D3 (3-epi-1α,25(OH)2D3), a natural metabolite of 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3), exhibits potent vitamin D receptor (VDR)-mediated actions such as inhibition of keratinocyte growth or suppression of parathyroid hormone secretion. These VDR-mediated actions of 3-epi-1α,25(OH)2D3 needed an explanation as 3-epi-1α,25(OH)2D3, unlike 1α,25(OH)2D3, exhibits low affinity towards VDR. Metabolic stability of 3-epi-1α,25(OH)2D3 over 1α,25(OH)2D3 has been hypothesized as a possible explanation. To provide further support for this hypothesis, we now performed comparative metabolism studies between 3-epi-1α,25(OH)2D3 and 1α,25(OH)2D3 using both the technique of isolated rat kidney perfusion and purified rat CYP24A1 in a cell-free reconstituted system. For the first time, these studies resulted in the isolation and identification of 3-epi-calcitroic acid as the final inactive metabolite of 3-epi-1α,25(OH)2D3 produced by rat CYP24A1. Furthermore, under identical experimental conditions, it was noted that the amount of 3-epi-calcitroic acid produced from 3-epi-1α,25(OH)2D3 is threefold less than that of calcitroic acid, the analogous final inactive metabolite produced from 1α,25(OH)2D3 . This key observation finally led us to conclude that the rate of overall side-chain oxidation of 3-epi-1α,25(OH)2D3 by rat CYP24A1 leading to its final inactivation is slower than that of 1α,25(OH)2D3. To elucidate the mechanism responsible for this important finding, we performed a molecular docking analysis using the crystal structure of rat CYP24A1. Docking results suggest that 3-epi-1α,25(OH)2D3, unlike 1α,25(OH)2D3, binds to CYP24A1 in an alternate configuration that destabilizes the formation of the enzyme-substrate complex sufficiently to slow the rate at which 3-epi-1α,25(OH)2D3 is inactivated by CYP24A1 through its metabolism into 3-epi-calcitroic acid.

Nanoscale plasmonic interferometers for multispectral, high-throughput biochemical sensing.

In this work, we report the design, fabrication, and characterization of novel biochemical sensors consisting of nanoscale grooves and slits milled in a metal film to form two-arm, three-beam, planar plasmonic interferometers. By integrating thousands of plasmonic interferometers per square millimeter with a microfluidic system, we demonstrate a sensor able to detect physiological concentrations of glucose in water over a broad wavelength range (400-800 nm). A wavelength sensitivity between 370 and 630 nm/RIU (RIU, refractive index units), a relative intensity change between ~10(3) and 10(6) %/RIU, and a resolution of ~3 × 10(-7) in refractive index change were experimentally measured using typical sensing volumes as low as 20 fL. These results show that multispectral plasmonic interferometry is a promising approach for the development of high-throughput, real-time, and extremely compact biochemical sensors.

Grazing incidence X-Ray diffraction: identifying the dominant facet in copper foams that electrocatalyze the reduction of carbon dioxide to formate.

Copper foams have been shown to electrocatalyze the carbon dioxide reduction reaction (CO2RR) to formate (HCOO-) with significant faradaic efficiency (FE) at low overpotentials. Unlike the CO2RR electrocatalyzed at copper foils, the CO2RR electrocatalyzed at porous copper foams selects for HCOO- essentially to the exclusion of hydrocarbon products. Formate is an environmentally friendly organic acid with many applications such as food preservation, textile processing, de-icing, and fuel in fuel cells. Thus, HCOO- is an attractive product from the CO2RR if it is produced at an overpotential lower than that at other electrocatalysts. In this study, grazing incidence X-ray diffraction (GIXRD) was used to identify the dominant surface facet of porous copper foams that accounts for its selectivity for HCOO- during the CO2RR. Included are data from the CO2RR at different temperatures using copper foams as the electrocatalyst. Under optimal reaction conditions at 2 °C, the FE for converting CO2 to HCOO- at Cu foams approaches 50% while the FE for hydrogen gas (H2) falls below 40%, a significant departure from that obtained at polycrystalline Cu foils. Computational studies by others have proposed (200) and (111) facets of Cu foils thermodynamically favour methane and other hydrocarbons, CO, HCOO- from the CO2RR. Results from the GIXRD studies indicate Cu foams are dominated by the (111) facet, which accounts for the selectivity of Cu foams toward HCOO- regardless of temperature used for the CO2RR.

Antioxidant deactivation on graphenic nanocarbon surfaces.

This article reports a direct chemical pathway for antioxidant deactivation on the surfaces of carbon nanomaterials. In the absence of cells, carbon nanotubes are shown to deplete the key physiological antioxidant glutathione (GSH) in a reaction involving dissolved dioxygen that yields the oxidized dimer, GSSG, as the primary product. In both chemical and electrochemical experiments, oxygen is only consumed at a significant steady-state rate in the presence of both nanotubes and GSH. GSH deactivation occurs for single- and multi-walled nanotubes, graphene oxide, nanohorns, and carbon black at varying rates that are characteristic of the material. The GSH depletion rates can be partially unified by surface area normalization, are accelerated by nitrogen doping, and suppressed by defect annealing or addition of proteins or surfactants. It is proposed that dioxygen reacts with active sites on graphenic carbon surfaces to produce surface-bound oxygen intermediates that react heterogeneously with glutathione to restore the carbon surface and complete a catalytic cycle. The direct catalytic reaction between nanomaterial surfaces and antioxidants may contribute to oxidative stress pathways in nanotoxicity, and the dependence on surface area and structural defects suggest strategies for safe material design.

A new insight into the role of rat cytochrome P450 24A1 in metabolism of selective analogs of 1α,25-dihydroxyvitamin D3.

We examined the metabolism of two synthetic analogs of 1α,25-dihydroxyvitamin D3 (1), namely 1α,25-dihydroxy-16-ene-23-yne-vitamin D3 (2) and 1α,25-dihydroxy-16-ene-23-yne-26,27-dimethyl-vitamin D3 (4) using rat cytochrome P450 24A1 (CYP24A1) in a reconstituted system. We noted that 2 is metabolized into a single metabolite identified as C26-hydroxy-2 while 4 is metabolized into two metabolites, identified as C26-hydroxy-4 and C26a-hydroxy-4. The structural modification of adding methyl groups to the side chain of 1 as in 4 is also featured in another analog, 1α,25-dihydroxy-22,24-diene-24,26,27-trihomo-vitamin D3 (6). In a previous study, 6 was shown to be metabolized exactly like 4, however, the enzyme responsible for its metabolism was found to be not CYP24A1. To gain a better insight into the structural determinants for substrate recognition of different analogs, we performed an in silico docking analysis using the crystal structure of rat CYP24A1 that had been solved for the substrate-free open form. Whereas analogs 2 and 4 docked similar to 1, 6 showed altered interactions for both the A-ring and side chain, despite prototypical recognition of the CD-ring. These findings hint that CYP24A1 metabolizes selectively different analogs of 1, based on their ability to generate discrete recognition cues required to close the enzyme and trigger the catalytic mechanism.

A scalable method for preparing Cu electrocatalysts that convert CO2 into C2+ products.

Development of efficient catalysts for selective electroreduction of CO2 to high-value products is essential for the deployment of carbon utilization technologies. Here we present a scalable method for preparing Cu electrocatalysts that favor CO2 conversion to C2+ products with faradaic efficiencies up to 72%. Grazing-incidence X-ray diffraction data confirms that anodic halogenation of electropolished Cu foils in aqueous solutions of KCl, KBr, or KI creates surfaces of CuCl, CuBr, or CuI, respectively. Scanning electron microscopy and energy dispersive X-ray spectroscopy studies show that significant changes to the morphology of Cu occur during anodic halogenation and subsequent oxide-formation and reduction, resulting in catalysts with a high density of defect sites but relatively low roughness. This work shows that efficient conversion of CO2 to C2+ products requires a Cu catalyst with a high density of defect sites that promote adsorption of carbon intermediates and C-C coupling reactions while minimizing roughness.

Cu nanowire-catalyzed electrochemical reduction of CO or CO2.

We prepared micrometer long Cu nanowires (NWs) of 25 and 50 nm diameters and studied their electrocatalysis for electrochemical reduction of CO/CO2 in 0.1 M KHCO3 at room temperature. The 50 nm NWs showed better selectivity than the 25 nm NWs, and catalyzed CO reduction to C2-hydrocarbons (C2H4 + C2H6) with a combined faradaic efficiency (FE) of 60% (C2H4 FE of 35% and mass activity of 4.25 A g-1 Cu) at -1.1 V (vs. reversible hydrogen electrode). The NW-catalyzed CO2 reduction is less efficient due to the extra CO2 to CO step required for the formation of C2-hydrocarbons. This experimental evidence combined with DFT calculations suggests that CO is an important intermediate and NWs provide a large Cu(100) surface for *CO hydrogenation (to *CHO) and *CO-*CHO coupling, leading to more selective reduction of CO than CO2 towards C2-hydrocarbons.

Enhanced fibronectin adsorption on carbon nanotube/poly(carbonate) urethane: independent role of surface nano-roughness and associated surface energy.

The contribution of nanoscale surface roughness on the adsorption of one key cell adhesive protein, fibronectin, on carbon nanotube/poly(carbonate) urethane composites of different surface energies was evaluated. Systematic control of various surface energies by creating different nanosurface roughness features was performed by mixing two promising biomaterials: multi-wall carbon nanotubes and poly(carbonate) urethane. High ratios of carbon nanotubes coated with poly(carbonate) urethane provided for greater hydrophilic surfaces because of higher nanosurface roughness although pure carbon nanotube surfaces were extremely hydrophobic. Fabrication methods followed in this study generated various homogenous nanosurface roughness values (ranging from 2 to 20nm root mean square (RMS) AFM roughness). With the aid of such nanosurface roughness values in composites, a model was developed that linearly correlated nanosurface roughness and associated nanosurface energy to fibronectin adsorption. Specifically, independent contributions of surface chemistry (70%) and surface nano-roughness (30%) were found to mediate fibronectin adsorption. The results of the present study showed why carbon nanotube/poly(carbonate) urethane composites enhance cellular functions and tissue growth by delineating the importance of their physical nano-roughness on promoting the adsorption of a protein well known to be critical for mediating the adhesion of anchorage-dependent cells.

Microfluidic biofuel cells: the influence of electrode diffusion layer on performance.

Microfluidic biofuel cells exploit the lack of convective mixing at low Reynolds number to eliminate the need for a physical membrane to separate fuel from oxidant. This paper demonstrates how the length and spacing of electrodes within a microchannel, and thus thickness of the diffusion layer, affects the performance of a microfluidic biofuel cell. It was found that splitting a single electrode into two (or more) smaller electrodes and separating them by a distance equal to three times their length prevents the continuous increase in thickness of a diffusion layer. This change results in a 25% increase in maximum power density compared to a single electrode device with identical electroactive area. Furthermore, we found that the maximum current density of a microfluidic biofuel cell operated with different electrode configurations (i.e., length of cathode) closely matches that predicted by theory.

Combined topographical and chemical micropatterns for templating neuronal networks.

In vitro neuronal networks with geometrically defined features are desirable for studying long-term electrical activity within the neuron assembly and for interfacing with external microelectronic circuits. In standard cultures, the random spatial distribution and overlap of neurites makes this aim difficult; hence, many recent efforts have been made on creating patterned cellular circuits. Here, we present a novel method for creating a planar neural network that is compatible with optical devices. This method combines both topographical and chemical micropatterns onto which neurons can be cultured. Compared to other reported patterning techniques, our approach and choice of template appears to show both geometrical control over the formation of specific neurite connections at low plating density and compatibility with microelectronic circuits that stimulate and record neural activity.

Two polymerizable derivatives of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid).

[structure: see text] A versatile strategy for the synthesis of polymerizable derivatives of the redox-active indicator dye 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) is reported. Two products are shown to illustrate how the final step in the synthetic strategy can be used to alter the physical properties of the product. Both products were characterized spectroscopically and electrochemically. The hydrophilic monomer (sABTS) was polymerized, and the utility of this polymer (polyABTS) is demonstrated in the context of bioelectrocatalysis.

Axon Outgrowth of Rat Embryonic Hippocampal Neurons in the Presence of an Electric Field.

Application of an electric field (EF) has long been used to induce axon outgrowth following nerve injuries. The response of mammalian neurons (e.g., axon length, axon guidance) from the central nervous system (CNS) to an EF, however, remains unclear, whereas those from amphibian or avian neuron models have been well characterized. Thus, to determine an optimal EF for axon outgrowth of mammalian CNS neurons, we applied a wide range of EF to rat hippocampal neurons. Our results showed that EF with either a high magnitude (100 mV/mm or higher) or long exposure time (10 h or longer) with low magnitude (10-30 mV/mm) caused a neurite collapse and cell death. We also investigated whether neuronal response to an EF is altered depending on the growth stage of neuron cultures by applying 30 mV/mm to cells from 1 to 11 days in vitro (DIV). Neurons showed the turnover of axon outgrowth pattern when electrically stimulated between 4-5 DIV at which point neurons have both axonal and dendritic formation. The findings of this study suggest that the developmental stage of neurons is an important factor to consider when using EF as a potential method for axon regeneration in mammalian CNS neurons.

In Situ Measurement of Voltage-Induced Stress in Conducting Polymers with Redox-Active Dopants.

Minimization of stress-induced mechanical rupture and delamination of conducting polymer (CP) films is desirable to prevent failure of devices based on these materials. Thus, precise in situ measurement of voltage-induced stress within these films should provide insight into the cause of these failure mechanisms. The evolution of stress in films of polypyrrole (pPy), doped with indigo carmine (IC), was measured in different electrochemical environments using the multibeam optical stress sensor (MOSS) technique. The stress in these films gradually increases to a constant value during voltage cycling, revealing an initial break-in period for CP films. The nature of the ions involved in charge compensation of pPy[IC] during voltage cycling was determined from electrochemical quartz crystal microbalance (EQCM) data. The magnitude of the voltage-induced stress within pPy[IC] at neutral pH correlated with the radius of the hydrated mobile ion in the order Li(+) > Na(+) > K(+). At acidic pH, the IC dopant in pPy[IC] undergoes reversible oxidation and reduction within the range of potentials investigated, providing a secondary contribution to the observed voltage-induced stress. We report on the novel stress response of these polymers due to the presence of pH-dependent redox-active dopants and how it can affect material performance.

Conductive polypyrrole via enzyme catalysis.

Laccase catalyzes the polymerization of pyrrole into a conducting polymer using dioxygen as the terminal oxidant. This finding is significant, because it identifies an enzymatic route, and thus an environmentally benign method, for preparing a technologically important polymer. In addition, the rate of oxidation of pyrrole increases when the redox molecule, ABTS [2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonate)], is included in a reaction medium that contains laccase. This increase in rate occurs because laccase catalyzes the oxidation of ABTS to ABTS*. In addition to laccase, the biocatalytically generated ABTS* oxidizes pyrrole to its corresponding radical cation to yield polypyrrole. Moreover, oxidation of pyrrole by ABTS* regenerates ABTS for subsequent biocatalytic turnover. Including ABTS in the reaction medium has two important consequences for the final product: (a) The reaction proceeds rapidly enough to form polymeric films instead of oligomeric precipitates, and (b) ABTS remains within the polymeric film as a redox-active dopant. The charge transport properties of the resulting polymers, both with and without ABTS as the counteranion, are compared to those of other conducting materials including polypyrrole prepared electrochemically or chemically.

Neuron Image Analyzer: Automated and Accurate Extraction of Neuronal Data from Low Quality Images.

Image analysis software is an essential tool used in neuroscience and neural engineering to evaluate changes in neuronal structure following extracellular stimuli. Both manual and automated methods in current use are severely inadequate at detecting and quantifying changes in neuronal morphology when the images analyzed have a low signal-to-noise ratio (SNR). This inadequacy derives from the fact that these methods often include data from non-neuronal structures or artifacts by simply tracing pixels with high intensity. In this paper, we describe Neuron Image Analyzer (NIA), a novel algorithm that overcomes these inadequacies by employing Laplacian of Gaussian filter and graphical models (i.e., Hidden Markov Model, Fully Connected Chain Model) to specifically extract relational pixel information corresponding to neuronal structures (i.e., soma, neurite). As such, NIA that is based on vector representation is less likely to detect false signals (i.e., non-neuronal structures) or generate artifact signals (i.e., deformation of original structures) than current image analysis algorithms that are based on raster representation. We demonstrate that NIA enables precise quantification of neuronal processes (e.g., length and orientation of neurites) in low quality images with a significant increase in the accuracy of detecting neuronal changes post-stimulation.

Crumpled graphene nanoreactors.

Nanoreactors are material structures that provide engineered internal cavities that create unique confined nanoscale environments for chemical reactions. Crumpled graphene nanoparticles or "nanosacks" may serve as nanoreactors when filled with reactive or catalytic particles and engineered for a specific chemical function. This article explores the behavior of crumpled graphene nanoreactors containing nanoscale ZnO, Ag, Ni, Cu, Fe, or TiO2 particles, either alone or in combination, in a series of case studies designed to reveal their fundamental behaviors. The first case study shows that ZnO nanoparticles undergo rapid dissolution inside the nanoreactor cavity accompanied by diffusive release of soluble products to surrounding aqueous media through the irregular folded shell. This behavior demonstrates the open nature of the sack structure, which facilitates rapid small-molecule exchange between inside and outside that is a requirement for nanoreactor function. In a case study on copper and silver nanoparticles, encapsulation in graphene nanoreactors is shown in some cases to enhance their oxidation rate in aqueous media, which is attributed to electron transfer from the metal core to graphene that bypasses surface oxides and allows reduction of molecular oxygen on the high-area graphene shell. Nanoreactors also allow particle-particle electron transfer interactions that are mediated by the connecting conductive graphene, which give rise to novel behaviors such as galvanic protection of Ag nanoparticles in Ag/Ni-filled nanoreactors, and the photochemical control of Ag-ion release in Ag/TiO2-filled nanoreactors. It is also shown that internal graphene structures within the sacks provide pockets that reduce particle mobility and inhibit particle sintering during thermal treatment. Finally, these novel behaviors are used to suggest and demonstrate several potential applications for graphene nanoreactors in catalysts, controlled release, and environmental remediation.

Enhanced CO2 electroreduction efficiency through secondary coordination effects on a pincer iridium catalyst.

An iridium(III) trihydride complex supported by a pincer ligand with a hydrogen bond donor in the secondary coordination sphere promotes the electrocatalytic reduction of CO2 to formate in water/acetonitrile with excellent Faradaic efficiency and low overpotential. Preliminary mechanistic experiments indicate formate formation is facile while product release is a kinetically difficult step.

The potential of apolipoprotein E4 to act as a substrate for primary cultures of hippocampal neurons.

The E4 isoform of apolipoprotein (apoE4) is known to be a major risk factor for Alzheimer's Disease (AD). Previous in vitro studies have shown apoE4 to have a negative effect on neuronal outgrowth when incubated with lipids. The effect of apoE4 itself on the development of neurons from the central nervous system (CNS), however, has not been well characterized. Consequently, apoE4 alone has not been pursued as a substrate for neuronal cultures. In this study, the effect of surface-bound apoE4 on developmental features of rat hippocampal neurons was examined. We show that apoE4 substrates elicit significantly enhanced values in all developmental features at day 2 of culture when compared to laminin (LN) substrates, which is the current substrate-of-choice for neuronal cultures. Interestingly, the adhesion of hippocampal neurons was found to be significantly lower on LN substrates than on glass substrates, but the axon lengths on both substrates were similar. In addition, this study demonstrates that the adhesion- and growth-enhancing effects of apoE4 substrates are not mediated by heparan sulfate proteoglycans (HSPGs), proteins that have been indicated to function as receptors or co-receptors for apoE4. In the absence of lipids, apoE4 appears to use an unknown pathway for up-regulating neuronal adhesion and neurite outgrowth. Our results indicate that apoE4 is better than LN as a substrate for primary cultures of CNS neurons and should be considered in the design of tissue engineered CNS.

Viologens as charge carriers in a polymer-based battery anode.

Viologens, either as anions in solution or as pendant substituents to pyrrole, were incorporated as dopants to electrodeposited films of polypyrrole. The resulting polymer films exhibited redox activity at -0.5 V vs Ag/AgCl. The film consisting of polypyrrole with pendant viologens exhibited the best charge-discharge behavior with a maximum capacity of 55 mAh/g at a discharge current of 0.25 mA/cm(2). An anode consisting of polypyrrole (pPy) doped with viologen (V) was coupled to a cathode consisting of pPy doped with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) to yield a polymer-based battery with a cell electromotive force (emf) of 1.0 V, maximum capacity of 16 mAh/g, and energy density of 15 Wh/kg.

WITHDRAWN: Corrigendum to "A novel method for analyzing images of live nerve cells" [J. Neurosci. Methods 201 (2011) 98-105].

This article has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy.