Periodically Sequenced Peptides: A New Tool for Nanoscale Materials Synthesis

Periodically sequenced peptides can be confined to interfaces and assembled into patterns that present chemical functionalities with exceptional spatial precision. The role of dynamics during the assembly of these peptides appears to be very important for inorganic nucleation and growth. Our work applies periodically sequenced sheet-forming peptides at interfaces to explore the fundamental dynamics of assembly. The peptide molecules are rationally designed to have amphiphilic properties and a propensity for sheet-like secondary structure. These designed peptides are deposited at the air-water interface to explore the dynamics of self-assembly and investigate their 2D order. To characterize the phase behavior, we apply Langmuir Blodgett techniques and Brewster angle microscopy. Thermodynamic analysis of structure formation with increasing pressure allows us to understand the nature of self- assembly with iterative changes in the peptide sequence. Additionally, we look at the dynamics of the self-assembled state, where the organic phase switches between short- and long-range order as a function of surface pressure. This model system allows us to explore our underlying hypothesis that the time scale of the confined peptide phase-transitions defines the length-scale of the crystalline phase. This is in contrast to a system that starts with a well-ordered preformed template that defines the inorganic phase. We have shown that our model peptides can effectively be used to control the polycrystallinity in gold by controlling the surface pressure and diffusive time scales at the interface

Odorant Receptor C-Terminal Motifs in Divergent Insect Species

Insect odorant receptors are a large family of seven transmembrane proteins believed to be G-protein coupled receptors. The peptide sequences of two odorant receptors within a given species may share as little as 17% identity, and there is limited similarity between receptors of divergent species. One exception is DmOr83b, which is found in Drosophila melanogaster and is highly conserved in at least ten other insect species. DmOr83b is broadly expressed in most of the olfactory sensory neurons of D. melanogaster at most developmental stages, while other odorant receptors tend to have more restricted and specific expression patterns. DmOr83b is critical for D. melanogaster olfaction, and it is involved in properly localizing other odorant receptors possibly by forming heterodimers with these receptors. The C-terminal region has been implicated as sites for such heterodimer formation. Multiple em for motif elicitation (MEME), a hidden markov model based program, was used to uncover three conserved motifs in the C-termini of a vast majority of the odorant receptor peptides from Anopheles gambiae, D. melanogaster, and Apis mellifera. These motifs are also found in DmOr83b and its orthologs and the order of these motifs is conserved as well. The conservation of these motifs among divergent odorant receptors in divergent species suggests functional importance. We propose that these motifs are involved in receptor- receptor protein interactions, contributing to the heterodimer formation between DmOr83b (or its orthologs) and other odorant receptors

Neuroimaging of Unusual Glioblastoma Using Diffusion Tensor Imaging

Background: Gliosarcoma refers to an uncommon astrocytic tumor of the central nervous system. These tumors include both glial and mesenchymal components by definition, and are extremely malignant. Gliosarcomas are particular to tumors with distinct gliomatous and sarcomatous constituents, and are distinguished from gliobastomas which have undergone mesenchymal metaplasia. Gliosarcomas encompass 2-8% of all glioblastoma cases and tend to occupy the supratentorial regions of the brain, especially the temporal lobe. Rare infratentorial lesions including the cerebellar hemisphere, intraventricular, and multi-focal tumors have also been reported. Accurate neuroimaging diagnosis is critical and diffusion tensor imaging (DTI) and spectroscopy can be useful to differentiate from inflammatory disease. Methods: We report a 38 year-old Caucasian male with a right parietal lobe glioblastoma. The patient presented with new onset tonic-clonic seizures lasting approximately five minutes associated with postictal confusion and incontinence. The patient had conventional brain MRI scans including DTI. MRI data was processed to obtain tractography and fractional anisotropy (FA) maps. MR images were examined for location and extent of tumor as well as invasion, destruction or displacement of brain parenchyma and white matter tracts. Results: Noncontract CT revealed no abnormality and emergent MR imaging shows a ring-enhancing lesion measuring 3.1x2.2x3.1 cm at the junction of the right parieto-occipital region. The lesion abuts the dural surface, characteristic of gliosarcoma. Extensive surrounding edema causing complete effacement of the posterior horn of the right lateral ventricle, parietal effacement of the anterior floor of the right ventricle, and a 1.2 cm right to left midline shift were observed. Mild diffuse enhancement in the region of the splenium of the corpus callosum was likely compatible with seizure activity. Increased signal intensity of axial FLAIR image was seen in this area after four weeks. Fractional anisotropy is reduced at the tumor site suggesting an aggressive and invasive lesion. Diffusion tensor tractography shows destruction of white matter tracts compatible with destruction rather than invasion of parenchyma. Histopathology confirms gliobastoma multiforme, demonstrating mixed glial and sarcomatous components. However, GFAP was strongly positive in both areas and a reticulin stain was not increased in the sarcomatorus areas excluding the sarcomatous variant of glioblastoma. Conclusion: We report the imaging findings of a rare gliosblastoma radiographically presenting as a gliosarcoma due to its location along the dural surface, but with lack of pathologic findings. Gross total tumor resection was performed and the patient and is undergoing adjuvant radiation therapy with concurrent chemotherapy

Association between protein-bound uremic toxins and asymptomatic cardiac dysfunction in patients with chronic kidney disease

Although the relationship between protein-bound uremic toxins (PBUTs) and cardiac structure and cardiac mortality in chronic kidney disease (CKD) has been studied in the past, the association between cardiac dysfunction and PBUTs has not yet been studied. We therefore evaluated the association between impaired peak cardiac performance and the serum free and total concentrations of potentially cardiotoxic PBUTs. In a cross-sectional study of 56 male CKD patients (stages 2⁻5 (pre-dialysis)) who were asymptomatic with no known cardiac diseases or diabetes we measured peak cardiac power (CPOmax), aerobic exercise capacity (VO2max), and echocardiographic parameters of cardiac morphology and evaluated their association with PBUTs. The serum total and free concentrations of indoxyl sulfate (IXS), p-cresyl sulfate (PCS), p-cresyl glucuronide, indole acetic acid, and hippuric acid showed significant negative correlation with CPOmax and VO2max. IXS and PCS were independently associated with CPOmax and VO2max even after controlling for eGFR. No correlation between left ventricular mass index (LVMI) and PBUTs was seen. The present study for the first time has demonstrated the association between subclinical cardiac dysfunction in CKD and serum levels of a panel of PBUTs. Further studies are required to evaluate the mechanism of cardiotoxicity of the individual uremic toxins

Circular Dichroistic Impacts of 1‑(3-Dimethylaminopropyl)-3- ethylurea: Secondary Structure Artifacts Arising from Bioconjugation Using 1‑Ethyl-3-[3-dimethylaminopropyl]carbodiimide

1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide (EDC) is a commonly used reagent for bioconjugation and peptide synthesis. Both EDC and the corresponding urea derivative, 1-(3-dimethylaminopropyl)-3-ethylurea (EDU), are achiral. As the reagent is active in aqueous solutions, it is a common choice for the study of evolving secondary structural changes via circular dichroism. This work highlights the effect of EDU on spectropolarimetric measurements, namely, the problematic absorption profile at low wavelengths (190−220 nm). We demonstrate that EDU is capable of erroneously indicating structural changes, particularly loss of α-helical character, through masking of the characteristic minimum at 208 nm. However, if the concentrations of the EDU in the sample are known, then this effect can be anticipated and calculations of secondary structure can be adjusted to avoid the impacted wavelengths. Impacts of EDU in a sample are compared to those of standard urea, which, by contrast, is commonly used as a denaturant in circular dichroism studies without issue

Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana.

To optimize the expression, extraction and purification of plant-derived tetrameric recombinant human butyrylcholinesterase (prBChE), we describe the development and use of plant viral amplicon-based gene expression system; Tobacco Mosaic Virus (TMV) RNA-based overexpression vector (TRBO) to express enzymatically active FLAG-tagged plant made recombinant butyrylcholinesterase (rBChE) in Nicotiana benthamiana leaves using transient agroinfiltration. Two gene expression cassettes were designed to express the recombinant protein in either the ER or to the apoplastic compartment. Leaf homogenization was used to isolate ER-retained recombinant butyrylcholinesterase (prBChE-ER) while apoplast-targeted rBChE was isolated by either leaf homogenization (prBChE) or vacuum-extraction of apoplastic wash fluid (prBChE-AWF). rBChE from apoplast wash fluid had a higher specific activity but lower enzyme yield than leaf homogenate. To optimize the isolation and purification of total recombinant protein from leaf homogenates, an acidic extraction buffer was used. The acidic extraction buffer yielded >95% enzymatically active tetrameric rBChE as verified by Coomassie stained and native gel electrophoresis. Furthermore, when compared to human butyrylcholinesterase, the prBChE was found to be similar in terms of tetramerization and enzyme kinetics. The N-linked glycan profile of purified prBChE-ER was found to be mostly high mannose structures while the N-linked glycans on prBChE-AWF were primarily complex. The glycan profile of the prBChE leaf homogenates showed a mixture of high mannose, complex and paucimannose type N-glycans. These findings demonstrate the ability of plants to produce rBChE that is enzymatically active and whose oligomeric state is comparable to mammalian butyrylcholinesterase. The process of plant made rBChE tetramerization and strategies for improving its pharmacokinetics properties are also discussed

Quantification of transient increase of the blood–brain barrier permeability to macromolecules by optimized focused ultrasound combined with microbubbles

Radioimmunotherapy using a radiolabeled monoclonal antibody that targets tumor cells has been shown to be efficient for the treatment of many malignant cancers, with reduced side effects. However, the blood–brain barrier (BBB) inhibits the transport of intravenous antibodies to tumors in the brain. Recent studies have demonstrated that focused ultrasound (FUS) combined with microbubbles (MBs) is a promising method to transiently disrupt the BBB for the drug delivery to the central nervous system. To find the optimal FUS and MBs that can induce reversible increase in the BBB permeability, we employed minimally invasive multiphoton microscopy to quantify the BBB permeability to dextran-155 kDa with similar molecular weight to an antibody by applying different doses of FUS in the presence of MBs with an optimal size and concentration. The cerebral microcirculation was observed through a section of frontoparietal bone thinned with a micro-grinder. About 5 minutes after applying the FUS on the thinned skull in the presence of MBs for 1 minute, TRITC (tetramethylrhodamine isothiocyanate)-dextran-155 kDa in 1% bovine serum albumin in mammalian Ringer’s solution was injected into the cerebral circulation via the ipsilateral carotid artery by a syringe pump. Simultaneously, the temporal images were collected from the brain parenchyma ~100–200 μm below the pia mater. Permeability was determined from the rate of tissue solute accumulation around individual microvessels. After several trials, we found the optimal dose of FUS. At the optimal dose, permeability increased by ~14-fold after 5 minutes post-FUS, and permeability returned to the control level after 25 minutes. FUS without MBs or MBs injected without FUS did not change the permeability. Our method provides an accurate in vivo assessment for the transient BBB permeability change under the treatment of FUS. The optimal FUS dose found for the reversible BBB permeability increase without BBB disruption is reliable and can be applied to future clinical trials

Effect of Cardiac and Noncardiac Conditions on Survival After Defibrillator Implantation

ObjectivesWe sought to examine outcomes in recipients of implantable cardioverter-defibrillators (ICDs) and the effect of age, gender, and comorbidities on survival.BackgroundAge, gender, and comorbidities may significantly affect outcomes in ICD recipients.MethodsWe examined factors associated with mortality in 2,467 ICD recipients in Ontario, Canada, using a province-wide database. Comorbidities were identified retrospectively by examining all diagnosis codes within the 3 years before implant.ResultsMean ages at ICD implant were 63.2 ± 12.5 years (1,944 men) and 59.8 ± 15.9 years (523 women). Mortality rates at one and 2 years were 7.8% and 14.0%. Older age at implant increased the risk of death with hazard ratios (HR) of 2.05 (95% confidence interval [CI] 1.70 to 2.47) and 3.00 (95% CI 2.43 to 3.71) for those 65 to 74 years and ≥75 years, respectively (both p < 0.001), but gender was not a predictor of death. Common noncardiac conditions associated with death included peripheral vascular disease (adjusted HR 1.50, 95% CI 1.18 to 1.91), pulmonary disease (adjusted HR 1.35, 95% CI 1.10 to 1.66), and renal disease (adjusted HR 1.57, 95% CI 1.25 to 1.99). Many ICD recipients had prior heart failure (46.2%) with an increased HR of 2.33 for death (95% CI 1.96 to 2.76; p < 0.001). Greater comorbidity burden conferred increased risk, with HRs adjusted for age, gender, and heart failure of 1.72 (95% CI 1.44 to 2.05), 2.79 (95% CI 2.15 to 3.62), and 2.98 (95% CI 1.74 to 5.10) for those with 1, 2, and 3 or more noncardiac comorbidities, respectively (all p < 0.001).ConclusionsAge, noncardiac comorbidities, and prior heart failure influence survival outcomes in ICD recipients. These factors should be considered in the care of ICD recipients