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Earwax is a readily accessible biological matrix that has the potential to be used in disease diagnostics. However, its semisolid nature and high chemical complexity have hampered efforts to investigate its potential to reveal disease markers. This is because more conventional methods of analysis such as gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry yield unsatisfactory results due to the presence of many nonvolatile and/or coeluting compounds, which in some cases have very similar mass spectrometric profiles. In addition, these routine methods often require the sample to be saponified, which dramatically increases the complexity of the analysis and makes it difficult to determine which compounds are actually present versus those that are produced by saponification. In this study, two-dimensional GC mass spectrometry (GC × GC-MS) was successfully applied for the characterization of the chemical components of earwax from healthy donors using nonpolar (primary) and midpolar (secondary) columns without saponification. Over 35 of the compounds that were identified are reported for the first time to be detected in unsaponified earwax. The resulting GC × GC-MS contour plots revealed visually recognizable compound class clusters of previously reported groups including alkanes, alkenes, fatty acids, esters, triglycerides, and cholesterol esters, as well as cholesterol and squalene. The application of GC × GC-MS revealed results that provide a foundation upon which future studies aimed at comparing healthy donor earwax to that from individuals exhibiting various disease states can be accomplished.

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Analysis of wood transects in a manner that preserves the spatial distribution of the metabolites present is highly desirable to among other things: (1) facilitate ecophysiology studies that reveal the association between chemical make-up and environmental factors or climatic events over time; and (2) investigate the mechanisms of the synthesis and trafficking of small molecules within specialised tissues. While a variety of techniques could be applied to achieve these goals, most remain challenging and impractical. Laser ablation direct analysis in real time imaging-mass spectrometry (LADI-MS) was successfully used to survey the chemical profile of wood, while also preserving the small-molecule spatial distributions. The tree species Entandrophragma candollei Harms, Millettia laurentii DeWild., Pericopsis elata (Harms) Meeuwen, Dalbergia nigra (Vell.) Benth. and Dalbergia normandii Bosser & R.Rabev were analysed. Several compounds were associated with anatomical features. A greater diversity was detected in the vessels and parenchyma compared with the fibres. Analysis of single vessels revealed that the chemical fingerprint used for timber identification is mainly determined by vessel content. Laser ablation direct analysis in real time imaging-mass spectrometry offers unprecedented opportunities to investigate the distribution of metabolites within wood samples, while circumventing the issues associated with previous methods. This technique opens up new vistas for the discovery of small-molecule biomarkers that are linked to environmental events.

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The rapid and accurate identification of condom-derived lubricant traces takes on heightened importance in sexual assault cases where the assailant has used a condom in order to avoid leaving behind incriminating DNA evidence. Previous reports have demonstrated that a variety of techniques can be used to confirm that a given residue is condom-derived, based on the detection of spermicides, slip agents and/or other common additives. However, limited success has been achieved in differentiating brands from among a broad range of condom types. In this study, the utility of direct analysis in real time-high resolution mass spectrometry (DART-HRMS) combined with chemometrics, for the rapid and accurate attribution of brands to condom residues of various types, was explored and developed. A database of condom residue spectra comprised of 110 different condom types representing 16 brands was generated, with the spectra serving as representative fingerprints for each brand. The spectral fingerprints were subjected to pre-processing prior to the application of Partial Least Squares-Discriminant Analysis (PLS-DA) which was used to generate a classifier that permitted identification of condom brands with an accuracy of 97.4%. An additional criterion was imposed on the PLS-DA to provide the confidence level and credibility of each prediction. The effect of time since deposition, the presence of contaminants and the influence of residue transfer on the prediction accuracy of the model were also assessed. The results from Sparse Discriminant Analysis (SDA) and PLS-DA were followed by application of the Student's t-test to determine m/z values representative of small-molecule markers that were most important for defining brand classes. The m/z values revealed by the two methods were found to be consistent in indicating which masses were representative of markers. The SDA method also provided low-dimensional views of the discriminative directions for classification of condom residues, thereby enabling easy visualization of the relationship between the indicated m/z values and brand discrimination. The results further revealed a subset of 14 m/z values that were observed in all 110 condoms representing the 16 brands, and some of these may serve as potential universal small-molecule condom markers. Overall, the results show that the DART-HRMS database of condom residue spectra can be used to identify residues based on differences in chemical components peculiar to each brand. The database can be readily expanded to include more condoms.

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A novel hemostatic and absorbent wound dressing material compatible with 3D printing is developed to address deficiencies in current wound dressing protocol. The design involves an open celled, microporous hydrogel foam via a high internal phase emulsion (HIPE) template with biocompatible components and tunable hemostatic character by kaolin loading, the viscosity and cure kinetics of which are tailored for 3D printing applications. The use of nontoxic mineral oil organic phase results in cytocompatability with human dermal fibroblasts. Kaolin distribution is shown by X-ray diffraction and elemental dispersive spectroscopy to be exfoliated and dispersed in the hydrogel dressing. In addition to demonstrating high fluid absorption and noncytotoxicity of relevant cell lines, the high internal phase emulsion polymers (polyHIPEs) also match the hemostatic performance of commercial wound dressing materials. Furthermore, the polyHIPEs display the requisite rheological properties for 3D printing that result in the fabrication of a prototype dressing with hierarchical porosity and a large number of controllable form factors.

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In recent years, there has been dramatic growth in the study of RNA. RNA has gone from being known as an intermediate in the central dogma of molecular biology to a molecule with a large diversity of structure and function that is involved in all aspects of biology. As new functions are rapidly discovered, it has become clear that there is a need for RNA-targeting small molecule probes to investigate RNA biology and clarify the potential for therapeutics based on RNA-small molecule interactions. While a host of techniques exist to measure RNA-small molecule interactions, many of these have drawbacks that make them intractable for routine use and are often not broadly applicable. A newer technology called microscale thermophoresis (MST), which measures the directed migration of a molecule and/or molecule-ligand complex along a temperature gradient, can be used to measure binding affinities using very small amounts of sample. The high sensitivity of this technique enables measurement of affinity constants in the nanomolar and micromolar range. Here, we demonstrate how MST can be used to study a range of biologically relevant RNA interactions, including peptide-RNA interactions, RNA-small molecule interactions, and displacement of an RNA-bound peptide by a small molecule.

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INTRODUCTION: Learning how to perform a speculum examination is a key component of the medical student curriculum, yet there is a paucity of data on the validity of available speculum examination models. This purpose of this study is to design, evaluate, and improve a low-cost speculum examination model. METHODS: A speculum examination training model was created using low-cost or recycled materials from other simulators. A total of 54 medical students, residents, and faculty in the obstetrics and gynecology department of a single academic institution performed speculum examinations on the model. Each participant completed a survey to provide qualitative and quantitative data. Using this feedback from participants, adjustments were made to the model and a similar survey was repeated with a total of 35 medical students and residents. RESULTS: The first iteration of the model was viewed positively by most participants. Eighty-three percent gave the model either a very realistic or realistic rating. Ninety-four percent thought the model was a very useful or useful teaching device. There were few significant differences in quantitative data based on experience level. Qualitative feedback yielded generally positive remarks with areas for improvement. The second iteration of the model was successful in differentiating between novice and skilled participants: residents were significantly better at identifying cervical position compared with students. Eighty-nine percent of participants thought the model was very useful or useful, whereas 49% thought the model was very realistic or realistic. DISCUSSION: The first iteration of the model demonstrated realism and usefulness; however, it lacked construct validity. Participant feedback yielded several helpful suggestions to improve the model. The second and final iteration of the model differentiated between novice and skilled participants at the cost of realism. This low-cost model is a useful tool to aid in teaching the speculum examination. Further development and study of the model could lead to a valid tool to evaluate speculum examination skills.

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We present a completely solid-phase synthetic strategy to create three- and four-fold peptide-appended π-electron molecules, where the multivalent oligopeptide presentation is dictated by the symmetries of reactive handles placed on discotic π-conjugated cores. Carboxylic acid and anhydride groups were viable amidation and imidation partners, respectively, and oligomeric π-electron discotic cores were prepared through Pd-catalyzed cross-couplings. Due to intermolecular hydrogen bonding between the three or four peptide axes, these π-peptide hybrids self-assemble into robust one-dimensional nanostructures with high aspect ratios in aqueous solution. The preparation of these systems via solid-phase methods will be detailed along with their self-assembly properties, as revealed by steady-state spectroscopy and transmission electron microscopy and electrical characterization using field-effect transistor measurements.

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We report the synthesis, self-assembly, and electron transfer capabilities of peptide-based electron donor-acceptor molecules and supramolecular nanostructures. These modified peptides contain π-conjugated oligothiophene electron donor cores that are peripherally substituted with naphthalene diimide electron acceptors installed via imidation of site-specific lysine residues. These molecules self-assemble into one-dimensional nanostructures in aqueous media, as shown through steady-state absorption, photoluminescence, and circular dichroism spectra, as well as transmission electron microscopy. Excitation of the oligothiophene donor moieties results in electron transfer to the acceptor units, ultimately creating polar, charge-separated states that persist for over a nanosecond as observed with transient absorption spectroscopy. This study demonstrates how transient electric fields can be engineered into aqueous nanomaterials of biomedical relevance through external, temporally controlled photonic inputs.

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We present a systematic study of the photophysical properties of one-dimensional electronically delocalized nanostructures assembled from π-conjugated subunits embedded within oligopeptide backbones. The nature of the excited states within these nanostructures is studied as a function of primary amino acid sequence utilizing steady-state and time-resolved spectroscopies, and their atomistic structure is probed by molecular simulation. Variations introduced into the amino acid side chains at specific residue locations along the molecular peptide backbone lead to pronounced changes in the observed photophysical behavior of the fibrillar structures (spanning H-like excitonic coupling and disordered excimeric coupling) that arise from subtle changes in the π-stacking within them. These results indicate that residue modification-in terms of relative size, solvation properties, and with respect to the distance from the central π-electron core-enables the ability to tune chromophore packing and the resulting photophysics of supramolecular assemblies of π-conjugated bioelectronic materials in a rational and systematic manner.

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We report a streamlined method for the synthesis of peptides embedded with complex and easily variable π-conjugated oligomeric subunits from commercially available precursors. These modified peptides self-assemble under aqueous conditions to form one-dimensional nanomaterials containing networks of π-stacked conduits, despite the inclusion of π-conjugated oligomers with quadrupoles extended over larger areas. The procedure has circumvented solubility and other synthetic issues to allow for the facile formation of a diverse library of bioelectronic nanomaterials, including a complex sexithiophene-containing peptide whose nanostructures display gate-induced conductivity within field effect transistors.

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INTRODUCTION: The principal aim of this work was to determine the prevalence of antinuclear antibodies and antinucleosomes antibodies during a treatment by interferon alpha with low dose for 18 months among patients with a melanoma stage I. The secondary objective consisted to seek the existence or not of a correlation with the clinical relapse, to determine the prevalence of appearance of clinical signs of autoimmune diseases and dysthyroidie. PATIENT AND METHODS: It was an exploratory study. The patients included in the study had a melanoma stage I (French classification), whose excision was realized for 6 weeks maximum, with a Breslow index equal or higher than 1,5 mm. The statistical model of logistic regression was used. RESULTS: Eighty-forth patients were included (38 women and 46 men) old from 21 to 75 years. The prevalence of antinuclear antibodies was 39%. None of the following variables: age, sex, phototype, localisation of melanoma in exposed photo zone, index of Breslow or Clark, were significantly associated with the presence of antinuclear antibodies. As the percentage of patients with anti-nucleosomes was low (5%), no statistical study was carried out. The prevalence of clinical and/or biological dysthyroidie was 37%. 60% of the patients presented at a moment in the evolution antinuclear antibodies or a dysthyroidie. The prevalence of relapses and death different was not correlated significantly with antinuclear antibodies and/or a dysthyroidie. DISCUSSION: Many studies report the appearance of antinuclear antibodies, generally without clinical lesions during the treatment by interferon alpha for cancers (tumours carcinoids, hemopathies) and viral chronic hepatitis. Our study is, to our knowledge, the first evaluating the induction of an autoimmunity during the adjuvant treatment by interferon alpha of melanoma stage I. The induction of autoantibody during the treatment by interferon alpha could constitute a marker of effectiveness of the treatment with improvement of the survival of these patients. In our study, however auto immunity markers do not appear as factors of severity of evolution of the melanoma or predictive factors.

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Oxygen free radicals and nitric oxide (NO) have been proposed to be involved in the cascade of injury elicited by traumatic brain injury. However, the mechanism(s) of injury remain to be explored. Since superoxide generation is triggered by traumatic brain injury, the cytotoxic peroxynitrite could be formed, but it is not known if this actually occurs. Dot blot and immunohistochemistry studies were performed to quantify tyrosine nitration and identify cell types in which such reactions occur in the brain of mice submitted to traumatic brain injury. Nitrotyrosine formation increased from 4 to 24 h after traumatic brain injury and was primarily observed in degenerating neurons, in areas corresponding to the sites of direct impact (frontal cortex) and diffuse impact (frontoparietal cortex and ventromedial hypothalamic nucleus). Furthermore, N omega-nitro-L-arginine-methylester (L-NAME), a NO-synthase inhibitor which has previously been shown to promote neurological recovery in traumatic brain injury, reduced nitrotyrosine formation and the number of nitrotyrosine-positive neurons. These results indicate that traumatic brain injury induces peroxynitrite formation which may contribute to cell damage.

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The pineal hormone melatonin has recently been shown to exert neuroprotective activity in a variety of experimental neuropathologies in which free radicals are involved. This neuroprotective effect has been attributed to the antioxidant properties of melatonin. Considering that free radicals also play a deleterious role in traumatic brain injury (TBI), the purpose of the present study was to determine whether melatonin would have a beneficial effect in this pathology. Head injury was induced in mice and the neurological deficit was evaluated at 24 hr by a grip test. In this model, the free radical scavenger, alpha-phenyl-tert-butyl-nitrone (2 x 100 mg/kg, i.p.) given 5 min and repeated at 4 hr after TBI was neuroprotective. Melatonin (1.25 mg/kg, i.p.) given 5 min and repeated at 1, 2, and 3 hr after head trauma also significantly reduced the neurological deficit. This beneficial effect was not due to melatonin-induced hypothermia since repeated treatment with melatonin did not modify the colonic temperature of mice. This study shows that melatonin exerts a beneficial effect on the neurological deficit induced by traumatic brain injury in mice. The mechanisms of this neuroprotection remains to be established, and more particularly, the contribution of the antioxidant activity of melatonin.

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1. In this study the effect of the dose and administration time of NG-nitro-L-arginine methyl ester (L-NAME), an NO-synthase inhibitor, in a model of transient focal cerebral ischaemia in rats was investigated. 2. Two injections of L-NAME were given, of 1, 3 and 10 mg kg-1, 5 min and 3 h after the onset of ischaemia. None of the doses gave any striatal neuroprotection, but 1 and 3 mg kg-1 L-NAME reduced the infarcted volume in the cortex (by 26%, P < 0.01 for 1 mg kg-1 and 21%, P < 0.05 for 3 mg kg-1), whereas 10 mg kg-1 had no neuroprotective effect. 3. Single injections of L-NAME 1 mg kg-1, given 5 min or 3 h after ischaemia onset, had similar neutoprotective effects on the cortical infarction as did the repeated injections. 4. L-NAME 1 mg kg-1 given 3, 6 or 9 h after ischaemia induction reduced the cortical infarct volume by 19% (P < 0.01) when given 3 h after ischaemia, by 21% (P < 0.01) when given at 6 h, and by 16% (P < 0.05) when given at 9 h, but had no neuroprotective activity when given 12 h after ischaemia. 5. Thus a low dose of L-NAME is neuroprotective in a model of transient focal ischaemia, with a wide therapeutic window, much larger than that found or MK-801.

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1. The temporal changes in constitutive NO-synthase (cNOS) and in calcium-independent NO-synthase activities were studied in mice subjected to 2 h of transient focal cerebral ischaemia. The changes in brain nitrites/nitrates (NOx) content were also studied. 2. NOS activities were measured by the conversion of L-[14C]-arginine to L-[14C]-citrulline. Brain NOx contents were investigated by the Griess colourimetric method. 3. cNOS activity in the infarcted cortical area was significantly reduced after 6 h of reperfusion and this activity remained attenuated for up to 10 days after ischaemia. A calcium-independent NOS activity began to increase 48 h after reperfusion, reached a maximum at 7 days and returned to baseline at 10 days. 4. There was a significant increase of brain NOx content beginning after 3 days of reperfusion. This increase was maximal at 7 days and returned to baseline at 10 days. 5. Thus, ischaemia followed by recirculation leads to a rapid, prolonged drop in cNOS activity in the infarcted cortex. There is also a substantial appearance of calcium-independent NOS activity in the later phase of transient ischaemia, leading to an important increase of NOx production.

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The beneficial effects of exogenous kappa receptor agonists in preventing neuronal damage elicited by brain ischemia suggest a role for endogenous dynorphins. In agreement prodynorphin (PDYN) gene expression in granule cells of the dentate gyrus detected by in situ hybridization was drastically but transiently decreased 18-32 h after four-vessel cerebral ischemia for 20 min in rats. We propose that decreased dynorphin synthesis and release could contribute to the delayed neuronal death of hippocampal pyramidal cells in this model.

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This study investigates the effect of the NO synthase inhibitors, NG-nitro-L-arginine methyl ester (L-NAME) and 7-nitroindazole (7-NI), on the neurological deficit 24 h after a moderate closed head injury in mice. Low doses of L-NAME or 7-NI given soon after the injury significantly reduced the neurological deficit compared to the vehicle-treated group. L-Arginine (300 mg/kg) did not alter the neurological deficit, but reversed the protective effects of both L-NAME and 7-NI when given at the same time. Both L-NAME and 7-NI had dose-related effects. The neuroprotective effects of L-NAME and 7-NI occurred when the drugs were given 5, 30, or 60 min after brain injury, but not when treatment was begun 2 h after brain injury, suggesting a short therapeutic window for both drugs. These results suggest that NO synthesis by neuronal NO synthase plays an important role in the early neurotoxic cascade leading to neurological deficit following traumatic brain injury.

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