Limited Macrophage Positional Dynamics in Progressing or Regressing Murine Atherosclerotic PlaquesBrief Report

Objective Macrophages play important roles in the pathogenesis of atherosclerosis, but their dynamics within plaques remain obscure. We aimed to quantify macrophage positional dynamics within progressing and regressing atherosclerotic plaques. Approach and Results In a stable intravital preparation, large asymmetrical foamy macrophages in the intima of carotid artery plaques were sessile, but smaller rounded cells nearer plaque margins, possibly newly recruited monocytes, mobilized laterally along plaque borders. Thus, to test macrophage dynamics in plaques over a longer period of time in progressing and regressing disease, we quantified displacement of nondegradable phagocytic particles within macrophages for up to 6 weeks. In progressing plaques, macrophage-associated particles appeared to mobilize to deeper layers in plaque, whereas in regressing plaques, the label was persistently located near the lumen. By measuring the distance of the particles from the floor of the plaque, we discovered that particles remained at the same distance from the floor regardless of plaque progression or regression. The apparent deeper penetration of labeled cells in progressing conditions could be attributed to monocyte recruitment that generated new superficial layers of macrophages over the labeled phagocytes. Conclusion: s Although there may be individual exceptions, as a population, newly differentiated macrophages fail to penetrate significantly deeper than the limited depth they reside on initial entry, regardless of plaque progression, or regression. These limited dynamics may prevent macrophages from escaping areas with unfavorable conditions (such as hypoxia) and pose a challenge for newly recruited macrophages to clear debris through efferocytosis deep within plaque

Forefoot pathology in rheumatoid arthritis identified with ultrasound may not localise to areas of highest pressure: cohort observations at baseline and twelve months

BackgroundPlantar pressures are commonly used as clinical measures, especially to determine optimum foot orthotic design. In rheumatoid arthritis (RA) high plantar foot pressures have been linked to metatarsophalangeal (MTP) joint radiological erosion scores. However, the sensitivity of foot pressure measurement to soft tissue pathology within the foot is unknown. The aim of this study was to observe plantar foot pressures and forefoot soft tissue pathology in patients who have RA.Methods A total of 114 patients with established RA (1987 ACR criteria) and 50 healthy volunteers were assessed at baseline. All RA participants returned for reassessment at twelve months. Interface foot-shoe plantar pressures were recorded using an F-Scan® system. The presence of forefoot soft tissue pathology was assessed using a DIASUS musculoskeletal ultrasound (US) system. Chi-square analyses and independent t-tests were used to determine statistical differences between baseline and twelve months. Pearson’s correlation coefficient was used to determine interrelationships between soft tissue pathology and foot pressures.ResultsAt baseline, RA patients had a significantly higher peak foot pressures compared to healthy participants and peak pressures were located in the medial aspect of the forefoot in both groups. In contrast, RA participants had US detectable soft tissue pathology in the lateral aspect of the forefoot. Analysis of person specific data suggests that there are considerable variations over time with more than half the RA cohort having unstable presence of US detectable forefoot soft tissue pathology. Findings also indicated that, over time, changes in US detectable soft tissue pathology are out of phase with changes in foot-shoe interface pressures both temporally and spatially.Conclusions We found that US detectable forefoot soft tissue pathology may be unrelated to peak forefoot pressures and suggest that patients with RA may biomechanically adapt to soft tissue forefoot pathology. In addition, we have observed that, in patients with RA, interface foot-shoe pressures and the presence of US detectable forefoot pathology may vary substantially over time. This has implications for clinical strategies that aim to offload peak plantar pressures

Quantum Monte Carlo calculations of the one-body density matrix and excitation energies of silicon

Quantum Monte Carlo (QMC) techniques are used to calculate the one-body density matrix and excitation energies for the valence electrons of bulk silicon. The one-body density matrix and energies are obtained from a Slater-Jastrow wave function with a determinant of local density approximation (LDA) orbitals. The QMC density matrix evaluated in a basis of LDA orbitals is strongly diagonally dominant. The natural orbitals obtained by diagonalizing the QMC density matrix resemble the LDA orbitals very closely. Replacing the determinant of LDA orbitals in the wave function by a determinant of natural orbitals makes no significant difference to the quality of the wave function's nodal surface, leaving the diffusion Monte Carlo energy unchanged. The Extended Koopmans' Theorem for correlated wave functions is used to calculate excitation energies for silicon, which are in reasonable agreement with the available experimental data. A diagonal approximation to the theorem, evaluated in the basis of LDA orbitals, works quite well for both the quasihole and quasielectron states. We have found that this approximation has an advantageous scaling with system size, allowing more efficient studies of larger systems.Comment: 13 pages, 4 figures. To appear in Phys. Rev.

Finite size errors in quantum many-body simulations of extended systems

Further developments are introduced in the theory of finite size errors in quantum many-body simulations of extended systems using periodic boundary conditions. We show that our recently introduced Model Periodic Coulomb interaction [A. J. Williamson et al., Phys. Rev. B 55, R4851 (1997)] can be applied consistently to all Coulomb interactions in the system. The Model Periodic Coulomb interaction greatly reduces the finite size errors in quantum many-body simulations. We illustrate the practical application of our techniques with Hartree-Fock and variational and diffusion quantum Monte Carlo calculations for ground and excited state calculations. We demonstrate that the finite size effects in electron promotion and electron addition/subtraction excitation energy calculations are very similar.Comment: 15 pages, 6 figures. To appear in Phys. Rev.

Reproducible copy number variation patterns among single circulating tumor cells of lung cancer patients

Circulating tumor cells (CTCs) enter peripheral blood from primary tumors and seed metastases. The genome sequencing of CTCs could offer noninvasive prognosis or even diagnosis, but has been hampered by low single-cell genome coverage of scarce CTCs. Here, we report the use of the recently developed multiple annealing and looping-based amplification cycles for whole-genome amplification of single CTCs from lung cancer patients. We observed characteristic cancer-associated single-nucleotide variations and insertions/deletions in exomes of CTCs. These mutations provided information needed for individualized therapy, such as drug resistance and phenotypic transition, but were heterogeneous from cell to cell. In contrast, every CTC from an individual patient, regardless of the cancer subtypes, exhibited reproducible copy number variation (CNV) patterns, similar to those of the metastatic tumor of the same patient. Interestingly, different patients with the same lung cancer adenocarcinoma (ADC) shared similar CNV patterns in their CTCs. Even more interestingly, patients of small-cell lung cancer have CNV patterns distinctly different from those of ADC patients. Our finding suggests that CNVs at certain genomic loci are selected for the metastasis of cancer. The reproducibility of cancer-specific CNVs offers potential for CTC-based cancer diagnostics.Chemistry and Chemical Biolog

Transcription factors that mediate epithelial–mesenchymal transition lead to multidrug resistance by upregulating ABC transporters

Development of multidrug resistance (MDR) is a major deterrent in the effective treatment of metastatic cancers by chemotherapy. Even though MDR and cancer invasiveness have been correlated, the molecular basis of this link remains obscure. We show here that treatment with chemotherapeutic drugs increases the expression of several ATP binding cassette transporters (ABC transporters) associated with MDR, as well as epithelial–mesenchymal transition (EMT) markers, selectively in invasive breast cancer cells, but not in immortalized or non-invasive cells. Interestingly, the mere induction of an EMT in immortalized and non-invasive cell lines increased their expression of ABC transporters, migration, invasion, and drug resistance. Conversely, reversal of EMT in invasive cells by downregulating EMT-inducing transcription factors reduced their expression of ABC transporters, invasion, and rendered them more chemosensitive. Mechanistically, we demonstrate that the promoters of ABC transporters carry several binding sites for EMT-inducing transcription factors, and overexpression of Twist, Snail, and FOXC2 increases the promoter activity of ABC transporters. Furthermore, chromatin immunoprecipitation studies revealed that Twist binds directly to the E-box elements of ABC transporters. Thus, our study identifies EMT inducers as novel regulators of ABC transporters, thereby providing molecular insights into the long-standing association between invasiveness and MDR. Targeting EMT transcription factors could hence serve as novel strategies to curb both metastasis and the associated drug resistance

Crystallisation route map

A route map for the assessment of crystallisation processes is presented. A theoretical background on solubility, meta-stable zone width, nucleation and crystal growth kinetics is presented with practical examples. The concepts of crystallisation hydrodynamics and the application of population balances and computational fluid dynamics for modelling crystallisation processes and their scaling up are also covered

Cruciferous vegetable supplementation in a controlled diet study alters the serum peptidome in a GSTM1-genotype dependent manner

<p>Abstract</p> <p>Background</p> <p>Cruciferous vegetable intake is inversely associated with the risk of several cancers. Isothiocyanates (ITC) are hypothesized to be the major bioactive constituents contributing to these cancer-preventive effects. The polymorphic glutathione-<it>S</it>-transferase (GST) gene family encodes several enzymes which catalyze ITC degradation <it>in vivo</it>.</p> <p>Methods</p> <p>We utilized high throughput proteomics methods to examine how human serum peptides (the "peptidome") change in response to cruciferous vegetable feeding in individuals of different <it>GSTM1 </it>genotypes. In two randomized, crossover, controlled feeding studies (EAT and 2EAT) participants consumed a fruit- and vegetable-free basal diet and the basal diet supplemented with cruciferous vegetables. Serum samples collected at the end of the feeding period were fractionated and matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry spectra were obtained. Peak identification/alignment computer algorithms and mixed effects models were used to analyze the data.</p> <p>Results</p> <p>After analysis of spectra from EAT participants, 24 distinct peaks showed statistically significant differences associated with cruciferous vegetable intake. Twenty of these peaks were driven by their <it>GSTM1 </it>genotype (i.e., <it>GSTM1+ </it>or <it>GSTM1- </it>null). When data from EAT and 2EAT participants were compared by joint processing of spectra to align a common set, 6 peaks showed consistent changes in both studies in a genotype-dependent manner. The peaks at 6700 <it>m/z </it>and 9565 <it>m/z </it>were identified as an isoform of transthyretin (TTR) and a fragment of zinc α2-glycoprotein (ZAG), respectively.</p> <p>Conclusions</p> <p>Cruciferous vegetable intake in <it>GSTM1+ </it>individuals led to changes in circulating levels of several peptides/proteins, including TTR and a fragment of ZAG. TTR is a known marker of nutritional status and ZAG is an adipokine that plays a role in lipid mobilization. The results of this study present evidence that the <it>GSTM1</it>-genotype modulates the physiological response to cruciferous vegetable intake.</p

Stable Expression of Antibiotic-Resistant Gene ble from Streptoalloteichus hindustanus in the Mitochondria of Chlamydomonas reinhardtii

The mitochondrial expression of exogenous antibiotic resistance genes has not been demonstrated successfully to date, which has limited the development of antibiotic resistance genes as selectable markers for mitochondrial site-directed transformation in Chlamydomonas reinhardtii. In this work, the plasmid pBSLPNCB was constructed by inserting the gene ble of Streptoalloteichus hindustanus (Sh ble), encoding a small (14-kilodalton) protective protein into the site between TERMINVREP-Left repeats and the cob gene in a fragment of mitochondrial DNA (mtDNA) of C. reinhardtii. The fusion DNA-construct, which contained TERMINVREP-Left, Sh ble, cob, and partial nd4 sequence, were introduced into the mitochondria of the respiratory deficient dum-1 mutant CC-2654 of C. reinhardtii by biolistic particle delivery system. A large number of transformants were obtained after eight weeks in the dark. Subsequent subculture of the transformants on the selection TAP media containing 3 ìg/mL Zeomycin for 12 months resulted in genetically modified transgenic algae MT-Bs. Sequencing and Southern analyses on the mitochondrial genome of the different MT-B lines revealed that Sh ble gene had been integrated into the mitochondrial genome of C. reinhardtii. Both Western blot, using the anti-BLE monoclonal antibody, and Zeomycin tolerance analysis confirmed the presence of BLE protein in the transgenic algal cells. It indicates that the Sh ble gene can be stably expressed in the mitochondria of C. reinhardtii

The genome-wide dynamics of purging during selfing in maize

Self-fertilization (also known as selfing) is an important reproductive strategy in plants and a widely applied tool for plant genetics and plant breeding. Selfing can lead to inbreeding depression by uncovering recessive deleterious variants, unless these variants are purged by selection. Here we investigated the dynamics of purging in a set of eleven maize lines that were selfed for six generations. We show that heterozygous, putatively deleterious single nucleotide polymorphisms are preferentially lost from the genome during selfing. Deleterious single nucleotide polymorphisms were lost more rapidly in regions of high recombination, presumably because recombination increases the efficacy of selection by uncoupling linked variants. Overall, heterozygosity decreased more slowly than expected, by an estimated 35% to 40% per generation instead of the expected 50%, perhaps reflecting pervasive associative overdominance. Finally, three lines exhibited marked decreases in genome size due to the purging of transposable elements. Genome loss was more likely to occur for lineages that began with larger genomes with more transposable elements and chromosomal knobs. These three lines purged an average of 398 Mb from their genomes, an amount equivalent to three Arabidopsis thaliana genomes per lineage, in only a few generations