Costunolide causes mitotic arrest and enhances radiosensitivity in human hepatocellular carcinoma cells

<p>Abstract</p> <p>Purpose</p> <p>This work aimed to investigate the effect of costunolide, a sesquiterpene lactone isolated from <it>Michelia compressa</it>, on cell cycle distribution and radiosensitivity of human hepatocellular carcinoma (HCC) cells.</p> <p>Methods</p> <p>The assessment used in this study included: cell viability assay, cell cycle analysis by DNA histogram, expression of phosphorylated histone H3 (Ser 10) by flow cytometer, mitotic index by Liu's stain and morphological observation, mitotic spindle alignment by immunofluorescence of alpha-tubulin, expression of cell cycle-related proteins by Western blotting, and radiation survival by clonogenic assay.</p> <p>Results</p> <p>Our results show that costunolide reduced the viability of HA22T/VGH cells. It caused a rapid G2/M arrest at 4 hours shown by DNA histogram. The increase in phosphorylated histone H3 (Ser 10)-positive cells and mitotic index indicates costunolide-treated cells are arrested at mitosis, not G2, phase. Immunofluorescence of alpha-tubulin for spindle formation further demonstrated these cells are halted at metaphase. Costunolide up-regulated the expression of phosphorylated Chk2 (Thr 68), phosphorylated Cdc25c (Ser 216), phosphorylated Cdk1 (Tyr 15) and cyclin B1 in HA22T/VGH cells. At optimal condition causing mitotic arrest, costunolide sensitized HA22T/VGH HCC cells to ionizing radiation with sensitizer enhancement ratio up to 1.9.</p> <p>Conclusions</p> <p>Costunolide could reduce the viability and arrest cell cycling at mitosis in hepatoma cells. Logical exploration of this mitosis-arresting activity for cancer therapeutics shows costunolide enhanced the killing effect of radiotherapy against human HCC cells.</p

Discovery of selective inhibitors of Glutaminase-2, which inhibit mTORC1, activate autophagy and inhibit proliferation in cancer cells

Glutaminase, which converts glutamine to glutamate, is involved in Warburg effect in cancer cells. Two human glutaminase genes have been identified, GLS (GLS1) and GLS2. Two alternative transcripts arise from each glutaminase gene: first, the kidney isoform (KGA) and glutaminase C (GAC) for GLS; and, second, the liver isoform (LGA) and glutaminase B (GAB) for GLS2. While GLS1 is considered as a cancer therapeutic target, the potential role of GLS2 in cancer remains unclear. Here, we discovered a series of alkyl benzoquinones that preferentially inhibit glutaminase B isoform (GAB, GLS2) rather than the kidney isoform of glutaminase (KGA, GLS1). We identified amino acid residues in an allosteric binding pocket responsible for the selectivity. Treatment with the alkyl benzoquinones decreased intracellular glutaminase activity and glutamate levels. GLS2 inhibition by either alkyl benzoquinones or GLS2 siRNA reduced carcinoma cell proliferation and anchorage-independent colony formation, and induced autophagy via AMPK mediated mTORC1 inhibition. Our findings demonstrate amino acid sequences for selective inhibition of glutaminase isozymes and validate GLS2 as a potential anti-cancer target

Introduction of a strong temperature-sensitive phenotype into enterovirus 71 by altering an amino acid of virus 3D polymerase

AbstractIn 1998, an enterovirus 71 (EV71) epidemic in Taiwan resulted in 78 deaths; however, the molecular basis of EV71 pathogenicity remains poorly understood. Comparison of the deduced amino acid sequences in 3D polymerases of EV71clinical isolates showed the T251V or T251I substitution from 1986 and 1998 outbreaks. An EV71 replicon system showed that introducing an I251T mutation did not affect luciferase activities at 35 °C when compared with wild type; however, lower luciferase activities were observed when they were incubated at 39.5 °C. In addition, the I251T mutation in the EV71 infectious clone not only reduced viral replication at 39.5 °C in vitro but also decreased the virulence of the mouse adaptive strain MP4 in neonatal mice in an i.p. infection model. Therefore, these results suggested that the threonine at position 251 results in a temperature sensitivity phenotype of EV71 which may contribute to the attenuation of circulating strains

TSR: A storage and cooling ring for HIE-ISOLDE

It is planned to install the heavy-ion, low-energy ring TSR, currently at the Max-Planck-Institute for Nuclear Physics in Heidelberg, at the HIE-ISOLDE facility in CERN, Geneva. Such a facility will provide a capability for experiments with stored, cooled secondary beams that is rich and varied, spanning from studies of nuclear ground-state properties and reaction studies of astrophysical relevance, to investigations with highly-charged ions and pure isomeric beams. In addition to experiments performed using beams recirculating within the ring, the cooled beams can be extracted and exploited by external spectrometers for high-precision measurements. The capabilities of the ring facility as well as some physics cases will be presented, together with a brief report on the status of the project

Genomic diversity of citrate fermentation in Klebsiella pneumoniae

<p>Abstract</p> <p>Background</p> <p>It has long been recognized that <it>Klebsiella pneumoniae </it>can grow anaerobically on citrate. Genes responsible for citrate fermentation of <it>K. pneumoniae </it>were known to be located in a 13-kb gene cluster on the chromosome. By whole genome comparison of the available <it>K. pneumoniae </it>sequences (MGH 78578, 342, and NTUH-K2044), however, we discovered that the fermentation gene cluster was present in MGH 78578 and 342, but absent in NTUH-K2044. In the present study, the previously unknown genome diversity of citrate fermentation among <it>K. pneumoniae </it>clinical isolates was investigated.</p> <p>Results</p> <p>Using a genomic microarray containing probe sequences from multiple <it>K. pneumoniae </it>strains, we investigated genetic diversity among <it>K. pneumoniae </it>clinical isolates and found that a genomic region containing the citrate fermentation genes was not universally present in all strains. We confirmed by PCR analysis that the gene cluster was detectable in about half of the strains tested. To demonstrate the metabolic function of the genomic region, anaerobic growth of <it>K. pneumoniae </it>in artificial urine medium (AUM) was examined for ten strains with different clinical histories and genomic backgrounds, and the citrate fermentation potential was found correlated with the genomic region. PCR detection of the genomic region yielded high positive rates among a variety of clinical isolates collected from urine, blood, wound infection, and pneumonia. Conserved genetic organizations in the vicinity of the citrate fermentation gene clusters among <it>K. pneumoniae</it>, <it>Salmonella enterica</it>, and <it>Escherichia coli </it>suggest that the13-kb genomic region were not independently acquired.</p> <p>Conclusion</p> <p>Not all, but nearly half of the <it>K. pneumoniae </it>clinical isolates carry the genes responsible for anaerobic growth on citrate. Genomic variation of citrate fermentation genes in <it>K. pneumoniae </it>may contribute to metabolic diversity and adaptation to variable nutrient conditions in different environments.</p

Cross-protection against European swine influenza viruses in the context of infection immunity against the 2009 pandemic H1N1 virus : studies in the pig model of influenza

Pigs are natural hosts for the same influenza virus subtypes as humans and are a valuable model for cross-protection studies with influenza. In this study, we have used the pig model to examine the extent of virological protection between a) the 2009 pandemic H1N1 (pH1N1) virus and three different European H1 swine influenza virus (SIV) lineages, and b) these H1 viruses and a European H3N2 SIV. Pigs were inoculated intranasally with representative strains of each virus lineage with 6- and 17-week intervals between H1 inoculations and between H1 and H3 inoculations, respectively. Virus titers in nasal swabs and/or tissues of the respiratory tract were determined after each inoculation. There was substantial though differing cross-protection between pH1N1 and other H1 viruses, which was directly correlated with the relatedness in the viral hemagglutinin (HA) and neuraminidase (NA) proteins. Cross-protection against H3N2 was almost complete in pigs with immunity against H1N2, but was weak in H1N1/pH1N1-immune pigs. In conclusion, infection with a live, wild type influenza virus may offer substantial cross-lineage protection against viruses of the same HA and/or NA subtype. True heterosubtypic protection, in contrast, appears to be minimal in natural influenza virus hosts. We discuss our findings in the light of the zoonotic and pandemic risks of SIVs

Enhanced Generation of Induced Pluripotent Stem Cells from a Subpopulation of Human Fibroblasts

BACKGROUND: The derivation of induced pluripotent stem cells (iPSCs) provides new possibilities for basic research and novel cell-based therapies. Limitations, however, include our current lack of understanding regarding the underlying mechanisms and the inefficiency of reprogramming. METHODOLOGY/PRINCIPAL FINDINGS: Here, we report identification and isolation of a subpopulation of human dermal fibroblasts that express the pluripotency marker stage specific embryonic antigen 3 (SSEA3). Fibroblasts that expressed SSEA3 demonstrated an enhanced iPSC generation efficiency, while no iPSC derivation was obtained from the fibroblasts that did not express SSEA3. Transcriptional analysis revealed NANOG expression was significantly increased in the SSEA3 expressing fibroblasts, suggesting a possible mechanistic explanation for the differential reprogramming. CONCLUSIONS/SIGNIFICANCE: To our knowledge, this study is the first to identify a pluripotency marker in a heterogeneous population of human dermal fibroblasts, to isolate a subpopulation of cells that have a significantly increased propensity to reprogram to pluripotency and to identify a possible mechanism to explain this differential reprogramming. This discovery provides a method to significantly increase the efficiency of reprogramming, enhancing the feasibility of the potential applications based on this technology, and a tool for basic research studies to understand the underlying reprogramming mechanisms

Panorama Phylogenetic Diversity and Distribution of Type A Influenza Virus

Type A influenza virus is one of important pathogens of various animals, including humans, pigs, horses, marine mammals and birds. Currently, the viral type has been classified into 16 hemagglutinin and 9 neuraminidase subtypes, but the phylogenetic diversity and distribution within the viral type largely remain unclear from the whole view.The panorama phylogenetic trees of influenza A viruses were calculated with representative sequences selected from approximately 23000 candidates available in GenBank using web servers in NCBI and the software MEGA 4.0. Lineages and sublineages were classified according to genetic distances, topology of the phylogenetic trees and distributions of the viruses in hosts, regions and time.Here, two panorama phylogenetic trees of type A influenza virus covering all the 16 hemagglutinin subtypes and 9 neuraminidase subtypes, respectively, were generated. The trees provided us whole views and some novel information to recognize influenza A viruses including that some subtypes of avian influenza viruses are more complicated than Eurasian and North American lineages as we thought in the past. They also provide us a framework to generalize the history and explore the future of the viral circulation and evolution in different kinds of hosts. In addition, a simple and comprehensive nomenclature system for the dozens of lineages and sublineages identified within the viral type was proposed, which if universally accepted, will facilitate communications on the viral evolution, ecology and epidemiology

A prospective study of serum insulin-like growth factor-I (IGF-I), IGF-II, IGF-binding protein-3 and breast cancer risk.

The associations between serum concentrations of insulin-like growth factor-I (IGF-I), IGF-II and IGF-binding proteins (IGFBP)-3 and risk of breast cancer were investigated in a nested case-control study involving 117 cases (70 premenopausal and 47 postmenopausal at blood collection) and 350 matched controls within a cohort of women from the island of Guernsey, UK. Women using exogenous hormones at the time of blood collection were excluded. Premenopausal women in the top vs bottom third of serum IGF-I concentration had a nonsignificantly increased risk for breast cancer after adjustment for IGFBP-3 (odds ratio (OR) 1.71; 95% confidence interval (CI): 0.74-3.95; test for linear trend, P=0.21). Serum IGFBP-3 was associated with a reduction in risk in premenopausal women after adjustment for IGF-I (top third vs the bottom third: OR 0.49; 95% CI: 0.21-1.12, P for trend=0.07). Neither IGF-I nor IGFBP-3 was associated with risk in postmenopausal women and serum IGF-II concentration was not associated with risk in pre- or postmenopausal women. These data are compatible with the hypothesis that premenopausal women with a relatively high circulating concentration of IGF-I and low IGFBP-3 are at an increased risk of developing breast cancer

Inferring structural connectivity using Ising couplings in models of neuronal networks

Functional connectivity metrics have been widely used to infer the underlying structural connectivity in neuronal networks. Maximum entropy based Ising models have been suggested to discount the effect of indirect interactions and give good results in inferring the true anatomical connections. However, no benchmarking is currently available to assess the performance of Ising couplings against other functional connectivity metrics in the microscopic scale of neuronal networks through a wide set of network conditions and network structures. In this paper, we study the performance of the Ising model couplings to infer the synaptic connectivity in in silico networks of neurons and compare its performance against partial and cross-correlations for different correlation levels, firing rates, network sizes, network densities, and topologies. Our results show that the relative performance amongst the three functional connectivity metrics depends primarily on the network correlation levels. Ising couplings detected the most structural links at very weak network correlation levels, and partial correlations outperformed Ising couplings and cross-correlations at strong correlation levels. The result was consistent across varying firing rates, network sizes, and topologies. The findings of this paper serve as a guide in choosing the right functional connectivity tool to reconstruct the structural connectivity