The aim of the study was to determine the effect of single whole-body cryotherapy (WBC) session applied prior to submaximal exercise on the activity of antioxidant enzymes, the concentration of lipid peroxidation products, total oxidative status, and the level of cytokines in blood of volleyball players. The study group consisted of 18 male professional volleyball players, who were subjected to extremely cold air (-130°C) prior to exercise performed on cycloergometer. Blood samples were taken five times: before WBC, after WBC procedure, after exercise preceded by cryotherapy (WBC exercise), and before and after exercise without WBC (control exercise). The activity of catalase statistically significantly increased after control exercise. Moreover, the activity of catalase and superoxide dismutase was lower after WBC exercise than after control exercise (P < 0.001). After WBC exercise, the level of IL-6 and IL-1β was also lower (P < 0.001) than after control exercise. The obtained results may suggest that cryotherapy prior to exercise may have some antioxidant and anti-inflammatory properties. The relations between the level of studied oxidative stress and inflammatory markers may testify to the contribution of reactive oxygen species in cytokines release into the blood system in response to exercise and WBC.


UNLABELLED The contribution of whole-genome doubling to chromosomal instability (CIN) and tumor evolution is unclear. We use long-term culture of isogenic tetraploid cells from a stable diploid colon cancer progenitor to investigate how a genome-doubling event affects genome stability over time. Rare cells that survive genome doubling demonstrate increased tolerance to chromosome aberrations. Tetraploid cells do not exhibit increased frequencies of structural or numerical CIN per chromosome. However, the tolerant phenotype in tetraploid cells, coupled with a doubling of chromosome aberrations per cell, allows chromosome abnormalities to evolve specifically in tetraploids, recapitulating chromosomal changes in genomically complex colorectal tumors. Finally, a genome-doubling event is independently predictive of poor relapse-free survival in early-stage disease in two independent cohorts in multivariate analyses [discovery data: hazard ratio (HR), 4.70, 95% confidence interval (CI), 1.04-21.37; validation data: HR, 1.59, 95% CI, 1.05-2.42]. These data highlight an important role for the tolerance of genome doubling in driving cancer genome evolution. SIGNIFICANCE Our work sheds light on the importance of whole-genome–doubling events in colorectal cancer evolution. We show that tetraploid cells undergo rapid genomic changes and recapitulate the genetic alterations seen in chromosomally unstable tumors. Furthermore, we demonstrate that a genome-doubling event is prognostic of poor relapse-free survival in this disease type.


The transmembrane protein CD33 is a sialic acid-binding immunoglobulin-like lectin that regulates innate immunity but has no known functions in the brain. We have previously shown that the CD33 gene is a risk factor for Alzheimer's disease (AD). Here, we observed increased expression of CD33 in microglial cells in AD brain. The minor allele of the CD33 SNP rs3865444, which confers protection against AD, was associated with reductions in both CD33 expression and insoluble amyloid beta 42 (Aβ42) levels in AD brain. Furthermore, the numbers of CD33-immunoreactive microglia were positively correlated with insoluble Aβ42 levels and plaque burden in AD brain. CD33 inhibited uptake and clearance of Aβ42 in microglial cell cultures. Finally, brain levels of insoluble Aβ42 as well as amyloid plaque burden were markedly reduced in APP(Swe)/PS1(ΔE9)/CD33(-/-) mice. Therefore, CD33 inactivation mitigates Aβ pathology and CD33 inhibition could represent a novel therapy for AD.


HIV causes a chronic infection characterized by depletion of CD4(+) T lymphocytes and the development of opportunistic infections. Despite drugs that inhibit viral spread, HIV infection has been difficult to cure because of uncharacterized reservoirs of infected cells that are resistant to highly active antiretroviral therapy (HAART) and the immune response. Here we used CD34(+) cells from infected people as well as in vitro studies of wild-type HIV to show infection and killing of CD34(+) multipotent hematopoietic progenitor cells (HPCs). In some HPCs, we detected latent infection that stably persisted in cell culture until viral gene expression was activated by differentiation factors. A unique reporter HIV that directly detects latently infected cells in vitro confirmed the presence of distinct populations of active and latently infected HPCs. These findings have major implications for understanding HIV bone marrow pathology and the mechanisms by which HIV causes persistent infection.


It is well known that upon stress, the level of the tumor suppressor p53 is remarkably elevated. However, despite extensive studies, the underlying mechanism involving important inter-players for stress-induced p53 regulation is still not fully understood. We present evidence that the human lincRNA-RoR (RoR) is a strong negative regulator of p53. Unlike MDM2 that causes p53 degradation through the ubiquitin-proteasome pathway, RoR suppresses p53 translation through direct interaction with the heterogeneous nuclear ribonucleoprotein I (hnRNP I). Importantly, a 28-base RoR sequence carrying hnRNP I binding motifs is essential and sufficient for p53 repression. We further show that RoR inhibits p53-mediated cell cycle arrest and apoptosis. Finally, we demonstrate a RoR-p53 autoregulatory feedback loop where p53 transcriptionally induces RoR expression. Together, these results suggest that the RoR-hnRNP I-p53 axis may constitute an additional surveillance network for the cell to better respond to various stresses.


The increase in lactate (L) and pyruvate (P) content of arterial blood during experimental and clinical shock states and the extent to which such increases serve as measures of oxygen deficit and irreversible injury were investigated on an empirical basis. A standardized method for production of hemorrhagic shock in the Wistar rat was employed. During a 4-hour bleeding period, oxygen consumption of the rat was reduced to approximately 40% of control value, pH was reduced from 7.39 to 7.08, and a concurrent increase in L from 0.80 to 6.06 mm and in P from 0.07 to 0.18 mm were observed. Cumulative oxygen debt correlated with log L (r = 0.50; P < 0.0005) and both were significantly related to survival. Correlation of cumulative oxygen debt and survival, both with P and with computed values of the lactate pyruvate ratio (L/P) and excess lactate (XL), were of no higher magnitude. Partial correlation analysis demonstrated that neither the measurement of P nor the computation of L/P or XL improved predictability...


Pancreatic ductal adenocarcinoma (PDAC) remains a lethal disease with a 5-year survival rate of 4%. A key hallmark of PDAC is extensive stromal involvement, which makes capturing precise tumor-specific molecular information difficult. Here we have overcome this problem by applying blind source separation to a diverse collection of PDAC gene expression microarray data, including data from primary tumor, metastatic and normal samples. By digitally separating tumor, stromal and normal gene expression, we have identified and validated two tumor subtypes, including a 'basal-like' subtype that has worse outcome and is molecularly similar to basal tumors in bladder and breast cancers. Furthermore, we define 'normal' and 'activated' stromal subtypes, which are independently prognostic. Our results provide new insights into the molecular composition of PDAC, which may be used to tailor therapies or provide decision support in a clinical setting where the choice and timing of therapies are critical.


Remarkable advances have been made in recent years towards therapeutics for cognitive impairment in individuals with Down syndrome (DS) by using mouse models. In this review, we briefly describe the phenotypes of mouse models that represent outcome targets for drug testing, the behavioral tests used to assess impairments in cognition and the known mechanisms of action of several drugs that are being used in preclinical studies or are likely to be tested in clinical trials. Overlaps in the distribution of targets and in the pathways that are affected by these diverse drugs in the trisomic brain suggest new avenues for DS research and drug development.


The proteolytic cleavages elicited by activation of the Notch receptor release an intracellular fragment, Notch intracellular domain, which enters the nucleus to activate the transcription of targets. Changes in transcription are therefore a major output of this pathway. However, the Notch outputs clearly differ from cell type to cell type. In this review we discuss current understanding of Notch targets, the mechanisms involved in their transcriptional regulation, and what might underlie the activation of different sets of targets in different cell types.


Allergic airway inflammation is characterized by marked in situ changes in gene and protein expression, yet the role of microRNAs (miRNAs), a new family of key mRNA regulatory molecules, in this process has not yet been reported. Using a highly sensitive microarray-based approach, we identified 21 miRNAs with differential expression between doxycycline-induced lung-specific IL-13 transgenic mice (with allergic airway inflammation) and control mice. In particular, we observed overexpression of miR-21 and underexpression of miR-1 in the induced IL-13 transgenic mice compared with control mice. These findings were validated in two independent models of allergen-induced allergic airway inflammation and in IL-4 lung transgenic mice. Although IL-13-induced miR-21 expression was IL-13Ralpha1 dependent, allergen-induced miR-21 expression was mediated mainly independent of IL-13Ralpha1 and STAT6. Notably, predictive algorithms identified potential direct miR-21 targets among IL-13-regulated lung transcripts, such as IL-12p35 mRNA, which was decreased in IL-13 transgenic mice. Introduction of pre-miR-21 dose dependently inhibited cellular expression of a reporter vector harboring the 3'-untranslated region of IL-12p35. Moreover, mutating miR-21 binding sites in IL-12p35 3'-untranslated region abrogated miR-21-mediated repression. In summary, we have identified a miRNA signature in allergic airway inflammation, which includes miR-21 that modulates IL-12, a molecule germane to Th cell polarization.


COPD continues to cause a heavy health and economic burden both in the United States and around the world. Some of the risk factors for COPD are well-known and include smoking, occupational exposures, air pollution, airway hyperresponsiveness, asthma, and certain genetic variations, although many questions, such as why < 20% of smokers develop significant airway obstruction, remain. Precise definitions of COPD vary and are frequently dependent on an accurate diagnosis of the problem by a physician. These differences in the definition of COPD can have large effects on the estimates of COPD in the population. Furthermore, evidence that COPD represents several different disease processes with potentially different interventions continues to emerge. In most of the world, COPD prevalence and mortality are still increasing and likely will continue to rise in response to increases in smoking, particularly by women and adolescents. Resources aimed at smoking cessation and prevention, COPD education and early detection, and better treatment will be of the most benefit in our continuing efforts against this important cause of morbidity and mortality.


The geographic ranges of ticks and tick-borne pathogens are changing due to global and local environmental (including climatic) changes. In this review we explore current knowledge of the drivers for changes in the ranges of ticks and tick-borne pathogen species and strains via effects on their basic reproduction number (R 0), and the mechanisms of dispersal that allow ticks and tick-borne pathogens to invade suitable environments. Using the expanding geographic distribution of the vectors and agent of Lyme disease as an example we then investigate what could be expected of the diversity of tick-borne pathogens during the process of range expansion, and compare this with what is currently being observed. Lastly we explore how historic population and range expansions and contractions could be reflected in the phylogeography of ticks and tick-borne pathogens seen in recent years, and conclude that combined study of currently changing tick and tick-borne pathogen ranges and diversity, with phylogeographic analysis, may help us better predict future patterns of invasion and diversity.


Successful cancer treatment requires understanding host immune response against tumor cells. PD-1 belongs to the CD28 superfamily of receptors that work as “checkpoints” of immune activation. PD-1 maintains immune self-tolerance to prevent autoimmunity and controls T-cell reaction during infection to prevent excessive tissue damage. Tumor cells that arise from normal tissue acquire mutations that can be targeted by lymphocytes. Accumulating lines of evidence suggest that tumor cells evade host immune attack by expressing physiological PD-1 ligands and stimulating PD-1 on the lymphocytes. Based on this idea, researchers have successfully demonstrated that systemic administration of monoclonal antibodies that inhibit the binding of PD-1 to the ligands reactivated T cells and augmented the anti-cancer immune response. In this review, I summarize the basics of T-cell biology and its regulation by PD-1 and discuss the current understanding and questions about this multifaceted molecule.


To initiate adaptive immunity, dendritic cells (DCs) move from parenchymal tissues to lymphoid organs by migrating along stromal scaffolds that display the glycoprotein podoplanin (PDPN). PDPN is expressed by lymphatic endothelial and fibroblastic reticular cells and promotes blood-lymph separation during development by activating the C-type lectin receptor, CLEC-2, on platelets. Here, we describe a role for CLEC-2 in the morphodynamic behavior and motility of DCs. CLEC-2 deficiency in DCs impaired their entry into lymphatics and trafficking to and within lymph nodes, thereby reducing T cell priming. CLEC-2 engagement of PDPN was necessary for DCs to spread and migrate along stromal surfaces and sufficient to induce membrane protrusions. CLEC-2 activation triggered cell spreading via downregulation of RhoA activity and myosin light-chain phosphorylation and triggered F-actin-rich protrusions via Vav signaling and Rac1 activation. Thus, activation of CLEC-2 by PDPN rearranges the actin cytoskeleton in DCs to promote efficient motility along stromal surfaces.


Although the entire coronary tree is exposed to the atherogenic effect of the systemic risk factors, atherosclerotic lesions form at specific arterial regions, where low and oscillatory endothelial shear stress (ESS) occur. Low ESS modulates endothelial gene expression through complex mechanoreception and mechanotransduction processes, inducing an atherogenic endothelial phenotype and formation of an early atherosclerotic plaque. Each early plaque exhibits an individual natural history of progression, regression, or stabilization, which is dependent not only on the formation and progression of atherosclerosis but also on the vascular remodeling response. Although the pathophysiologic mechanisms involved in the remodeling of the atherosclerotic wall are incompletely understood, the dynamic interplay between local hemodynamic milieu, low ESS in particular, and the biology of the wall is likely to be important. In this review, we explore the molecular, cellular, and vascular processes supporting the role of low ESS in the natural history of coronary atherosclerosis and vascular remodeling and indicate likely mechanisms concerning the different natural history trajectories of individual coronary lesions. Atherosclerotic plaques associated with excessive expansive remodeling evolve to high-risk plaques, because low ESS conditions persist, thereby promoting continued local lipid accumulation, inflammation, oxidative stress, matrix breakdown, and eventually further plaque progression and excessive expansive remodeling. An enhanced understanding of the pathobiologic processes responsible for atherosclerosis and vascular remodeling might allow for early identification of a high-risk coronary plaque and thereby provide a rationale for innovative diagnostic and/or therapeutic strategies for the management of coronary patients and prevention of acute coronary syndromes.


T cell homeostasis is crucial for a functional immune system, as the accumulation of T cells resulting from lack of regulatory T cells or an inability to shut down immune responses can lead to inflammation and autoimmune pathology. Here we show that Blimp-1, a transcriptional repressor that is a 'master regulator' of terminal B cell differentiation, was expressed in a subset of antigen-experienced CD4+ and CD8+ T cells. Mice reconstituted with fetal liver stem cells expressing a mutant Blimp-1 lacking the DNA-binding domain developed a lethal multiorgan inflammatory disease caused by an accumulation of effector and memory T cells. These data identify Blimp-1 as an essential regulator of T cell homeostasis and suggest that Blimp-1 regulates both B cell and T cell differentiation.


BACKGROUND Readmission soon after hospital discharge is an expensive and often preventable event for patients with heart failure. We present a model approved by the National Quality Forum for the purpose of public reporting of hospital-level readmission rates by the Centers for Medicare & Medicaid Services. METHODS AND RESULTS We developed a hierarchical logistic regression model to calculate hospital risk-standardized 30-day all-cause readmission rates for patients hospitalized with heart failure. The model was derived with the use of Medicare claims data for a 2004 cohort and validated with the use of claims and medical record data. The unadjusted readmission rate was 23.6%. The final model included 37 variables, had discrimination ranging from 15% observed 30-day readmission rate in the lowest predictive decile to 37% in the upper decile, and had a c statistic of 0.60. The 25th and 75th percentiles of the risk-standardized readmission rates across 4669 hospitals were 23.1% and 24.0%, with 5th and 95th percentiles of 22.2% and 25.1%, respectively. The odds of all-cause readmission for a hospital 1 standard deviation above average was 1.30 times that of a hospital 1 standard deviation below average. State-level adjusted readmission rates developed with the use of the claims model are similar to rates produced for the same cohort with the use of a medical record model (correlation, 0.97; median difference, 0.06 percentage points). CONCLUSIONS This claims-based model of hospital risk-standardized readmission rates for heart failure patients produces estimates that may serve as surrogates for those derived from a medical record model.


Reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) resets the epigenome to an embryonic-like state. Vitamin C enhances the reprogramming process, but the underlying mechanisms are unclear. Here we show that the histone demethylases Jhdm1a/1b are key effectors of somatic cell reprogramming downstream of vitamin C. We first observed that vitamin C induces H3K36me2/3 demethylation in mouse embryonic fibroblasts in culture and during reprogramming. We then identified Jhdm1a/1b, two known vitamin-C-dependent H3K36 demethylases, as potent regulators of reprogramming through gain- and loss-of-function approaches. Furthermore, we found that Jhdm1b accelerates cell cycle progression and suppresses cell senescence during reprogramming by repressing the Ink4/Arf locus. Jhdm1b also cooperates with Oct4 to activate the microRNA cluster 302/367, an integral component of the pluripotency machinery. Our results therefore reveal a role for H3K36me2/3 in cell fate determination and establish a link between histone demethylases and vitamin-C-induced reprogramming.


Tuberculosis and helminthic infections coexist in many parts of the world, yet the impact of helminth-elicited Th2 responses on the ability of the host to control Mycobacterium tuberculosis (Mtb) infection has not been fully explored. We show that mice infected with the intestinal helminth Nippostrongylus brasiliensis (Nb) exhibit a transitory impairment of resistance to airborne Mtb infection. Furthermore, a second dose of Nb infection substantially increases the bacterial burden in the lungs of co-infected mice. Interestingly, the Th2 response in the co-infected animals did not impair the onset and development of the protective Mtb-specific Th1 cellular immune responses. However, the helminth-induced Th2 environment resulted in the accumulation of alternatively activated macrophages (AAMs) in the lung. Co-infected mice lacking interleukin (IL) 4Rα exhibited improved ability to control Mtb infection, which was accompanied by significantly reduced accumulation of AAMs. Moreover, IL-4Rα(-/-) mice adoptively transferred with wild-type macrophages had a significantly higher Mtb load in their lungs compared with those that received IL-4Rα(-/-) macrophages, suggesting a direct contribution for the IL-4R pathway to the heightened susceptibility of co-infected animals. The Th2 response can thus enhance the intracellular persistence of Mtb, in part by mediating the alternative activation of macrophages via the IL-4Rα signaling pathway.


Cancer has been recognized for thousands of years. Egyptians believed that cancer occurred at the will of the gods. Hippocrates believed human disease resulted from an imbalance of the four humors: blood, phlegm, yellow bile, and black bile with cancer being caused by excess black bile. The lymph theory of cancer replaced the humoral theory and the blastema theory replaced the lymph theory. Rudolph Virchow was the first to recognize that cancer cells like all cells came from other cells and believed chronic irritation caused cancer. At the same time there was a belief that trauma caused cancer, though it never evolved after many experiments inducing trauma. The birth of virology occurred in 1892 when Dimitri Ivanofsky demonstrated that diseased tobacco plants remained infective after filtering their sap through a filter that trapped bacteria. Martinus Beijerinck would call the tiny infective agent a virus and both Dimitri Ivanofsky and Marinus Beijerinck would become the fathers of virology. Not to long thereafter, Payton Rous founded the field of tumor virology in 1911 with his discovery of a transmittable sarcoma of chickens by what would come to be called Rous sarcoma virus or RSV for short. The first identified human tumor virus was the Epstein-Barr virus (EBV), named after Tony Epstein and Yvonne Barr who visualized the virus particles in Burkitt's lymphoma cells by electron microscopy in 1965. Since that time, many viruses have been associated with carcinogenesis including the most studied, human papilloma virus associated with cervical carcinoma, many other anogenital carcinomas, and oropharyngeal carcinoma. The World Health Organization currently estimates that approximately 22% of worldwide cancers are attributable to infectious etiologies, of which viral etiologies is estimated at 15-20%. The field of tumor virology/viral carcinogenesis has not only identified viruses as etiologic agents of human cancers, but has also given molecular insights to all human cancers including the oncogene activation and tumor suppressor gene inactivation.


Many cellular proteins perform their roles within macromolecular assemblies. Hence, an understanding of how these multiprotein complexes form is a fundamental question in cell biology. We developed a translation-controlled pulse-chase system that allows time-resolved isolation of newly forming multiprotein complexes in chemical quantities suitable for biochemical and cell biological analysis. The "pulse" is triggered by an unnatural amino acid, which induces immediate translation of an amber stop codon repressed mRNA encoding the protein of interest with a built-in tag for detection and purification. The "chase" is elicited by stopping translation of this bait via a riboswitch in the respective mRNA. Over the course of validating our method, we discovered a distinct time-resolved assembly step during NPC biogenesis and could directly monitor the spatiotemporal maturation of preribosomes via immunofluorescence detection and purification of a pulse-labeled ribosomal protein. Thus, we provide an innovative strategy to study dynamic protein assembly within cellular networks.


Ever since the first demonstration of their repetitive sequence and unique replication pathway, telomeres have beguiled researchers with how they function in protecting chromosome ends. Of course much has been learned over the years, and we now appreciate that telomeres are comprised of the multimeric protein/DNA shelterin complex and that the formation of t-loops provides protection from DNA damage machinery. Deriving their name from D-loops, t-loops are generated by the insertion of the 3' overhang into telomeric repeats facilitated by the binding of TRF2. Recent studies have uncovered novel forms of chromosome end-structure that may implicate telomere organization in cellular processes beyond its essential role in telomere protection and homeostasis. In particular, we have recently described that t-loops form in a TRF2-dependent manner at interstitial telomere repeat sequences, which we termed interstitial telomere loops (ITLs). These structures are also dependent on association of lamin A/C, a canonical component of the nucleoskeleton that is mutated in myriad human diseases, including human segmental progeroid syndromes. Since ITLs are associated with telomere stability and require functional lamin A/C, our study suggests a mechanistic link between cellular aging (replicative senescence induced by telomere shortening) and organismal aging (modeled by Hutchinson Gilford Progeria Syndrome). Here we speculate on other potential ramifications of ITL formation, from gene expression to genome stability to chromosome structure.


Chromatin remodelers regulate genes by organizing nucleosomes around promoters, but their individual contributions are obfuscated by the complex in vivo milieu of factor redundancy and indirect effects. Genome-wide reconstitution of promoter nucleosome organization with purified proteins resolves this problem and is therefore a critical goal. Here, we reconstitute four stages of nucleosome architecture using purified components: yeast genomic DNA, histones, sequence-specific Abf1/Reb1, and remodelers RSC, ISW2, INO80, and ISW1a. We identify direct, specific, and sufficient contributions that in vivo observations validate. First, RSC clears promoters by translating poly(dA:dT) into directional nucleosome removal. Second, partial redundancy is recapitulated where INO80 alone, or ISW2 at Abf1/Reb1sites, positions +1 nucleosomes. Third, INO80 and ISW2 each align downstream nucleosomal arrays. Fourth, ISW1a tightens the spacing to canonical repeat lengths. Such a minimal set of rules and proteins establishes core mechanisms by which promoter chromatin architecture arises through a blend of redundancy and specialization.


A current paradigm states that monocytes circulate freely and patrol blood vessels but differentiate irreversibly into dendritic cells (DCs) or macrophages upon tissue entry. Here we show that bona fide undifferentiated monocytes reside in the spleen and outnumber their equivalents in circulation. The reservoir monocytes assemble in clusters in the cords of the subcapsular red pulp and are distinct from macrophages and DCs. In response to ischemic myocardial injury, splenic monocytes increase their motility, exit the spleen en masse, accumulate in injured tissue, and participate in wound healing. These observations uncover a role for the spleen as a site for storage and rapid deployment of monocytes and identify splenic monocytes as a resource that the body exploits to regulate inflammation.


The dentate gyrus has an important role in learning and memory, and adult neurogenesis in the subgranular zone of the dentate gyrus may play a role in the acquisition of new memories. The homeobox gene Prox1 is expressed in the dentate gyrus during embryonic development and adult neurogenesis. Here we show that Prox1 is necessary for the maturation of granule cells in the dentate gyrus during development and for the maintenance of intermediate progenitors during adult neurogenesis. We also demonstrate that Prox1-expressing intermediate progenitors are required for adult neural stem cell self-maintenance in the subgranular zone; thus, we have identified a previously unknown non-cell autonomous regulatory feedback mechanism that controls adult neurogenesis in this region of the mammalian brain. Finally, we show that the ectopic expression of Prox1 induces premature differentiation of neural stem cells.


Human mesenchymal stem cells (hMSCs) have been widely studied as a source of primary adult stem cells for cell therapy because of their multidifferentiation potential; however, the growth arrest (also known as "premature senescence") often found in hMSCs cultured in vitro has been a major obstacle to the in-depth characterization of these cells. In addition, the inability to maintain constant cell growth hampers the development of additional genetic modifications aimed at achieving desired levels of differentiation to specific tissues; however, the molecular mechanisms that govern this phenomenon remain unclear, with the exception of a few studies demonstrating that induction of p16INK4a is responsible for this senescence-like event. Here, we observed that the premature growth arrest in hMSCs occurs in parallel with the induction of p16INK4a, following abrogation of inhibitory phosphorylation of retinoblastoma protein. These stress responses were concurrent with increased formation of reactive oxygen species (ROSs) from mitochondria and increased p38 mitogen-activated protein kinase (MAPK) activity. The introduction of Wip1 (wild-type p53 inducible phosphatase-1), a well-studied stress modulator, significantly lowered p16INK4a expression and led to p38 MAPK inactivation, although it failed to affect the levels of ROSs. Moreover, the suppression of stress responses by Wip1 apparently extended the life span of hMSCs, compared with control conditions, while maintaining their multilineage differentiation potential. Based on these results, we suggest that senescent growth arrest in hMSCs may result from activation of stress signaling pathways and consequent onset of stress responses, due in part to ROS production during prolonged in vitro culture.


Evolutionary considerations suggest aging is caused not by active gene programming but by evolved limitations in somatic maintenance, resulting in a build-up of damage. Ecological factors such as hazard rates and food availability influence the trade-offs between investing in growth, reproduction, and somatic survival, explaining why species evolved different life spans and why aging rate can sometimes be altered, for example, by dietary restriction. To understand the cell and molecular basis of aging is to unravel the multiplicity of mechanisms causing damage to accumulate and the complex array of systems working to keep damage at bay.


Mononuclear phagocytes, including monocytes, macrophages, and dendritic cells, contribute to tissue integrity as well as to innate and adaptive immune defense. Emerging evidence for labor division indicates that manipulation of these cells could bear therapeutic potential. However, specific ontogenies of individual populations and the overall functional organization of this cellular network are not well defined. Here we report a fate-mapping study of the murine monocyte and macrophage compartment taking advantage of constitutive and conditional CX(3)CR1 promoter-driven Cre recombinase expression. We have demonstrated that major tissue-resident macrophage populations, including liver Kupffer cells and lung alveolar, splenic, and peritoneal macrophages, are established prior to birth and maintain themselves subsequently during adulthood independent of replenishment by blood monocytes. Furthermore, we have established that short-lived Ly6C(+) monocytes constitute obligatory steady-state precursors of blood-resident Ly6C(-) cells and that the abundance of Ly6C(+) blood monocytes dynamically controls the circulation lifespan of their progeny.


BACKGROUND Dealing with heterogeneity in meta-analyses is often tricky, and there is only limited advice for authors on what to do. We investigated how authors addressed different degrees of heterogeneity, in particular whether they used a fixed effect model, which assumes that all the included studies are estimating the same true effect, or a random effects model where this is not assumed. METHODS We sampled randomly 60 Cochrane reviews from 2008, which presented a result in its first meta-analysis with substantial heterogeneity (I2 greater than 50%, i.e. more than 50% of the variation is due to heterogeneity rather than chance). We extracted information on choice of statistical model, how the authors had handled the heterogeneity, and assessed the methodological quality of the reviews in relation to this. RESULTS The distribution of heterogeneity was rather uniform in the whole I2 interval, 50-100%. A fixed effect model was used in 33 reviews (55%), but there was no correlation between I2 and choice of model (P = 0.79). We considered that 20 reviews (33%), 16 of which had used a fixed effect model, had major problems. The most common problems were: use of a fixed effect model and lack of rationale for choice of that model, lack of comment on even severe heterogeneity and of reservations and explanations of its likely causes. The problematic reviews had significantly fewer included trials than other reviews (4.3 vs. 8.0, P = 0.024). The problems became less pronounced with time, as those reviews that were most recently updated more often used a random effects model. CONCLUSION One-third of Cochrane reviews with substantial heterogeneity had major problems in relation to their handling of heterogeneity. More attention is needed to this issue, as the problems we identified can be essential for the conclusions of the reviews.


Polarized transport in neurons is fundamental for the formation of neuronal circuitry. A motor domain-containing truncated KIF5 (a kinesin-1) recognizes axonal microtubules, which are enriched in EB1 binding sites, and selectively accumulates at the tips of axons. However, it remains unknown what cue KIF5 recognizes to result in this selective accumulation. We found that axonal microtubules were preferentially stained by the anti-GTP-tubulin antibody hMB11. Super-resolution microscopy combined with EM immunocytochemistry revealed that hMB11 was localized at KIF5 attachment sites. In addition, EB1, which binds preferentially to guanylyl-methylene-diphosphate (GMPCPP) microtubules in vitro, recognized hMB11 binding sites on axonal microtubules. Further, expression of hMB11 antibody in neurons disrupted the selective accumulation of truncated KIF5 in the axon tips. In vitro studies revealed approximately threefold stronger binding of KIF5 motor head to GMPCPP microtubules than to GDP microtubules. Collectively, these data suggest that the abundance of GTP-tubulin in axonal microtubules may underlie selective KIF5 localization and polarized axonal vesicular transport.


The pancreas is an organ containing two distinct populations of cells, the exocrine cells that secrete enzymes into the digestive tract, and the endocrine cells that secrete hormones into the bloodstream. It arises from the endoderm as a dorsal and a ventral bud which fuse together to form the single organ. Mammals, birds, reptiles and amphibians have a pancreas with similar histology and mode of development, while in some fish, the islet cells are segregated as Brockmann bodies. Invertebrates do not have a pancreas, but comparable endocrine cells may be found in the gut or the brain. The early pancreatic bud shows uniform expression of the homeobox gene IPF-1 (also known as IDX-1, STF-1 or PDX), which when mutated to inactivity leads to total absence of the organ. The occurrence of heterotopic pancreas in the embryo, and also the metaplasias that can be displayed by a regenerating pancreas in the adult, both suggest that only a few gene products distinguish the pancreatic cell state from that of the surrounding tissues of duodenum, gall bladder and liver. In the developing pancreatic buds, the endocrine cells start to differentiate before the exocrine cells, and co-expression of different hormones by the same cell is often observed at early stages. Although pancreatic endocrine cells produce many gene products also characteristic of neurons, evidence from in vitro cultures and from quailchick grafts shows that they are of endogenous and not of neural crest origin. Observational studies suggest strongly that both endocrine and exocrine cells arise from the same endodermal rudiment. Development of the pancreas in embryonic life requires a trophic stimulus from the associated mesenchyme. In postnatal life, all cell types in the pancreas continue to grow. Destruction of acinar tissue by duct ligation or ethionine treatment is followed by rapid regeneration. Surgical removal of parts of the pancreas is followed by moderate but incomplete regeneration of both acini and islets. Poisoning with alloxan or streptozotocin can lead to permanent depletion of beta cells. Although the cell kinetics of the pancreas are not understood, it seems likely that there is a continuous slow turnover of cells, fed from a stem cells population in the ducts, and that the controls on the production rate of each cell type are local rather than systemic.


OBJECTIVE To summarise the long term efficacy of anti-obesity drugs in reducing weight and improving health status. DESIGN Updated meta-analysis of randomised trials. DATA SOURCES Medline, Embase, the Cochrane controlled trials register, the Current Science meta-register of controlled trials, and reference lists of identified articles. All data sources were searched from December 2002 (end date of last search) to December 2006. STUDIES REVIEWED Double blind randomised placebo controlled trials of approved anti-obesity drugs used in adults (age over 18) for one year or longer. RESULTS 30 trials of one to four years' duration met the inclusion criteria: 16 orlistat (n=10 631 participants), 10 sibutramine (n=2623), and four rimonabant (n=6365). Of these, 14 trials were new and 16 had previously been identified. Attrition rates averaged 30-40%. Compared with placebo, orlistat reduced weight by 2.9 kg (95% confidence interval 2.5 kg to 3.2 kg), sibutramine by 4.2 kg (3.6 kg to 4.7 kg), and rimonabant by 4.7 kg (4.1 kg to 5.3 kg). Patients receiving active drug treatment were significantly more likely to achieve 5% and 10% weight loss thresholds. Orlistat reduced the incidence of diabetes and improved concentrations of total cholesterol and low density lipoprotein cholesterol, blood pressure, and glycaemic control in patients with diabetes but increased rates of gastrointestinal side effects and slightly lowered concentrations of high density lipoprotein. Sibutramine improved [corrected] concentrations of high density lipoprotein cholesterol and triglycerides [corrected] Rimonabant improved concentrations of high density lipoprotein cholesterol and triglycerides, blood pressure, and glycaemic control in patients with diabetes but increased the risk of mood disorders. CONCLUSIONS Orlistat, sibutramine, and rimonabant modestly reduce weight, have differing effects on cardiovascular risk profiles, and have specific adverse effects.


TRPM7 encodes a Ca2+-permeable nonselective cation channel with kinase activity. TRPM7 has been implicated in control of cell adhesion and migration, but whether TRPM7 activity contributes to cancer progression has not been established. Here we report that high levels of TRPM7 expression independently predict poor outcome in breast cancer patients and that it is functionally required for metastasis formation in a mouse xenograft model of human breast cancer. Mechanistic investigation revealed that TRPM7 regulated myosin II-based cellular tension, thereby modifying focal adhesion number, cell-cell adhesion and polarized cell movement. Our findings therefore suggest that TRPM7 is part of a mechanosensory complex adopted by cancer cells to drive metastasis formation.


Frailty has long been considered synonymous with disability and comorbidity, to be highly prevalent in old age and to confer a high risk for falls, hospitalization and mortality. However, it is becoming recognized that frailty may be a distinct clinical syndrome with a biological basis. The frailty process appears to be a transitional state in the dynamic progression from robustness to functional decline. During this process, total physiological reserves decrease and become less likely to be sufficient for the maintenance and repair of the ageing body. Central to the clinical concept of frailty is that no single altered system alone defines it, but that multiple systems are involved. Clinical consensus regarding the phenotype which constitutes frailty, drawing upon the opinions of numerous authors, shows the characteristics to include wasting (loss of both muscle mass and strength and weight loss), loss of endurance, decreased balance and mobility, slowed performance, relative inactivity and, potentially, decreased cognitive function. Frailty is a distinct entity easily recognized by clinicians, with multiple manifestations and with no single symptom being sufficient or essential in its presentation. Manifestations include appearance (consistent or not with age), nutritional status (thin, weight loss), subjective health rating (health perception), performance (cognition, fatigue), sensory/physical impairments (vision, hearing, strength) and current care (medication, hospital). Although the early stages of the frailty process may be clinically silent, when depleted reserves reach an aggregate threshold leading to serious vulnerability, the syndrome may become detectable by looking at clinical, functional, behavioral and biological markers. Thus, a better understanding of these clinical changes and their underlying mechanisms, beginning in the pre-frail state, may confirm the impression held by many geriatricians that increasing frailty is distinguishable from ageing and in consequence is potentially reversible. We therefore provide an update of the physiopathology and clinical and biological characteristics of the frailty process and speculate on possible preventative approaches.


BACKGROUND Preclinical diastolic dysfunction (PDD) has been defined as subjects with normal systolic function, diastolic dysfunction but no symptoms of heart failure (HF). The clinical phenotype and natural history of the syndrome remains poorly defined. This study's objective was to determine the clinical phenotype and progression to HF in a group of patients with normal systolic function and moderate or severe diastolic dysfunction as determinate by Doppler criteria without any clinical diagnosis of HF according to the Framingham criteria or any symptoms of HF, specifically dyspnoea, oedema or fatigue at the time of echocardiography. METHODS The authors used resources of the Mayo Clinic echocardiography database to consecutively select among patients who had an echocardiogram in 2005, a cohort with moderate or severe diastolic dysfunction by Doppler criteria and EF >or=50%. Patients could not have a diagnosis of HF, or any HF symptoms-specifically dyspnoea, oedema or fatigue-at the time of echocardiography; nor grade 3 or greater valvular dysfunction (except tricuspid valve). A total of 82 patients had their medical chart reviewed. Primary endpoint was the time to the development of (1) HF according to the Framingham criteria or (2) any symptoms of dyspnoea, oedema or fatigue. RESULTS The mean age of the cohort of PDD subjects was 69+/-10 years with a female (67%) preponderance. Presence of hypertension was 76%, coronary artery disease was 29%, paroxysmal atrial fibrillation was 26%, estimated creatinine clearance <60 ml/min was 51%. The 2-year cumulative probability of development of HF according to the Framingham criteria was 1.9%; however, the 2-year cumulative probability of development of any symptoms was 31.1%. The 2-year cumulative probability for cardiac hospitalisation was 21.2%. Peripheral vascular disease and hypertension were independently associated with increased likelihood for the development of symptoms. CONCLUSION The study demonstrates that hypertension, hyperlipidaemia, CAD and renal dysfunction are prevalent in patients with PDD. More importantly, although the progression to the development of clinical HF over 2 years was low, there was a moderate degree of progression to development of symptoms and cardiac hospitalisations over 2 years. Based on the finding that only PVD and hypertension were independently associated with the progression to the development of symptoms in subject with PDD, the authors speculate that ventricular-arterial interaction may be important to the progression of diastolic dysfunction to the development of symptoms.


Considerable evidence supports the effectiveness of aspirin for chemoprevention of colorectal cancer (CRC) in addition to its well-established benefits in the prevention of vascular disease. Epidemiologic studies have consistently observed an inverse association between aspirin use and risk of CRC. A recent pooled analysis of a long-term posttrial follow-up of nearly 14,000 patients from four randomized, cardiovascular disease prevention trials showed that daily aspirin treatment for about five years was associated with a 34% reduction in 20-year CRC mortality. A separate metaanalysis of nearly 3,000 patients with a history of colorectal adenoma or cancer in four randomized adenoma prevention trials showed that aspirin reduced the occurrence of advanced adenomas by 28% and any adenoma by 17%. Aspirin has also been shown to be beneficial in a clinical trial of patients with Lynch syndrome, a hereditary CRC syndrome; in those treated with aspirin for at least two years, there was a 50% or more reduction in the risk of CRC commencing five years after randomization and after aspirin had been discontinued. A few observational studies have shown an increase in survival among patients with CRC who use aspirin. Taken together, these findings strengthen the case for consideration of long-term aspirin use in CRC prevention. Despite these compelling data, there is a lack of consensus about the balance of risks and benefits associated with long-term aspirin use, particularly in low-risk populations. The optimal dose to use for cancer prevention and the precise mechanism underlying aspirin's anticancer effect require further investigation.


Although chemokines are well established to function in immunity and endothelial cell activation and proliferation, a rapidly growing literature suggests that CXC Chemokine receptors CXCR3, CXCR4 and CXCR7 are critical in the development and progression of solid tumors. The effect of these chemokine receptors in tumorigenesis is mediated via interactions with shared ligands I-TAC (CXCL11) and SDF-1 (CXCL12). Over the last decade, CXCR4 has been extensively reported to be overexpressed in most human solid tumors and has earned considerable attention toward elucidating its role in cancer metastasis. To enrich the existing armamentarium of anti-cancerous agents, many inhibitors of CXCL12-CXCR4 axis have emerged as additional or alternative agents for neo-adjuvant treatments and even many of them are in preclinical and clinical stages of their development. However, the discovery of CXCR7 as another receptor for CXCL12 with rather high binding affinity and recent reports about its involvement in cancer progression, has questioned the potential of "selective blockade" of CXCR4 as cancer chemotherapeutics. Interestingly, CXCR7 can also bind another chemokine CXCL11, which is an established ligand for CXCR3. Recent reports have documented that CXCR3 and their ligands are overexpressed in different solid tumors and regulate tumor growth and metastasis. Therefore, it is important to consider the interactions and crosstalk between these three chemokine receptors and their ligand mediated signaling cascades for the development of effective anti-cancer therapies. Emerging evidence also indicates that these receptors are differentially expressed in tumor endothelial cells as well as in cancer stem cells, suggesting their direct role in regulating tumor angiogenesis and metastasis. In this review, we will focus on the signals mediated by this receptor trio via their shared ligands and their role in tumor growth and progression.


We have used microinjection and time-lapse video microscopy to study the role of cyclin A in mitosis. We have injected purified, active cyclin A/cyclin-dependent kinase 2 (CDK2) into synchronized cells at specific points in the cell cycle and assayed its effect on cell division. We find that cyclin A/CDK2 will drive G2 phase cells into mitosis within 30 min of microinjection, up to 4 h before control cells enter mitosis. Often this premature mitosis is abnormal; the chromosomes do not completely condense and daughter cells fuse. Remarkably, microinjecting cyclin A/CDK2 into S phase cells has no effect on progress through the following G2 phase or mitosis. In complementary experiments we have microinjected the amino terminus of p21Cip1/Waf1/Sdi1 (p21N) into cells to inhibit cyclin A/CDK2 activity. We find that p21N will prevent S phase or G2 phase cells from entering mitosis, and will cause early prophase cells to return to interphase. These results suggest that cyclin A/CDK2 is a rate-limiting component required for entry into mitosis, and for progress through mitosis until late prophase. They also suggest that cyclin A/CDK2 may be the target of the recently described prophase checkpoint.


The cloning of animals from adult cells has demonstrated that the developmental state of adult cells can be reprogrammed into that of embryonic cells by uncharacterized factors within the oocyte. More recently, transcription factors have been identified that can induce pluripotency in somatic cells without the use of oocytes, generating induced pluripotent stem (iPS) cells. iPS cells provide a unique platform to dissect the molecular mechanisms that underlie epigenetic reprogramming. Moreover, iPS cells can teach us about principles of normal development and disease, and might ultimately facilitate the treatment of patients by custom-tailored cell therapy.


CONTEXT Conflicting information exists about whether sex differences modulate short-term mortality following acute coronary syndromes (ACS). OBJECTIVES To investigate the relationship between sex and 30-day mortality in ACS, and to determine whether this relationship was modified by clinical syndrome or coronary anatomy using a large database across the spectrum of ACS and adjusting for potentially confounding clinical covariates. DESIGN, SETTING, AND PARTICIPANTS A convenience sample of patients pooled from 11 independent, international, randomized ACS clinical trials between 1993 and 2006 whose databases are maintained at the Duke Clinical Research Institute, Durham, North Carolina. Of 136 247 patients, 38 048 (28%) were women; 102 004 (26% women) with ST-segment elevation myocardial infarction (STEMI), 14 466 (29% women) with non-STEMI (NSTEMI), and 19 777 (40% women) with unstable angina. MAIN OUTCOME MEASURE Thirty-day mortality following ACS. RESULTS Thirty-day mortality was 9.6% in women and 5.3% in men (odds ratio [OR], 1.91; 95% confidence interval [CI], 1.83-2.00). After multivariable adjustment, mortality was not significantly different between women and men (adjusted OR, 1.06; 95% CI, 0.99-1.15). A significant sex by type of ACS interaction was demonstrated (P < .001). In STEMI, 30-day mortality was higher among women (adjusted OR, 1.15; 95% CI, 1.06-1.24), whereas in NSTEMI (adjusted OR, 0.77; 95% CI, 0.63-0.95) and unstable angina, mortality was lower among women (adjusted OR, 0.55; 95% CI, 0.43-0.70). In a cohort of 35 128 patients with angiographic data, women more often had nonobstructive (15% vs 8%) and less often had 2-vessel (25% vs 28%) and 3-vessel (23% vs 26%) coronary disease, regardless of ACS type. After additional adjustment for angiographic disease severity, 30-day mortality among women was not significantly different than men, regardless of ACS type. The relationship between sex and 30-day mortality was similar across the levels of angiographic disease severity (P for interaction = .70). CONCLUSIONS Sex-based differences existed in 30-day mortality among patients with ACS and vary depending on clinical presentation. However, these differences appear to be largely explained by clinical differences at presentation and severity of angiographically documented disease.


Hepatitis B X protein (HBx) plays an essential role in the hepatitis B virus (HBV) replication cycle, but the function of HBx has been elusive until recently. It was recently shown that transcription from the HBV genome (covalently-closed circular DNA, cccDNA) is inhibited by the structural maintenance of chromosome 5/6 complex (Smc5/6), and that a key function of HBx is to redirect the DNA-damage binding protein 1 (DDB1) E3 ubiquitin ligase to target this complex for degradation. By doing so, HBx alleviates transcriptional repression by Smc5/6 and stimulates HBV gene expression. In this review, we discuss in detail how the interplay between HBx and Smc5/6 was identified and characterized. We also discuss what is known regarding the repression of cccDNA transcription by Smc5/6, the timing of HBx expression, and the potential role of HBx in promoting hepatocellular carcinoma (HCC).


Background. Hepatitis C virus (HCV) is an underappreciated cause of pediatric liver disease, most frequently acquired by vertical transmission (VT). Current guidelines that include the option of screening infants for HCV RNA at 1-2 months are based on data prior to current real-time polymerase chain reaction (PCR)-based testing. Previous studies have demonstrated VT rates of 4%-15% and an association with high maternal viral load. We evaluated HCV RNA in infants with HCV VT and assessed maternal risk factors in a prospective cohort in Cairo, Egypt. Methods. Pregnant women were screened for HCV from December 2012 to March 2014. For those with HCV viremia, their infants were tested at 12 months for HCV RNA using real-time PCR. Maternal risk factors assessed for HCV VT association included HCV RNA levels, mode of delivery, and maternal IL28B genotype. Results. Of 2514 women screened, a total of 54 women were viremic (2.1%) and delivered 56 infants. Of those, 51 infants of 49 women were tested at 12 months of age. Only 7 infants were viremic, with an HCV VT rate of 14.3% (7 of 49). Median HCV RNA in the infants was 2100 IU/mL. None of the maternal risk factors analyzed were associated with transmission. Conclusions. In Egypt where HCV is highly endemic, we observed an overall 12-month HCV VT rate of 14.3%. Further studies should focus on better identification of pregnant women more likely to vertically transmit HCV and earlier testing of infants to identify those likely to develop chronicity.


BACKGROUND Several studies have shown that testing for high-risk human papillomavirus (HPV) types results in an improved sensitivity for CIN2+, compared with cytology, although with a somewhat lower specificity. METHODS We obtained follow-up results, with at least one smear after participation in the HART study, which compared HPV testing (HC-II) with cytology as a primary screening modality. RESULTS With a median follow-up of 6 years, 42 additional cases of CIN2+ were identified; women who were HPV positive at baseline were more likely to develop CIN2+ than those who were HPV negative (hazard ratio (HR) 17.2; 95% confidence interval (CI) (9.3-31.6)) and the risk increased with increasing viral load. Compared with HPV-negative women (relative light unit (RLU) <1), the HR (95% CI) was 5.4 (1.6, 18.2) for 1-10 RLU and 25.5 (13.6, 47.9) for RLU > or = 10. Positive cytology (borderline or worse compared with negative) was also predictive of developing CIN2, although to a lesser extent (HR 8.7; 95% CI (4.5-17.1)). Only one case of CIN3 and three cases of CIN2 were found in women who showed a positive cytology result but were HPV negative at baseline. CONCLUSION After 5 years of follow-up, CIN2+ occurred in 0.23% of women who were HPV negative at baseline compared with 0.48% of women who showed a negative cytology result, indicating a much longer low-risk interval for CIN2+ after HPV testing.


BACKGROUND Blindness due to trachoma is avoidable through Surgery, Antibiotics, Facial hygiene and Environmental improvements (SAFE). Recent surveys have shown trachoma to be a serious cause of blindness in Southern Sudan. We conducted this survey in Ayod County of Jonglei State to estimate the need for intervention activities to eliminate blinding trachoma. METHODOLOGY AND FINDINGS A cross-sectional two-stage cluster random survey was conducted in November 2006. All residents of selected households were clinically assessed for trachoma using the World Health Organization (WHO) simplified grading scheme. A total of 2,335 people from 392 households were examined, of whom 1,107 were over 14 years of age. Prevalence of signs of active trachoma in children 1-9 years of age was: trachomatous inflammation follicular (TF) = 80.1% (95% confidence interval [CI], 73.9-86.3); trachomatous inflammation intense (TI) = 60.7% (95% CI, 54.6-66.8); and TF and/or TI (active trachoma) = 88.3% (95% CI, 83.7-92.9). Prevalence of trachomatous trichiasis (TT) was 14.6% (95% CI, 10.9-18.3) in adults over 14 years of age; 2.9% (95% CI, 0.4-5.3) in children 1-14 years of age; and 8.4% (95% CI, 5.5-11.3) overall. The prevalence of corneal opacity in persons over 14 years of age with TT was 6.4% (95% CI, 4.5-8.3). No statistically significant difference was observed in the prevalence of trachoma signs between genders. Trachoma affected almost all households surveyed: 384/392 (98.0%) had at least one person with active trachoma and 130 (33.2%) had at least one person with trichiasis. CONCLUSIONS Trachoma is an unnecessary public health problem in Ayod. The high prevalence of active trachoma and trichiasis confirms the severe burden of blinding trachoma found in other post-conflict areas of Southern Sudan. Based on WHO recommended thresholds, all aspects of the SAFE strategy are indicated to eliminate blinding trachoma in Ayod.


The E2F family of transcription factors are essential for the regulation of genes required for appropriate progression through the cell cycle. Five members of the E2F family have been previously reported, namely E2F1-5. All five are key elements in transcriptional regulation of essential genes, and they can be divided into two functional groups, those that induce S-phase progression when overexpressed in quiescent cells (E2Fs 1–3), and those that do not (E2Fs 4–5). Here, we describe the identification of a novel member of this family, which we refer to as E2F-6. E2F-6 shares significant homology with E2Fs 1–5, especially within the DNA binding, heterodimerization and marked box domains. Unlike E2Fs 1–5, E2F-6 lacks a transactivation and a pocket protein binding domain, hence, forms a unique third group within the E2F family. E2F-6 is a nuclear protein that can form heterodimers with the DP proteins (both DP-1 and DP-2) in vitro and in vivo. Our results show that the complex formed between E2F-6 and the DP proteins, possesses high DNA binding activity, displaying a preference for a TTTCCCGC E2F recognition site, which is slightly different to the E2F consensus site derived from the E2 promoter (TTTCGCGC). In contrast to the other members of the E2F family, ectopic expression of E2F-6 inhibits transcription from promoters possessing E2F recognition sites rather than activating transcription. In addition, overexpression of E2F-6 suppresses the transactivational effects of co-expression of E2F-1 and DP-1. The inhibitory effect of E2F-6 is dependent on its DNA binding activity and its ability to form heterodimers with the DPs. Interestingly, ectopic expression of E2F-6 leads to accumulation of cells in S-phase. Our data suggest that E2F-6 expression delays the exit from S-phase rather than inducing S-phase, which further emphasizes the functional difference between E2F-6 and the previously known E2F family members.


This review will reconsider the current paradigm for understanding the critical, final steps in the progression of atherosclerotic lesions. That scheme, largely an outgrowth of observations of autopsy tissues by Davies and colleagues,1 2 asserts that the cause of death in atherosclerotic coronary artery disease is rupture of an advanced atherosclerotic lesion. Although this assumption may be partially true, recent autopsy studies suggest that it is incomplete. To reconsider this paradigm, we reexamined the morphological classification scheme for lesions proposed by the American Heart Association (AHA).3 4 This scheme is difficult to use for 2 reasons. First, it uses a very long list of roman numerals modified by letter codes that are difficult to remember. Second, it implies an orderly, linear pattern of lesion progression. This tends to be ambiguous, because it is not clear whether there is a single sequence of events during the progression of all lesions. We have therefore tried to devise a simpler classification scheme that is consistent with the AHA categories but is easier to use, able to deal with a wide array of morphological variations, and not overly burdened by mechanistic implications. The current paradigm is based on the belief that type IV lesions, or “atheromas,” described by the AHA are stable because the fatty, necrotic core is contained by a smooth muscle cell–rich fibrous cap. Virchow’s analysis5 in 1858 pointed out that historically, the term “atheroma” refers to a dermal cyst (“Grutzbalg”), a fatty …


Microbial survival in a host is usually dependent on the ability of a pathogen to undergo changes that promote escape from host defense mechanisms. The human-pathogenic fungus Cryptococcus neoformans undergoes phenotypic switching in vivo that promotes persistence in tissue. By microarray and real-time PCR analyses, the allergen 1 gene (ALL1) was found to be downregulated in the hypervirulent mucoid switch variant, both during logarithmic growth and during intracellular growth in macrophages. The ALL1 gene encodes a small cytoplasmic protein that is involved in capsule formation. Growth of an all1Delta gene deletion mutant was normal. Similar to cells of the mucoid switch variant, all1Delta cells produced a larger polysaccharide capsule than cells of the smooth parent and the complemented strain produced, and the enlarged capsule inhibited macrophage phagocytosis. The mutant exhibited a modest defect in capsule induction compared to all of the other variants. In animal models the phenotype of the all1Delta mutant mimicked the hypervirulent phenotype of the mucoid switch variant, which is characterized by decreased host survival and elevated intracranial pressure. Decreased survival is likely the result of both an ineffective cell-mediated immune response and impaired phagocytosis by macrophages. Consequently, we concluded that, unlike loss of most virulence-associated genes, where loss of gene function results in attenuated virulence, loss of the ALL1 gene enhances virulence by altering the host-pathogen interaction and thereby impairing clearance. Our data identified the first cryptococcal gene associated with elevated intracranial pressure and support the hypothesis that an environmental opportunistic pathogen has modified its virulence in vivo by epigenetic downregulation of gene function.


Ionotropic glutamate receptors (iGluRs) constitute a family of ligand-gated ion channels that are essential for mediating fast synaptic transmission in the central nervous system. These receptors play an important role for the development and function of the nervous system, and are essential in learning and memory. However, iGluRs are also implicated in or have causal roles for several brain disorders, e.g. epilepsy, Alzheimer's disease, Parkinson's disease and schizophrenia. Their involvement in neurological diseases has stimulated widespread interest in their structure and function. Since the first publication in 1998 of the structure of a recombinant soluble protein comprising the ligand-binding domain of GluA2 extensive studies have afforded numerous crystal structures of wildtype and mutant proteins including different ligands. The structural information obtained combined with functional data have led to models for receptor activation and desensitization by agonists, inhibition by antagonists and block of desensitization by positive allosteric modulators. Furthermore, the structural and functional studies have formed a powerful platform for the design of new selective compounds.


Store-operated Ca(2+) entry (SOCE) is the principal Ca(2+) entry mechanism in nonexcitable cells. Stromal-interaction molecule 1 (STIM1) is an endoplasmic reticulum Ca(2+) sensor that triggers SOCE activation. However, the role of STIM1 in regulating cancer progression remains controversial and its clinical relevance is unclear. Here we show that STIM1-dependent signaling is important for cervical cancer cell proliferation, migration, and angiogenesis. STIM1 overexpression in tumor tissue is noted in 71% cases of early-stage cervical cancer. In tumor tissues, the level of STIM1 expression is significantly associated with the risk of metastasis and survival. EGF-stimulated cancer cell migration requires STIM1 expression and EGF increases the interaction between STIM1 and Orai1 in juxta-membrane areas, and thus induces Ca(2+) influx. STIM1 involves the activation of Ca(2+)-regulated protease calpain, as well as Ca(2+)-regulated cytoplasmic kinase Pyk2, which regulate the focal-adhesion dynamics of migratory cervical cancer cells. Because of an increase of p21 protein levels and a decrease of Cdc25C protein levels, STIM1-silencing in cervical cancer cells significantly inhibits cell proliferation by arresting the cell cycle at the S and G2/M phases. STIM1 also regulates the production of VEGF in cervical cancer cells. Interference with STIM1 expression or blockade of SOCE activity inhibits tumor angiogenesis and growth in animal models, confirming the crucial role of STIM1-mediated Ca(2+) influx in aggravating tumor development in vivo. These results make STIM1-dependent signaling an attractive target for therapeutic intervention.


Both eukaryotic and prokaryotic cells release small, phospholipid-enclosed vesicles into their environment. Why do cells release vesicles? Initial studies showed that eukaryotic vesicles are used to remove obsolete cellular molecules. Although this release of vesicles is beneficial to the cell, the vesicles can also be a danger to their environment, for instance in blood, where vesicles can provide a surface supporting coagulation. Evidence is accumulating that vesicles are cargo containers used by eukaryotic cells to exchange biomolecules as transmembrane receptors and genetic information. Because also bacteria communicate to each other via extracellular vesicles, the intercellular communication via extracellular cargo carriers seems to be conserved throughout evolution, and therefore vesicles are likely to be a highly efficient, robust, and economic manner of exchanging information between cells. Furthermore, vesicles protect cells from accumulation of waste or drugs, they contribute to physiology and pathology, and they have a myriad of potential clinical applications, ranging from biomarkers to anticancer therapy. Because vesicles may pass the blood-brain barrier, they can perhaps even be considered naturally occurring liposomes. Unfortunately, pathways of vesicle release and vesicles themselves are also being used by tumors and infectious diseases to facilitate spreading, and to escape from immune surveillance. In this review, the different types, nomenclature, functions, and clinical relevance of vesicles will be discussed.


BACKGROUND It is not yet known whether DNA methylation levels can be used to accurately predict age across a broad spectrum of human tissues and cell types, nor whether the resulting age prediction is a biologically meaningful measure. RESULTS I developed a multi-tissue predictor of age that allows one to estimate the DNA methylation age of most tissues and cell types. The predictor, which is freely available, was developed using 8,000 samples from 82 Illumina DNA methylation array datasets, encompassing 51 healthy tissues and cell types. I found that DNA methylation age has the following properties: first, it is close to zero for embryonic and induced pluripotent stem cells; second, it correlates with cell passage number; third, it gives rise to a highly heritable measure of age acceleration; and, fourth, it is applicable to chimpanzee tissues. Analysis of 6,000 cancer samples from 32 datasets showed that all of the considered 20 cancer types exhibit significant age acceleration, with an average of 36 years. Low age-acceleration of cancer tissue is associated with a high number of somatic mutations and TP53 mutations, while mutations in steroid receptors greatly accelerate DNA methylation age in breast cancer. Finally, I characterize the 353 CpG sites that together form an aging clock in terms of chromatin states and tissue variance. CONCLUSIONS I propose that DNA methylation age measures the cumulative effect of an epigenetic maintenance system. This novel epigenetic clock can be used to address a host of questions in developmental biology, cancer and aging research.


Coronary heart disease (CHD) patients often show atherosclerotic vascular disease in other vascular territories. We evaluated how often whole-body MR imaging detects concomitant arterial pathologies in CHD patients, and how often these pathologies were not known to the patients previously. Of 4,814 participants in the population-based Heinz Nixdorf Recall Study, 327 reported CHD (i.e., previous coronary bypass surgery, angioplasty); of those, 160 patients (mean age 66.4 years) were examined using MR of the brain, the heart (excluding the coronary arteries), and whole-body MR angiography. The prevalence of each vascular pathology was assessed, correlated to the others and compared to patients’ histories. Of the 160 CHD patients, 16 (10%) showed MR signs of stroke, and 77 (48.1%) had a stenosis >50% in at least one extracerebral peripheral artery (other than the coronaries), including 28 (17.5%) with relevant renal artery stenoses, and 20 (12.5%) with relevant extracerebral internal carotid artery stenoses. False negative histories were reported in 12 of 81 cases with myocardial infarctions, and in 11 of 16 cases with cerebrovascular infarctions. This whole-body atherosclerosis MR screening program allows previously unknown concomitant vascular disease to be detected in coronary heart disease patients. Its prospective value should be assessed in further studies.


The mitochondrial antiviral signaling protein (MAVS; also known as IPS-1, VISA, and CARDIF) is essential for innate immune response against RNA viruses. MAVS transduces signals from the cytosolic RIG-I-like receptors, which bind to viral RNAs. But how MAVS activates downstream transcription factors such as IRF3 to induce type-I interferons is not well understood. We have established a cell-free system in which mitochondria derived from virus-infected cells activate IRF3 in the cytosol. Fractionation of the cytosol led to the identification of Ubc5 as a ubiquitin-conjugating enzyme (E2) required for IRF3 activation. Using an inducible RNAi strategy, we demonstrate that catalytically active Ubc5 is required for IRF3 activation by viral infection. The activation of IRF3 also requires two ubiquitin-binding domains of NEMO. Furthermore, we show that replacement of endogenous ubiquitin with its K63R mutant abolishes viral activation of IRF3, demonstrating that K63 polyubiquitination plays a key role in IRF3 activation.


Neoadjuvant chemotherapy (NAC) induces a pathological complete response (pCR) in ∼30% of patients with breast cancer. However, many patients have residual cancer after chemotherapy, which correlates with a higher risk of metastatic recurrence and poorer outcome than those who achieve a pCR. We hypothesized that molecular profiling of tumors after NAC would identify genes associated with drug resistance. Digital transcript counting was used to profile surgically resected breast cancers after NAC. Low concentrations of dual specificity protein phosphatase 4 (DUSP4), an ERK phosphatase, correlated with high post-NAC tumor cell proliferation and with basal-like breast cancer (BLBC) status. BLBC had higher DUSP4 promoter methylation and gene expression patterns of Ras-ERK pathway activation relative to other breast cancer subtypes. DUSP4 overexpression increased chemotherapy-induced apoptosis, whereas DUSP4 depletion dampened the response to chemotherapy. Reduced DUSP4 expression in primary tumors after NAC was associated with treatment-refractory high Ki-67 scores and shorter recurrence-free survival. Finally, inhibition of mitogen-activated protein kinase kinase (MEK) synergized with docetaxel treatment in BLBC xenografts. Thus, DUSP4 downregulation activates the Ras-ERK pathway in BLBC, resulting in an attenuated response to anti-cancer chemotherapy.


There has been a great deal of interest in identifying potential biomarkers of aging. Biomarkers of aging would be useful to predict potential vulnerabilities in an individual that may arise well before they are chronologically expected, due to idiosyncratic aging rates that occur between individuals. Prior attempts to identify biomarkers of aging have often relied on the comparisons of long-lived animals to a wild-type control. However, the effect of interventions in model systems that prolong lifespan (such as single gene mutations or caloric restriction) can sometimes be difficult to interpret due to the manipulation itself having multiple unforeseen consequences on physiology, unrelated to aging itself. The search for predictive biomarkers of aging therefore is problematic, and the identification of metrics that can be used to predict either physiological or chronological age would be of great value. One methodology that has been used to identify biomarkers for numerous pathologies is gene expression profiling. Here, we report whole-genome expression profiles of individual wild-type Caenorhabditis elegans covering the entire wild-type nematode lifespan. Individual nematodes were scored for either age-related behavioral phenotypes, or survival, and then subsequently associated with their respective gene expression profiles. This facilitated the identification of transcriptional profiles that were highly associated with either physiological or chronological age. Overall, our approach serves as a paradigm for identifying potential biomarkers of aging in higher organisms that can be repeatedly sampled throughout their lifespan.


The pathway of gene expression in higher eukaryotes involves a highly complex network of physical and functional interactions among the different machines involved in each step of the pathway. Here we established an efficient in vitro system to determine how RNA polymerase II (RNAP II) transcription is functionally coupled to pre-mRNA splicing. Strikingly, our data show that nascent pre-messenger RNA (pre-mRNA) synthesized by RNAP II is immediately and quantitatively directed into the spliceosome assembly pathway. In contrast, nascent pre-mRNA synthesized by T7 RNA polymerase is quantitatively assembled into the nonspecific H complex, which consists of heterogeneous nuclear ribonucleoprotein (hnRNP) proteins and is inhibitory for spliceosome assembly. Consequently, RNAP II transcription results in a dramatic increase in both the kinetics of splicing and overall yield of spliced mRNA relative to that observed for T7 transcription. We conclude that RNAP II mediates the functional coupling of transcription to splicing by directing the nascent pre-mRNA into spliceosome assembly, thereby bypassing interaction of the pre-mRNA with the inhibitory hnRNP proteins.


Ca2+ messages are broadly important in cellular signal transduction. In immune cells, Ca2+ signaling is an essential step in many forms of activation. Neutrophil-mediated antibody-dependent cell-mediated cytotoxicity (ADCC) is one form of leukocyte activation that plays an important role in tumor cell killing in vitro and in patient care. Using fluorescence methodologies, we found that neutrophils exhibit Ca2+ signals during ADCC directed against breast fibrosarcoma cells. Importantly, these signals were localized to Ca2+ microdomains at the neutrophil-to-tumor cell interface where they display dynamic features such as movement, fusion, and fission. These signals were blocked by the intracellular Ca2+ buffer BAPTA. At the neutrophil–tumor cell synapse, the neutrophil’s cytoplasm was enriched in STIM1, a crucial mediator of Ca2+ signaling, whereas the Ca2+-binding proteins calbindin and parvalbumin were not affected. Our findings suggest that Ca2+ microdomains are due to an active signaling process. As Ca2+ signals within neutrophils were necessary for specific tumor cell apoptosis, a central role of microdomains in leukocyte-mediated tumor cell destruction is indicated.


BACKGROUND Diabetes mellitus is a major risk factor for adverse outcomes after acute coronary syndromes (ACS). Because this disease may be associated with increased platelet aggregation, we investigated whether diabetic patients with ACS derive particular benefit from platelet glycoprotein (GP) IIb/IIIa receptor inhibition. METHODS AND RESULTS We performed a meta-analysis of the diabetic populations enrolled in the 6 large-scale platelet GP IIb/IIIa inhibitor ACS trials: PRISM, PRISM-PLUS, PARAGON A, PARAGON B, PURSUIT, and GUSTO IV. Among 6458 diabetic patients, platelet GP IIb/IIIa inhibition was associated with a significant mortality reduction at 30 days, from 6.2% to 4.6% (OR 0.74; 95% CI 0.59 to 0.92; P=0.007). Conversely, 23 072 nondiabetic patients had no survival benefit (3.0% versus 3.0%). The interaction between platelet GP IIb/IIIa inhibition and diabetic status was statistically significant (P=0.036). Among 1279 diabetic patients undergoing percutaneous coronary intervention (PCI) during index hospitalization, the use of these agents was associated with a mortality reduction at 30 days from 4.0% to 1.2% (OR 0.30; 95% CI 0.14 to 0.69; P=0.002). CONCLUSIONS This meta-analysis, including the entire large-scale trial experience of intravenous platelet GP IIb/IIIa inhibitors for the medical management of non-ST-segment-elevation ACS, shows that these agents may significantly reduce mortality at 30 days in diabetic patients. Although not based on a randomized assessment, the survival benefit appears to be of greater magnitude in patients undergoing PCI. Therefore, the use of platelet GP IIb/IIIa inhibitors should be strongly considered in diabetic patients with ACS.


We previously observed that glucose deprivation induces cell death in multidrug-resistant human breast carcinoma cells (MCF-7/ADR). As a follow up we wished to test the hypothesis that metabolic oxidative stress was the causative process or at least the link between causative processes behind the cytotoxicity. In the studies described here, we demonstrate that mitogen-activated protein kinase (MAPK) was activated within 3 min of being in glucose-free medium and remained activated for 3 h. Glucose deprivation for 2-4 h also caused oxidative stress as evidenced by a 3-fold greater steady state concentration of oxidized glutathione and a 3-fold increase in pro-oxidant production. Glucose and glutamate treatment rapidly suppressed MAPK activation and rescued cells from cytotoxicity. Glutamate and the peroxide scavenger, pyruvate, rescued the cells from cell killing as well as suppressed pro-oxidant production. In addition the thiol antioxidant, N-acetyl-L-cysteine, rescued cells from glucose deprivation-induced cytotoxicity and suppressed MAPK activation. These results suggest that glucose deprivation-induced cytotoxicity and alterations in MAPK signal transduction are mediated by oxidative stress in MCF-7/ADR. These results also support the speculation that a common mechanism of glucose deprivation-induced cytotoxicity in mammalian cells may involve metabolic oxidative stress.


Allelic exclusion underpins antigenic variation and immune evasion in African trypanosomes. These bloodstream parasites use RNA polymerase-I (pol-I) to transcribe just one telomeric variant surface glycoprotein (VSG) gene at a time, producing superabundant and switchable VSG coats. We identified trypanosome VSG exclusion-1 (VEX1) using a genetic screen for defects in telomere-exclusive expression. VEX1 was sequestered by the active VSG and silencing of other VSGs failed when VEX1 was either ectopically expressed or depleted, indicating positive and negative regulation, respectively. Positive regulation affected VSGs and nontelomeric pol-I-transcribed genes, whereas negative regulation primarily affected VSGs. Negative regulation by VEX1 also affected telomeric pol-I-transcribed reporter constructs, but only when they contained blocks of sequence sharing homology with a pol-I-transcribed locus. We conclude that restricted positive regulation due to VEX1 sequestration, combined with VEX1-dependent, possibly homology-dependent silencing, drives a "winner-takes-all" mechanism of allelic exclusion.


Kruppel-like factor 2 (KLF2) plays an important role in the regulation of a variety of immune cells, including monocytes. We have previously shown that KLF2 inhibits proinflammatory activation of monocytes. However, the role of KLF2 in arthritis is yet to be investigated. In the current study, we show that recruitment of significantly greater numbers of inflammatory subset of CD11b(+)F4/80(+)Ly6C+ monocytes to the inflammatory sites in KLF2 hemizygous mice compared to the wild type littermate controls. In parallel, inflammatory mediators, MCP-1, Cox-2 and PAI-1 were significantly up-regulated in bone marrow-derived monocytes isolated from KLF2 hemizygous mice, in comparison to wild-type controls. Methylated-BSA and IL-1β-induced arthritis was more severe in KLF2 hemizygous mice as compared to the littermate wild type controls. Consistent with this observation, monocytes isolated from KLF2 hemizygous mice showed an increased number of cells matured and differentiated towards osteoclastic lineage, potentially contributing to the severity of cartilage and bone damage in induced arthritic mice. The severity of arthritis was associated with the higher expression of proteins such as HSP60, HSP90 and MMP13 and attenuated levels of pPTEN, p21, p38 and HSP25/27 molecules in bone marrow cells of arthritic KLF2 hemizygous mice compared to littermate wild type controls. The data provide new insights and evidences of KLF2-mediated transcriptional regulation of arthritis via modulation of monocyte differentiation and function.


IMPORTANCE Concussion and subconcussive impacts have been associated with short-term disrupted cognitive performance in collegiate athletes, but there are limited data on their long-term neuroanatomic and cognitive consequences. OBJECTIVE To assess the relationships of concussion history and years of football experience with hippocampal volume and cognitive performance in collegiate football athletes. DESIGN, SETTING, AND PARTICIPANTS Cross-sectional study conducted between June 2011 and August 2013 at a US psychiatric research institute specializing in neuroimaging among collegiate football players with a history of clinician-diagnosed concussion (n = 25), collegiate football players without a history of concussion (n = 25), and non-football-playing, age-, sex-, and education-matched healthy controls (n = 25). EXPOSURES History of clinician-diagnosed concussion and years of football experience. MAIN OUTCOMES AND MEASURES High-resolution anatomical magnetic resonance imaging was used to quantify brain volumes. Baseline scores on a computerized concussion-related cognitive battery were used for cognitive assessment in athletes. RESULTS Players with and without a history of concussion had smaller hippocampal volumes relative to healthy control participants (with concussion: t48 = 7.58; P < .001; mean difference, 1788 μL; 95% CI, 1317-2258 μL; without concussion: t48 = 4.35; P < .001, mean difference, 1027 μL; 95% CI, 556-1498 μL). Players with a history of concussion had smaller hippocampal volumes than players without concussion (t48 = 3.15; P < .001; mean difference, 761 μL; 95% CI, 280-1242 μL). In both athlete groups, there was a statistically significant inverse relationship between left hippocampal volume and number of years of football played (t46 = -3.62; P < .001; coefficient = -43.54; 95% CI, -67.66 to -19.41). Behavioral testing demonstrated no differences between athletes with and without a concussion history on 5 cognitive measures but did show an inverse correlation between years of playing football and reaction time (ρ42 = -0.43; 95% CI, -0.46 to -0.40; P = .005). CONCLUSIONS AND RELEVANCE Among a group of collegiate football athletes, there was a significant inverse relationship of concussion and years of football played with hippocampal volume. Years of football experience also correlated with slower reaction time. Further research is needed to determine the temporal relationships of these findings.


Cytosolic detection of pathogen-derived nucleic acids is critical for the initiation of innate immune defense against diverse bacterial, viral and eukaryotic pathogens. Conversely, inappropriate responses to cytosolic nucleic acids can produce severe autoimmune pathology. The host protein STING has been identified as a central signaling molecule in the innate immune response to cytosolic nucleic acids. STING seems to be especially critical for responses to cytosolic DNA and the unique bacterial nucleic acids called 'cyclic dinucleotides'. Here we discuss advances in the understanding of STING and highlight the many unresolved issues in the field.


Identifying genomic elements required for viability is central to our understanding of the basic physiology of bacterial pathogens. Recently, the combination of high-density mutagenesis and deep sequencing has allowed for the identification of required and conditionally required genes in many bacteria. Genes, however, make up only a part of the complex genomes of important bacterial pathogens. Here, we use an unbiased analysis to comprehensively identify genomic regions, including genes, domains, and intergenic elements, required for the optimal growth of Mycobacterium tuberculosis, a major global health pathogen. We found that several proteins jointly contain both domains required for optimal growth and domains that are dispensable. In addition, many non-coding regions, including regulatory elements and non-coding RNAs, are critical for mycobacterial growth. Our analysis shows that the genetic requirements for growth are more complex than can be appreciated using gene-centric analysis.


The mechanisms by which melanoma and other cancer cells evade anti-tumor immunity remain incompletely understood. Here, we show that the growth of tumors formed by mutant Braf(V600E) mouse melanoma cells in an immunocompetent host requires their production of prostaglandin E2, which suppresses immunity and fuels tumor-promoting inflammation. Genetic ablation of cyclooxygenases (COX) or prostaglandin E synthases in Braf(V600E) mouse melanoma cells, as well as in Nras(G12D) melanoma or in breast or colorectal cancer cells, renders them susceptible to immune control and provokes a shift in the tumor inflammatory profile toward classic anti-cancer immune pathways. This mouse COX-dependent inflammatory signature is remarkably conserved in human cutaneous melanoma biopsies, arguing for COX activity as a driver of immune suppression across species. Pre-clinical data demonstrate that inhibition of COX synergizes with anti-PD-1 blockade in inducing eradication of tumors, implying that COX inhibitors could be useful adjuvants for immune-based therapies in cancer patients.


We previously reported that intetumumab (CNTO 95), a fully human anti-αv integrin monoclonal antibody, is a radiosensitizer in mice with xenograft tumors. Because intetumumab does not cross-react with mouse integrins, but has cross-reactivity with rat integrins, we next studied the potential combined use of radiation therapy and intetumumab in human cancer xenograft models in nude rats to assess effects on both tumor cells and the tumor microenvironment. Nude rats bearing human head and neck cancer and non-small cell lung cancer (NSCLC) xenografts were treated with intetumumab and fractionated local tumor radiotherapy. Effects on tumor growth and metastasis, blood perfusion, oxygenation, and gastrointestinal toxicity were studied. Intetumumab alone had a moderate effect on tumor growth. When combined with fractionated radiation therapy, intetumumab significantly inhibited tumor growth and produced a tumor response rate that was significantly better than with radiation therapy alone. Treatment with intetumumab also significantly reduced lung metastasis in the A549 NSCLC xenograft model. The oxygenation and blood perfusion in xenograft tumors measured by microbubble-enhanced ultrasound imaging were substantially increased after treatment with intetumumab. The combined use of intetumumab and radiation therapy reduced the microvessel density and increased apoptosis in tumor cells and the tumor microenvironment. Toxicity studies showed that treatment with intetumumab did not cause the histopathologic changes in the lungs and did not sensitize the sensitive gastrointestinal epithelium to the effect of radiation therapy. Intetumumab can potentiate the efficacy of fractionated radiation therapy in human cancer xenograft tumors in nude rats without increased toxicity.


Epigenetic processes play a key role in orchestrating transcriptional regulation during development. The importance of DNA methylation in fetal brain development is highlighted by the dynamic expression of de novo DNA methyltransferases during the perinatal period and neurodevelopmental deficits associated with mutations in the methyl-CpG binding protein 2 (MECP2) gene. However, our knowledge about the temporal changes to the epigenome during fetal brain development has, to date, been limited. We quantified genome-wide patterns of DNA methylation at ∼ 400,000 sites in 179 human fetal brain samples (100 male, 79 female) spanning 23 to 184 d post-conception. We identified highly significant changes in DNA methylation across fetal brain development at >7% of sites, with an enrichment of loci becoming hypomethylated with fetal age. Sites associated with developmental changes in DNA methylation during fetal brain development were significantly underrepresented in promoter regulatory regions but significantly overrepresented in regions flanking CpG islands (shores and shelves) and gene bodies. Highly significant differences in DNA methylation were observed between males and females at a number of autosomal sites, with a small number of regions showing sex-specific DNA methylation trajectories across brain development. Weighted gene comethylation network analysis (WGCNA) revealed discrete modules of comethylated loci associated with fetal age that are significantly enriched for genes involved in neurodevelopmental processes. This is, to our knowledge, the most extensive study of DNA methylation across human fetal brain development to date, confirming the prenatal period as a time of considerable epigenomic plasticity.


There is mounting evidence showing that the structural and molecular organization of synaptic connections is affected both in human patients and in animal models of neurological and psychiatric diseases. As a consequence of these experimental observations, it has been introduced the concept of synapsopathies, a notion describing brain disorders of synaptic function and plasticity. A close correlation between neurological diseases and synaptic abnormalities is especially relevant for those syndromes including also mental retardation in their symptomatology, such as Rett syndrome (RS). RS (MIM312750) is an X-linked dominant neurological disorder that is caused in the majority of cases by mutations in methyl-CpG-binding protein 2 (MeCP2). This review will focus on the current knowledge of the synaptic alterations produced by mutations of the gene MeCP2 in mouse models of RS and will highlight prospects experimental therapies currently in use. Different experimental approaches have revealed that RS could be the consequence of an impairment in the homeostasis of synaptic transmission in specific brain regions. Indeed, several forms of experience-induced neuronal plasticity are impaired in the absence of MeCP2. Based on the results presented in this review, it is reasonable to propose that understanding how the brain is affected by diseases such as RS is at reach. This effort will bring us closer to identify the neurobiological bases of human cognition.


CONTEXT Significant symptoms of depression are common in the older community-dwelling population. Although depressive symptoms and disability may commonly occur in the same person, whether depressive symptoms contribute to subsequent functional decline has not been elucidated. OBJECTIVE To determine whether depressive symptoms in older persons increase the risk of subsequent decline in physical function as measured by objective performance-based tests. DESIGN A 4-year prospective cohort study. SETTING The communities of Iowa and Washington counties, Iowa. PARTICIPANTS A total of 1286 persons aged 71 years and older who completed a short battery of physical performance tests in 1988 and again 4 years later. MAIN OUTCOME MEASURES Baseline depressive symptoms were assessed by the Center for Epidemiological Studies Depression Scale. Physical performance tests included an assessment of standing balance, a timed 2.4-m (8-ft) walk, and a timed test of 5 repetitions of rising from a chair and sitting down. RESULTS After adjustment for baseline performance score, health status, and sociodemographic factors, increasing levels of depressive symptoms were predictive of greater decline in physical performance over 4 years (odds ratio for decline in those with depressed mood vs those without, 1.55; 95% confidence interval [CI], 1.02-2.34). Even among those at the high end of the functional spectrum, who reported no disability, the severity of depressive symptoms predicted subsequent decline in physical performance (odds ratio for decline, 1.03; 95% CI, 1.00-1.08). CONCLUSIONS This study provides evidence that older persons who report depressive symptoms are at higher risk of subsequent physical decline. These results suggest that prevention or reduction of depressed mood could play a role in reducing functional decline in older persons.


Autophagy is the primary catabolic process triggered in response to starvation. 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. Nevertheless, a thorough understanding of the mechanisms by which sequence-specific transcription factors modulate expression of genes required for autophagy is lacking. Here, we identify Foxk proteins (Foxk1 and Foxk2) as transcriptional repressors of autophagy in muscle cells and fibroblasts. Interestingly, Foxk1/2 serve to counter-balance another forkhead transcription factor, Foxo3, which induces an overlapping set of autophagic and atrophic targets in muscle. Foxk1/2 specifically recruits Sin3A-HDAC complexes to restrict acetylation of histone H4 and expression of critical autophagy genes. Remarkably, mTOR promotes the transcriptional activity of Foxk1 by facilitating nuclear entry to specifically limit basal levels of autophagy in nutrient-rich conditions. 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.


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. PRRSV infection of the host cells actually is a receptor-mediated endocytosis and replication process. The presence and absence of the cellular receptors decide whether the cell lines are permissive or non-permissive to PRRSV infection. 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. 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. 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. Joint forces of the receptors recently attract attentions due to the specific function. 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. In this paper, the role of PRRSV receptors and the molecular mechanism of the interaction between the virus and the receptors are reviewed.