Mechanism of graded persistent cellular activity of entorhinal cortex layer v neurons

Working memory is an emergent property of neuronal networks, but its cellular basis remains elusive. Recent data show that principal neurons of the entorhinal cortex display persistent firing at graded firing rates that can be shifted up or down in response to brief excitatory or inhibitory stimuli. Here, we present a model of a potential mechanism for graded firing. Our multicompartmental model provides stable plateau firing generated by a nonspecific calcium-sensitive cationic (CAN) current. Sustained firing is insensitive to small variations in Ca2+ concentration in a neutral zone. However, both high and low Ca2+ levels alter firing rates. Specifically, increases in persistent firing rate are triggered only during high levels of calcium, while decreases in rate occur in the presence of low levels of calcium. The model is consistent with detailed experimental observations and provides a mechanism for maintenance of memory-related activity in individual neurons.     

Crystal structure of the his-tagged saccharopine reductase from Saccharomyces cerevisiae at 1.7-Å resolution

The three-dimensional structure of the saccharopine reductase enzyme from the budding yeast Saccharomyces cerevisiae was determined to 1.7-A resolution in the apo form by using molecular replacement. The enzyme monomer consists of three domains: domain I is a variant of the Rossmann fold, domain II folds into a alpha/beta structure containing a mixed seven-stranded beta-sheet as the central core, and domain III has an all-helical fold. Comparative fold alignment with the enzyme from Magnaporthe grisea suggests that domain I binds to NADPH, and domain II binds to saccharopine and is involved in dimer formation. Domain III is involved in closing the active site of the enzyme once substrates are bound. Structural comparison of the saccharopine reductase enzymes from S. cerevisiae and M. grisea indicates that domain II has the highest number of conserved residues, suggesting that it plays an important role in substrate binding and in spatially orienting domains I and III.     

The MHC2TA -168A>G gene polymorphism is not associated with rheumatoid arthritis in Austrian patients

An association between susceptibility to rheumatoid arthritis (RA) and a common -168A>G polymorphism in the MHC2TA gene with differential major histocompatibility complex (MHC) II molecule expression was recently reported in a Swedish population. The objective of the present study was to replicate this finding by examining the -168A>G polymorphism in an Austrian case-control study. Three hundred and sixty-two unrelated RA cases and 351 sex-matched and age-matched controls as well as 1,709 Austrian healthy individuals were genotyped. All participants were from the same ethnic background. Genotyping was performed using 5' allelic discrimination assays. The association between susceptibility to RA and the -168A>G single nucleotide polymorphism was examined by chi-square test. Comparison was made assuming a dominant effect (AG + GG genotypes versus AA genotype). In contrast to the primary report, the frequency of MHC2TA -168G allele carriers was not significantly different between patients and controls in the Austrian cohort. The homozygous MHC2TA -168 GG genotype was more frequent in matched controls than in Austrian RA patients. There was no association between the presence of RA-specific autoantibodies and the MHC2TA -168 GG genotype. In this cohort of Austrian patients, no association between the MHC2TA polymorphism and RA was found.     

Toll-like receptor deficiency worsens inflammation and lymphedema after lymphatic injury

Mechanisms regulating lymphedema pathogenesis remain unknown. Recently, we have shown that lymphatic fluid stasis increases endogenous danger signal expression, and these molecules influence lymphatic repair (Zampbell JC, et al. Am J Physiol Cell Physiol 300: C1107-C1121, 2011). Endogenous danger signals activate Toll-like receptors (TLR) 2, 4, and 9 and induce homeostatic or harmful responses, depending on physiological context. The purpose of this study was to determine the role of TLRs in regulating tissue responses to lymphatic fluid stasis. A surgical model of lymphedema was used in which wild-type or TLR2, 4, or 9 knockout (KO) mice underwent tail lymphatic excision. Six weeks postoperatively, TLR KOs demonstrated markedly increased tail edema compared with wild-type animals (50-200% increase; P < 0.01), and this effect was most pronounced in TLR4 KOs (P < 0.01). TLR deficiency resulted in decreased interstitial and lymphatic transport, abnormal lymphatic architecture, and fewer capillary lymphatics (40-50% decrease; P < 0.001). Lymphedematous tissues of TLR KOs demonstrated increased leukocyte infiltration (P < 0.001 for TLR4 KOs), including higher numbers of infiltrating CD3+ cells (P < 0.05, TLR4 and TLR9 KO), yet decreased infiltrating F4/80+ macrophages (P < 0.05, all groups). Furthermore, analysis of isolated macrophages revealed twofold reductions in VEGF-C (P < 0.01) and LYVE-1 (P < 0.05) mRNA from TLR2-deficient animals. Finally, TLR deficiency was associated with increased collagen type I deposition and increased transforming growth factor-β1 expression (P < 0.01, TLR4 and TLR9 KO), contributing to dermal fibrosis. In conclusion, TLR deficiency worsens tissue responses to lymphatic fluid stasis and is associated with decreased lymphangiogenesis, increased fibrosis, and reduced macrophage infiltration. These findings suggest a role for innate immune responses, including TLR signaling, in lymphatic repair and lymphedema pathogenesis.     

Lymphatic function is regulated by a coordinated expression of lymphangiogenic and anti-lymphangiogenic cytokines

Lymphangiogenic cytokines such as vascular endothelial growth factor-C (VEGF-C) are critically required for lymphatic regeneration; however, in some circumstances, lymphatic function is impaired despite normal or elevated levels of these cytokines. The recent identification of anti-lymphangiogenic molecules such as interferon-γ (IFN-γ), transforming growth factor-β1, and endostatin has led us to hypothesize that impaired lymphatic function may represent a dysregulated balance in the expression of pro/anti-lymphangiogenic stimuli. We observed that nude mice have significantly improved lymphatic function compared with wild-type mice in a tail model of lymphedema. We show that gradients of lymphatic fluid stasis regulate the expression of lymphangiogenic cytokines (VEGF-A, VEGF-C, and hepatocyte growth factor) and that paradoxically the expression of these molecules is increased in wild-type mice. More importantly, we show that as a consequence of T-cell-mediated inflammation, these same gradients also regulate expression patterns of anti-lymphangiogenic molecules corresponding temporally and spatially with impaired lymphatic function in wild-type mice. We show that neutralization of IFN-γ significantly increases inflammatory lymph node lymphangiogenesis independently of changes in VEGF-A or VEGF-C expression, suggesting that alterations in the balance of pro- and anti-lymphangiogenic cytokine expression can regulate lymphatic vessel formation. In conclusion, we show that gradients of lymphatic fluid stasis regulate not only the expression of pro-lymphangiogenic cytokines but also potent suppressors of lymphangiogenesis as a consequence of T-cell inflammation and that modulation of the balance between these stimuli can regulate lymphatic function.     

Quantification of atherosclerotic plaque activity and vascular inflammation using [18-F] fluorodeoxyglucose positron emission tomography/computed tomography (FDG-PET/CT)

Conventional non-invasive imaging modalities of atherosclerosis such as coronary artery calcium (CAC) and carotid intimal medial thickness (C-IMT) provide information about the burden of disease. However, despite multiple validation studies of CAC, and C-IMT, these modalities do not accurately assess plaque characteristics, and the composition and inflammatory state of the plaque determine its stability and, therefore, the risk of clinical events. [(18)F]-2-fluoro-2-deoxy-D-glucose (FDG) imaging using positron-emission tomography (PET)/computed tomography (CT) has been extensively studied in oncologic metabolism. Studies using animal models and immunohistochemistry in humans show that FDG-PET/CT is exquisitely sensitive for detecting macrophage activity, an important source of cellular inflammation in vessel walls. More recently, we and others have shown that FDG-PET/CT enables highly precise, novel measurements of inflammatory activity of activity of atherosclerotic plaques in large and medium-sized arteries. FDG-PET/CT studies have many advantages over other imaging modalities: 1) high contrast resolution; 2) quantification of plaque volume and metabolic activity allowing for multi-modal atherosclerotic plaque quantification; 3) dynamic, real-time, in vivo imaging; 4) minimal operator dependence. Finally, vascular inflammation detected by FDG-PET/CT has been shown to predict cardiovascular (CV) events independent of traditional risk factors and is also highly associated with overall burden of atherosclerosis. Plaque activity by FDG-PET/CT is modulated by known beneficial CV interventions such as short term (12 week) statin therapy as well as longer term therapeutic lifestyle changes (16 months). The current methodology for quantification of FDG uptake in atherosclerotic plaque involves measurement of the standardized uptake value (SUV) of an artery of interest and of the venous blood pool in order to calculate a target to background ratio (TBR), which is calculated by dividing the arterial SUV by the venous blood pool SUV. This method has shown to represent a stable, reproducible phenotype over time, has a high sensitivity for detection of vascular inflammation, and also has high inter-and intra-reader reliability. Here we present our methodology for patient preparation, image acquisition, and quantification of atherosclerotic plaque activity and vascular inflammation using SUV, TBR, and a global parameter called the metabolic volumetric product (MVP). These approaches may be applied to assess vascular inflammation in various study samples of interest in a consistent fashion as we have shown in several prior publications.     

Cryosectioning Yeast Communities for Examining Fluorescence Patterns

Microbes typically live in communities. The spatial organization of cells within a community is believed to impact the survival and function of the community¹. Optical sectioning techniques, including confocal and two-photon microscopy, have proven useful for observing spatial organization of bacterial and archaeal communities^2,3. A combination of confocal imaging and physical sectioning of yeast colonies has revealed internal organization of cells⁴. However, direct optical sectioning using confocal or two-photon microscopy has been only able to reach a few cell layers deep into yeast colonies. This limitation is likely because of strong scattering of light from yeast cells⁴.Here, we present a method based on fixing and cryosectioning to obtain spatial distribution of fluorescent cells within Saccharomyces cerevisiae communities. We use methanol as the fixative agent to preserve the spatial distribution of cells. Fixed communities are infiltrated with OCT compound, frozen, and cryosectioned in a cryostat. Fluorescence imaging of the sections reveals the internal organization of fluorescent cells within the community.Examples of yeast communities consisting of strains expressing red and green fluorescent proteins demonstrate the potentials of the cryosectioning method to reveal the spatial distribution of fluorescent cells as well as that of gene expression within yeast colonies^2,3. Even though our focus has been on Saccharomyces cerevisiae communities, the same method can potentially be applied to examine other microbial communities.     

Estimation of trunk mechanical properties using system identification: effects of experimental setup and modelling assumptions

The use of system identification to quantify trunk mechanical properties is growing in biomechanics research. The effects of several experimental and modelling factors involved in the system identification of trunk mechanical properties were investigated. Trunk kinematics and kinetics were measured in six individuals when exposed to sudden trunk perturbations. Effects of motion sensor positioning and properties of elements between the perturbing device and the trunk were investigated by adopting different models for system identification. Results showed that by measuring trunk kinematics at a location other than the trunk surface, the deformation of soft tissues is erroneously included into trunk kinematics and results in the trunk being predicted as a more damped structure. Results also showed that including elements between the trunk and the perturbing device in the system identification model did not substantially alter model predictions. Other important parameters that were found to substantially affect predictions were the cut-off frequency used when low-pass filtering raw data and the data window length used to estimate trunk properties.     

Carbonized Chicken Eggshell Membranes with 3D Architectures as High‐Performance Electrode Materials for Supercapacitors

Supercapacitor electrode materials are synthesized by carbonizing a common livestock biowaste in the form of chicken eggshell membranes. The carbonized eggshell membrane (CESM) is a three‐dimensional macroporous carbon film composed of interwoven connected carbon fibers containing around 10 wt% oxygen and 8 wt% nitrogen. Despite a relatively low surface area of 221 m² g^?1, exceptional specific capacitances of 297 F g^?1 and 284 F g^?1 are achieved in basic and acidic electrolytes, respectively, in a 3‐electrode system. Furthermore, the electrodes demonstrate excellent cycling stability: only 3% capacitance fading is observed after 10 000 cycles at a current density of 4 A g^?1. These very attractive electrochemical properties are discussed in the context of the unique structure and chemistry of the material.