Foxk proteins repress the initiation of starvation-induced atrophy and autophagy programs

Abstract: Autophagy is the primary catabolic process triggered in response to starvation.

Abstract: Although autophagic regulation within the cytosolic compartment is well established, it is becoming clear that nuclear events also regulate the induction or repression of autophagy.

Abstract: Nevertheless, a thorough understanding of the mechanisms by which sequence-specific transcription factors modulate expression of genes required for autophagy is lacking.

Abstract: Here, we identify Foxk proteins (Foxk1 and Foxk2) as transcriptional repressors of autophagy in muscle cells and fibroblasts.

Abstract: Interestingly, Foxk1/2 serve to counter-balance another forkhead transcription factor, Foxo3, which induces an overlapping set of autophagic and atrophic targets in muscle.

Abstract: Foxk1/2 specifically recruits Sin3A-HDAC complexes to restrict acetylation of histone H4 and expression of critical autophagy genes.

Abstract: Remarkably, mTOR promotes the transcriptional activity of Foxk1 by facilitating nuclear entry to specifically limit basal levels of autophagy in nutrient-rich conditions.

Abstract: Our study highlights an ancient, conserved mechanism whereby nutritional status is interpreted by mTOR to restrict autophagy by repressing essential autophagy genes through Foxk-Sin3-mediated transcriptional control.


PRRSV receptors and their roles in virus infection

Abstract: Porcine reproductive and respiratory syndrome virus (PRRSV) has a restricted cell tropism and prefers to invade well-differentiated cells of the monocyte/macrophage lineage, such as pulmonary alveolar macrophages and African green monkey kidney cell line MA-104 and its derivatives, such as Marc-145, Vero and CL-2621.

Abstract: PRRSV infection of the host cells actually is a receptor-mediated endocytosis and replication process.

Abstract: The presence and absence of the cellular receptors decide whether the cell lines are permissive or non-permissive to PRRSV infection.

Abstract: Several PRRSV non-permissive cell lines, such as BHK-21, PK-15 and CHO-K1, have been shown to become sensitive to the virus infection upon expression of the recombinant receptor proteins.

Abstract: Up to now, heparin sulfate, sialoadhesin, CD163, CD151 and vimentin have been identified as the important PRRSV receptors via their involvement in virus attachment, internalization or uncoating.

Abstract: Each receptor is characterized by the distribution in different cells, the function in virus different infection stages and the interaction model with the viral proteins or genes.

Abstract: Joint forces of the receptors recently attract attentions due to the specific function.

Abstract: PRRSV receptors have become the targets for designing the new anti-viral reagents or the recombinant cell lines used for isolating the viruses or developing more effective vaccines due to their more conserved sequences compared with the genetic variation of the virus.

Abstract: In this paper, the role of PRRSV receptors and the molecular mechanism of the interaction between the virus and the receptors are reviewed.


Kinetic Analysis Reveals the Fate of a MicroRNA following Target Regulation in Mammalian Cells

Abstract: Considerable details about microRNA (miRNA) biogenesis and regulation have been uncovered, but little is known about the fate of the miRNA subsequent to target regulation.

Abstract: To gain insight into this process, we carried out kinetic analysis of a miRNA's turnover following termination of its biogenesis and during regulation of a target that is not subject to Ago2-mediated catalytic cleavage.

Abstract: By quantitating the number of molecules of the miRNA and its target in steady state and in the course of its decay, we found that each miRNA molecule was able to regulate at least two target transcripts, providing in vivo evidence that the miRNA is not irreversibly sequestered with its target and that the nonslicing pathway of miRNA regulation is multiple-turnover.

Abstract: Using deep sequencing, we further show that miRNA recycling is limited by target regulation, which promotes posttranscriptional modifications to the 3' end of the miRNA and accelerates the miRNA's rate of decay.

Abstract: These studies provide new insight into the efficiency of miRNA regulation that help to explain how a miRNA can regulate a vast number of transcripts and that identify one of the mechanisms that impart specificity to miRNA decay in mammalian cells.


Estimation of nuclear DNA content in plants using flow cytometry

Abstract: Flow cytometry (FCM) using DNA-selective fluorochromes is now the prevailing method for the measurement of nuclear DNA content in plants.

Abstract: Ease of sample preparation and high sample throughput make it generally better suited than other methods such as Feulgen densitometry to estimate genome size, level of generative polyploidy, nuclear replication state and endopolyploidy (polysomaty).

Abstract: Here we present four protocols for sample preparation (suspensions of intact cell nuclei) and describe the analysis of nuclear DNA amounts using FCM.

Abstract: We consider the chemicals and equipment necessary, the measurement process, data analysis, and describe the most frequent problems encountered with plant material such as the interference of secondary metabolites.

Abstract: The purpose and requirement of internal and external standardization are discussed.

Abstract: The importance of using a correct terminology for DNA amounts and genome size is underlined, and its basic principles are explained.


Predictive value of bone resorption and formation markers in cancer patients with bone metastases receiving the bisphosphonate zoledronic acid.

Abstract: PURPOSE Three large, randomized trials of patients with bone metastases recently demonstrated that zoledronic acid reduces the risk of skeletal-related events.

Abstract: These trials provide an opportunity for investigating the correlation between bone metabolism and clinical outcome during bisphosphonate therapy.

Abstract: PATIENTS AND METHODS Urinary measurements of N-telopeptide (Ntx) and serum bone alkaline phosphatase (BAP) were obtained in 1,824 bisphosphonate-treated patients-1,462 with zoledronic acid (breast, 490; prostate, 411; myeloma, 210; non-small-cell lung, 183; other, 168) and 362 with pamidronate (breast, 254; myeloma, 108).

Abstract: This exploratory cohort analysis grouped patients by baseline and most recent levels of Ntx as low (< 50 nmol/mmol creatinine), moderate (50 to 99 nmol/mmol creatinine), or high (> or = 100 nmol/mmol creatinine), and BAP as low (< 146 U/L) or high (> or = 146 U/L).

Abstract: The relative risks for negative clinical outcomes were estimated for each group using multiple-event and Cox regression models with time-varying covariates.

Abstract: RESULTS Patients with high and moderate Ntx levels had 2-fold increases in their risk of skeletal complications and disease progression compared with patients with low Ntx levels (P < .001 for all).

Abstract: High Ntx levels in each solid tumor category were associated with a 4- to 6-fold increased risk of death on study, and moderate Ntx levels a 2- to 4-fold increased risk compared with low Ntx levels (P < .001 for all).

Abstract: Bone alkaline phosphatase also showed some correlation with risk of negative clinical outcomes.

Abstract: CONCLUSION The bone resorption marker Ntx provides valuable prognostic information in patients with bone metastases receiving bisphosphonates.


Role of CD28-B7 interactions in generation and maintenance of CD8 T cell memory.

Abstract: Although the role of CD28-B7 interaction in the activation of naive T cells is well established, its importance in the generation and maintenance of T cell memory is not well understood.

Abstract: In this study, we examined the requirement for CD28-B7 interactions in primary T cell activation and immune memory.

Abstract: Ag-specific CD8 T cell responses were compared between wild-type (+/+) and CD28-deficient (CD28(-/-)) mice following an acute infection with lymphocytic choriomeningitis virus (LCMV).

Abstract: During the primary response, there was a substantial activation and expansion of LCMV-specific CD8 T cells in both +/+ and CD28(-/-) mice.

Abstract: However, the magnitude of the primary CD8 T cell response to both dominant and subdominant LCMV CTL epitopes was approximately 2- to 3-fold lower in CD28(-/-) mice compared with +/+ mice; the lack of CD28-mediated costimulation did not lead to preferential suppression of CD8 T cell responses to the weaker subdominant epitopes.

Abstract: As seen in CD28(-/-) mice, blockade of B7-mediated costimulation by CTLA4-Ig treatment of +/+ mice also resulted in a 2-fold reduction in the anti-LCMV CD8 T cell responses.

Abstract: Loss of CD28/B7 interactions did not significantly affect the generation and maintenance of CD8 T cell memory; the magnitude of CD8 T cell memory was approximately 2-fold lower in CD28(-/-) mice as compared with +/+ mice.

Abstract: Further, in CD28(-/-) mice, LCMV-specific memory CD8 T cells showed normal homeostatic proliferation in vivo and also conferred protective immunity.

Abstract: Therefore, CD28 signaling is not necessary for the proliferative renewal and maintenance of memory CD8 T cells.


Systematic reviews from astronomy to zoology: myths and misconceptions.

Abstract: Systematic literature reviews are widely used as an aid to evidence based decision making.

Abstract: For example, reviews of randomised controlled trials are regularly used to answer questions about the effectiveness of healthcare interventions.

Abstract: The high profile of systematic reviews as a cornerstone of evidence based medicine, however, has led to several misconceptions about their purpose and methods.

Abstract: Among these is the belief that systematic reviews are applicable only to randomised controlled trials and that they are incapable of dealing with other forms of evidence, such as from non-randomised studies or qualitative research.

Abstract: The systematic literature review is a method of locating, appraising, and synthesising evidence.

Abstract: The value of regularly updated systematic reviews in the assessment of effectiveness of healthcare interventions was dramatically illustrated by Antman and colleagues, who showed that review articles failed to mention advances in treatment identified by an updated systematic review.1  It is nearly a quarter of a century since Gene Glass coined the term “meta-analysis” to refer to the quantitative synthesis of the results of primary studies.2 The importance of making explicit efforts to limit bias in the review of literature, however, has been emphasised by social scientists at least since the 1960s.3 In recent years systematic reviews have found an important role in health services research, and the growing interest in evidence based approaches to decision making makes it likely that their use will increase.

Abstract: Not everybody accepts that systematic reviews are necessary or desirable, and as one moves further away from the clinical applications of systematic reviews cynicism about their utility grows.

Abstract: Several arguments are commonly used to reject a wider role for systematic reviews, and these arguments are often based on major misconceptions about the history, purpose, methods, and uses of systematic reviews.

Abstract: I have examined eight common myths about …


Acellular pertussis vaccines protect against disease but fail to prevent infection and transmission in a nonhuman primate model.

Abstract: Pertussis is a highly contagious respiratory illness caused by the bacterial pathogen Bordetella pertussis.

Abstract: Pertussis rates in the United States have been rising and reached a 50-y high of 42,000 cases in 2012.

Abstract: Although pertussis resurgence is not completely understood, we hypothesize that current acellular pertussis (aP) vaccines fail to prevent colonization and transmission.

Abstract: To test our hypothesis, infant baboons were vaccinated at 2, 4, and 6 mo of age with aP or whole-cell pertussis (wP) vaccines and challenged with B. pertussis at 7 mo.

Abstract: Infection was followed by quantifying colonization in nasopharyngeal washes and monitoring leukocytosis and symptoms.

Abstract: Baboons vaccinated with aP were protected from severe pertussis-associated symptoms but not from colonization, did not clear the infection faster than naïve animals, and readily transmitted B. pertussis to unvaccinated contacts.

Abstract: Vaccination with wP induced a more rapid clearance compared with naïve and aP-vaccinated animals.

Abstract: By comparison, previously infected animals were not colonized upon secondary infection.

Abstract: Although all vaccinated and previously infected animals had robust serum antibody responses, we found key differences in T-cell immunity.

Abstract: Previously infected animals and wP-vaccinated animals possess strong B. pertussis-specific T helper 17 (Th17) memory and Th1 memory, whereas aP vaccination induced a Th1/Th2 response instead.

Abstract: The observation that aP, which induces an immune response mismatched to that induced by natural infection, fails to prevent colonization or transmission provides a plausible explanation for the resurgence of pertussis and suggests that optimal control of pertussis will require the development of improved vaccines.


Specific and cooperative binding of E. coli single-stranded DNA binding protein to mRNA.

Abstract: Fluorometric titration of E. coli single-stranded DNA binding protein with various RNAs showed that the protein specifically and cooperatively binds to its own mRNA.

Abstract: The binding inhibited in vitro expression of ssb and bla but not nusA. This inhibition takes place at a physiological concentration of SSB.

Abstract: The function of the protein in gene regulation is discussed.


ACAM2000™: The new smallpox vaccine for United States Strategic National Stockpile

Abstract: Smallpox was eradicated more than 30 years ago, but heightened concerns over bioterrorism have brought smallpox and smallpox vaccination back to the forefront.

Abstract: The previously licensed smallpox vaccine in the United States, Dryvax (Wyeth Laboratories, Inc.), was highly effective, but the supply was insufficient to vaccinate the entire current US population.

Abstract: Additionally, Dryvax had a questionable safety profile since it consisted of a pool of vaccinia virus strains with varying degrees of virulence, and was grown on the skin of calves, an outdated technique that poses an unnecessary risk of contamination.

Abstract: The US government has therefore recently supported development of an improved live vaccinia virus smallpox vaccine.

Abstract: This initiative has resulted in the development of ACAM2000 (Acambis, Inc.), a single plaque-purified vaccinia virus derivative of Dryvax, aseptically propagated in cell culture.

Abstract: Preclinical and clinical trials reported in 2008 demonstrated that ACAM2000 has comparable immunogenicity to that of Dryvax, and causes a similar frequency of adverse events.

Abstract: Furthermore, like Dryvax, ACAM2000 vaccination has been shown by careful cardiac screening to result in an unexpectedly high rate of myocarditis and pericarditis.

Abstract: ACAM2000 received US Food and Drug Administration (FDA) approval in August 2007, and replaced Dryvax for all smallpox vaccinations in February 2008.

Abstract: Currently, over 200 million doses of ACAM2000 have been produced for the US Strategic National Stockpile.

Abstract: This review of ACAM2000 addresses the production, characterization, clinical trials, and adverse events associated with this new smallpox vaccine.


Risk of malignant neoplasms in patients with pulmonary sarcoidosis.

Abstract: BACKGROUND For over 20 years the association between sarcoidosis and malignancy, particularly lymphoma and lung cancer, has been disputed with misclassification being the major concern.

Abstract: The aim of the present study was to analyse the incidence of malignancies in a cohort of patients with sarcoidosis by linkage to a nationwide population based cancer register.

Abstract: METHODS The cohort comprised 254 patients followed for a median of 25 years until death, emigration, or 31 December 1992, whichever came first.

Abstract: The expected number of cancer cases was calculated using the annual age and sex specific cancer rates from the Danish Cancer Registry.

Abstract: RESULTS Thirty six cancers were registered, three of which were misclassified as sarcoidosis, leaving 33 cancers compared with 23 expected (standardised incidence ratio (SIR) = 1.4; 95% CI 0.99 to 2.0).

Abstract: Five lung cancers were observed compared with 2.5 expected, yielding an SIR of 2.0 (95% CI 0.7 to 4.7).

Abstract: There was no incidence of lymphoma and only one case of leukaemia.

Abstract: There was a significant excess number of pharyngeal cancers based on two cases (SIR = 15.4; 95% CI 1.7 to 56).

Abstract: CONCLUSIONS This study does not support the theory of an association between sarcoidosis and malignancy, and the main reason other studies have shown such an association is most likely to have been due to selection bias and misclassification.


Rps14 haploinsufficiency causes a block in erythroid differentiation mediated by S100A8 and S100A9

Abstract: Impaired erythropoiesis in the deletion 5q (del(5q)) subtype of myelodysplastic syndrome (MDS) has been linked to heterozygous deletion of RPS14, which encodes the ribosomal protein small subunit 14.

Abstract: We generated mice with conditional inactivation of Rps14 and demonstrated an erythroid differentiation defect that is dependent on the tumor suppressor protein p53 (encoded by Trp53 in mice) and is characterized by apoptosis at the transition from polychromatic to orthochromatic erythroblasts.

Abstract: This defect resulted in age-dependent progressive anemia, megakaryocyte dysplasia and loss of hematopoietic stem cell (HSC) quiescence.

Abstract: As assessed by quantitative proteomics, mutant erythroblasts expressed higher levels of proteins involved in innate immune signaling, notably the heterodimeric S100 calcium-binding proteins S100a8 and S100a9.

Abstract: S100a8—whose expression was increased in mutant erythroblasts, monocytes and macrophages—is functionally involved in the erythroid defect caused by the Rps14 deletion, as addition of recombinant S100a8 was sufficient to induce a differentiation defect in wild-type erythroid cells, and genetic inactivation of S100a8 expression rescued the erythroid differentiation defect of Rps14-haploinsufficient HSCs.

Abstract: Our data link Rps14 haploinsufficiency in del(5q) MDS to activation of the innate immune system and induction of S100A8-S100A9 expression, leading to a p53-dependent erythroid differentiation defect.


A Microtubule Interactome: Complexes with Roles in Cell Cycle and Mitosis

Abstract: The microtubule (MT) cytoskeleton is required for many aspects of cell function, including the transport of intracellular materials, the maintenance of cell polarity, and the regulation of mitosis.

Abstract: These functions are coordinated by MT-associated proteins (MAPs), which work in concert with each other, binding MTs and altering their properties.

Abstract: We have used a MT cosedimentation assay, combined with 1D and 2D PAGE and mass spectrometry, to identify over 250 MAPs from early Drosophila embryos.

Abstract: We have taken two complementary approaches to analyse the cellular function of novel MAPs isolated using this approach.

Abstract: First, we have carried out an RNA interference (RNAi) screen, identifying 21 previously uncharacterised genes involved in MT organisation.

Abstract: Second, we have undertaken a bioinformatics analysis based on binary protein interaction data to produce putative interaction networks of MAPs.

Abstract: By combining both approaches, we have identified and validated MAP complexes with potentially important roles in cell cycle regulation and mitosis.

Abstract: This study therefore demonstrates that biologically relevant data can be harvested using such a multidisciplinary approach, and identifies new MAPs, many of which appear to be important in cell division.


Structure of the Guanine Nucleotide Exchange Factor Sec7 Domain of Human Arno and Analysis of the Interaction with ARF GTPase

Abstract: Sec7-related guanine nucleotide exchange factors (GEFs) initiate vesicle budding from the Golgi membrane surface by converting the GTPase ARF to a GTP-bound, membrane-associated form.

Abstract: Here we report the crystal structure of the catalytic Sec7 homology domain of Arno, a human GEF for ARF1, determined at 2.2 angstroms resolution.

Abstract: The Sec7 domain is an elongated, all-helical protein with a distinctive hydrophobic groove that is phylogenetically conserved.

Abstract: Structure-based mutagenesis identifies the groove and an adjacent conserved loop as the ARF-interacting surface.

Abstract: The sites of Sec7 domain interaction on ARF1 have subsequently been mapped, by protein footprinting experiments, to the switch 1 and switch 2 GTPase regions, leading to a model for the interaction between ARF GTPases and Sec7 domain exchange factors.


Brief Communication Repeated Cocaine Administration Increases Voltage-Sensitive Calcium Currents in Response to Membrane Depolarization in Medial Prefrontal Cortex Pyramidal Neurons

Abstract: The medial prefrontal cortex (mPFC) plays a critical role in cocaine addiction.

Abstract: However, evidence to elucidate how the mPFC is functionally involved in cocaine addiction remains incomplete.

Abstract: Recent studies have revealed that repeated cocaine administration induces various neuroadaptations in pyramidal mPFC neurons, including a reduction in voltage-gated K+ currents (VGKCs) and a possible increase in voltage-sensitive Ca2+ currents (I(Ca)).

Abstract: Here, we performed both current-clamp recordings in brain slices and voltage-clamp recordings in freshly dissociated cells to determine whether I(Ca) is altered in mPFC pyramidal neurons after chronic cocaine treatment with a short-term or long-term withdrawal.

Abstract: In addition, a critical role of VGKCs in regulating the generation of Ca2+ plateau potential was also studied in mPFC neurons.

Abstract: Repeated cocaine administration significantly prolonged the duration of evoked Ca2+ plateau potentials and increased the whole-cell I(Ca) in mPFC neurons after a 3 d withdrawal.

Abstract: Selective blockade of L-type Ca2+ channels by nifedipine not only significantly increased the threshold but also reduced the duration and amplitude of Ca2+ plateau potentials in both saline- and cocaine-withdrawn mPFC neurons.

Abstract: However, there was no significant difference in the increased threshold, reduced duration, and decreased amplitude of Ca2+ potentials between saline- and cocaine-withdrawn neurons after blockade of L-type Ca2+ channels.

Abstract: Moreover, an increase in amplitude was also observed, whereas the prolonged duration persisted, in Ca2+ potentials after 2-3 weeks of withdrawal.

Abstract: These findings indicate that chronic exposure to cocaine facilitates the responsiveness of I(Ca), particularly via the activated L-type Ca2+ channels, to excitatory stimuli in rat mPFC pyramidal neurons.


Rational Extension of the Ribosome Biogenesis Pathway Using Network-Guided Genetics

Abstract: Biogenesis of ribosomes is an essential cellular process conserved across all eukaryotes and is known to require >170 genes for the assembly, modification, and trafficking of ribosome components through multiple cellular compartments.

Abstract: Despite intensive study, this pathway likely involves many additional genes.

Abstract: Here, we employ network-guided genetics-an approach for associating candidate genes with biological processes that capitalizes on recent advances in functional genomic and proteomic studies-to computationally identify additional ribosomal biogenesis genes.

Abstract: We experimentally evaluated >100 candidate yeast genes in a battery of assays, confirming involvement of at least 15 new genes, including previously uncharacterized genes (YDL063C, YIL091C, YOR287C, YOR006C/TSR3, YOL022C/TSR4).

Abstract: We associate the new genes with specific aspects of ribosomal subunit maturation, ribosomal particle association, and ribosomal subunit nuclear export, and we identify genes specifically required for the processing of 5S, 7S, 20S, 27S, and 35S rRNAs.

Abstract: These results reveal new connections between ribosome biogenesis and mRNA splicing and add >10% new genes-most with human orthologs-to the biogenesis pathway, significantly extending our understanding of a universally conserved eukaryotic process.


Quantifying Transmission Investment in Malaria Parasites

Abstract: Many microparasites infect new hosts with specialized life stages, requiring a subset of the parasite population to forgo proliferation and develop into transmission forms.

Abstract: Transmission stage production influences infectivity, host exploitation, and the impact of medical interventions like drug treatment.

Abstract: Predicting how parasites will respond to public health efforts on both epidemiological and evolutionary timescales requires understanding transmission strategies.

Abstract: These strategies can rarely be observed directly and must typically be inferred from infection dynamics.

Abstract: Using malaria as a case study, we test previously described methods for inferring transmission stage investment against simulated data generated with a model of within-host infection dynamics, where the true transmission investment is known.

Abstract: We show that existing methods are inadequate and potentially very misleading.

Abstract: The key difficulty lies in separating transmission stages produced by different generations of parasites.

Abstract: We develop a new approach that performs much better on simulated data.

Abstract: Applying this approach to real data from mice infected with a single Plasmodium chabaudi strain, we estimate that transmission investment varies from zero to 20%, with evidence for variable investment over time in some hosts, but not others.

Abstract: These patterns suggest that, even in experimental infections where host genetics and other environmental factors are controlled, parasites may exhibit remarkably different patterns of transmission investment.


Roles of induced expression of MAPK phosphatase-2 in tumor development in RET-MEN2A transgenic mice

Abstract: Germline mutations in the RET tyrosine kinase gene are responsible for the development of multiple endocrine neoplasia 2A and 2B (MEN2A and MEN2B).

Abstract: However, knowledge of the fundamental principles that determine the mutant RET-mediated signaling remains elusive.

Abstract: Here, we report increased expression of mitogen-activated protein kinase phosphatase-2 (MKP-2) in carcinomas developed in transgenic mice carrying RET with the MEN2A mutation (RET-MEN2A).

Abstract: The expression of MKP-2 was not only induced by RET-MEN2A or RET-MEN2B mutant proteins but also by the activation of endogenous RET by its ligand, glial cell line-derived neurotrophic factor (GDNF).

Abstract: MKP-2 expression was also evident in the MKK-f cell line, which was established from a mammary tumor developed in a RET-MEN2A transgenic mouse.

Abstract: Inhibition of MKP-2 attenuated the in vitro and in vivo proliferation of MKK-f cells, which was mediated by the suppression of cyclin B1 expression.

Abstract: Furthermore, we found that MKP-2 is highly expressed in medullary thyroid carcinomas derived from MEN2A patients.

Abstract: These findings suggest that the increased expression of MKP-2 may play a crucial role in oncogenic signaling downstream of mutant RET, leading to deregulation of cell cycle.


Effects of repeated social stress on leukocyte distribution in bone marrow, peripheral blood and spleen

Abstract: Leukocyte trafficking between the various body compartments has an important surveillance function that ensures the detection of antigen and enables the immune system to initiate a rapid and effective response.

Abstract: Repeated social defeat of group-housed male mice induced by daily, acute encounters with an aggressive conspecific substantially altered leukocyte trafficking and led to a gradual redistribution of immune cells in bone marrow, peripheral blood and spleen.

Abstract: Recurrent exposure to the stressor over a period of 2, 4 or 6 consecutive days was associated with cell mobilization and increased myelopoiesis in the bone marrow that was paralleled by an accumulation of neutrophils and monocytes in circulation and spleen.

Abstract: Substantial depletion of B cells in bone marrow and blood was associated with an increase in splenic B cells indicating a redirection of this cell type to the spleen.

Abstract: In contrast, T cells were markedly reduced in these immune compartments.

Abstract: The recruitment of CD11b+ leukocytes (i.e., monocytes/macrophages and neutrophils) from the bone marrow to the spleen might play a critical role in the development of functional glucocorticoid resistance in the murine spleen that was reported in context with repeated social defeat.


Lentivirally transduced dendritic cells as a tool for cancer immunotherapy.

Abstract: BACKGROUND Dendritic cells (DC) are the professional antigen-presenting cells of the immune system, fully equipped to prime naive T cells and thus essential components for cancer immunotherapy.

Abstract: METHODS We tested the influence of several elements (cPPT, trip, WPRE, SIN) on the transduction efficiency of human DC.

Abstract: Human and murine DC were transduced with tNGFR-encoding lentiviruses to assess the effect of transduction on phenotype and function.

Abstract: Human DC were transduced with lentiviruses encoding huIi80MAGE-A3 and murine DC with huIi80tOVA to test antigen presentation.

Abstract: RESULTS A self-inactivating (SIN) lentiviral vector containing the trip element was most efficient in transducing human DC.

Abstract: The transduction of DC with trip/SIN tNGFR encoding lentiviral vectors at MOI 15 resulted in stable gene expression in up to 94.6% (murine) and 88.2% (human) of the mature DC, without perturbing viability, phenotype and function.

Abstract: Human huIi80MAGE-A3-transduced DC were able to stimulate MAGE-A3-specific CD4(+) and CD8(+) T cell clones and could prime both MAGE-A3-specific CD4(+) and CD8(+) T cells in vitro.

Abstract: Murine huIi80tOVA-transduced DC were able to present OVA peptides in the context of MHC class I and class II in vitro and induced a strong OVA-specific cytotoxic T lymphocyte response in vivo, that was protective against subsequent challenge with OVA-expressing tumor cells.

Abstract: CONCLUSIONS We show that, using lentiviral vectors, efficient gene transfer in human and murine DC can be obtained and that these DC can elicit antigen-specific immune responses in vitro and in vivo.

Abstract: The composition of the transfer vector has a major impact on the transduction efficiency.


Global Reorganization of Replication Domains During Embryonic Stem Cell Differentiation

Abstract: DNA replication in mammals is regulated via the coordinate firing of clusters of replicons that duplicate megabase-sized chromosome segments at specific times during S-phase.

Abstract: Cytogenetic studies show that these "replicon clusters" coalesce as subchromosomal units that persist through multiple cell generations, but the molecular boundaries of such units have remained elusive.

Abstract: Moreover, the extent to which changes in replication timing occur during differentiation and their relationship to transcription changes has not been rigorously investigated.

Abstract: We have constructed high-resolution replication-timing profiles in mouse embryonic stem cells (mESCs) before and after differentiation to neural precursor cells.

Abstract: We demonstrate that chromosomes can be segmented into multimegabase domains of coordinate replication, which we call "replication domains," separated by transition regions whose replication kinetics are consistent with large originless segments.

Abstract: The molecular boundaries of replication domains are remarkably well conserved between distantly related ESC lines and induced pluripotent stem cells.

Abstract: Unexpectedly, ESC differentiation was accompanied by the consolidation of smaller differentially replicating domains into larger coordinately replicated units whose replication time was more aligned to isochore GC content and the density of LINE-1 transposable elements, but not gene density.

Abstract: Replication-timing changes were coordinated with transcription changes for weak promoters more than strong promoters, and were accompanied by rearrangements in subnuclear position.

Abstract: We conclude that replication profiles are cell-type specific, and changes in these profiles reveal chromosome segments that undergo large changes in organization during differentiation.

Abstract: Moreover, smaller replication domains and a higher density of timing transition regions that interrupt isochore replication timing define a novel characteristic of the pluripotent state.


Molecular characterization of the yeast meiotic regulatory gene RIM1.

Abstract: In the yeast Saccharomyces cerevisiae, genetic studies suggest that the RIM1 gene encodes a positive regulator of meiosis.

Abstract: rim1 mutations cause reduced expression of IME1, which is required for expression of many meiotic genes, and thus lead to a partial defect in meiosis and spore formation.

Abstract: We report the sequence of RIM1 and functional analysis of its coding region.

Abstract: The RIM1 gene product (RIM1) contains three regions similar to C2H2 zinc fingers.

Abstract: Serine substitutions for cysteine in each of the putative zinc fingers abolish RIM1 function.

Abstract: The carboxyl-terminus of RIM1 is enriched in acidic amino acids and is required for full RIM1 activity.

Abstract: RIM1 also contains two putative cAMP-dependent protein kinase (cAPK) phosphorylation sites.

Abstract: At one site, substitution of alanine for serine does not affect RIM1 activity; at the other site, this substitution impairs activity.

Abstract: This analysis of RIM1 suggests that the protein may function as a transcriptional activator.

Abstract: We have used the cloned RIM1 gene to create a complete rim1 deletion.

Abstract: This null allele, like previously isolated rim1 mutations, causes a partial meiotic defect.

Abstract: In addition to RIM1, maximum IME1 expression requires the MCK1 and IME4 gene products.

Abstract: Defects associated with rim1, mck1, and ime4 mutations in expression of a meiotic reporter gene (ime2-lacZ) and in sporulation are additive.

Abstract: These findings suggest that RIM1 acts independently of MCK1 and IME4 to stimulate IME1 expression.


Butyrate greatly enhances derivation of human induced pluripotent stem cells by promoting epigenetic remodeling and the expression of pluripotency-associated genes.

Abstract: We report here that butyrate, a naturally occurring fatty acid commonly used as a nutritional supplement and differentiation agent, greatly enhances the efficiency of induced pluripotent stem (iPS) cell derivation from human adult or fetal fibroblasts.

Abstract: After transient butyrate treatment, the iPS cell derivation efficiency is enhanced by 15- to 51-fold using either retroviral or piggyBac transposon vectors expressing 4 to 5 reprogramming genes.

Abstract: Butyrate stimulation is more remarkable (>100- to 200-fold) on reprogramming in the absence of either KLF4 or MYC transgene.

Abstract: Butyrate treatment did not negatively affect properties of iPS cell lines established by either 3 or 4 retroviral vectors or a single piggyBac DNA transposon vector.

Abstract: These characterized iPS cell lines, including those derived from an adult patient with sickle cell disease by either the piggyBac or retroviral vectors, show normal karyotypes and pluripotency.

Abstract: To gain insights into the underlying mechanisms of butyrate stimulation, we conducted genome-wide gene expression and promoter DNA methylation microarrays and other epigenetic analyses on established iPS cells and cells from intermediate stages of the reprogramming process.

Abstract: By days 6 to 12 during reprogramming, butyrate treatment enhanced histone H3 acetylation, promoter DNA demethylation, and the expression of endogenous pluripotency-associated genes, including DPPA2, whose overexpression partially substitutes for butyrate stimulation.

Abstract: Thus, butyrate as a cell permeable small molecule provides a simple tool to further investigate molecular mechanisms of cellular reprogramming.

Abstract: Moreover, butyrate stimulation provides an efficient method for reprogramming various human adult somatic cells, including cells from patients that are more refractory to reprogramming.


Nanoparticle conjugation of CpG enhances adjuvancy for cellular immunity and memory recall at low dose.

Abstract: In subunit vaccines, strong CD8(+) T-cell responses are desired, yet they are elusive at reasonable adjuvant doses.

Abstract: We show that targeting adjuvant to the lymph node (LN) via ultrasmall polymeric nanoparticles (NPs), which rapidly drain to the LN after intradermal injection, greatly enhances adjuvant efficacy at low doses.

Abstract: Coupling CpG-B or CpG-C oligonucleotides to NPs led to better dual-targeting of adjuvant and antigen (codelivered on separate NPs) in cross-presenting dendritic cells compared with free adjuvant.

Abstract: This led to enhanced dendritic cell maturation and T helper 1 (Th1)-cytokine secretion, in turn driving stronger effector CD8(+) T-cell activation with enhanced cytolytic profiles and, importantly, more powerful memory recall.

Abstract: With only 4 μg CpG, NP-CpG-B could substantially protect mice from syngeneic tumor challenge, even after 4 mo of vaccination, compared with free CpG-B. Together, these results show that nanocarriers can enhance vaccine efficacy at a low adjuvant dose for inducing potent and long-lived cellular immunity.


Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds

Abstract: Reprogramming of mouse and human somatic cells can be achieved by ectopic expression of transcription factors, but with low efficiencies.

Abstract: We report that DNA methyltransferase and histone deacetylase (HDAC) inhibitors improve reprogramming efficiency.

Abstract: In particular, valproic acid (VPA), an HDAC inhibitor, improves reprogramming efficiency by more than 100-fold, using Oct4-GFP as a reporter.

Abstract: VPA also enables efficient induction of pluripotent stem cells without introduction of the oncogene c-Myc.


Noninvasive ventilation for treatment of acute respiratory failure in patients undergoing solid organ transplantation: a randomized trial.

Abstract: CONTEXT Noninvasive ventilation (NIV) has been associated with lower rates of endotracheal intubation in populations of patients with acute respiratory failure.

Abstract: OBJECTIVE To compare NIV with standard treatment using supplemental oxygen administration to avoid endotracheal intubation in recipients of solid organ transplantation with acute hypoxemic respiratory failure.

Abstract: DESIGN AND SETTING Prospective randomized study conducted at a 14-bed, general intensive care unit of a university hospital.

Abstract: PATIENTS Of 238 patients who underwent solid organ transplantation from December 1995 to October 1997, 51 were treated for acute respiratory failure.

Abstract: Of these, 40 were eligible and 20 were randomized to each group.

Abstract: INTERVENTION Noninvasive ventilation vs standard treatment with supplemental oxygen administration.

Abstract: MAIN OUTCOME MEASURES The need for endotracheal intubation and mechanical ventilation at any time during the study, complications not present on admission, duration of ventilatory assistance, length of hospital stay, and intensive care unit mortality.

Abstract: RESULTS The 2 groups were similar at study entry.

Abstract: Within the first hour of treatment, 14 patients (70%) in the NIV group, and 5 patients (25%) in the standard treatment group improved their ratio of the PaO2 to the fraction of inspired oxygen (FIO2).

Abstract: Over time, a sustained improvement in PaO2 to FIO2 was noted in 12 patients (60%) in the NIV group, and in 5 patients (25%) randomized to standard treatment (P = .03).

Abstract: The use of NIV was associated with a significant reduction in the rate of endotracheal intubation (20% vs 70%; P = .002), rate of fatal complications (20% vs 50%; P = .05), length of stay in the intensive care unit by survivors (mean [SD] days, 5.5 [3] vs 9 [4]; P = .03), and intensive care unit mortality (20% vs 50%; P = .05).

Abstract: Hospital mortality did not differ.

Abstract: CONCLUSIONS These results indicate that transplantation programs should consider NIV in the treatment of selected recipients of transplantation with acute respiratory failure.


BIOINFORMATICS APPLICATIONS NOTE

Abstract: SUMMARY MADE4, microarray ade4, is a software package that facilitates multivariate analysis of microarray gene-expression data.

Abstract: MADE4 accepts a wide variety of gene-expression data formats.

Abstract: MADE4 takes advantage of the extensive multivariate statistical and graphical functions in the R package ade4, extending these for application to microarray data.

Abstract: In addition, MADE4 provides new graphical and visualization tools that aid in interpretation of multivariate analysis of microarray data.


Dual and opposing roles of primary cilia in medulloblastoma development

Abstract: Recent work has shown that primary cilia are essential for Hedgehog (Hh) signaling during mammalian development.

Abstract: It is also known that aberrant Hh signaling can lead to cancer, but the role of primary cilia in oncogenesis is not known.

Abstract: Cerebellar granule neuron precursors (GNPs) can give rise to medulloblastomas, the most common malignant brain tumor in children.

Abstract: The primary cilium and Hh signaling are required for GNP proliferation.

Abstract: We asked whether primary cilia in GNPs have a role in medulloblastoma growth in mice.

Abstract: Genetic ablation of primary cilia blocked medulloblastoma formation when this tumor was driven by a constitutively active Smoothened protein (Smo), an upstream activator of Hh signaling.

Abstract: In contrast, removal of cilia was required for medulloblastoma growth by a constitutively active glioma-associated oncogene family zinc finger-2 (GLI2), a downstream transcription factor.

Abstract: Thus, primary cilia are either required for or inhibit medulloblastoma formation, depending on the initiating oncogenic event.

Abstract: Remarkably, the presence or absence of cilia was associated with specific variants of human medulloblastomas; primary cilia were found in medulloblastomas with activation in HH or WNT signaling but not in most medulloblastomas in other distinct molecular subgroups.

Abstract: Primary cilia could serve as a diagnostic tool and provide new insights into the mechanism of tumorigenesis.


RAS genes in Saccharomyces cerevisiae: signal transduction in search of a pathway.

Abstract: Ras proteins in budding yeasts initially appeared to regulate initiation of the cell cycle in response to nutrient availability.

Abstract: More recent work, while clarifying the mechanism of Ras-mediated signal transduction, has undermined our notion of the signal Ras transmits.

Abstract: We now suspect that Ras helps to coordinate cellular metabolism and mass accumulation, but what Ras responds to is not clear.


Use of synthetic peptides to locate novel integrin alpha2beta1-binding motifs in human collagen III.

Abstract: A set of 57 synthetic peptides encompassing the entire triplehelical domain of human collagen III was used to locate binding sites for the collagen-binding integrin alpha(2)beta(1).

Abstract: The capacity of the peptides to support Mg(2+)-dependent binding of several integrin preparations was examined.

Abstract: Wild-type integrins (recombinant alpha(2) I-domain, alpha(2)beta(1) purified from platelet membranes, and recombinant soluble alpha(2)beta(1) expressed as an alpha(2)-Fos/beta(1)-Jun heterodimer) bound well to only three peptides, two containing GXX'GER motifs (GROGER and GMOGER, where O is hydroxyproline) and one containing two adjacent GXX'GEN motifs (GLKGEN and GLOGEN).

Abstract: Two mutant alpha(2) I-domains were tested: the inactive T221A mutant, which recognized no peptides, and the constitutively active E318W mutant, which bound a larger subset of peptides.

Abstract: Adhesion of activated human platelets to GER-containing peptides was greater than that of resting platelets, and HT1080 cells bound well to more of the peptides compared with platelets.

Abstract: Binding of cells and recombinant proteins was abolished by anti-alpha(2) monoclonal antibody 6F1 and by chelation of Mg(2+).

Abstract: We describe two novel high affinity integrin-binding motifs in human collagen III (GROGER and GLOGEN) and a third motif (GLKGEN) that displays intermediate activity.

Abstract: Each motif was verified using shorter synthetic peptides.


Locations of cerebral infarctions in tuberculous meningitis

Abstract: The locations of cerebral infarctions were studied in 14 patients with tuberculous meningitis (TBM) and 173 patients with noninflammatory ischemic stroke (IS).

Abstract: In patients with TBM, 75% of infarctions occurred in the “TB zone” supplied by medial striate and thalamoperforating arteries; only 11% occurred in the “IS zone” supplied by lateral striate, anterior choroidal and thalamogeniculate arteries.

Abstract: In patients with IS 29% of infarctions occurred in the IS zone, 29% in the subcortical white matter, and 24% in (or involving) the cerebral cortex.

Abstract: Only 11% occurred in the TB zone.

Abstract: Bilaterally symmetrical infarctions of the TB zone were common with TMB (71%) but rare with IS (5%).


TRP channels of intracellular membranes.

Abstract: Ion channels are classically understood to regulate the flux of ions across the plasma membrane in response to a variety of environmental and intracellular cues.

Abstract: Ion channels serve a number of functions in intracellular membranes as well.

Abstract: These channels may be temporarily localized to intracellular membranes as a function of their biosynthetic or secretory pathways, i.e., en route to their destination location.

Abstract: Intracellular membrane ion channels may also be located in the endocytic pathways, either being recycled back to the plasma membrane or targeted to the lysosome for degradation.

Abstract: Several channels do participate in intracellular signal transduction; the most well known example is the inositol 1,4,5-trisphosphate receptor (IP(3)R) in the endoplasmic reticulum.

Abstract: Some organellar intracellular membrane channels are required for the ionic homeostasis of their residing organelles.

Abstract: Several newly-discovered intracellular membrane Ca(2+) channels actually play active roles in membrane trafficking.

Abstract: Transient receptor potential (TRP) proteins are a superfamily (28 members in mammal) of Ca(2+)-permeable channels with diverse tissue distribution, subcellular localization, and physiological functions.

Abstract: Almost all mammalian TRP channels studied thus far, like their ancestor yeast TRP channel (TRPY1) that localizes to the vacuole compartment, are also (in addition to their plasma membrane localization) found to be localized to intracellular membranes.

Abstract: Accumulated evidence suggests that intracellularly-localized TRP channels actively participate in regulating membrane traffic, signal transduction, and vesicular ion homeostasis.

Abstract: This review aims to provide a summary of these recent works.

Abstract: The discussion will also be extended to the basic membrane and electrical properties of the TRP-residing compartments.


the Development of Autoimmune Myocarditis in Mice by an

Abstract: BACKGROUND Interleukin (IL)-12 exerts a potent proinflammatory effect by stimulating T-helper (Th) 1 responses.

Abstract: This effect is believed to be mediated primarily through the activation of STAT4 and subsequent production of interferon (IFN)-gamma.

Abstract: Methods and Results- We examined the role of IL-12 receptor (IL-12R) signaling in the development of murine experimental autoimmune myocarditis (EAM) induced by cardiac myosin immunization.

Abstract: Both IL-12Rbeta1-deficient mice and STAT4-deficient mice were resistant to the induction of myocarditis.

Abstract: Treatment with exogenous IL-12 exacerbated disease.

Abstract: We questioned whether IFN-gamma is required for the disease-promoting activity of IL-12.

Abstract: On the contrary, we found that IFN-gamma suppresses EAM.

Abstract: Lack of IFN-gamma due to either depletion with an antibody or a genetic deficiency exacerbated myocarditis.

Abstract: Spleens from IFN-gamma-deficient mice immunized with cardiac myosin showed increased cellularity; greater numbers of CD3+, CD4+, CD8+, and IL-2-producing cells; and heightened ability to produce cytokines on stimulation in vitro.

Abstract: Treatment of mice with recombinant IFN-gamma suppressed the development of myocarditis.

Abstract: CONCLUSIONS IL-12/IL-12R/STAT4 signaling promotes the development of EAM.

Abstract: In contrast, IFN-gamma plays a protective role.

Abstract: The disease-limiting effects of IFN-gamma might be explained by its ability to control the expansion of activated T lymphocytes.


The Viral and Cellular MicroRNA Targetome in Lymphoblastoid Cell Lines

Abstract: Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus linked to a number of B cell cancers and lymphoproliferative disorders.

Abstract: During latent infection, EBV expresses 25 viral pre-microRNAs (miRNAs) and induces the expression of specific host miRNAs, such as miR-155 and miR-21, which potentially play a role in viral oncogenesis.

Abstract: To date, only a limited number of EBV miRNA targets have been identified; thus, the role of EBV miRNAs in viral pathogenesis and/or lymphomagenesis is not well defined.

Abstract: Here, we used photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) combined with deep sequencing and computational analysis to comprehensively examine the viral and cellular miRNA targetome in EBV strain B95-8-infected lymphoblastoid cell lines (LCLs).

Abstract: We identified 7,827 miRNA-interaction sites in 3,492 cellular 3'UTRs.

Abstract: 531 of these sites contained seed matches to viral miRNAs.

Abstract: 24 PAR-CLIP-identified miRNA:3'UTR interactions were confirmed by reporter assays.

Abstract: Our results reveal that EBV miRNAs predominantly target cellular transcripts during latent infection, thereby manipulating the host environment.

Abstract: Furthermore, targets of EBV miRNAs are involved in multiple cellular processes that are directly relevant to viral infection, including innate immunity, cell survival, and cell proliferation.

Abstract: Finally, we present evidence that myc-regulated host miRNAs from the miR-17/92 cluster can regulate latent viral gene expression.

Abstract: This comprehensive survey of the miRNA targetome in EBV-infected B cells represents a key step towards defining the functions of EBV-encoded miRNAs, and potentially, identifying novel therapeutic targets for EBV-associated malignancies.


Mitochondrial Dysfunction Leads to Nuclear Genome Instability via an Iron-Sulfur Cluster Defect

Abstract: Mutations and deletions in the mitochondrial genome (mtDNA), as well as instability of the nuclear genome, are involved in multiple human diseases.

Abstract: Here, we report that in Saccharomyces cerevisiae, loss of mtDNA leads to nuclear genome instability, through a process of cell-cycle arrest and selection we define as a cellular crisis.

Abstract: This crisis is not mediated by the absence of respiration, but instead correlates with a reduction in the mitochondrial membrane potential.

Abstract: Analysis of cells undergoing this crisis identified a defect in iron-sulfur cluster (ISC) biogenesis, which requires normal mitochondrial function.

Abstract: We found that downregulation of nonmitochondrial ISC protein biogenesis was sufficient to cause increased genomic instability in cells with intact mitochondrial function.

Abstract: These results suggest mitochondrial dysfunction stimulates nuclear genome instability by inhibiting the production of ISC-containing protein(s), which are required for maintenance of nuclear genome integrity.

Abstract: For a video summary of this article, see the PaperFlick file available with the online Supplemental Data.


Replisome stall events have shaped the distribution of replication origins in the genomes of yeasts

Abstract: During S phase, the entire genome must be precisely duplicated, with no sections of DNA left unreplicated.

Abstract: Here, we develop a simple mathematical model to describe the probability of replication failing due to the irreversible stalling of replication forks.

Abstract: We show that the probability of complete genome replication is maximized if replication origins are evenly spaced, the largest inter-origin distances are minimized, and the end-most origins are positioned close to chromosome ends.

Abstract: We show that origin positions in the yeast Saccharomyces cerevisiae genome conform to all three predictions thereby maximizing the probability of complete replication if replication forks stall.

Abstract: Origin positions in four other yeasts-Kluyveromyces lactis, Lachancea kluyveri, Lachancea waltii and Schizosaccharomyces pombe-also conform to these predictions.

Abstract: Equating failure rates at chromosome ends with those in chromosome interiors gives a mean per nucleotide fork stall rate of ∼5 × 10(-8), which is consistent with experimental estimates.

Abstract: Using this value in our theoretical predictions gives replication failure rates that are consistent with data from replication origin knockout experiments.

Abstract: Our theory also predicts that significantly larger genomes, such as those of mammals, will experience a much greater probability of replication failure genome-wide, and therefore will likely require additional compensatory mechanisms.


Regulation of chromatin by histone modifications

Abstract: Chromatin is not an inert structure, but rather an instructive DNA scaffold that can respond to external cues to regulate the many uses of DNA.

Abstract: A principle component of chromatin that plays a key role in this regulation is the modification of histones.

Abstract: There is an ever-growing list of these modifications and the complexity of their action is only just beginning to be understood.

Abstract: However, it is clear that histone modifications play fundamental roles in most biological processes that are involved in the manipulation and expression of DNA.

Abstract: Here, we describe the known histone modifications, define where they are found genomically and discuss some of their functional consequences, concentrating mostly on transcription where the majority of characterisation has taken place.


Single-Cell Expression Analyses during Cellular Reprogramming Reveal an Early Stochastic and a Late Hierarchic Phase

Abstract: During cellular reprogramming, only a small fraction of cells become induced pluripotent stem cells (iPSCs).

Abstract: Previous analyses of gene expression during reprogramming were based on populations of cells, impeding single-cell level identification of reprogramming events.

Abstract: We utilized two gene expression technologies to profile 48 genes in single cells at various stages during the reprogramming process.

Abstract: Analysis of early stages revealed considerable variation in gene expression between cells in contrast to late stages.

Abstract: Expression of Esrrb, Utf1, Lin28, and Dppa2 is a better predictor for cells to progress into iPSCs than expression of the previously suggested reprogramming markers Fbxo15, Fgf4, and Oct4.

Abstract: Stochastic gene expression early in reprogramming is followed by a late hierarchical phase with Sox2 being the upstream factor in a gene expression hierarchy.

Abstract: Finally, downstream factors derived from the late phase, which do not include Oct4, Sox2, Klf4, c-Myc, and Nanog, can activate the pluripotency circuitry.


Scaffold-based three-dimensional human fibroblast culture provides a structural matrix that supports angiogenesis in infarcted heart tissue.

Abstract: BACKGROUND We have developed techniques to implant angiogenic patches onto the epicardium over regions of infarcted cardiac tissue to stimulate revascularization of the damaged tissue.

Abstract: These experiments used a scaffold-based 3D human dermal fibroblast culture (3DFC) as an epicardial patch.

Abstract: The 3DFC contains viable cells that secrete angiogenic growth factors and has previously been shown to stimulate angiogenic activity.

Abstract: The hypothesis tested was that a viable 3DFC cardiac patch would stimulate an angiogenic response within an area of infarcted cardiac tissue.

Abstract: METHODS AND RESULTS A coronary occlusion of a branch of the left anterior descending coronary artery was performed by thermal ligation in severe combined immunodeficient mice.

Abstract: 3DFCs with or without viable cells were sized to the damaged area, implanted in replicate mice onto the epicardium at the site of tissue injury, and compared with animals that received infarct surgery but no implant.

Abstract: Fourteen and 30 days after surgery, hearts were exposed and photographed, and tissue samples were prepared for histology and cytochemistry.

Abstract: Fourteen and 30 days after surgery, the damaged myocardium receiving viable 3DFC exhibited a significantly greater angiogenic response (including arterioles, venules, and capillaries) than nonviable and untreated control groups.

Abstract: CONCLUSIONS In this animal model, viable 3DFC stimulates angiogenesis within a region of cardiac infarction and can augment a repair response in damaged tissue.

Abstract: Therefore, a potential use for 3DFC is the repair of myocardial tissue damaged by infarction.


Global tuberculosis control: lessons learnt and future prospects

Abstract: Tuberculosis (TB) is an ancient disease, but not a disease of the past.

Abstract: After disappearing from the world public health agenda in the 1960s and 1970s, TB returned in the early 1990s for several reasons, including the emergence of the HIV/AIDS pandemic and increases in drug resistance.

Abstract: More than 100 years after the discovery of the tubercle bacillus by Robert Koch, what is the status of TB control worldwide?

Abstract: Here, we review the evolution of global TB control policies, including DOTS (directly observed therapy, short course) and the Stop TB Strategy, and assess whether the challenges and obstacles faced by the public health community worldwide in developing and implementing this strategy can aid future action towards the elimination of TB.


Global prevalence of glaucoma and projections of glaucoma burden through 2040: a systematic review and meta-analysis.

Abstract: PURPOSE Glaucoma is the leading cause of global irreversible blindness.

Abstract: Present estimates of global glaucoma prevalence are not up-to-date and focused mainly on European ancestry populations.

Abstract: We systematically examined the global prevalence of primary open-angle glaucoma (POAG) and primary angle-closure glaucoma (PACG), and projected the number of affected people in 2020 and 2040.

Abstract: DESIGN Systematic review and meta-analysis.

Abstract: PARTICIPANTS Data from 50 population-based studies (3770 POAG cases among 140,496 examined individuals and 786 PACG cases among 112 398 examined individuals).

Abstract: METHODS We searched PubMed, Medline, and Web of Science for population-based studies of glaucoma prevalence published up to March 25, 2013.

Abstract: Hierarchical Bayesian approach was used to estimate the pooled glaucoma prevalence of the population aged 40-80 years along with 95% credible intervals (CrIs).

Abstract: Projections of glaucoma were estimated based on the United Nations World Population Prospects.

Abstract: Bayesian meta-regression models were performed to assess the association between the prevalence of POAG and the relevant factors.

Abstract: MAIN OUTCOME MEASURES Prevalence and projection numbers of glaucoma cases.

Abstract: RESULTS The global prevalence of glaucoma for population aged 40-80 years is 3.54% (95% CrI, 2.09-5.82).

Abstract: The prevalence of POAG is highest in Africa (4.20%; 95% CrI, 2.08-7.35), and the prevalence of PACG is highest in Asia (1.09%; 95% CrI, 0.43-2.32).

Abstract: In 2013, the number of people (aged 40-80 years) with glaucoma worldwide was estimated to be 64.3 million, increasing to 76.0 million in 2020 and 111.8 million in 2040.

Abstract: In the Bayesian meta-regression model, men were more likely to have POAG than women (odds ratio [OR], 1.36; 95% CrI, 1.23-1.52), and after adjusting for age, gender, habitation type, response rate, and year of study, people of African ancestry were more likely to have POAG than people of European ancestry (OR, 2.80; 95% CrI, 1.83-4.06), and people living in urban areas were more likely to have POAG than those in rural areas (OR, 1.58; 95% CrI, 1.19-2.04).

Abstract: CONCLUSIONS The number of people with glaucoma worldwide will increase to 111.8 million in 2040, disproportionally affecting people residing in Asia and Africa.

Abstract: These estimates are important in guiding the designs of glaucoma screening, treatment, and related public health strategies.


Gastrointestinal tract as a major site of CD4+ T cell depletion and viral replication in SIV infection.

Abstract: Human and simian immunodeficiency virus (HIV and SIV) replicate optimally in activated memory CD4(+) T cells, a cell type that is abundant in the intestine.

Abstract: SIV infection of rhesus monkeys resulted in profound and selective depletion of CD4+ T cells in the intestine within days of infection, before any such changes in peripheral lymphoid tissues.

Abstract: The loss of CD4+ T cells in the intestine occurred coincident with productive infection of large numbers of mononuclear cells at this site.

Abstract: The intestine appears to be a major target for SIV replication and the major site of CD4+ T cell loss in early SIV infection.


Interaction of streptavidin-based peptide-MHC oligomers (tetramers) with cell-surface TCRs.

Abstract: The binding of oligomeric peptide-MHC (pMHC) complexes to cell surface TCR can be considered to approximate TCR-pMHC interactions at cell-cell interfaces.

Abstract: In this study, we analyzed the equilibrium binding of streptavidin-based pMHC oligomers (tetramers) and their dissociation kinetics from CD8(pos) T cells from 2C-TCR transgenic mice and from T cell hybridomas that expressed the 2C TCR or a high-affinity mutant (m33) of this TCR.

Abstract: Our results show that the tetramers did not come close to saturating cell-surface TCR (binding only 10-30% of cell-surface receptors), as is generally assumed in deriving affinity values (K(D)), in part because of dissociative losses from tetramer-stained cells.

Abstract: Guided by a kinetic model, the oligomer dissociation rate and equilibrium constants were seen to depend not only on monovalent association and dissociation rates (k(off) and k(on)), but also on a multivalent association rate (μ) and TCR cell-surface density.

Abstract: Our results suggest that dissociation rates could account for the recently described surprisingly high frequency of tetramer-negative, functionally competent T cells in some T cell responses.


Human monocyte characteristics are altered in hypercholesterolaemia.

Abstract: Peripheral blood monocytes are involved during atherogenesis in adhering to endothelium, migrating into the subendothelial space and taking-up lipoproteins to become macrophage/foam cells.

Abstract: We have assessed whether peripheral blood monocyte characteristics are altered in human hyperlipidaemia in age/sex/smoking status matched pairs of patients and controls.

Abstract: Monocytes from the hypercholesterolaemic patients, as opposed to the controls, were more sensitive to stimulation by the agonist, N-formyl-methionyl-leucyl-phenylalanine, with respect to chemokinesis (stimulation index 1.48 +/-

Abstract: 0.17 vs. 1.10 +/-

Abstract: 0.14), chemotaxis (4.05 +/- 0.55 vs. 2.72 +/- 0.24) and adhesion to porcine aortic endothelial monolayers (1.26 +/- 0.05 vs. 1.17 +/- 0.06).

Abstract: The patients' monocyte total surface expression of the adhesion glycoprotein CD11b/CD18 (37.5 +/- 7.1 vs. 36.0 +/-

Abstract: 7.1), but not CD11c/CD18 (31.6 +/- 7.2 vs. 31.4 +/-

Abstract: 6.8), was increased; however, the monocytes in hyperlipidaemia were larger (9.15 +/-

Abstract: 0.11 microns vs. 8.98 +/-

Abstract: 0.11 microns) such that the surface density of CD11b/CD18 was not altered (0.144 +/- 0.029 vs. 0.142 +/-

Abstract: The data suggest that circulating monocytes are functionally different in hypercholesterolaemia.

Abstract: This may explain the increased involvement by monocytes in hypercholesterolaemia-related atherogenesis.


Adult Mammalian Neural Stem Cells and Neurogenesis: Five Decades Later.

Abstract: Adult somatic stem cells in various organs maintain homeostatic tissue regeneration and enhance plasticity.

Abstract: Since its initial discovery five decades ago, investigations of adult neurogenesis and neural stem cells have led to an established and expanding field that has significantly influenced many facets of neuroscience, developmental biology, and regenerative medicine.

Abstract: Here we review recent progress and focus on questions related to adult mammalian neural stem cells that also apply to other somatic stem cells.

Abstract: We further discuss emerging topics that are guiding the field toward better understanding adult neural stem cells and ultimately applying these principles to improve human health.


Relationship of ambulatory blood pressure and the heart rate profile with renal function parameters in hypertensive patients with chronic kidney disease.

Abstract: Strict blood pressure (BP) control is reportedly important for the management of hypertensive patients with chronic kidney disease (CKD).

Abstract: The purpose of this cross-sectional study was to examine whether the variables of ambulatory BP and the heart rate (HR) profile, central hemodynamics, and arterial stiffness were closely related to the renal function parameters (urine albumin excretion rate [UACR] and estimated glomerular filtration rate [eGFR]) observed in 25 consecutive hospitalized hypertensive patients with CKD.

Abstract: There were significant positive relationships between UACR and 24-hour, daytime, and nighttime ambulatory systolic BP.

Abstract: In addition, there were significant negative relationships between UACR and 24-hour and daytime HR variability.

Abstract: The circulating B-type natriuretic peptide level and hemoglobin A1c were also positively related to UACR.

Abstract: With respect to eGFR, although the 24-hour and nighttime HR variability were positively associated with eGFR, the circulating pentosidine and nighttime HR had a negative relationship with eGFR.

Abstract: On the other hand, central hemodynamics and arterial stiffness did not exhibit any significant association with renal function parameters.

Abstract: These results indicate that ambulatory BP and the HR profile are closely modulated by renal function deterioration.

Abstract: Further studies are needed to investigate the causal relationship between ambulatory BP and the HR profile and renal function parameters in hypertensive patients with CKD.


The frequency and immunodominance of islet-specific CD8+ T-cell responses change after type 1 diabetes diagnosis and treatment.

Abstract: OBJECTIVE Islet-reactive CD8(+) T-cells play a key role in the pathogenesis of type 1 diabetes in the NOD mouse.

Abstract: The predominant T-cell specificities change over time, but whether similar shifts also occur after clinical diagnosis and insulin treatment in type 1 diabetic patients is unknown.

Abstract: RESEARCH DESIGN AND METHODS

Abstract: We took advantage of a recently validated islet-specific CD8(+) T-cell gamma-interferon enzyme-linked immunospot (ISL8Spot) assay to follow responses against preproinsulin (PPI), GAD, insulinoma-associated protein 2 (IA-2), and islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) epitopes in 15 HLA-A2(+) adult type 1 diabetic patients close to diagnosis and at a second time point 7-16 months later.

Abstract: RESULTS CD8(+) T-cell reactivities were less frequent at follow-up, as 28.6% of responses tested positive at type 1 diabetes diagnosis vs. 13.2% after a median of 11 months (P = 0.003).

Abstract: While GAD and IA-2 autoantibody (aAb) titers were unchanged in 75% of cases, the fraction of patients responding to PPI and/or GAD epitopes by ISL8Spot decreased from 60-67 to 20% (P < 0.02).

Abstract: The previously subdominant IA-2(206-214) and IGRP(265-273) peptides were newly targeted, thus becoming the immunodominant epitopes.

Abstract: CONCLUSIONS Shifts both in frequency and in immunodominance of CD8(+) T-cell responses occur more rapidly than do changes in aAb titers.

Abstract: These different kinetics may suggest complementary clinical applications for T-cell and aAb measurements.


A Protein Inventory of Human Ribosome Biogenesis Reveals an Essential Function of Exportin 5 in 60S Subunit Export

Abstract: The assembly of ribosomal subunits in eukaryotes is a complex, multistep process so far mostly studied in yeast.

Abstract: In S. cerevisiae, more than 200 factors including ribosomal proteins and trans-acting factors are required for the ordered assembly of 40S and 60S ribosomal subunits.

Abstract: To date, only few human homologs of these yeast ribosome synthesis factors have been characterized.

Abstract: Here, we used a systematic RNA interference (RNAi) approach to analyze the contribution of 464 candidate factors to ribosomal subunit biogenesis in human cells.

Abstract: The screen was based on visual readouts, using inducible, fluorescent ribosomal proteins as reporters.

Abstract: By performing computer-based image analysis utilizing supervised machine-learning techniques, we obtained evidence for a functional link of 153 human proteins to ribosome synthesis.

Abstract: Our data show that core features of ribosome assembly are conserved from yeast to human, but differences exist for instance with respect to 60S subunit export.

Abstract: Unexpectedly, our RNAi screen uncovered a requirement for the export receptor Exportin 5 (Exp5) in nuclear export of 60S subunits in human cells.

Abstract: We show that Exp5, like the known 60S exportin Crm1, binds to pre-60S particles in a RanGTP-dependent manner.

Abstract: Interference with either Exp5 or Crm1 function blocks 60S export in both human cells and frog oocytes, whereas 40S export is compromised only upon inhibition of Crm1.

Abstract: Thus, 60S subunit export is dependent on at least two RanGTP-binding exportins in vertebrate cells.


Binding of the winged-helix transcription factor HNF3 to a linker histone site on the nucleosome.

Abstract: The transcription factor HNF3 and linker histones H1 and H5 possess winged-helix DNA-binding domains, yet HNF3 and other fork head-related proteins activate genes during development whereas linker histones compact DNA in chromatin and repress gene expression.

Abstract: We compared how the two classes of factors interact with chromatin templates and found that HNF3 binds DNA at the side of nucleosome cores, similarly to what has been reported for linker histone.

Abstract: A nucleosome structural binding site for HNF3 is occupied at the albumin transcriptional enhancer in active and potentially active chromatin, but not in inactive chromatin in vivo.

Abstract: While wild-type HNF3 protein does not compact DNA extending from the nucleosome, as does linker histone, site-directed mutants of HNF3 can compact nucleosomal DNA if they contain basic amino acids at positions previously shown to be essential for nucleosomal DNA compaction by linker histones.

Abstract: The results illustrate how transcription factors can possess special nucleosome-binding activities that are not predicted from studies of factor interactions with free DNA.


Rapid recent growth and divergence of rice nuclear genomes.

Abstract: By employing the nuclear DNA of the African rice Oryza glaberrima as a reference genome, the timing, natures, mechanisms, and specificities of recent sequence evolution in the indica and japonica subspecies of Oryza sativa were identified.

Abstract: The data indicate that the genome sizes of both indica and japonica have increased substantially, >2% and >6%, respectively, since their divergence from a common ancestor, mainly because of the amplification of LTR-retrotransposons.

Abstract: However, losses of all classes of DNA sequence through unequal homologous recombination and illegitimate recombination have attenuated the growth of the rice genome.

Abstract: Small deletions have been particularly frequent throughout the genome.

Abstract: In >1 Mb of orthologous regions that we analyzed, no cases of complete gene acquisition or loss from either indica or japonica were found, nor was any example of precise transposon excision detected.

Abstract: The sequences between genes were observed to have a very high rate of divergence, indicating a molecular clock for transposable elements that is at least 2-fold more rapid than synonymous base substitutions within genes.

Abstract: We found that regions prone to frequent insertions and deletions also exhibit higher levels of point mutation.

Abstract: These results indicate a highly dynamic rice genome with competing processes for the generation and removal of genetic variation.


Detection of an autoreactive T-cell population within the polyclonal repertoire that undergoes distinct autoimmune regulator (Aire)-mediated selection.

Abstract: The autoimmune regulator (Aire) plays a critical role in central tolerance by promoting the display of tissue-specific antigens in the thymus.

Abstract: To study the influence of Aire on thymic selection in a physiological setting, we used tetramer reagents to detect autoreactive T cells specific for the Aire-dependent tissue-specific antigen interphotoreceptor retinoid-binding protein (IRBP), in the polyclonal repertoire.

Abstract: Two class II tetramer reagents were designed to identify T cells specific for two different peptide epitopes of IRBP.

Abstract: Analyses of the polyclonal T-cell repertoire showed a high frequency of activated T cells specific for both IRBP tetramers in Aire(-/-) mice, but not in Aire(+/+) mice.

Abstract: Surprisingly, although one tetramer-binding T-cell population was efficiently deleted in the thymus in an Aire-dependent manner, the second tetramer-binding population was not deleted and could be detected in both the Aire(-/-) and Aire(+/+) T-cell repertoires.

Abstract: We found that Aire-dependent thymic deletion of IRBP-specific T cells relies on intercellular transfer of IRBP between thymic stroma and bone marrow-derived antigen-presenting cells.

Abstract: Furthermore, our data suggest that Aire-mediated deletion relies not only on thymic expression of IRBP, but also on proper antigen processing and presentation of IRBP by thymic antigen-presenting cells.


Spatial and temporal heterogeneity of Anopheles mosquitoes and Plasmodium falciparum transmission along the Kenyan coast.

Abstract: The seasonal dynamics and spatial distributions of Anopheles mosquitoes and Plasmodium falciparum parasites were studied for one year at 30 villages in Malindi, Kilifi, and Kwale Districts along the coast of Kenya.

Abstract: Anopheline mosquitoes were sampled inside houses at each site once every two months and malaria parasite prevalence in local school children was determined at the end of the entomologic survey.

Abstract: A total of 5,476 Anopheles gambiae s.l.

Abstract: and 3,461 An.

Abstract: funestus were collected.

Abstract: Species in the An.

Abstract: gambiae complex, identified by a polymerase chain reaction, included 81.9% An.

Abstract: gambiae s.s., 12.8% An.

Abstract: arabiensis, and 5.3% An.

Abstract: merus.

Abstract: Anopheles gambiae s.s.

Abstract: contributed most to the transmission of P. falciparum along the coast as a whole, while An.

Abstract: funestus accounted for more than 50% of all transmission in Kwale District.

Abstract: Large spatial heterogeneity of transmission intensity (< 1 up to 120 infective bites per person per year) resulted in correspondingly large and significantly related variations in parasite prevalence (range = 38-83%).

Abstract: Thirty-two percent of the sites (7 of 22 sites) with malaria prevalences ranging from 38% to 70% had annual entomologic inoculation rates (EIR) less than five infective bites per person per year.

Abstract: Anopheles gambiae s.l.

Abstract: and An.

Abstract: funestus densities in Kwale were not significantly influenced by rainfall.

Abstract: However, both were positively correlated with rainfall one and three months previously in Malindi and Kilifi Districts, respectively.

Abstract: These unexpected variations in the relationship between mosquito populations and rainfall suggest environmental heterogeneity in the predominant aquatic habitats in each district.

Abstract: One important conclusion is that the highly non-linear relationship between EIRs and prevalence indicates that the consistent pattern of high prevalence might be governed by substantial variation in transmission intensity measured by entomologic surveys.

Abstract: The field-based estimate of entomologic parameters on a district level does not provide a sensitive indicator of transmission intensity in this study.


Myelodysplasia and leukemia of Fanconi anemia are associated with a specific pattern of genomic abnormalities that includes cryptic RUNX1/AML1 lesions.

Abstract: Fanconi anemia (FA) is a genetic condition associated with bone marrow (BM) failure, myelodysplasia (MDS), and acute myeloid leukemia (AML).

Abstract: We studied 57 FA patients with hypoplastic or aplastic anemia (n = 20), MDS (n = 18), AML (n = 11), or no BM abnormality (n = 8).

Abstract: BM samples were analyzed by karyotype, high-density DNA arrays with respect to paired fibroblasts, and by selected oncogene sequencing.

Abstract: A specific pattern of chromosomal abnormalities was found in MDS/AML, which included 1q+ (44.8%), 3q+ (41.4%), -7/7q (17.2%), and 11q- (13.8%).

Abstract: Moreover, cryptic RUNX1/AML1 lesions (translocations, deletions, or mutations) were observed for the first time in FA (20.7%).

Abstract: Rare mutations of NRAS, FLT3-ITD, MLL-PTD, ERG amplification, and ZFP36L2-PRDM16 translocation, but no TP53, TET2, CBL, NPM1, and CEBPα mutations were found.

Abstract: Frequent homozygosity regions were related not to somatic copy-neutral loss of heterozygosity but to consanguinity, suggesting that homologous recombination is not a common progression mechanism in FA.

Abstract: Importantly, the RUNX1 and other chromosomal/genomic lesions were found at the MDS/AML stages, except for 1q+, which was found at all stages.

Abstract: These data have implications for staging and therapeutic managing in FA patients, and also to analyze the mechanisms of clonal evolution and oncogenesis in a background of genomic instability and BM failure.


Molecular and Cellular Approaches for Diversifying and Extending Optogenetics

Abstract: Optogenetic technologies employ light to control biological processes within targeted cells in vivo with high temporal precision.

Abstract: Here, we show that application of molecular trafficking principles can expand the optogenetic repertoire along several long-sought dimensions.

Abstract: Subcellular and transcellular trafficking strategies now permit (1) optical regulation at the far-red/infrared border and extension of optogenetic control across the entire visible spectrum, (2) increased potency of optical inhibition without increased light power requirement (nanoampere-scale chloride-mediated photocurrents that maintain the light sensitivity and reversible, step-like kinetic stability of earlier tools), and (3) generalizable strategies for targeting cells based not only on genetic identity, but also on morphology and tissue topology, to allow versatile targeting when promoters are not known or in genetically intractable organisms.

Abstract: Together, these results illustrate use of cell-biological principles to enable expansion of the versatile fast optogenetic technologies suitable for intact-systems biology and behavior.


An analysis of the Caenorhabditis elegans lipid raft proteome using geLC-MS/MS.

Abstract: Lipid rafts are microdomains of the phospholipid bilayer, proposed to form semi-stable "islands" that act as a platform for several important cellular processes; major classes of raft-resident proteins include signalling proteins and glycosylphosphatidylinositol (GPI)-anchored proteins.

Abstract: Proteomic studies into lipid rafts have been mainly carried out in mammalian cell lines and single cell organisms.

Abstract: The nematode Caenorhabditis elegans, the model organism with a well-defined developmental profile, is ideally suited for the study of this subcellular locale in a complex developmental context.

Abstract: A study of the lipid raft proteome of C. elegans is presented here.

Abstract: A total of 44 proteins were identified from the lipid raft fraction using geLC-MS/MS, of which 40 have been determined to be likely raft proteins after analysis of predicted functions.

Abstract: Prediction of GPI-anchoring of the proteins found 21 to be potentially modified in this way, two of which were experimentally confirmed to be GPI-anchored.

Abstract: This work is the first reported study of the lipid raft proteome in C. elegans.

Abstract: The results show that raft proteins, including numerous GPI-anchored proteins, may have a variety of potentially important roles within the nematode, and will hopefully lead to C. elegans becoming a useful model for the study of lipid rafts.


Geographic distribution of community-acquired methicillin-resistant Staphylococcus aureus soft tissue infections.

Abstract: PURPOSE The goal of this study is to look at the geographic growth patterns of community-acquired methicillin-resistant Staphylococcus aureus (MRSA) infections in our local region and to determine if specific geographic areas are at increased risk.

Abstract: METHODS After Institution Review Board approval (132603-3), a retrospective chart review was conducted of 614 patients who underwent incision and drainage of an abscess by a single pediatric surgical practice from January 2004 to December 2008.

Abstract: In addition, previously published data from 195 patients who underwent incision and drainage of an abscess from January 2000 to December 2003 were reviewed.

Abstract: RESULTS The most commonly cultured organism found in the pediatric population undergoing incision and drainage was S aureus (n = 388), of which 258 (66%) were methicillin resistant.

Abstract: This is a 21% increase from the rate of MRSA cultures identified from 2000 to 2003.

Abstract: Geographic information system space-time analysis showed that a cluster of 14 MRSA cases was located within a 1.44-km radius between 2000 and 2003, and 5 separate clusters of more than 20 MRSA infection cases each were identified in 3 separate cities over the 8-year time span using geographic information system spatial analysis (P value = .001).

Abstract: CONCLUSION Methicillin-resistant S aureus has now become the most prevalent organism isolated from cultures of community-acquired abscesses requiring incision and drainage in the pediatric population in our local region.

Abstract: Significant clustering of MRSA infections has appeared in several different cities within our geographic region.


The right place at the right time: chaperoning core histone variants.

Abstract: Histone proteins dynamically regulate chromatin structure and epigenetic signaling to maintain cell homeostasis.

Abstract: These processes require controlled spatial and temporal deposition and eviction of histones by their dedicated chaperones.

Abstract: With the evolution of histone variants, a network of functionally specific histone chaperones has emerged.

Abstract: Molecular details of the determinants of chaperone specificity for different histone variants are only slowly being resolved.

Abstract: A complete understanding of these processes is essential to shed light on the genuine biological roles of histone variants, their chaperones, and their impact on chromatin dynamics.


Phosphorylation site mutations in heterochromatin protein 1 (HP1) reduce or eliminate silencing activity.

Abstract: HP1 is an essential heterochromatin-associated protein in Drosophila.

Abstract: HP1 has dosage-dependent effects on the silencing of euchromatic genes that are mislocalized to heterochromatin and is required for the normal expression of at least two heterochromatic genes.

Abstract: HP1 is multiply phosphorylated in vivo, and HP1 hyperphosphorylation is correlated with heterochromatin assembly during development.

Abstract: The purpose of this study was to test whether HP1 phosphorylation modifies biological activity and biochemical properties of HP1.

Abstract: To determine sites of HP1 phosphorylation in vivo and whether phosphorylation affects any biochemical properties of HP1, we expressed Drosophila HP1 in lepidopteran cultured cells using a recombinant baculovirus vector.

Abstract: Phosphopeptides were identified by matrix-assisted laser desorption ionization/time of flight mass spectroscopy; these peptides contain target sites for casein kinase II, protein tyrosine kinase, and PIM-1 kinase.

Abstract: Purified HP1 from bacterial (unphosphorylated) and lepidopteran (phosphorylated) cells has similar secondary structure.

Abstract: Phosphorylation has no effect on HP1 self-association but alters the DNA binding properties of HP1, suggesting that phosphorylation could differentially regulate HP1-dependent interactions.

Abstract: Serine-to-alanine and serine-to-glutamate substitutions at consensus protein kinase motifs resulted in reduction or loss of silencing activity of mutant HP1 in transgenic flies.

Abstract: These results suggest that dynamic phosphorylation/dephosphorylation regulates HP1 activity in heterochromatic silencing.


Elimination of paternal mitochondria in mouse embryos occurs through autophagic degradation dependent on PARKIN and MUL1

Abstract: A defining feature of mitochondria is their maternal mode of inheritance.

Abstract: However, little is understood about the cellular mechanism through which paternal mitochondria, delivered from sperm, are eliminated from early mammalian embryos.

Abstract: Autophagy has been implicated in nematodes, but whether this mechanism is conserved in mammals has been disputed.

Abstract: Here, we show that cultured mouse fibroblasts and pre-implantation embryos use a common pathway for elimination of mitochondria.

Abstract: Both situations utilize mitophagy, in which mitochondria are sequestered by autophagosomes and delivered to lysosomes for degradation.

Abstract: The E3 ubiquitin ligases PARKIN and MUL1 play redundant roles in elimination of paternal mitochondria.

Abstract: The process is associated with depolarization of paternal mitochondria and additionally requires the mitochondrial outer membrane protein FIS1, the autophagy adaptor P62, and PINK1 kinase.

Abstract: Our results indicate that strict maternal transmission of mitochondria relies on mitophagy and uncover a collaboration between MUL1 and PARKIN in this process.


Ribosome Profiling Provides Evidence that Large Noncoding RNAs Do Not Encode Proteins

Abstract: Large noncoding RNAs are emerging as an important component in cellular regulation.

Abstract: Considerable evidence indicates that these transcripts act directly as functional RNAs rather than through an encoded protein product.

Abstract: However, a recent study of ribosome occupancy reported that many large intergenic ncRNAs (lincRNAs) are bound by ribosomes, raising the possibility that they are translated into proteins.

Abstract: Here, we show that classical noncoding RNAs and 5' UTRs show the same ribosome occupancy as lincRNAs, demonstrating that ribosome occupancy alone is not sufficient to classify transcripts as coding or noncoding.

Abstract: Instead, we define a metric based on the known property of translation whereby translating ribosomes are released upon encountering a bona fide stop codon.

Abstract: We show that this metric accurately discriminates between protein-coding transcripts and all classes of known noncoding transcripts, including lincRNAs.

Abstract: Taken together, these results argue that the large majority of lincRNAs do not function through encoded proteins.


Isolation of a Fusion Transcript, AMLl IETO, With Similarity to

Abstract: We have developed a restriction map of the chromosome 21 breakpoint region involved in t(8;21)(q22;q22.3) acute myelogenous leukemia (AML) and have isolated a genomic junction clone containing chromosome 8 and 21 material.

Abstract: Using probes from these regions, rearrangements have been identified in each of nine cases of t(8;21) AML examined.

Abstract: In addition, we have isolated cDNA clones from a t(8;21) AML cDNA library that contain fused sequences from chromosome 8 and 21.

Abstract: The chromosome 8 component, referred to as ETO (for eight twenty-one), is encoded over a large genomic region, as suggested by the analysis of corresponding yeast artificial chromosomes (YACs).

Abstract: The DNA sequence of the chromosome 21 portion of the fusion transcript is derived from the normal AML1 gene.

Abstract: A striking similarity (67% identity over 387 bp, with a corresponding 69% amino acid identity) was detected between AML1 and the Drosophila segmentation gene, runt.

Abstract: The critical consequence of the translocation is the juxtaposition of 5' sequences of AML1 to 3' sequences of ETO, oriented telomere to centromere on the der(8) chromosome.


Treatment of spondyloarthropathy with 5-aminosalicylic acid (mesalazine): an open trial.

Abstract: OBJECTIVE Ankylosing spondylitis (AS) and spondyloarthropathy (SpA) are inflammatory diseases of unknown etiology.

Abstract: Various exogenous and endogenous (inherited) factors play a role in their development.

Abstract: Sulfasalazine (SSZ) is generally accepted as a disease modifying drug in the treatment of AS and SpA. Which part of SSZ, 5-acetylsalicylic acid (5-ASA, mesalazine) or sulfapyridine (SP), is the effective moiety is unknown.

Abstract: As the bowel, colon, and the ileum play an important role in the development of AS and SpA, it may be possible that 5-ASA is the effective moiety, with a similar mode of action as in the treatment of inflammatory bowel disease.

Abstract: To determine the efficacy of 5-ASA an open pilot study was done in 2 groups of patients with SpA.   METHODS Twenty patients with SpA, who were taking SSZ, were switched to 5-ASA (Pentasa), and 19 patients with active SpA were treated with 5-ASA without previous administration of SSZ.

Abstract: RESULTS In the first group, 17 (85%) patients responded with respect to the physician global clinical assessment compared to the previous SSZ treatment period; whereas in the second patient group a statistically significant improvement was obtained in erythrocyte sedimentation rate.

Abstract: CONCLUSION The results support our hypothesis that 5-ASA might be the active moiety of SSZ in the treatment of SpA.


Relationship of physical activity and television watching with body weight and level of fatness among children: results from the Third National Health and Nutrition Examination Survey.

Abstract: CONTEXT Physical inactivity contributes to weight gain in adults, but whether this relationship is true for children of different ethnic groups is not well established.

Abstract: OBJECTIVE To assess participation in vigorous activity and television watching habits and their relationship to body weight and fatness in US children.

Abstract: DESIGN Nationally representative cross-sectional survey with an in-person interview and medical examination.

Abstract: SETTING AND PARTICIPANTS Between 1988 and 1994, 4063 children aged 8 through 16 years were examined as part of the National Health and Nutrition Examination Survey III.

Abstract: Mexican Americans and non-Hispanic blacks were oversampled to produce reliable estimates for these groups.

Abstract: MAIN OUTCOME MEASURES Episodes of weekly vigorous activity and daily hours of television watched, and their relationship to body mass index and body fatness.

Abstract: RESULTS Eighty percent of US children reported performing 3 or more bouts of vigorous activity each week.

Abstract: This rate was lower in non-Hispanic black and Mexican American girls (69% and 73%, respectively).

Abstract: Twenty percent of US children participated in 2 or fewer bouts of vigorous activity perweek, and the rate was higher in girls (26%) than in boys (17%).

Abstract: Overall, 26% of US children watched 4 or more hours of television per day and 67% watched at least 2 hours per day.

Abstract: Non-Hispanic black children had the highest rates of watching 4 or more hours of television per day (42%).

Abstract: Boys and girls who watch 4 or more hours of television each day had greater body fat (P<.001) and had a greater body mass index (P<.001) than those who watched less than 2 hours per day.

Abstract: CONCLUSIONS Many US children watch a great deal of television and are inadequately vigorously active.

Abstract: Vigorous activity levels are lowest among girls, non-Hispanic blacks, and Mexican Americans.

Abstract: Intervention strategies to promote lifelong physical activity among US children are needed to stem the adverse health consequences of inactivity.


Vascular Calcification in Chronic Kidney Disease is Induced by Bone Morphogenetic Protein-2 via a Mechanism Involving the Wnt/β-Catenin Pathway

Abstract: Background: Vascular calcification (VC), in which vascular smooth muscle cells (VSMCs) undergo a phenotypic transformation into osteoblast-like cells, is one of the emergent risk factors for the accelerated atherosclerosis process characteristic of chronic kidney disease (CKD).

Abstract: Phosphate is an important regulator of VC.

Abstract: Methods: The expression of different smooth muscle cell or osteogenesis markers in response to high concentrations of phosphate or exogenous bone morphogenetic protein 2 (BMP-2) was examined by qRT-PCR and western blotting in rat VSMCs.

Abstract: Osteocalcin secretion was measured by radioimmunoassay.

Abstract: Differentiation and calcification of VSMCs were examined by alkaline phosphatase (ALP) activity assay and Alizarin staining.

Abstract: Short hairpin RNA-mediated silencing of β-catenin was performed to examine the involvement of Wnt/β-catenin signaling in VSMC calcification and osteoblastic differentiation induced by high phosphate or BMP-2.

Abstract: Apoptosis was determined by TUNEL assay and immunofluorescence imaging.

Abstract: Results: BMP-2 serum levels were significantly higher in CKD patients than in controls.

Abstract: High phosphate concentrations and BMP-2 induced VSMC apoptosis and upregulated the expression of β-catenin, Msx2, Runx2 and the phosphate cotransporter Pit1, whereas a BMP-2 neutralization antibody reversed these effects.

Abstract: Knockdown of β-catenin abolished the effect of high phosphate and BMP-2 on VSMC apoptosis and calcification.

Abstract: Conclusions: BMP-2 plays a crucial role in calcium deposition in VSMCs and VC in CKD patients via a mechanism involving the Wnt/β-catenin pathway.


Advances in brain tumor surgery.

Abstract: Advances in the fields of molecular and translational research, oncology, and surgery have emboldened the medical community to believe that intrinsic brain tumors may be treatable.

Abstract: Intraoperative imaging and brain mapping allow operations adjacent to eloquent cortex and more radical resection of tumors with increased confidence and safety.

Abstract: Despite these advances, the infiltrating edge of a neoplasm and distant microscopic satellite lesions will never be amendable to a surgical cure.

Abstract: Indeed, it is continued research into the delivery of an efficacious chemobiologic agent that will eventually allows us to manage this primary cause of treatment failure.


Systematic review of controlled clinical trials of gastric lavage in acute organophosphorus pesticide poisoning.

Abstract: BACKGROUND Organophosphorus pesticide (OP) self-poisoning is a major problem in the developing rural world.

Abstract: There is little clinical trial data to guide therapy, hindering the identification of best therapy.

Abstract: Despite the recognition of adverse effects, gastric lavage is commonly done in Asia.

Abstract: We aimed to identify studies assessing its effectiveness.

Abstract: METHOD We systematically searched the literature for controlled clinical studies that assessed the effect of gastric lavage in OP pesticide self-poisoning.

Abstract: RESULTS All 56 studies identified were Chinese and reported benefit from the intervention studied, including multiple gastric lavages, use of norepinephrine or pralidoxime in the lavage fluid, concurrent treatment with naloxone or scopolamine, insertion of the gastric tube via a laparotomy incision, and lavage later than 12 h post-ingestion.

Abstract: However, only 23 were RCTs and none presented adequate methodology for their quality to be assessed.

Abstract: The patient population and study treatment protocol were not defined - large variation in case fatality in the control arm of the studies (from 4.5 to 93%) suggests marked variation between studies and likely between study arms.

Abstract: No study compared an intervention against a control group receiving no gastric lavage or provided any data to indicate whether a significant quantity of poison was removed.

Abstract: CONCLUSION Despite widespread use of multiple gastric lavages for OP pesticide poisoning across Asia, there is currently no high-quality evidence to support its clinical effectiveness.

Abstract: There is a need for studies to identify in which patients and for what duration gastric lavage is able to remove significant quantities of poison.

Abstract: Following these studies, large clinical trials will be required to address the effectiveness and safety of gastric lavage (either single or multiple) in acute OP pesticide poisoning.