Triglyceride-Rich Lipoproteins Modulate the Distribution and Extravasation of Ly6C/Gr1low Monocytes

Monocytes are heterogeneous effector cells involved in the maintenance and restoration of tissue integrity. However, their response to hyperlipidemia remains poorly understood. Here, we report that in the presence of elevated levels of triglyceride-rich lipoproteins, induced by administration of poloxamer 407, the blood numbers of non-classical Ly6C/Gr1(low) monocytes drop, while the number of bone marrow progenitors remains similar. We observed an increased crawling and retention of the Gr1(low) monocytes at the endothelial interface and a marked accumulation of CD68(+) macrophages in several organs. Hypertriglyceridemia was accompanied by an increased expression of tissue, and plasma CCL4 and blood Gr1(low) monocyte depletion involved a pertussis-toxin-sensitive receptor axis. Collectively, these findings demonstrate that a triglyceride-rich environment can alter blood monocyte distribution, promoting the extravasation of Gr1(low) cells. The behavior of these cells in response to dyslipidemia highlights the significant impact that high levels of triglyceride-rich lipoproteins may have on innate immune cells

Evaluation of a blocking ELISA for the detection of antibodies against Lawsonia intracellularis in pig sera

<p>Abstract</p> <p>Background</p> <p><it>Lawsonia intracellularis </it>is a common cause of chronic diarrhoea and poor performance in young growing pigs. Diagnosis of this obligate intracellular bacterium is based on the demonstration of the microbe or microbial DNA in tissue specimens or faecal samples, or the demonstration of <it>L. intracellularis</it>-specific antibodies in sera. The aim of the present study was to evaluate a blocking ELISA in the detection of serum antibodies to <it>L. intracellularis</it>, by comparison to the previously widely used immunofluorescent antibody test (IFAT).</p> <p>Methods</p> <p>Sera were collected from 176 pigs aged 8-12 weeks originating from 24 herds with or without problems with diarrhoea and poor performance in young growing pigs. Sera were analyzed by the blocking ELISA and by IFAT. Bayesian modelling techniques were used to account for the absence of a gold standard test and the results of the blocking ELISA was modelled against the IFAT test with a "2 dependent tests, 2 populations, no gold standard" model.</p> <p>Results</p> <p>At the finally selected cut-off value of percent inhibition (PI) 35, the diagnostic sensitivity of the blocking ELISA was 72% and the diagnostic specificity was 93%. The positive predictive value was 0.82 and the negative predictive value was 0.89, at the observed prevalence of 33.5%.</p> <p>Conclusion</p> <p>The sensitivity and specificity as evaluated by Bayesian statistic techniques differed from that previously reported. Properties of diagnostic tests may well vary between countries, laboratories and among populations of animals. In the absence of a true gold standard, the importance of validating new methods by appropriate statistical methods and with respect to the target population must be emphasized.</p

Monodisperse α-Fe2O3 Mesoporous Microspheres: One-Step NaCl-Assisted Microwave-Solvothermal Preparation, Size Control and Photocatalytic Property

A simple one-step NaCl-assisted microwave-solvothermal method has been developed for the preparation of monodisperse α-Fe2O3 mesoporous microspheres. In this approach, Fe(NO3)3 · 9H2O is used as the iron source, and polyvinylpyrrolidone (PVP) acts as a surfactant in the presence of NaCl in mixed solvents of H2O and ethanol. Under the present experimental conditions, monodisperse α-Fe2O3 mesoporous microspheres can form via oriented attachment of α-Fe2O3 nanocrystals. One of the advantages of this method is that the size of α-Fe2O3 mesoporous microspheres can be adjusted in the range from ca. 170 to ca. 260 nm by changing the experimental parameters. High photocatalytic activities in the degradation of salicylic acid are observed for α-Fe2O3 mesoporous microspheres with different specific surface areas

Measurement of the top quark mass using the matrix element technique in dilepton final states

We present a measurement of the top quark mass in pp¯ collisions at a center-of-mass energy of 1.96 TeV at the Fermilab Tevatron collider. The data were collected by the D0 experiment corresponding to an integrated luminosity of 9.7  fb−1. The matrix element technique is applied to tt¯ events in the final state containing leptons (electrons or muons) with high transverse momenta and at least two jets. The calibration of the jet energy scale determined in the lepton+jets final state of tt¯ decays is applied to jet energies. This correction provides a substantial reduction in systematic uncertainties. We obtain a top quark mass of mt=173.93±1.84  GeV

The zinc finger domain of Wilms' tumor 1 suppressor gene (WT1) behaves as a dominant negative, leading to abrogation of WT1 oncogenic potential in breast cancer cells

Abstract Introduction There is growing evidence that the Wilms' tumor 1 suppressor gene (WT1) behaves as an oncogene in some forms of breast cancer. Previous studies have demonstrated that the N-terminal domain of WT1 can act as a dominant negative through self-association. In the studies presented here we have explored the potential for the zinc finger domain (ZF) of WT1 to also have dominant-negative effects, and thus further our understanding of this protein. Methods Using full-length and ZF-only forms of WT1 we assessed their effect on the WT1 and c-myc promoter using luciferase and chromatin immunoprecipitation assays. The gene expression levels were determined by quantitative real-time RT-PCR, northern blot and western blot. We also assessed the effect of the ZF-only form on the growth of breast cancer cell lines in culture. Results Transfection with WT1–ZF plasmids resulted in a stronger inhibition of WT1 promoter than full-length WT1 in breast cancer cells. The WT1–ZF form lacking the lysine–threonine–serine (KTS) insert (ZF - KTS) can bind to the majority of WT1 consensus sites throughout the WT1 promoter region, while the ZF containing the insert (ZF + KTS) form only binds to sites in the proximal promoter. The abundances of endogenous WT1 mRNA and protein were markedly decreased following the stable expression of ZF - KTS in breast cancer cells. The expressions of WT1 target genes, including c-myc, Bcl-2, amphiregulin and TERT, were similarly suppressed by ZF - KTS. Moreover, WT1–ZF - KTS abrogated the transcriptional activation of c-myc mediated by all four predominant isoforms of WT1 (including or lacking alternatively spliced exons 5 and 9). Finally, WT1–ZF - KTS inhibited colony formation and cell division, but induced apoptosis in MCF-7 cells. Conclusion Our observations strongly argue that the WT1–ZF plasmid behaves as a dominant-negative regulator of the endogenous WT1 in breast cancer cells. The inhibition on proliferation of breast cancer cells by WT1–ZF - KTS provides a potential candidate of gene therapy for breast cancer

Adaptive Mutations in the JC Virus Protein Capsid Are Associated with Progressive Multifocal Leukoencephalopathy (PML)

PML is a progressive and mostly fatal demyelinating disease caused by JC virus infection and destruction of infected oligodendrocytes in multiple brain foci of susceptible individuals. While JC virus is highly prevalent in the human population, PML is a rare disease that exclusively afflicts only a small percentage of immunocompromised individuals including those affected by HIV (AIDS) or immunosuppressive drugs. Viral- and/or host-specific factors, and not simply immune status, must be at play to account for the very large discrepancy between viral prevalence and low disease incidence. Here, we show that several amino acids on the surface of the JC virus capsid protein VP1 display accelerated evolution in viral sequences isolated from PML patients but not in sequences isolated from healthy subjects. We provide strong evidence that at least some of these mutations are involved in binding of sialic acid, a known receptor for the JC virus. Using statistical methods of molecular evolution, we performed a comprehensive analysis of JC virus VP1 sequences isolated from 55 PML patients and 253 sequences isolated from the urine of healthy individuals and found that a subset of amino acids found exclusively among PML VP1 sequences is acquired via adaptive evolution. By modeling of the 3-D structure of the JC virus capsid, we showed that these residues are located within the sialic acid binding site, a JC virus receptor for cell infection. Finally, we go on to demonstrate the involvement of some of these sites in receptor binding by demonstrating a profound reduction in hemagglutination properties of viral-like particles made of the VP1 protein carrying these mutations. Collectively, these results suggest that a more virulent PML causing phenotype of JC virus is acquired via adaptive evolution that changes viral specificity for its cellular receptor(s)

CHMP1A encodes an essential regulator of BMI1-INK4A in cerebellar development

Charged multivesicular body protein 1A (CHMP1A; also known as chromatin-modifying protein 1A) is a member of the ESCRT-III (endosomal sorting complex required for transport-III) complex but is also suggested to localize to the nuclear matrix and regulate chromatin structure. Here, we show that loss-of-function mutations in human CHMP1A cause reduced cerebellar size (pontocerebellar hypoplasia) and reduced cerebral cortical size (microcephaly). CHMP1A-mutant cells show impaired proliferation, with increased expression of INK4A, a negative regulator of stem cell proliferation. Chromatin immunoprecipitation suggests loss of the normal INK4A repression by BMI in these cells. Morpholino-based knockdown of zebrafish chmp1a resulted in brain defects resembling those seen after bmi1a and bmi1b knockdown, which were partially rescued by INK4A ortholog knockdown, further supporting links between CHMP1A and BMI1-mediated regulation of INK4A. Our results suggest that CHMP1A serves as a critical link between cytoplasmic signals and BMI1-mediated chromatin modifications that regulate proliferation of central nervous system progenitor cells

Improvement of Cardiac Function in Mouse Myocardial Infarction after Transplantation of Epigenetically-Modified Bone Marrow Progenitor Cells

OBJECTIVE: To study usefulness of bone marrow progenitor cells (BPCs) epigenetically altered by chromatin modifying agents in mediating heart repair after myocardial infarction in mice. METHODS AND RESULTS: We tested the therapeutic efficacy of bone marrow progenitor cells treated with the clinically-used chromatin modifying agents Trichostatin A (TSA, histone deacetylase inhibitor) and 5Aza-2-deoxycytidine (Aza, DNA methylation inhibitor) in a mouse model of acute myocardial infarction (AMI). Treatment of BPCs with Aza and TSA induced expression of pluripotent genes Oct4, Nanog, Sox2, and thereafter culturing these cells in defined cardiac myocyte-conditioned medium resulted in their differentiation into cardiomyocyte progenitors and subsequently into cardiac myocytes. Their transition was deduced by expression of repertoire of markers: Nkx2.5, GATA4, cardiotroponin T, cardiotroponin I, α-sarcomeric actinin, Mef2c and MHC-α. We observed that the modified BPCs had greater AceH3K9 expression and reduced histone deacetylase1 (HDAC1) and lysine-specific demethylase1 (LSD1) expression compared to untreated BPCs, characteristic of epigenetic changes. Intra-myocardial injection of modified BPCs after AMI in mice significantly improved left ventricular function. These changes were ascribed to differentiation of the injected cells into cardiomyocytes and endothelial cells. CONCLUSION: Treatment of BPCs with Aza and TSA converts BPCs into multipotent cells, which can then be differentiated into myocyte progenitors. Transplantation of these modified progenitor cells into infarcted mouse hearts improved left ventricular function secondary to differentiation of cells in the niche into myocytes and endothelial cells

Synaptic Maturation at Cortical Projections to the Lateral Amygdala in a Mouse Model of Rett Syndrome

Rett syndrome (RTT) is a neuro-developmental disorder caused by loss of function of Mecp2 - methyl-CpG-binding protein 2 - an epigenetic factor controlling DNA transcription. In mice, removal of Mecp2 in the forebrain recapitulates most of behavioral deficits found in global Mecp2 deficient mice, including amygdala-related hyper-anxiety and lack of social interaction, pointing a role of Mecp2 in emotional learning. Yet very little is known about the establishment and maintenance of synaptic function in the adult amygdala and the role of Mecp2 in these processes. Here, we performed a longitudinal examination of synaptic properties at excitatory projections to principal cells of the lateral nucleus of the amygdala (LA) in Mecp2 mutant mice and their wild-type littermates. We first show that during animal life, Cortico-LA projections switch from a tonic to a phasic mode, whereas Thalamo-LA synapses are phasic at all ages. In parallel, we observed a specific elimination of Cortico-LA synapses and a decrease in their ability of generating presynaptic long term potentiation. In absence of Mecp2, both synaptic maturation and synaptic elimination were exaggerated albeit still specific to cortical projections. Surprisingly, associative LTP was unaffected at Mecp2 deficient synapses suggesting that synaptic maintenance rather than activity-dependent synaptic learning may be causal in RTT physiopathology. Finally, because the timing of synaptic evolution was preserved, we propose that some of the developmental effects of Mecp2 may be exerted within an endogenous program and restricted to synapses which maturate during animal life

OzDES Reverberation Mapping Programme: Mg II lags and R−L relation

The correlation between the broad line region radius and continuum luminosity (R-L relation) of active galactic nuclei (AGNs) is critical for single-epoch mass estimates of supermassive black holes (SMBHs). At z ∼ 1-2, where AGN activity peaks, the R-L relation is constrained by the reverberation mapping (RM) lags of the Mg II line. We present 25 Mg II lags from the Australian Dark Energy Survey RM project based on 6 yr of monitoring. We define quantitative criteria to select good lag measurements and verify their reliability with simulations based on both the damped random walk stochastic model and the rescaled, resampled versions of the observed light curves of local, well-measured AGN. Our sample significantly increases the number of Mg II lags and extends the R-L relation to higher redshifts and luminosities. The relative iron line strength RFe has little impact on the R-L relation. The best-fitting Mg II R-L relation has a slope α = 0.39 ± 0.08 with an intrinsic scatter σrl = 0.15+−000203. The slope is consistent with previous measurements and shallower than the H β R-L relation. The intrinsic scatter of the new R-L relation is substantially smaller than previous studies and comparable to the intrinsic scatter of the H β R-L relation. Our new R-L relation will enable more precise single-epoch mass estimates and SMBH demographic studies at cosmic noon