bluuebunny/crossref_metadata_2025 huggingface dataset:
Cell cycle regulation of the Escherichia coli nrd operon: requirement for a cis-acting upstream AT-rich sequence
Authors: Sun L , Jacobson B A , Dien B S , Srienc F , Fuchs J A
Published: November 2016

The expression of the nrd operon encoding ribonucleotide reductase in Escherichia coli has been shown to be cell cycle regulated. To identify the cis-acting elements required for the cell cycle regulation of the nrd promoter, different 5' deletions as well as site-directed mutations were translationally fused to a lacZ reporter gene. The expression of beta-galactosidase from these nrd-lacZ fusions in single-copy plasmids was determined with synchronously growing cultures obtained by repeated phosphate starvation as well as with exponentially growing cultures by flow cytometry analysis. Although Fis and DnaA, two regulatory proteins that bind at multiple sites on the E. coli chromosome, have been found to regulate the nrd promoter, the results in this study demonstrated that neither Fis nor DnaA was required for nrd cell cycle regulation. A cis-acting upstream AT-rich sequence was found to be required for the cell cycle regulation. This sequence could be replaced by a different sequence that maintained the AT richness. A flow cytometry analysis that combined specific immunofluorescent staining of beta-galactosidase with a DNA-specific stain was developed and employed to study the nrd promoter activity in cells at specific cell cycle positions. The results of the flow cytometry analysis confirmed the results obtained from studies with synchronized cells.
Characterization of the sigD transcription unit of Bacillus subtilis
Authors: Márquez-Magaña L M , Chamberlin M J
Published: November 2016

The sigma D factor of Bacillus subtilis is required for the transcription of the flagellin and motility genes as well as for wild-type chemotaxis. Southern blot and sequence analyses demonstrate that the structural gene for sigma D, sigD, is located immediately downstream of a region of DNA originally identified as the chemotaxis (che) locus and now renamed the fla/che region. In fact, sigD appears to be part of a very large operon (> 26 kb) containing genes which encode structural proteins that form the hook-basal body complex as well as regulatory proteins required for chemotaxis. Transposon insertions up to 24 kb upstream of sigD, within several of the genes for the hook-basal body components, give rise to only a moderate decrease in sigD expression. The transposon insertions, however, block sigma D activity as demonstrated by the lack of flagellin expression in strains bearing these insertions. These effects appear to arise from two types of regulation. In cis the transposon insertions appear to introduce a partial block to transcription of sigD from upstream promoter elements; in trans they disrupt genes whose gene products are required for sigma D activity. It appears that sigD transcription is initiated, at least in part, by a promoter many kilobases upstream of its translation start site and that transcription of the flagellin gene by sigma D is dependent on the formation of a functional hook-basal body complex. The possibility that sigD is part of the fla/che operon was further tested by the integration of an insertion plasmid, containing strong transcription terminators, 1.6 and 24 kb upstream of the sigD gene. In both cases, the introduction of the terminators resulted in a greater decrease of sigD expression than was caused by the plasmid sequences alone. These results indicate that wild-type transcription of sigD is dependent on promoter sequences > 24kb upstream of its structural gene and that the entire fla/che region forms a single operon.
Ylt1, a highly repetitive retrotransposon in the genome of the dimorphic fungus Yarrowia lipolytica
Authors: Schmid-Berger N , Schmid B , Barth G
Published: November 2016

A highly repetitive composite element, Ylt1, was detected in the genome of the dimorphic fungus Yarrowia lipolytica. Ylt1 resembles retrotransposons found in other eukaryotes. It is about 9.4 kb long and can transpose in the genome. The Ylt1 element is bounded by a long terminal repeat (LTR), the zeta element. Several copies of zeta were isolated and sequenced. The sequence of this element is well conserved. It is 714 bp long and is bounded by nucleotides 5'-TG...CA-3', which are part of a short inverted repeat, a feature conserved in the LTRs of retroviruses and retrotransposons. Sequence analysis revealed motifs commonly found in LTR elements, like signals for the start and termination of transcription. The zeta element exists as part of retrotransposon Ylt1, as well as a solo element in the genome. Ylt1 and solo zeta elements are flanked by a 4-bp directly repeated genomic sequence. The copy numbers of Ylt1 and solo zeta are dependent on the strain examined, but at least 35 copies of the composite Ylt1 element and more than 30 copies of the solo zeta element per haploid genome have been observed.
Mutations in pts cause catabolite-resistant sporulation and altered regulation of spo0H in Bacillus subtilis
Authors: Frisby D , Zuber P
Published: November 2016

A mutation in Bacillus subtilis, ggr-31, that relieves glucose-glutamine-dependent control of a spoVG-lacZ translational fusion was isolated and was subsequently found to confer a pleiotropic phenotype. Mutants cultured in glucose- and glutamine-rich media exhibited a Crs- (catabolite-resistant sporulation) phenotype; enhanced expression of the spo0H gene, encoding sigma H, as evidenced by immunoblot analysis with anti-sigma H antiserum; and derepression of srfA, an operon involved in surfactin biosynthesis and competence development. In addition, ggr-31 mutants exhibited a significant increase in generation time when they were cultured in minimal glucose medium. The mutant phenotype was restored to the wild type by Campbell integration of a plasmid containing part of the ptsG (encoding the enzyme II/III glucose permease) gene, indicating that the mutation probably resides within ptsG and adversely affects glucose uptake. A deletion mutation within ptsI exhibited a phenotype similar to that of ggr-31.
Location of a potassium tellurite resistance operon (tehA tehB) within the terminus of Escherichia coli K-12
Authors: Taylor D E , Hou Y , Turner R J , Weiner J H
Published: November 2016

A tellurite resistance determinant, believed to have been cloned from the IncHII plasmid pHH1508a (E. G. Walter, J. H. Weiner, and D. E. Taylor, Gene 101:1-7, 1991), was shown instead to have originated from the chromosome of Escherichia coli K-12. The two genes, tehA and tehB, constitute an operon located in the terminus at approximately 32.3 min.
Management and treatment of chronic kidney disease in cats
Authors: Caney Sarah
Published: November 2016

Making a diagnosis of chronic kidney disease (CKD) in cats marks the start of a new and complex journey to provide optimal treatment for the patient. A successful outcome depends on attention to detail and a good working relationship with the carer and patient. If it is possible to institute good treatment measures, many patients with CKD survive for several years following diagnosis, with an excellent quality of life.
Republished: Endovascular recanalization of the common carotid artery in a patient with radio induced chronic occlusion
Authors: Nico Lorena , Cester Giacomo , Viaro Federica , Baracchini Claudio , Causin Francesco
Published: November 2016

Recanalization of chronic occlusion of the common carotid artery (CCA) in patients with a history of neck irradiation is challenging, both for vascular surgeons and interventional neuroradiologists. We describe a case of successful stenting of radio induced chronic occlusion of the right CCA in a 41-year-old patient with neurological deterioration and minor stroke due to cerebral hypoperfusion caused by concomitant bilateral arterial occlusions. Direct surgery and surgical bypass were considered contraindicated. The endovascular approach was successful and required multiple precautions during the procedure. We describe particular solutions, not used in day to day practice, that allowed us to carry out the endovascular treatment in this unusual situation.
Systematic review and quality analysis of emerging diagnostic measures for calcium pyrophosphate crystal deposition disease
Authors: Wu Y , Chen K , Terkeltaub R
Published: November 2016

Objectives

Calcium pyrophosphate crystal deposition disease (CPPD) is common, yet prevalence and overall clinical impact remain unclear. Sensitivity and specificity of CPPD reference standards (conventional crystal analysis (CCA) and radiography (CR)) were meta-analysed by EULAR (published 2011). Since then, new diagnostic modalities are emerging. Hence, we updated 2009–2016 literature findings by systematic review and evidence grading, and assessed unmet needs.

Methods

We performed systematic search of full papers (PubMed, Scopus/EMBASE, Cochrane 2009–2016 databases). Search terms included CPPD, chondrocalcinosis, pseudogout, ultrasound, MRI, dual energy CT (DECT). Paper selection, data abstraction, EULAR evidence level, and Quality Assessment of Diagnostic Accuracy Studies (QUADAS)-2 bias and applicability grading were performed independently by 3 authors.

Results

We included 26 of 111 eligible papers, which showed emergence in CPPD diagnosis of ultrasound (U/S), and to lesser degree, DECT and Raman spectroscopy. U/S detected CPPD crystals in peripheral joints with sensitivity >80%, superior to CR. However, most study designs, though analytical, yielded low EULAR evidence level. DECT was marginally explored for CPPD, compared with 35 published DECT studies in gout. QUADAS-2 grading indicated strong applicability of U/S, DECT and Raman spectroscopy, but high study bias risk (in ∼30% of papers) due to non-controlled designs, and non-randomised subject selection.

Conclusions

Though CCA and CR remain reference standards for CPPD diagnosis, U/S, DECT and Raman spectroscopy are emerging U/S sensitivity appears to be superior to CR. We identified major unmet needs, including for randomised, blinded, controlled studies of CPPD diagnostic performance and rigorous analyses of 4 T MRI and other emerging modalities.
Modelling the impact of poaching on metapopulation viability for data-limited species
Authors: Camaclang Abbey E. , Curtis Janelle M.R. , Naujokaitis-Lewis Ilona , Poesch Mark S. , Koops Marten A.
Published: November 2016

We developed a spatially explicit simulation model of poaching behaviour to quantify the relative influence of the intensity, frequency, and spatial distribution of poaching on metapopulation viability. We integrated our model of poaching with a stochastic, habitat-based, spatially explicit population model, applied it to examine the impact of poaching on northern abalone (Haliotis kamtschatkana) metapopulation dynamics in Barkley Sound, British Columbia, Canada, and quantified model sensitivity to input parameters. While demographic parameters remained important in predicting extinction probabilities for northern abalone, our simulations indicate that the odds of extinction are twice as high when populations are subjected to poaching. Viability was influenced by poaching variables that affect the total number of individuals removed. Of these, poaching mortality was the most influential in predicting metapopulation viability, with each 0.1 increase in mortality rate resulting in 22.6% increase in the odds of extinction. By contrast, the location and spatial correlation of events were less important predictors of viability. When data are limited, simulation models of poaching combined with sensitivity analyses can be useful in informing management strategies and future research directions.
The essential activities of the bacterial sigma factor
Authors: Davis Maria C. , Kesthely Christopher A. , Franklin Emily A. , MacLellan Shawn R.
Published: November 2016

Transcription is the first and most heavily regulated step in gene expression. Sigma (σ) factors are general transcription factors that reversibly bind RNA polymerase (RNAP) and mediate transcription of all genes in bacteria. σ Factors play 3 major roles in the RNA synthesis initiation process: they (i) target RNAP holoenzyme to specific promoters, (ii) melt a region of double-stranded promoter DNA and stabilize it as a single-stranded open complex, and (iii) interact with other DNA-binding transcription factors to contribute complexity to gene expression regulation schemes. Recent structural studies have demonstrated that when σ factors bind promoter DNA, they capture 1 or more nucleotides that are flipped out of the helical DNA stack and this stabilizes the promoter open-complex intermediate that is required for the initiation of RNA synthesis. This review describes the structure and function of the σ70family of σ proteins and the essential roles they play in the transcription process.
THE RELATION BETWEEN MIGRATION AND FDI IN THE OECD FROM A COMPLEX NETWORK PERSPECTIVE
Authors: GARAS ANTONIOS , LAPATINAS ATHANASIOS , POULIOS KONSTANTINOS
Published: November 2016

We explore the relationship between human migration and OECD’s foreign direct investment (FDI) using a gravity equation enriched with variables that account for complex-network effects. Based on a panel data analysis, we find a strong positive correlation between the migration network and the FDI network, which can be mostly explained by countries’ economic/demographic sizes and geographical distance. We highlight the existence of a stronger positive FDI relationship in pairs of countries that are more central in the migration network. Both intensive and extensive forms of centrality are FDI enhancing. Illuminating this result, we show that bilateral FDI between any two countries is further affected positively by the complex web of ‘third party’ corridors/migration stocks of the international migration network (IMN). Our findings are consistent whether we consider bilateral FDI and bilateral migration figures, or we focus on the outward FDI and the respective inward migration of the OECD countries.
Quaternary glaciation in the Mediterranean mountains: a new synthesis
Authors: Hughes P. D. , Woodward J. C.
Published: November 2016

The Mediterranean mountains were repeatedly glaciated during the Pleistocene. Glaciers were present in most of the major mountains areas from Morocco in the west to the Black Sea coast of Turkey in the east. Some mountains supported extensive ice caps and ice fields with valley glaciers tens of kilometres long. Other massifs sustained only small-scale ice masses, although this was the exception rather than the norm. Glaciers still exist today and there is evidence that small glaciers were a common sight in many regions during the Little Ice Age. The Mediterranean mountains are important for palaeoclimate research because of their position in the mid-latitudes and sensitivity to changes in the climate regimes of adjacent areas including the North Atlantic. These mountains are also important areas of biodiversity and long-term biological change through the Quaternary ice age. All of this provided challenges and opportunities for Palaeolithic societies. This paper reviews the history of the study of glaciation in the Mediterranean mountains from pioneer nineteenth century observations through to the detailed geomorphological mapping and advanced geochronological datasets of recent times. We also review the current state of knowledge to frame the contributions presented in this volume. Lastly, this new synthesis then identifies outstanding research problems and assesses the prospects for new studies of glaciation in the Mediterranean mountains.
Toll-like receptor 4 activation attenuates profibrotic response in control lung fibroblasts but not in fibroblasts from patients with IPF
Authors: Ebener Simone , Barnowski Sandra , Wotzkow Carlos , Marti Thomas M. , Lopez-Rodriguez Elena , Crestani Bruno , Blank Fabian , Schmid Ralph A. , Geiser Thomas , Funke Manuela
Published: November 2016

Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with a median survival of 3 yr. IPF deteriorates upon viral or bacterial lung infection although pulmonary infection (pneumonia) in healthy lungs rarely induces fibrosis. Bacterial lipopolysaccharide (LPS) activates Toll-like receptor 4 (TLR4), initiating proinflammatory pathways. As TLR4 has already been linked to hepatic fibrosis and scleroderma, we now investigated the role of TLR4 in IPF fibroblasts. Lung tissue sections from patients with IPF were analyzed for TLR4 expression. Isolated normal human lung fibroblasts (NL-FB) and IPF fibroblasts (IPF-FB) were exposed to LPS and transforming growth factor-β (TGF-β) before expression analysis of receptors, profibrotic mediators, and cytokines. TLR4 is expressed in fibroblast foci of IPF lungs as well as in primary NL-FB and IPF-FB. As a model for a gram-negative pneumonia in the nonfibrotic lung, NL-FB and IPF-FB were coexposed to LPS and TGF-β. Whereas NL-FB produced significantly less connective tissue growth factor upon costimulation compared with TGF-β stimulation alone, IPF-FB showed significantly increased profibrotic markers compared with control fibroblasts after costimulation. Although levels of antifibrotic prostaglandin E2 were elevated after costimulation, they were not responsible for this effect. However, significant downregulation of TGF-β receptor type 1 in control fibroblasts seems to contribute to the reduced profibrotic response in our in vitro model. Normal and IPF fibroblasts thus differ in their profibrotic response upon LPS-induced TLR4 stimulation.
Partial deficiency of CTRP12 alters hepatic lipid metabolism
Authors: Tan Stefanie Y. , Little Hannah C. , Lei Xia , Li Shuoyang , Rodriguez Susana , Wong G. William
Published: November 2016

Secreted hormones play pivotal roles in tissue cross talk to maintain physiologic blood glucose and lipid levels. We previously showed that C1q/TNF-related protein 12 (CTRP12) is a novel secreted protein involved in regulating glucose metabolism whose circulating levels are reduced in obese and insulin-resistant mouse models. Its role in lipid metabolism, however, is unknown. Using a novel heterozygous mouse model, we show that the loss of a single copy of the Ctrp12 gene (also known as Fam132a and adipolin) affects whole body lipid metabolism. In Ctrp12 (+/−) male mice fed a control low-fat diet, hepatic fat oxidation was upregulated while hepatic VLDL-triglyceride secretion was reduced relative to wild-type (WT) littermates. When challenged with a high-fat diet, Ctrp12 (+/−) male mice had impaired lipid clearance in response to acute lipid gavage, reduced hepatic triglyceride secretion, and greater steatosis with higher liver triglyceride and cholesterol levels. Unlike male mice, Ctrp12 (+/−) female mice fed a control low-fat diet were indistinguishable from WT littermates. When obesity was induced by high-fat feeding, Ctrp12 (+/−) female mice developed mild insulin resistance with impaired insulin tolerance. In contrast to male mice, hepatic triglyceride secretion was increased in Ctrp12 (+/−) female mice fed a high-fat diet. Thus, in different dietary and metabolic contexts, loss of a single Ctrp12 allele affects glucose and lipid metabolism in a sex-dependent manner, highlighting the importance of genetic and environmental determinants of metabolic phenotypes.
MELAS Syndrome with Cardiac Involvement: A Multimodality Imaging Approach
Authors: Seitun Sara , Massobrio Laura , Rubegni Anna , Nesti Claudia , Castiglione Morelli Margherita , Boccalini Sara , Galletto Pregliasco Athena , Budaj Irilda , Deferrari Luca , Rosa Gian Marco , Montecucco Fabrizio , Valbusa Alberto
Published: November 2016

A 49-year-old man presented with chest pain, dyspnea, and lactic acidosis. Left ventricular hypertrophy and myocardial fibrosis were detected. The sequencing of mitochondrial genome (mtDNA) revealed the presence of A to G mtDNA point mutation at position 3243 (m.3243A>G) in tRNALeu(UUR) gene. Diagnosis of cardiac involvement in a patient with Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke-like episodes syndrome (MELAS) was made. Due to increased risk of sudden cardiac death, cardioverter defibrillator was implanted.
Wormhole Detection Based on Ordinal MDS Using RTT in Wireless Sensor Network
Authors: Mukherjee Saswati , Chattopadhyay Matangini , Chattopadhyay Samiran , Kar Pragma
Published: November 2016

In wireless communication, wormhole attack is a crucial threat that deteriorates the normal functionality of the network. Invasion of wormholes destroys the network topology completely. However, most of the existing solutions require special hardware or synchronized clock or long processing time to defend against long path wormhole attacks. In this work, we propose a wormhole detection method using range-based topology comparison that exploits the local neighbourhood subgraph. The Round Trip Time (RTT) for each node pair is gathered to generate neighbour information. Then, the network is reconstructed by ordinal Multidimensional Scaling (MDS) followed by a suspicion phase that enlists the suspected wormholes based on the spatial reconstruction. Iterative computation of MDS helps to visualize the topology changes and can localize the potential wormholes. Finally, a verification phase is used to remove falsely accused nodes and identify real adversaries. The novelty of our algorithm is that it can detect both short path and long path wormhole links. Extensive simulations are executed to demonstrate the efficacy of our approach compared to existing ones.
Complex Perineal Trauma with Anorectal Avulsion
Authors: Cruceru Adelina Maria , Negoi Ionut , Paun Sorin , Hostiuc Sorin , Negoi Ruxandra Irina , Beuran Mircea
Published: November 2016

Introduction. The objective of this case report is to illustrate a severe perineal impalement injury, associated with anorectal avulsion and hemorrhagic shock.Results. A 32-year-old male patient was referred to our hospital for an impalement perineal trauma, associated with complex pelvic fracture and massive perineal soft tissue destruction and anorectal avulsion. On arrival, the systolic blood pressure was 85 mm Hg and the hemoglobin was 7.1 g/dL. The patient was transported to the operating room, and perineal lavage, hemostasis, and repacking were performed. After 12 hours in the Intensive Care Unit, the abdominal ultrasonography revealed free peritoneal fluid. We decided emergency laparotomy, and massive hemoperitoneum due to intraperitoneal rupture of pelvic hematoma was confirmed. Pelvic packing controlled the ongoing diffuse bleeding. After 48 hours, the relaparotomy with packs removal and loop sigmoid colostomy was performed. The postoperative course was progressive favorable, with discharge after 70 days and colostomy closure after four months, with no long-term complications.Conclusions. Severe perineal injuries are associated with significant morbidity and mortality. Their management in high volume centers, with experience in colorectal and trauma surgery, allocating significant human and material resources, decreases the early mortality and long-term complications, offering the best quality of life for patients.
Primary Diffuse Leptomeningeal Gliomatosis: Radiological/Pathological Features
Authors: Ahmad Ehtasham , Mohamed Mohamed , Vrettos Apostolos
Published: November 2016

We present the case of a 43-year-old lady who presented with headaches, visual impairment, and seizures, progressing rapidly over the course of a few weeks. Extensive workup excluded an inflammatory or infectious cause. Imaging studies revealed diffuse thickening of the leptomeninges and serial CSF analysis showed raised opening pressures and increased protein levels. A diagnostic biopsy of the lower thoracic dura confirmed the diagnosis of primary diffuse leptomeningeal gliomatosis (PDGL). She was managed supportively for her symptoms and unfortunately she passed away a few weeks later.
Fault Tolerant Controller Design for a Faulty UAV Using Fuzzy Modeling Approach
Authors: Qian Moshu , Xiong Ke , Wang Lili , Qian Zhongmin
Published: November 2016

We address a fault tolerant control (FTC) issue about an unmanned aerial vehicle (UAV) under possible simultaneous actuator saturation and faults occurrence. Firstly, the Takagi-Sugeno fuzzy models representing nonlinear flight control systems (FCS) for an UAV with unknown disturbances and actuator saturation are established. Then, a normal H-infinity tracking controller is presented using an online estimator, which is introduced to weaken the saturation effect. Based on the normal tracking controller, we propose an adaptive fault tolerant tracking controller (FTTC) to solve actuator loss of effectiveness (LOE) fault problem. Compared with previous work, this approach developed in our research need not rely on any fault diagnosis unit and is easily applied in engineering. Finally, these results in simulation indicate the efficiency of our presented FTC scheme.
World’s First Clinical Case of Gene-Activated Bone Substitute Application
Authors: Bozo I. Y. , Deev R. V. , Drobyshev A. Y. , Isaev A. A. , Eremin I. I.
Published: November 2016

Treatment of patients with large bone defects is a complex clinical problem. We have initiated the first clinical study of a gene-activated bone substitute composed of the collagen-hydroxyapatite scaffold and plasmid DNA encoding vascular endothelial growth factor. The first patient with two nonunions of previously reconstructed mandible was enrolled into the study. Scar tissues were excised; bone defects (5–14 mm) between the mandibular fragments and nonvascularized rib-bone autograft were filled in with the gene-activated bone substitute. No adverse events were observed during 12 months of follow-up. In 3 months, the average density of newly formed tissues within the implantation zone was 402.21 ± 84.40 and 447.68 ± 106.75 HU in the frontal and distal regions, respectively, which correlated with the density of spongy bone. Complete distal bone defect repair with vestibular and lingual cortical plates formation was observed in 6 and 12 months after surgery; thereby the posterior nonunion was successfully eliminated. However, there was partial resorption of the proximal edge of the autograft entailed to relapse of the anterior nonunion. Thus, the first clinical data on the safety and efficacy of the gene-activated bone substitute were obtained. Given a high complexity of the clinical situation the treatment, results might be considered as promising. NCT02293031.
Loss of the Methyl-CpG–Binding Protein ZBTB4 Alters Mitotic Checkpoint, Increases Aneuploidy, and Promotes Tumorigenesis
Authors: Roussel-Gervais Audrey , Naciri Ikrame , Kirsh Olivier , Kasprzyk Laetitia , Velasco Guillaume , Grillo Giacomo , Dubus Pierre , Defossez Pierre-Antoine
Published: November 2016

Chromosome segregation during mitosis is monitored by the mitotic checkpoint and is dependent upon DNA methylation. ZBTB4 is a mammalian epigenetic regulator with high affinity for methylated CpGs that localizes at pericentromeric heterochromatin and is frequently downregulated in cancer. Here, we report that decreased ZBTB4 expression correlates with high genome instability across many frequent human cancers. In human cell lines, ZBTB4 depletion was sufficient to increase the prevalence of micronuclei and binucleated cells in parallel with aberrant mitotic checkpoint gene expression, a weakened mitotic checkpoint, and an increased frequency of lagging chromosomes during mitosis. To extend these findings, we generated Zbtb4-deficient mice. Zbtb4−/− mice were smaller than their wild-type littermates. Primary cells isolated from Zbtb4−/− mice exhibited diminished mitotic checkpoint activity, increased mitotic defects, aneuploid cells marked by a specific transcriptional signature, and increased genomic instability. Zbtb4−/− mice were also more susceptible to 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate (DMBA/TPA)–induced skin carcinogenesis. Our results establish the epigenetic regulator ZBTB4 as an essential component in maintaining genomic stability in mammals. Cancer Res; 77(1); 62–73. ©2016 AACR.
Quantitative Fluorescence Microscopy Measures Vascular Pore Size in Primary and Metastatic Brain Tumors
Authors: Mittapalli Rajendar K. , Adkins Chris E. , Bohn Kaci. A. , Mohammad Afroz S. , Lockman Julie A. , Lockman Paul R.
Published: November 2016

Tumors residing in the central nervous system (CNS) compromise the blood–brain barrier (BBB) via increased vascular permeability, with the magnitude of changes dependent on the tumor type and location. Current studies determine penetrability of a cancer therapeutic by administering progressively larger molecules until cutoff is observed where little to no tumor accumulation occurs. However, decades-old experimental work and mathematical modeling document methods to calculate both the size of the vascular opening (pore) with solute permeability values. In this study, we updated this classic mathematical modeling approach with quantitative fluorescence microscopy in two preclinical tumor models, allowing simultaneous administration of multiple sized tracers to determine vascular permeability at a resolution of nearly one micron. We observed that three molecules ranging from 100 Da to 70 kDa permeated into a preclinical glioblastoma model at rates proportional to their diffusion in water. This suggests the solutes freely diffused from blood to glioma across vascular pores without steric restriction, which calculates to a pore size of >140 nm in diameter. In contrast, the calculated pore size of a brain metastasis of breast cancer was approximately 10-fold smaller than glioma vasculature. This difference explains why antibodies are effective against glioblastoma but generally fail in brain metastases of breast cancer. On the basis of our observations, we hypothesize that trastuzumab most likely fails in the treatment of brain metastases of breast cancer because of poor CNS penetration, while the similar sized antibody bevacizumab is effective in the same tumor type not because it penetrates the CNS degree better, but because it scavenges VEGF in the vascular compartment, which reduces edema and permeation. Cancer Res; 77(2); 238–46. ©2016 AACR.
Polycomb-Mediated Disruption of an Androgen Receptor Feedback Loop Drives Castration-Resistant Prostate Cancer
Authors: Fong Ka-wing , Zhao Jonathan C. , Kim Jung , Li Shangze , Yang Yeqing A. , Song Bing , Rittie Laure , Hu Ming , Yang Ximing , Perbal Bernard , Yu Jindan
Published: November 2016

The lethal phenotype of castration-resistant prostate cancer (CRPC) is generally caused by augmented signaling from the androgen receptor (AR). Here, we report that the AR-repressed gene CCN3/NOV inhibits AR signaling and acts in a negative feedback loop to block AR function. Mechanistically, a cytoplasmic form of CCN3 interacted with the AR N-terminal domain to sequester AR in the cytoplasm of prostate cancer cells, thereby reducing AR transcriptional activity and inhibiting cell growth. However, constitutive repression of CCN3 by the Polycomb group protein EZH2 disrupted this negative feedback loop in both CRPC and enzalutamide-resistant prostate cancer cells. Notably, restoring CCN3 was sufficient to effectively reduce CPRC cell proliferation in vitro and to abolish xenograft tumor growth in vivo. Taken together, our findings establish CCN3 as a pivotal regulator of AR signaling and prostate cancer progression and suggest a functional intersection between Polycomb and AR signaling in CRPC. Cancer Res; 77(2); 412–22. ©2016 AACR.
PTEN Signaling in the Postnatal Perivascular Progenitor Niche Drives Medulloblastoma Formation
Authors: Zhu Guo , Rankin Sherri L. , Larson Jon D. , Zhu Xiaoyan , Chow Lionel M.L. , Qu Chunxu , Zhang Jinghui , Ellison David W. , Baker Suzanne J.
Published: November 2016

Loss of the tumor suppressor gene PTEN exerts diverse outcomes on cancer in different developmental contexts. To gain insight into the effect of its loss on outcomes in the brain, we conditionally inactivated the murine Pten gene in neonatal neural stem/progenitor cells. Pten inactivation created an abnormal perivascular proliferative niche in the cerebellum that persisted in adult animals but did not progress to malignancy. Proliferating cells showed undifferentiated morphology and expressed the progenitor marker Nestin but not Math1, a marker of committed granule neuron progenitors. Codeletion of Pten and Trp53 resulted in fully penetrant medulloblastoma originating from the perivascular niche, which exhibited abnormal blood vessel networks and advanced neuronal differentiation of tumor cells. EdU pulse-chase experiments demonstrated a perivascular cancer stem cell population in Pten/Trp53 double mutant medulloblastomas. Genetic analyses revealed recurrent somatic inactivations of the tumor suppressor gene Ptch1 and a recapitulation of the sonic hedgehog subgroup of human medulloblastomas. Overall, our results showed that PTEN acts to prevent the proliferation of a progenitor niche in postnatal cerebellum predisposed to oncogenic induction of medulloblastoma. Cancer Res; 77(1); 123–33. ©2016 AACR.
Kv1.3 Channels Mark Functionally Competent CD8+ Tumor-Infiltrating Lymphocytes in Head and Neck Cancer
Authors: Chimote Ameet A. , Hajdu Peter , Sfyris Alexandros M. , Gleich Brittany N. , Wise-Draper Trisha , Casper Keith A. , Conforti Laura
Published: November 2016

Tumor-infiltrating lymphocytes (TIL) are potent mediators of an antitumor response. However, their function is attenuated in solid tumors. CD8+ T-cell effector functions, such as cytokine and granzyme production, depend on cytoplasmic Ca2+, which is controlled by ion channels. In particular, Kv1.3 channels regulate the membrane potential and Ca2+ influx in human effector memory T (TEM) cells. In this study, we assessed the contribution of reduced Kv1.3 and Ca2+ flux on TIL effector function in head and neck cancer (HNC). We obtained tumor samples and matched peripheral blood from 14 patients with HNC. CD3+ TILs were composed of 57% CD4+ (82% TEM and 20% Tregs) and 36% CD8+ cells. Electrophysiology revealed a 70% reduction in functional Kv1.3 channels in TILs as compared with peripheral blood T cells from paired patients, which was accompanied by a decrease in Ca2+ influx. Immunofluorescence analysis showed that CD8+ TILs expressing high Kv1.3 preferentially localized in the stroma. Importantly, high expression of Kv1.3 correlated with high Ki-67 and granzyme B expression. Overall, these data indicate that defective Kv1.3 channels and Ca2+ fluxes in TILs may contribute to reduced immune surveillance in HNC. Cancer Res; 77(1); 53–61. ©2016 AACR.
Multiplex Genome Editing to Generate Universal CAR T Cells Resistant to PD1 Inhibition
Authors: Ren Jiangtao , Liu Xiaojun , Fang Chongyun , Jiang Shuguang , June Carl H. , Zhao Yangbing
Published: November 2016

Purpose: Using gene-disrupted allogeneic T cells as universal effector cells provides an alternative and potentially improves current chimeric antigen receptor (CAR) T-cell therapy against cancers and infectious diseases.

Experimental Design: The CRISPR/Cas9 system has recently emerged as a simple and efficient way for multiplex genome engineering. By combining lentiviral delivery of CAR and electro-transfer of Cas9 mRNA and gRNAs targeting endogenous TCR, β-2 microglobulin (B2M) and PD1 simultaneously, to generate gene-disrupted allogeneic CAR T cells deficient of TCR, HLA class I molecule and PD1.

Results: The CRISPR gene–edited CAR T cells showed potent antitumor activities, both in vitro and in animal models and were as potent as non-gene–edited CAR T cells. In addition, the TCR and HLA class I double deficient T cells had reduced alloreactivity and did not cause graft-versus-host disease. Finally, simultaneous triple genome editing by adding the disruption of PD1 led to enhanced in vivo antitumor activity of the gene-disrupted CAR T cells.

Conclusions: Gene-disrupted allogeneic CAR and TCR T cells could provide an alternative as a universal donor to autologous T cells, which carry difficulties and high production costs. Gene-disrupted CAR and TCR T cells with disabled checkpoint molecules may be potent effector cells against cancers and infectious diseases. Clin Cancer Res; 23(9); 2255–66. ©2016 AACR.
Cytokine-Induced Killer Cells Kill Chemo-surviving Melanoma Cancer Stem Cells
Authors: Gammaitoni Loretta , Giraudo Lidia , Macagno Marco , Leuci Valeria , Mesiano Giulia , Rotolo Ramona , Sassi Francesco , Sanlorenzo Martina , Zaccagna Alessandro , Pisacane Alberto , Senetta Rebecca , Cangemi Michela , Cattaneo Giulia , Martin Valentina , Coha Valentina , Gallo Susanna , Pignochino Ymera , Sapino Anna , Grignani Giovanni , Carnevale-Schianca Fabrizio , Aglietta Massimo , Sangiolo Dario
Published: November 2016

Purpose: The MHC-unrestricted activity of cytokine-induced killer (CIK) cells against chemo-surviving melanoma cancer stem cells (mCSC) was explored, as CSCs are considered responsible for chemoresistance and relapses.

Experimental Design: Putative mCSCs were visualized by engineering patient-derived melanoma cells (MC) with a lentiviral vector encoding eGFP under expression control by stemness gene promoter oct4. Their stemness potential was confirmed in vivo by limiting dilution assays. We explored the sensitivity of eGFP+ mCSCs to chemotherapy (CHT), BRAF inhibitor (BRAFi) or CIK cells, as single agents or in sequence, in vitro. First, we treated MCs in vitro with fotemustine or dabrafenib (BRAF-mutated cases); then, surviving MCs, enriched in mCSCs, were challenged with autologous CIK cells. CIK cell activity against chemoresistant mCSCs was confirmed in vivo in two distinct immunodeficient murine models.

Results: We visualized eGFP+ mCSCs (14% ± 2.1%) in 11 MCs. The tumorigenic precursor rate in vivo was higher within eGFP+ MCs (1/42) compared with the eGFP− counterpart (1/4,870). In vitro mCSCs were relatively resistant to CHT and BRAFi, but killed by CIK cells (n = 11, 8/11 autologous), with specific lysis ranging from 95% [effector:tumor ratio (E:T), 40:1] to 20% (E:T 1:3). In vivo infusion of autologous CIK cells into mice bearing xenografts from three distinct melanomas demonstrated significant tumor responses involving CHT-spared eGFP+ mCSCs (P = 0.001). Sequential CHT–immunotherapy treatment retained antitumor activity (n = 12, P = 0.001) reducing mCSC rates (P = 0.01).

Conclusions: These findings are the first demonstration that immunotherapy with CIK cells is active against autologous mCSCs surviving CHT or BRAFi. An experimental platform for mCSC study and rationale for CIK cells in melanoma clinical study is provided. Clin Cancer Res; 23(9); 2277–88. ©2016 AACR.
Activated Phosphorylated STAT1 Levels as a Biologically Relevant Immune Signal in Schizophrenia
Authors: Sharma Rajiv P. , Rosen Cherise , Melbourne Jennifer K. , Feiner Benjamin , Chase Kayla A.
Published: November 2016

Objective: Activation of STAT1 is directly downstream of the cytokine receptors that signal from specific proinflammatory cytokines known to be dysregulated in schizophrenia (SZ), such as IFNγ, IL6, IL2 and IL10, as well as hypoxia, viral/bacterial infections and peptide growth factors. If the increased cytokine protein levels repeatedly observed in SZ have biological consequences, then the measurement of pSTAT1 is a logical step forward. Methods: Peripheral blood mononuclear cells (PBMCs) from controls (n = 13) and subjects with SZ (n = 22) were extracted using the Ficoll method. Participants with SZ were diagnosed using the SCID, clinical symptomatology was measured using the Positive and Negative Syndrome Scale (PANSS), and cognitive functioning was measured using the MATRICS Consensus Cognitive Battery. Levels of activated STAT1 (Y701), i.e. phosphorylated STAT1 (pSTAT1), were measured by a commercially available ELISA in nuclear extracts from PBMCs. Results: There was a significant bimodal distribution in the sample, with an SZ subgroup expressing significantly greater levels of activated pSTAT1 than the remainder of the participants. In this subgroup, levels of pSTAT1 were significantly higher than in the control group, as well as significantly higher than in the remainder of the SZ subjects. Furthermore, this subsample of patients manifested significantly poorer cognitive performance on several measures of the MATRICS. Discussion: pSTAT1 levels may provide a measure of the biological relevance of widely reported elevations in levels of cytokines in SZ over the past several decades. Activation of kinase cascades can be used to partition or disassemble the composite immune signal in patients living with SZ.
Estimating Longitudinal Risks and Benefits From Cardiovascular Preventive Therapies Among Medicare Patients: The Million Hearts Longitudinal ASCVD Risk Assessment Tool: A Special Report From the American Heart Association and American College of Cardiology
Authors: Lloyd-Jones Donald M. , Huffman Mark D. , Karmali Kunal N. , Sanghavi Darshak M. , Wright Janet S. , Pelser Colleen , Gulati Martha , Masoudi Frederick A. , Goff David C.
Published: November 2016

The Million Hearts Initiative has a goal of preventing 1 million heart attacks and strokes—the leading causes of mortality—through several public health and healthcare strategies by 2017. The American Heart Association and American College of Cardiology support the program. The Cardiovascular Risk Reduction Model was developed by Million Hearts and the Center for Medicare & Medicaid Services as a strategy to assess a value-based payment approach toward reduction in 10-year predicted risk of atherosclerotic cardiovascular disease (ASCVD) by implementing cardiovascular preventive strategies to manage the “ABCS” (aspirin therapy in appropriate patients, blood pressure control, cholesterol management, and smoking cessation). The purpose of this special report is to describe the development and intended use of the Million Hearts Longitudinal ASCVD Risk Assessment Tool. The Million Hearts Tool reinforces and builds on the “2013 ACC/AHA Guideline on the Assessment of Cardiovascular Risk” by allowing clinicians to estimate baseline and updated 10-year ASCVD risk estimates for primary prevention patients adhering to the appropriate ABCS over time, alone or in combination. The tool provides updated risk estimates based on evidence from high-quality systematic reviews and meta-analyses of the ABCS therapies. This novel approach to personalized estimation of benefits from risk-reducing therapies in primary prevention may help target therapies to those in whom they will provide the greatest benefit, and serves as the basis for a Center for Medicare & Medicaid Services program designed to evaluate the Million Hearts Cardiovascular Risk Reduction Model.
Role for Galectin‐3 in Calcific Aortic Valve Stenosis
Authors: Sádaba J. Rafael , Martínez‐Martínez Ernesto , Arrieta Vanessa , Álvarez Virginia , Fernández‐Celis Amaya , Ibarrola Jaime , Melero Amaia , Rossignol Patrick , Cachofeiro Victoria , López‐Andrés Natalia
Published: November 2016

Background

Aortic stenosis (
AS
) is a chronic inflammatory disease, and calcification plays an important role in the progression of the disease. Galectin‐3 (Gal‐3) is a proinflammatory molecule involved in vascular osteogenesis in atherosclerosis. Therefore, we hypothesized that Gal‐3 could mediate valve calcification in
AS
.

Methods and Results

Blood samples and aortic valves (
AV
s) from 77 patients undergoing
AV
replacement were analyzed. As controls, noncalcified human
AV
s were obtained at autopsy (n=11). Gal‐3 was spontaneously expressed in valvular interstitial cells (
VIC
s) from
AV
s and increased in
AS
as compared to control
AV
s. Positive correlations were found between circulating and valvular Gal‐3 levels. Valvular Gal‐3 colocalized with the
VIC
s markers, alpha‐smooth muscle actin and vimentin, and with the osteogenic markers, osteopontin, bone morphogenetic protein 2, runt‐related transcription factor 2, and SRY (sex‐determining region Y)‐box 9. Gal‐3 also colocalized with the inflammatory markers cd68, cd80 and tumor necrosis factor alpha. In vitro, in
VIC
s isolated from
AV
s, Gal‐3 induced expression of inflammatory, fibrotic, and osteogenic markers through the extracellular signal‐regulated kinase 1 and 2 pathway. Gal‐3 expression was blocked in
VIC
s undergoing osteoblastic differentiation using its pharmacological inhibitor, modified citrus pectin, or the clustered regularly interspaced short palindromic repeats/Cas9 knockout system. Gal‐3 blockade and knockdown decreased the expression of inflammatory, fibrotic, and osteogenic markers in differentiated
VIC
s.

Conclusions

Gal‐3, which is overexpressed in
AVs
from
AS
patients, appears to play a central role in calcification in
AS
. Gal‐3 could be a new therapeutic approach to delay the progression of
AV
calcification in
AS
.
YKL‐40 Level and Hypertension Incidence: A Population‐Based Nested Case‐Control Study in China
Authors: Xu Tian , Zhong Chongke , Wang Aili , Guo Zhirong , Bu Xiaoqing , Zhou Yipeng , Tian Yunfan , HuangFu Xinfeng , Zhu Zhengbao , Zhang Yonghong
Published: November 2016

Background

Human cartilage glycoprotein‐39 (YKL‐40) has been suggested to be a new marker of inflammation, atherosclerosis, and endothelial dysfunction. However, whether a higher level of YKL‐40 is an independent risk factor for hypertension incidence is still unknown.

Methods and Results

In a nested case‐control study within a prospective cohort of 12 423 initially healthy Chinese adults, we measured baseline plasma concentrations of YKL‐40 among 700 new‐onset hypertension cases and 700 age‐ and sex‐matched controls. Multiple conditional logistic regression analyses were used to calculate the odds ratios (95% CIs) of hypertension associated with higher levels of YKL‐40 both in the total population and in the age‐ (>55 and ≤55 years) and sex‐matched subgroups. Among the total population, YKL‐40 levels were not associated with hypertension risk. In the subgroup older than 55 years, odds ratios (95% CIs) of hypertension for those in the two higher tertiles of YKL‐40 were 1.23 (0.77–1.97) and 1.59 (0.99–2.55) (
P
for linear trend=0.05). In the male subgroup, odds ratios (95% CIs) of hypertension for those in the two higher tertiles of YKL‐40 were 1.55 (0.88–2.72) and 2.09 (1.14–3.82) (
P
for linear trend=0.02). An interaction effect was observed between YKL‐40 and sex (
P
for interaction <0.01) but not between YKL‐40 and age (
P
for interaction=0.21). High YKL‐40 level significantly increased hypertension risk in men but decreased hypertension risk with a trend although not significant in women.

Conclusions

This study suggests that YKL‐40 is associated with hypertension incidence only among men. The study findings need to be further verified by prospective cohort studies or clinical trials.
Smellmap: Amsterdam—Olfactory Art and Smell
Visualization
Authors: McLean Kate
Published: November 2016

Creating a smellmap of a city is a subjective, collaborative exercise. During a series of smellwalks local participants foreground their sense of smell and name perceived aromas emanating from the urban smell-scape. Data and conversations arising from the walks are “analyzed,” and a representative smellscape of the city is visualized as a map. Scents—the nasal stimuli and a catalyst for discussion—accompany the map. As a map of what we do not know, indications of geolocated smell possibilities and ephemeral scents combine visualization with the olfactory to place the emphasis on human interaction with sensory data to create meaning and an understanding of place.