Linear random knots and their scaling behavior

We present here a nonbiased probabilistic method that allows us to consistently analyze knottedness of linear random walks with up to several hundred noncorrelated steps. The method consists of analyzing the spectrum of knots formed by multiple closures of the same open walk through random points on a sphere enclosing the walk. Knottedness of individual "frozen" configurations of linear chains is therefore defined by a characteristic spectrum of realizable knots. We show that in the great majority of cases this method clearly defines the dominant knot type of a walk, i.e., the strongest component of the spectrum. In such cases, direct end-to-end closure creates a knot that usually coincides with the knot type that dominates the random closure spectrum. Interestingly, in a very small proportion of linear random walks, the knot type is not clearly defined. Such walks can be considered as residing in a border zone of the configuration space of two or more knot types. We also characterize the scaling behavior of linear random knots.

Differential expression of NLRP3 among hematopoietic cells.

Although the importance of the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome in health and disease is well appreciated, a precise characterization of NLRP3 expression is yet undetermined. To this purpose, we generated a knock-in mouse in which the Nlrp3 coding sequence was substituted for the GFP (enhanced GFP [egfp]) gene. In this way, the expression of eGFP is driven by the endogenous regulatory elements of the Nlrp3 gene. In this study, we show that eGFP expression indeed mirrors that of NLRP3. Interestingly, splenic neutrophils, macrophages, and, in particular, monocytes and conventional dendritic cells showed robust eGFP fluorescence, whereas lymphoid subsets, eosinophils, and plasmacytoid dendritic cells showed negligible eGFP levels. NLRP3 expression was highly inducible in macrophages, both by MyD88- and Trif-dependent pathways. In vivo, when mice were challenged with diverse inflammatory stimuli, differences in both the number of eGFP-expressing cells and fluorescence intensity were observed in the draining lymph node. Thus, NLRP3 levels at the site of adaptive response initiation are controlled by recruitment of NLRP3-expressing cells and by NLRP3 induction.

Comparison between a linear ion trap and a triple quadruple MS in the sensitive detection of large peptides at femtomole amounts on column.

In addition to the importance of sample preparation and extract separation, MS detection is a key factor in the sensitive quantification of large undigested peptides. In this article, a linear ion trap MS (LIT-MS) and a triple quadrupole MS (TQ-MS) have been compared in the detection of large peptides at subnanomolar concentrations. Natural brain natriuretic peptide, C-peptide, substance P and D-Junk-inhibitor peptide, a full D-amino acid therapeutic peptide, were chosen. They were detected by ESI and simultaneous MS(1) and MS(2) acquisitions. With direct peptide infusion, MS(2) spectra revealed that fragmentation was peptide dependent, milder on the LIT-MS and required high collision energies on the TQ-MS to obtain high-intensity product ions. Peptide adsorption on surfaces was overcome and peptide dilutions ranging from 0.1 to 25 nM were injected onto an ultra high-pressure LC system with a 1 mm id analytical column and coupled with the MS instruments. No difference was observed between the two instruments when recording in LC-MS(1) acquisitions. However, in LC-MS(2) acquisitions, a better sensitivity in the detection of large peptides was observed with the LIT-MS. Indeed, with the three longer peptides, the typical fragmentation in the TQ-MS resulted in a dramatic loss of sensitivity (> or = 10x).

Differential regulation of Na-K-ATPase isoform gene expression by T3 during rat brain development.

A fetal rat telencephalon organotypic cell culture system was found to reproduce the developmental pattern of Na-K-adenosinetriphosphatase (ATPase) gene expression observed in vivo [Am. J. Physiol. 258 (Cell Physiol. 27): C1062-C1069, 1990]. We have used this culture system to study the effects of triiodothyronine (T3; 0.003-30 nM) on mRNA abundance and basal transcription rates of Na-K-ATPase isoforms. Steady-state mRNA levels were low at culture day 6 (corresponding to the day of birth) but distinct for each isoform alpha 3 much greater than beta 1 = beta 2 greater than alpha 2 greater than alpha 1. At culture day 6, T3 did not modify mRNA abundance of any isoform. At culture day 12 (corresponding to day 7 postnatal), T3 increased the mRNA level of alpha 2 (4- to 7-fold), beta 2 (4- to 5-fold), alpha 1 (3- to 6-fold), and beta 1 (1.5-fold), whereas alpha 3 mRNA levels remained unchanged. Interestingly, the basal transcription rate for each isoform differed strikingly (alpha 2 greater than alpha 1 much greater than beta 1 = beta 2 greater than alpha 3) but remained stable throughout 12 days of culture and was not regulated by T3. Thus we observed an inverse relationship between rate of transcription and rate of mRNA accumulation for each alpha-isoform, suggesting that alpha 1- and alpha 2-mRNA are turning over rapidly whereas alpha 3-mRNA is turning over slowly. Our data indicate that one of the mechanisms by which T3 selectively controls Na-K-ATPase gene expression during brain development in vitro occurs at the posttranscriptional level.

Differential insertion of GPI-anchored GFPs into lipid rafts of live cells.

Partitioning of proteins in cholesterol and sphingolipid enriched plasma membrane microdomains, called lipid rafts, is critical for many signal transduction and protein sorting events. Although raft partitioning of many signaling molecules remains to be determined, glycosylphosphatidyl-inositol (GPI)-anchored proteins possess high affinity for lipid rafts and are currently exploited as markers to investigate fundamental mechanisms in protein sorting and signal transduction events. In this study, we demonstrate that two recombinant GPI-anchored green fluorescent proteins (GFP-GPIs) that differ in their GPI signal sequence confer distinct localization in plasma membrane microdomains. GFP fused to the GPI signal of the decay accelerating factor GFP-GPI(DAF) partitioned exclusively in lipid rafts, whereas GFP fused to the GPI signal of TRAIL-R3, GFP-GPI(TRAIL-R3), associated only minimally with microdomains. In addition, we investigated the unique ability of purified GFP-GPIs to insert into membrane microdomains of primary lymphocytes. This cell surface painting allows rapid, stable, and functional association of the GPI-anchored proteins with the target cell plasma membrane. The distinct membrane localization of the two GFP-GPIs was observed irrespective of whether the GPI-anchored molecules were painted or transfected. Furthermore, we show that painted GFP-GPI(DAF) was totally dependent on the GPI anchor and that the membrane insertion was increased by the addition of raft-associated lipids such as cholesterol, sphingomyelin, and dipalmitoyl-phosphatidylethanolamine. Thus, this study provides evidence that different GPI signal sequences lead to distinct membrane microdomain localization and that painted GFP-GPI(DAF) serves as an excellent fluorescent marker for lipid rafts in live cells.

A general linear relaxometry model of R1 using imaging data.

PURPOSE: The longitudinal relaxation rate (R1 ) measured in vivo depends on the local microstructural properties of the tissue, such as macromolecular, iron, and water content. Here, we use whole brain multiparametric in vivo data and a general linear relaxometry model to describe the dependence of R1 on these components. We explore a) the validity of having a single fixed set of model coefficients for the whole brain and b) the stability of the model coefficients in a large cohort. METHODS: Maps of magnetization transfer (MT) and effective transverse relaxation rate (R2 *) were used as surrogates for macromolecular and iron content, respectively. Spatial variations in these parameters reflected variations in underlying tissue microstructure. A linear model was applied to the whole brain, including gray/white matter and deep brain structures, to determine the global model coefficients. Synthetic R1 values were then calculated using these coefficients and compared with the measured R1 maps. RESULTS: The model's validity was demonstrated by correspondence between the synthetic and measured R1 values and by high stability of the model coefficients across a large cohort. CONCLUSION: A single set of global coefficients can be used to relate R1 , MT, and R2 * across the whole brain. Our population study demonstrates the robustness and stability of the model. Magn Reson Med, 2014. © 2014 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. Magn Reson Med 73:1309-1314, 2015. © 2014 Wiley Periodicals, Inc.

A two-phase composite in simple shear: Effective mechanical anisotropy development and localization potential

We present a combined shape and mechanical anisotropy evolution model for a two-phase inclusion-bearing rock subject to large deformation. A single elliptical inclusion embedded in a homogeneous but anisotropic matrix is used to represent a simplified shape evolution enforced on all inclusions. The mechanical anisotropy develops due to the alignment of elongated inclusions. The effective anisotropy is quantified using the differential effective medium (DEM) approach. The model can be run for any deformation path and an arbitrary viscosity ratio between the inclusion and host phase. We focus on the case of simple shear and weak inclusions. The shape evolution of the representative inclusion is largely insensitive to the anisotropy development and to parameter variations in the studied range. An initial hardening stage is observed up to a shear strain of gamma = 1 irrespective of the inclusion fraction. The hardening is followed by a softening stage related to the developing anisotropy and its progressive rotation toward the shear direction. The traction needed to maintain a constant shear rate exhibits a fivefold drop at gamma = 5 in the limiting case of an inviscid inclusion. Numerical simulations show that our analytical model provides a good approximation to the actual evolution of a two-phase inclusion-host composite. However, the inclusions develop complex sigmoidal shapes resulting in the formation of an S-C fabric. We attribute the observed drop in the effective normal viscosity to this structural development. We study the localization potential in a rock column bearing varying fraction of inclusions. In the inviscid inclusion case, a strain jump from gamma = 3 to gamma = 100 is observed for a change of the inclusion fraction from 20% to 33%.

Genotype-phenotype interactions in primary dystonias revealed by differential changes in brain structure.

Our understanding of how genotype determines phenotype in primary dystonia is limited. Familial young-onset primary dystonia is commonly due to the DYT1 gene mutation. A critical question, given the 30% penetrance of clinical symptoms in DYT1 mutation carriers, is why the same genotype leads to differential clinical expression and whether non-DYT1 adult-onset primary dystonia, with and without family history share pathophysiological mechanisms with DYT1 dystonia. This study examines the relationship between dystonic phenotype and the DYT1 gene mutation by monitoring whole-brain structure using voxel-based morphometry. We acquired magnetic resonance imaging data of symptomatic and asymptomatic DYT1 mutation carriers, of non-DYT1 primary dystonia patients, with and without family history and control subjects with normal DYT1 alleles. By crossing the factors genotype and phenotype we demonstrate a significant interaction in terms of brain anatomy confined to the basal ganglia bilaterally. The explanation for this effect differs according to both gene and dystonia status: non-DYT1 adult-onset dystonia patients and asymptomatic DYT1 carriers have significantly larger basal ganglia compared to healthy subjects and symptomatic DYT1 mutation carriers. There is a significant negative correlation between severity of dystonia and basal ganglia size in DYT1 mutation carriers. We propose that differential pathophysiological and compensatory mechanisms lead to brain structure changes in non-DYT1 primary adult-onset dystonias and DYT1 gene carriers. Given the range of age of onset, there may be differential genetic modulation of brain development that in turn determines clinical expression. Alternatively, a DYT1 gene dependent primary defect of motor circuit development may lead to stress-induced remodelling of the basal ganglia and hence dystonia.

Brain Spectrins 240/235 and 240/235E: Differential Expression During Development of Chicken Dorsal Root Ganglia in vivo and in vitro.

Brain spectrin, a membrane-related cytoskeletal protein, exists as two isoforms. Brain spectrin 240/235 is localized preferentially in the perikaryon and axon of neuronal cells and brain spectrin 240/235E is found essentially in the neuronal soma and dendrites and in glia (Riederer et al., 1986, J. Cell Biol., 102, 2088 - 2097). The sensory neurons in dorsal root ganglia, devoid of any dendrites, make a good tool to investigate such differential expression of spectrin isoforms. In this study expression and localization of both brain spectrin isoforms were analysed during early chicken dorsal root ganglia development in vivo and in culture. Both isoforms appeared at embryonic day 6. Brain spectrin 240/235 exhibited a transient increase during embryonic development and was first expressed in ventrolateral neurons. In ganglion cells in situ and in culture this spectrin type showed a somato - axonal distribution pattern. In contrast, brain spectrin 240/235E slightly increased between E6 and E15 and remained practically unchanged. It was localized mainly in smaller neurons of the mediodorsal area as punctate staining in the cytoplasm, was restricted exclusively to the ganglion cell perikarya and was absent from axons both in situ and in culture. This study suggests that brain spectrin 240/235 may contribute towards outgrowth, elongation and maintenance of axonal processes and that brain spectrin 240/235E seems to be exclusively involved in the stabilization of the cytoarchitecture of cell bodies in a selected population of ganglion cells.

Differential neuroglycan C expression during retinal degeneration in Rpe65-/- mice.

PURPOSE: An increased mRNA expression of the genes coding for the extracellular matrix proteins neuroglycan C (NGC), interphotoreceptor matrix proteoglycan 2 (IMPG2), and CD44 antigen (CD44) has been observed during retinal degeneration in mice with a targeted disruption of the Rpe65 gene (Rpe65-/- mouse). To validate these data, we analyzed this differential expression in more detail by characterizing retinal NGC mRNA isoform and protein expression during disease progression. METHODS: Retinas from C57/Bl6 wild-type and Rpe65-/- mice, ranging 2 to 18 months of age, were used. NGC, IMPG2, and CD44 mRNA expression was assessed by oligonucleotide microarray, quantitative PCR, and in situ hybridization. Retinal NGC protein expression was analyzed by western blot and immunohistochemistry. RESULTS: As measured by quantitative PCR, mRNA expression of NGC and CD44 was induced by about 2 fold to 3 fold at all time points in Rpe65-/- retinas, whereas initially 4 fold elevated IMPG2 mRNA levels progressively declined. NGC and IMPG2 mRNAs were expressed in the ganglion cell layer, the inner nuclear layer, and at the outer limiting membrane. NGC mRNA was also detected in retinal pigment epithelium cells (RPE), where its mRNA expression was not induced during retinal degeneration. NGC-I was the major isoform detected in the retina and the RPE, whereas NGC-III was barely detected and NGC-II could not be assessed. NGC protein expression was at its highest levels on the apical membrane of the RPE. NGC protein levels were induced in retinas from 2- and 4-month-old Rpe65-/- mice, and an increased amount of the activity-cleaved NGC ectodomain containing an epidermal growth factor (EGF)-like domain was detected. CONCLUSIONS: During retinal degeneration in Rpe65-/- mice, NGC expression is induced in the neural retina, but not in the RPE, where NGC is expressed at highest levels.

Differential impact of lacunes and microvascular lesions on poststroke depression.

BACKGROUND AND PURPOSE: Previous studies have postulated that poststroke depression (PSD) might be related to cumulative vascular brain pathology rather than to the location and severity of a single macroinfarct. We performed a detailed analysis of all types of microvascular lesions and lacunes in 41 prospectively documented and consecutively autopsied stroke cases. METHODS: Only cases with first-onset depression <2 years after stroke were considered as PSD in the present series. Diagnosis of depression was established prospectively using DSM-IV criteria for major depression. Neuropathological evaluation included bilateral semiquantitative assessment of microvascular ischemic pathology and lacunes; statistical analysis included Fisher exact test, Mann-Whitney U test, and regression models. RESULTS: Macroinfarct site was not related to the occurrence of PSD for any of the locations studied. Thalamic and basal ganglia lacunes occurred significantly more often in PSD cases. Higher lacune scores in basal ganglia, thalamus, and deep white matter were associated with an increased PSD risk. In contrast, microinfarct and diffuse or periventricular demyelination scores were not increased in PSD. The combined lacune score (thalamic plus basal ganglia plus deep white matter) explained 25% of the variability of PSD occurrence. CONCLUSIONS: The cumulative vascular burden resulting from chronic accumulation of lacunar infarcts within the thalamus, basal ganglia, and deep white matter may be more important than single infarcts in the prediction of PSD.

Differential protein labeling with thiol-reactive infrared DY-680 and DY-780 maleimides and analysis by two-dimensional gel electrophoresis.

Differential protein labeling with 2-DE separation is an effective method for distinguishing differences in the protein composition of two or more protein samples. Here, we report on a sensitive infrared-based labeling procedure, adding a novel tool to the many labeling possibilities. Defined amounts of newborn and adult mouse brain proteins and tubulin were exposed to maleimide-conjugated infrared dyes DY-680 and DY-780 followed by 1- and 2-DE. The procedure allows amounts of less than 5 microg of cysteine-labeled protein mixtures to be detected (together with unlabeled proteins) in a single 2-DE step with an LOD of individual proteins in the femtogram range; however, co-migration of unlabeled proteins and subsequent general protein stains are necessary for a precise comparison. Nevertheless, the most abundant thiol-labeled proteins, such as tubulin, were identified by MS, with cysteine-containing peptides influencing the accuracy of the identification score. Unfortunately, some infrared-labeled proteins were no longer detectable by Western blots. In conclusion, differential thiol labeling with infrared dyes provides an additional tool for detection of low-abundant cysteine-containing proteins and for rapid identification of differences in the protein composition of two sets of protein samples.

A re-evaluation of Raman as a tool for the determination of 12C and 13C in geological fluid inclusions

Raman spectroscopy has been used by fluid inclusionists to: 1) identify and quantitatively determine the relative abundances of gaseous species within fluid inclusions; 2) identify solid phases precipitating from, or accidentally trapped, within fluid inclusions; and 3) determine the detection limits of the C-13/C-12 ratio in the CO2 bearing phase of fluid inclusions.

Are niche-based species distribution models transferable in space?

Aim To assess the geographical transferability of niche-based species distribution models fitted with two modelling techniques. Location Two distinct geographical study areas in Switzerland and Austria, in the subalpine and alpine belts. Methods Generalized linear and generalized additive models (GLM and GAM) with a binomial probability distribution and a logit link were fitted for 54 plant species, based on topoclimatic predictor variables. These models were then evaluated quantitatively and used for spatially explicit predictions within (internal evaluation and prediction) and between (external evaluation and prediction) the two regions. Comparisons of evaluations and spatial predictions between regions and models were conducted in order to test if species and methods meet the criteria of full transferability. By full transferability, we mean that: (1) the internal evaluation of models fitted in region A and B must be similar; (2) a model fitted in region A must at least retain a comparable external evaluation when projected into region B, and vice-versa; and (3) internal and external spatial predictions have to match within both regions. Results The measures of model fit are, on average, 24% higher for GAMs than for GLMs in both regions. However, the differences between internal and external evaluations (AUC coefficient) are also higher for GAMs than for GLMs (a difference of 30% for models fitted in Switzerland and 54% for models fitted in Austria). Transferability, as measured with the AUC evaluation, fails for 68% of the species in Switzerland and 55% in Austria for GLMs (respectively for 67% and 53% of the species for GAMs). For both GAMs and GLMs, the agreement between internal and external predictions is rather weak on average (Kulczynski's coefficient in the range 0.3-0.4), but varies widely among individual species. The dominant pattern is an asymmetrical transferability between the two study regions (a mean decrease of 20% for the AUC coefficient when the models are transferred from Switzerland and 13% when they are transferred from Austria). Main conclusions The large inter-specific variability observed among the 54 study species underlines the need to consider more than a few species to test properly the transferability of species distribution models. The pronounced asymmetry in transferability between the two study regions may be due to peculiarities of these regions, such as differences in the ranges of environmental predictors or the varied impact of land-use history, or to species-specific reasons like differential phenotypic plasticity, existence of ecotypes or varied dependence on biotic interactions that are not properly incorporated into niche-based models. The lower variation between internal and external evaluation of GLMs compared to GAMs further suggests that overfitting may reduce transferability. Overall, a limited geographical transferability calls for caution when projecting niche-based models for assessing the fate of species in future environments.