bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression, purification, and characterization of protective MPT64 antigen protein and identification of its multimers isolated from nontoxic Mycobacterium tuberculosis H37Ra
Authors: Chu Teng‐Ping J. , Yuann Jeu‐Ming P.
Published: May 2011

MPT64, a secreted protein of Mycobacterium tuberculosis (MTB), stimulates the immune reactions within cells and is a protective antigen that is lost by the bacilli Calmette–Guérin (BCG) vaccine during propagation. To minimize the toxicity caused by MTB, we used the MPT64 gene encoded by nontoxic H37Ra MTB to carry out genetic expansion via polymerase chain reaction and gene clone MPT64. The plasmid DNA encoded MPT64 was expressed at 20°C for 22 H, and a large quantity of MPT64 was obtained. In the absence of urea, MPT64 multimers with subunits being covalently connected via disulfide bonds were detected by Western blot showing strong protein–protein interactions, as evidenced by the formation of MPT64 tetramers. Finally, with urea of decreasing concentrations, we refolded MPT64 purified in the presence of urea and determined its secondary structures using circular dichroism. MPT64 was found to contain 2.2% α‐helix, 50.9% β‐sheet, 19.5% turn, and 27.4% random coil. The molecular weight of MPT64 was determined by a matrix‐assisted laser desorption ionization‐time of flight mass spectrometer and found to be 23,497 Da, very close to the theoretical molecular weight of MPT64. The results presented here provide a sound basis for future biochemical and biophysical studies of MPT64 or any other proteins encoded by nontoxic H37Ra MTB.
An in silico approach for the discovery of CDK5/p25 interaction inhibitors
Authors: Zhang Bing , Corbel Caroline , Guéritte Françoise , Couturier Cyril , Bach Stéphane , Tan Vincent B. C.
Published: May 2011

The lack of selectivity of all existing ATP competitive inhibitors for a single cyclin‐dependent kinase (CDK) has led us to redirect the structure‐based molecule design from targeting the classic ATP‐binding pocket in CDK5 toward the CDK5/p25 interface. The aim was to seek novel inhibition mechanisms to interrupt protein‐protein interactions. A combined strategy of alanine‐scanning calculations for locating binding sites, virtual screening for small molecules, molecular dynamics simulations for examining the binding stability of virtual screening hits and bio‐assays for testing the level of inhibition was set up and used to explore novel inhibitors capable of interrupting the interactions between the proteins, and consequently of inhibiting the kinase activity. Two compounds were shown to inhibit the complex formation between CDK5 and p25 through p25 binding. They could open avenues for the discovery of new types of structures that prevent interactions between CDK5 and p25 or other CDK and activator proteins, and, more importantly, provide leads in the development of selective inhibitors among CDKs.

Accompanying article Corbel et al.
Promoter methylation and transgene copy numbers predict unstable protein production in recombinant chinese hamster ovary cell lines
Authors: Osterlehner Andrea , Simmeth Silke , Göpfert Ulrich
Published: May 2011

Mammalian cell lines for recombinant protein production need to maintain productivity over extended cultivation times. Long‐term stability studies are time and resource intensive, but are widely performed to identify and eliminate unstable candidates during cell line development. Production instability of manufacturing cell lines can be associated with methylation and silencing of the heterologous promoter. We have identified CpG dinucleotides within the human cytomegalovirus major immediate early promoter/enhancer (hCMV‐MIE) that are frequently methylated in unstable antibody‐producing Chinese hamster ovary (CHO) cell lines. We have established methylation‐specific real‐time qPCR for the rapid and sensitive measurement of hCMV‐MIE methylation in multiple cell lines and provide evidence that hCMV‐MIE methylation and transgene copy numbers can be used as early markers to predict production instability of recombinant CHO cell lines. These markers should provide the opportunity to enrich stable producers early in cell line development and allow developers to put more emphasis on other criteria, such as product quality and bioprocess robustness. Biotechnol. Bioeng. 2011;108: 2670–2681. © 2011 Wiley Periodicals, Inc.
Transforming growth factor‐β1 is constitutively secreted by chinese hamster ovary cells and is functional in human cells
Authors: Beatson Richard , Sproviero Daisy , Maher John , Wilkie Scott , Taylor‐Papadimitriou Joyce , Burchell Joy M.
Published: May 2011

Chinese hamster ovary (CHO) cells are widely used for the production of recombinant proteins for clinical use as well as academic research. They are particularly important for the production of glycoproteins where bacteria cannot be used. TGFβ1 is a potent cytokine highly conserved across species with multiple immunological and non‐immunological effects. We have discovered that CHOK1, the CHO clone most commonly used by the pharmaceutical industry, constitutively secretes latent TGFβ1 and that this hamster TGFβ1 is active on human cells inducing profound immunological effects. As far as we are aware, the production of TGFβ1 by CHOK1 cells has not been reported before in the literature. As TGFβ1 exerts powerful and pleiotropic effects on diverse cell types, and as CHO cells are used to produce a large number of clinical and non‐clinical products, our findings are highly relevant to studies that rely on recombinant proteins. Biotechnol. Bioeng. 2011;108: 2759–2764. © 2011 Wiley Periodicals, Inc.
Utilization of non‐AUG initiation codons in a flow cytometric method for efficient selection of recombinant cell lines
Authors: Cairns Victor R. , DeMaria Christine T. , Poulin Francis , Sancho José , Liu Ping , Zhang Jin , Campos‐Rivera Juanita , Karey Kenneth P. , Estes Scott
Published: May 2011

Here we describe a method that couples flow cytometric detection with the attenuated translation of a reporter protein to enable efficient selection of CHO clones producing high levels of recombinant proteins. In this system, a small cell surface reporter protein is expressed from an upstream open reading frame utilizing a non‐AUG initiation (alternate start) codon. Due to the low translation initiation efficiency of this alternate start codon, the majority of translation initiation events occur at the first AUG of the downstream open reading frame encoding the recombinant protein of interest. While translation of the reporter is significantly reduced, the levels are sufficient for detection using flow cytometric methods and, in turn, predictive of protein expression from the gene of interest since both ORFs are translated from the same mRNA. Using this system, CHO cells have been sorted to obtain enriched pools producing significantly higher levels of recombinant proteins than the starting cell population and clones with significantly better productivity than those generated from limiting dilution cloning. This method also serves as an effective screening tool during clone expansion to enable resources to be focused solely on clones with both high and stable expression. Biotechnol. Bioeng. 2011;108: 2611–2622. © 2011 Wiley Periodicals, Inc.
The role of polymer nanolayer architecture on the separation performance of anion‐exchange membrane adsorbers: Part II. DNA and virus separations
Authors: Bhut Bharat V. , Weaver Justin , Carter Andrew R. , Wickramasinghe S. Ranil , Husson Scott M.
Published: May 2011

The surface‐initiated polymerization protocol developed in part I was used to prepare strong anion‐exchange membranes with variable polymer chain graft densities and degrees of polymerization for DNA and virus particle separations. A focus of part II was to evaluate the role of polymer nanolayer architecture on DNA and virus binding. Salmon sperm‐DNA (SS‐DNA) was used as model nucleic acid to measure the dynamic‐binding capacities at 10% breakthrough. The dynamic‐binding capacity increases linearly with increasing poly ([2‐(methacryloyloxy)ethyl]trimethylammonium chloride) chain density up to the highest chain density used in this study. The new membranes yielded threefold higher SS‐DNA‐binding capacity (30 mg/mL) than a leading commercial membrane with the same functional group chemistry. Elution of bound DNA yielded a sharp peak, and resulted in a 13‐fold increase relative to the feed concentration. This concentration effect further demonstrates the highly favorable transport properties of the newly designed Q‐type membranes. However, unlike findings in part I on protein binding, SS‐DNA binding was not fully reversible. Minute virus of mice (MVM) was used as model virus to evaluate the virus clearance performance of newly designed Q‐type membranes. Log reduction of virus (LRV) of MVM increased with increasing polymer chain density. Membranes exhibited >4.5 LRV for the given MVM impurity load and may be capable of higher LRV values, as the MVM concentration in the flow‐through fraction of these samples was below the limit of detection of the assay. Biotechnol. Bioeng. 2011;108: 2654–2660. © 2011 Wiley Periodicals, Inc.
Autophagy induced by rapamycin and carbon‐starvation have distinct proteome profiles in Aspergillus nidulans
Authors: Kim Yonghyun , Islam Nazrul , Moss Bill J. , Nandakumar M. P. , Marten Mark R.
Published: May 2011

It is hypothesized that autophagy, a global catabolic pathway which is highly conserved from yeast to man, plays an important role in many bioprocesses. Though autophagy is known to be induced by either nutrient starvation or treatment with the drug rapamycin, it is not clear whether the two modes of induction have the same long‐term impact in the cell, particularly in the biotechnologically important filamentous fungi. Here, we compare the overall proteomes from the carbon‐starved (G−) and rapamycin treated (R+) model fungus Aspergillus nidulans. From about 1,100 visualized protein spots, we conservatively selected a total of 26 proteins with significant different expression. To highlight, increased levels of glucosidases and decreased levels of N‐acetylglucosamine pyrophosphorylase were observed, suggesting degradation of the fungal cell wall as an alternate carbon source for both modes of induction. Cdc37 was reduced in expression while 14‐3‐3 ArtA was increased, implying regulation of polar growth, while also potentially regulating autophagy negatively via PKA or Tor. Other proteins included aspartate transaminase, tryptophan synthase B (TrpB), glycylpeptide N‐tetradecanoyltransferase (Nmt1), and aldehyde dehydrogenase (aldA). More interestingly, the majority of the identified proteins (16 of 26) were uniquely expressed in elevated levels in G−. A novel predicted protein from AN8223 which has no sequence homology to other organisms is also implicated to be involved in carbon‐starvation. Thus, proteomic data here show that in A. nidulans, rapamycin‐induced autophagy and carbon‐starvation induced autophagy share some effectors for cell survival, but predominantly involve different long‐term effectors. Biotechnol. Bioeng. 2011;108: 2705–2715. © 2011 Wiley Periodicals, Inc.
Impact of anticoagulation on ionic and nonionic contrast media effect on thrombogenesis and fibrinolysis: The PEPCIT study
Authors: Bellemain‐Appaix Anne , Beygui Farzin , Lesty Claude , Gupta Satya , Silvain Johanne , Le Feuvre Claude , Cayla Guillaume , Allali Yann , Montalescot Gilles , Collet Jean‐Philippe
Published: May 2011

Objectives: The effect of ionic low osmolar contrast media (ICM) and nonionic iso‐osmolar CM (NICM) on acute thrombotic complications of percutaneous coronary intervention (PCI) is subject to controversies possibly related to a potential interaction with anticoagulation regimens. We sought to compare physical and morphological properties of fibrin clots made in the presence of ioxaglate (ICM), iodixanol (NICM) versus control and to evaluate the effect of four anticoagulants used in PCI. Methods and Results: Maximum platelet aggregation (MPA%), maximum elastic modulus (EM, dyne/cm2) fiber density (n/10−5/μm2), and lysis front velocity (nm/sec) of fibrin rich clot (FRC) were measured simultaneously using peripheral blood from 12 patients undergoing elective PCI. We compared the effects of adding iodixanol or ioxaglate or saline (control) to blood with enoxaparin, unfractionated heparin, fondaparinux, and bivalirudin. Iodixanol and ioxaglate led to nonsignificant reduction in MPA compared to control (33.6% ± 16.9%, 28.2% ± 18.9%, and 40.7% ± 13.9%, respectively, P = ns). Fibrin formed with iodixanol was stiffer (42.7 ± 41.9, 18.7 ± 3.7, and 15.9 ± 9 dyne/cm2, P < 0.01) and displayed more fibrin fibers (1089 ± 175, 260 ± 108, and 456 ± 131 n/10−5/μm2, respectively, P < 0.01) than with ioxaglate or control. This resulted in a profound reduction in the lysis front velocity (191 ± 95, 261 ± 112, and 360 ± 153 nm/sec). None of the four anticoagulants displayed any significant interaction on the effect of contrast media. Conclusions: The prothrombogenic effect of iodixanol is related primarily to an increase in fibrin stiffness with subsequent delayed fibrinolysis, something not seen with ioxaglate. Anticoagulation does not appear to have any impact on this fibrin clot abnormalities. © 2011 Wiley‐Liss, Inc.
Mechanistic Insights into Ring‐Closing Enyne Metathesis with the Second‐Generation Grubbs–Hoveyda Catalyst: A DFT Study
Authors: Nuñez‐Zarur Francisco , Solans‐Monfort Xavier , Rodríguez‐Santiago Luis , Pleixats Roser , Sodupe Mariona
Published: May 2011

The full catalytic process (precatalyst activation, propagating cycle and active‐species interconversion) of the ring‐closing enyne metathesis (RCEYM) reaction of 1‐allyloxy‐2‐propyne with the Grubbs–Hoveyda complex as catalyst was studied by B3LYP density functional theory. Both the ene‐then‐yne and yne‐then‐ene pathways are considered and, for the productive catalytic cycle, the feasibility of the endo‐yne‐then‐ene route is also explored. Calculations predict that the ene‐then‐yne and yne‐then‐ene pathways proceed through equivalent steps, the only major difference being the order in which they take place. In this way, all alkene metathesis processes studied here involve four steps: olefin coordination, cycloaddition, cycloreversion and olefin decoordination. Among them, the two more energetically demanding ones are the olefin coordination and decoordination steps. The reaction of the alkyne fragment consists of two steps: alkyne coordination and alkyne skeletal reorganization, the latter of which has the highest Gibbs energy barrier. Comparison between the ene‐then‐yne and yne‐then‐ene pathways shows that there is no clear energetic preference for either of the two processes, and thus both should be operative when unsubstituted enynes are involved. In addition, although the endo orientation is computed to be slightly disfavored, it is not ruled out for 1‐allyloxy‐2‐propyne, and thus calculations seem to indicate that the exo versus endo selectivity is strongly influenced by the presence of substituents in the reagent.
The Crystal Structure of the CeNA:RNA Hybrid ce(GCGTAGCG):r(CGCUACGC)
Authors: Ovaere Margriet , Herdewijn Piet , Van Meervelt Luc
Published: May 2011

Cyclohexenyl nucleic acids (CeNA) are characterised by the carbon–carbon double bond replacing the O4′‐oxygen atom of the natural D‐2′‐deoxyribose sugar ring in DNA. CeNAs exhibit a high conformational flexibility, are stable against nuclease activity and their hybridisation is RNA selective. Additionally, CeNA has been shown to induce an enhanced biological activity when incorporated in siRNA. This makes CeNA a good candidate for siRNA and synthetic aptamer applications. The crystal structure of the synthetic CeNA:RNA hybrid ce(GCGTAGCG):r(CGCUACGC) has been solved with a resolution of 2.50 Å. The CeNA:RNA duplex adopts an anti‐parallel, right‐handed double helix with standard Watson–Crick base pairing. Analyses of the helical parameters revealed the octamer to form an A‐like double helix. The cyclohexenyl rings mainly adopt the 3H2 conformation, which resembles the C3′‐endo conformation of RNA ribose ring. This C3′‐endo ring puckering was found in most of the RNA residues and is typical for A‐family helices. The crystal structure is stabilised by the presence of hexahydrated magnesium ions. The fact that the CeNA:RNA hybrid adopts an A‐type double helical conformation confirms the high potential of CeNAs for the construction of efficient siRNAs which can be used for therapeutical applications.
Light‐Induced Water Oxidation by a Ru complex Containing a Bio‐Inspired Ligand
Authors: Kärkäs Markus D. , Johnston Eric V. , Karlsson Erik A. , Lee Bao‐Lin , Åkermark Torbjörn , Shariatgorji Mohammadreza , Ilag Leopold , Hansson Örjan , Bäckvall Jan‐E. , Åkermark Björn
Published: May 2011

The new Ru complex 8 containing the bio‐inspired ligand 7 was successfully synthesized and characterized. Complex 8 efficiently catalyzes water oxidation using CeIV and RuIII as chemical oxidants. More importantly, this complex has a sufficiently low overpotential to utilize ruthenium polypyridyl‐type complexes as photosensitizers.
Effect of fenoterol stereochemistry on the β2 adrenergic receptor system: Ligand‐directed chiral recognition
Authors: Jozwiak Krzysztof , Plazinska Anita , Toll Lawrence , Jimenez Lucita , Woo Anthony Yiu‐Ho , Xiao Rui‐Ping , Wainer Irving W.
Published: May 2011

The β2 adrenergic receptor (β2‐AR) is a model system for studying the ligand recognition process in G protein‐coupled receptors. Fenoterol (FEN) is a β2‐AR selective agonist that has two centers of chirality and exists as four stereoisomers. Radioligand binding studies determined that stereochemistry greatly influences the binding affinity. Subsequent Van't Hoff analysis shows very different thermodynamics of binding depending on the stereoconfiguration of the molecule. The binding of (S,x′)‐isomers is almost entirely enthalpy controlled whereas binding of (R,x′)‐isomers is purely entropy driven. Stereochemistry of FEN molecule also affects the coupling of the receptor to different G proteins. In a rat cardiomyocyte contractility model, (R,R′)‐FEN was shown to selectively activate Gs protein signaling while the (S,R′)‐isomer activated both Gi and Gs protein. The overall data demonstrate that the chirality at the two chiral centers of the FEN molecule influences the magnitude of binding affinity, thermodynamics of local interactions within the binding site, and the global mechanism of β2‐AR activation. Differences in thermodynamic parameters and nonuniform G‐protein coupling suggest a mechanism of chiral recognition in which observed enantioselectivities arise from the interaction of the (R,x′)‐FEN stereoisomers with a different receptor conformation than the one with which the (S,x′)‐isomer interacts. Chirality 2011. © 2011 Wiley‐Liss, Inc.
MOCVD of ZnO Films from Bis(Ketoiminato)Zn(II) Precursors: Structure, Morphology and Optical Properties
Authors: Bekermann Daniela , Ludwig Arne , Toader Teodor , Maccato Chiara , Barreca Davide , Gasparotto Alberto , Bock Claudia , Wieck Andreas D. , Kunze Ulrich , Tondello Eugenio , Fischer Roland A. , Devi Anjana
Published: May 2011

Two closely related bis(ketoiminato) zinc precursors, which are air stable and possess favorable properties for metal‐organic (MO)CVD, are successfully employed for the growth of ZnO films on silicon and borosilicate glass substrates at temperatures between 400 and 700 °C. The as‐deposited films are investigated by X‐ray diffraction (XRD), field emission scanning electron microscopy (FESEM), atomic force microscopy (AFM), X‐ray photoelectron spectroscopy (XPS), nuclear reaction analysis (NRA), as well as by UV‐vis absorption spectroscopy and photoluminescence (PL) measurements. The structure, morphology, and composition of the as‐grown films show a strong dependence on the substrate temperature. The formation of pure and (001)‐oriented wurtzite‐type stoichiometric ZnO is observed. PL measurements are performed both at room temperature and 77 K, revealing a defect‐free emission of ZnO films.
Early transcriptional regulation by C‐peptide in freshly isolated rat proximal tubular cells
Authors: Lindahl Emma , Nordquist Lina , Müller Patrick , El Agha Eli , Friederich Malou , Dahlman‐Wright Karin , Palm Fredrik , Jörnvall Hans
Published: May 2011

Background

Clinical studies have shown that proinsulin C‐peptide exerts renoprotective effects in type 1 diabetes, although the underlying mechanisms are poorly understood. As C‐peptide has been shown to induce several intracellular events and to localize to nuclei, we aimed to determine whether gene transcription is affected in proximal tubular kidney cells, and if so, whether the genes with altered transcription include those related to protective mechanisms.

Methods

The effect of C‐peptide incubation (2 h) on gene expression was investigated in freshly isolated proximal tubular cells from streptozotocin‐diabetic Sprague‐Dawley rats using global gene expression profiling and real‐time quantitative polymerase chain reaction. Protein expression was assayed using western blotting. Different bioinformatic strategies were employed.

Results

Gene transcription profiling demonstrated differential transcription of 492 genes (p < 0.01) after 2 h of C‐peptide exposure, with the majority of these genes repressed (83%). Real‐time quantitative polymerase chain reaction validation supported a trend of several G protein‐coupled receptors being activated, and certain transcription factors being repressed. Also, C‐peptide repressed the transcription of genes associated with the pathways of circulatory and inflammatory diseases.

Conclusion

This study shows that C‐peptide exerts early effects on gene transcription in proximal tubular cells. The findings also bring further knowledge to the renoprotective mechanisms of C‐peptide in type 1 diabetes, and support a transcriptional activity for C‐peptide. It is suggested that C‐peptide may play a regulatory role in the gene expression of proximal tubular cells. Copyright © 2011 John Wiley & Sons, Ltd.
Phosphorus‐Centered and Phosphinidene‐Capped Tungsten Chloride Clusters
Authors: Ströbele Markus , Eichele Klaus , Meyer H.‐Jürgen
Published: May 2011

Black crystalline powders of W6PCl17 and W4(PCl)Cl10 were obtained after reducing WCl6 with red phosphorus at 370 and 500 °C. The crystal structures were determined by single‐crystal and powder X‐ray diffraction analyses. The structure of W6PCl17 is represented by a phosphorus‐centered hexanuclear tungsten cluster, whose (W6PCl11)Cl4aCl4/2a–a chains form a hexagonal stick packing structure. The structure of W4(PCl)Cl10 is represented by a Jahn–Teller distorted tetranuclear tungsten cluster that is interconnected into a layered [W4(μ4‐PCl)Cl6i]Cl8/2a–a structure containing a chloro‐phosphinidene ligand. Solid‐state 31P magic‐angle spinning (MAS) NMR spectra, electronic structures, and magnetic properties are reported.
Racial identification modulates default network activity for same and other races
Authors: Mathur Vani A. , Harada Tokiko , Chiao Joan Y.
Published: May 2011

Racial identification shapes self‐concept and how people share in and respond to the emotional states of others around them. Prior neuroimaging studies have demonstrated the role of the neural default network in self‐referential and empathic processing. However, how racial identification affects neural processing of social information remains unknown. Here, we examined the effect of racial identification on neural response related to social perception among African American and Caucasian American individuals using functional magnetic resonance imaging. Our results demonstrate that degree of racial identification predicts activity within cortical midline structures of the default network in response to viewing racial ingroup, relative to outgroup members, and activity within the medial temporal lobe subsystem of the default network in response to viewing racial outgroup, relative to ingroup members. Broadly, our findings suggest that the strength of racial identification is associated with differential recruitment of neural and cognitive processes to understand and respond to other people within and outside of one's racial group. Hum Brain Mapp, 2011. © 2011 Wiley‐Liss, Inc.
Elimination of hepatocellular carcinoma and acute hepatitis B in children 25 years after a hepatitis B newborn and catch-up immunization program
Authors: McMahon Brian J. , Bulkow Lisa R. , Singleton Rosalyn J. , Williams James , Snowball Mary , Homan Chriss , Parkinson Alan J.
Published: May 2011

Alaska Native people experience the highest rates of acute and chronic hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) in the United States. We examined the effect of a universal newborn immunization with hepatitis B vaccine and mass population screening immunization program initiated in 1984 on rates of HBV and HCC in children 25 years later. During this time, the population of Alaska Native people grew from an estimated 75,000 to 130,000 persons. A surveillance system to detect acute HBV infection in Alaska Native facilities was established in 1981. Cases of HCC in children under 20 years of age were identified using a National Cancer Institute (NCI)-funded Cancer Registry established in 1969 coupled with an active surveillance program of screening persons with chronic HBV semiannually for alpha-fetoprotein since 1982. The incidence of acute symptomatic HBV infection in persons <20 years of age fell from cases 19/100,000 in 1981-1982 to 0/100,000 in 1993-1994. No cases of acute HBV have occurred in children since 1992. The incidence of HCC in persons <20 years decreased from 3/100,000 in 1984-1988 to zero in 1995-1999 and no cases have occurred since 1999. The number of identified hepatitis B surface antigen-positive children <20 years in the Alaska Native population declined from 657 in 1987 to two in 2008.

Conclusion:

Universal newborn vaccination coupled with mass screening and immunization of susceptible Alaska Natives has eliminated HCC and acute symptomatic HBV infection among Alaska Native children and this approach is the best way to prevent HBV-related disease in children. (HEPATOLOGY 2011;)
Survival after Yttrium-90 resin microsphere radioembolization of hepatocellular carcinoma across Barcelona clinic liver cancer stages: A European evaluation
Authors: Sangro Bruno , Carpanese Livio , Cianni Roberto , Golfieri Rita , Gasparini Daniele , Ezziddin Samer , Paprottka Philipp M. , Fiore Francesco , Van Buskirk Mark , Bilbao Jose Ignacio , Ettorre Giuseppe Maria , Salvatori Rita , Giampalma Emanuela , Geatti Onelio , Wilhelm Kai , Hoffmann Ralf Thorsten , Izzo Francesco , Iñarrairaegui Mercedes , Maini Carlo Ludovico , Urigo Carlo , Cappelli Alberta , Vit Alessandro , Ahmadzadehfar Hojjat , Jakobs Tobias Franz , Lastoria Secondo
Published: May 2011

A multicenter analysis was conducted to evaluate the main prognostic factors driving survival after radioembolization using yttrium-90–labeled resin microspheres in patients with hepatocellular carcinoma at eight European centers. In total, 325 patients received a median activity of 1.6 GBq between September 2003 and December 2009, predominantly as whole-liver (45.2%) or right-lobe (38.5%) infusions. Typically, patients were Child-Pugh class A (82.5%), had underlying cirrhosis (78.5%), and had good Eastern Cooperative Oncology Group (ECOG) performance status (ECOG 0-1; 87.7%), but many had multinodular disease (75.9%) invading both lobes (53.1%) and/or portal vein occlusion (13.5% branch; 9.8% main). Over half had advanced Barcelona Clinic Liver Cancer (BCLC) staging (BCLC C, 56.3%) and one-quarter had intermediate staging (BCLC B, 26.8%). The median overall survival was 12.8 months (95% confidence interval, 10.9-15.7), which varied significantly by disease stage (BCLC A, 24.4 months [95% CI, 18.6-38.1 months]; BCLC B, 16.9 months [95% CI, 12.8-22.8 months]; BCLC C, 10.0 months [95% CI, 7.7-10.9 months]). Consistent with this finding, survival varied significantly by ECOG status, hepatic function (Child-Pugh class, ascites, and baseline total bilirubin), tumor burden (number of nodules, alpha-fetoprotein), and presence of extrahepatic disease. When considered within the framework of BCLC staging, variables reflecting tumor burden and liver function provided additional prognostic information. The most significant independent prognostic factors for survival upon multivariate analysis were ECOG status, tumor burden (nodules >5), international normalized ratio >1.2, and extrahepatic disease. Common adverse events were: fatigue, nausea/vomiting, and abdominal pain. Grade 3 or higher increases in bilirubin were reported in 5.8% of patients. All-cause mortality was 0.6% and 6.8% at 30 and 90 days, respectively.

Conclusion:

This analysis provides robust evidence of the survival achieved with radioembolization, including those with advanced disease and few treatment options. (HEPATOLOGY 2011;)
Human hepatic cancer stem cells are characterized by common stemness traits and diverse oncogenic pathways
Authors: Marquardt Jens U. , Raggi Chiara , Andersen Jesper B. , Seo Daekwan , Avital Itzhak , Geller David , Lee Yun-Han , Kitade Mitsuteru , Holczbauer Agnes , Gillen Matthew C. , Conner Elizabeth A. , Factor Valentina M. , Thorgeirsson Snorri S.
Published: May 2011

Epigenetic mechanisms play critical roles in stem cell biology by maintaining pluripotency of stem cells and promoting differentiation of more mature derivatives. If similar mechanisms are relevant for the cancer stem cell (CSC) model, then epigenetic modulation might enrich the CSC population, thereby facilitating CSC isolation and rigorous evaluation. To test this hypothesis, primary human cancer cells and liver cancer cell lines were treated with zebularine (ZEB), a potent DNA methyltransferase-1 inhibitor, and putative CSCs were isolated using the side population (SP) approach. The CSC properties of ZEB-treated and untreated subpopulations were tested using standard

in vitro

and

in vivo

assays. Whole transcriptome profiling of isolated CSCs was performed to generate CSC signatures. Clinical relevance of the CSC signatures was evaluated in diverse primary human cancers. Epigenetic modulation increased frequency of cells with CSC properties in the SP fraction isolated from human cancer cells as judged by self-renewal, superior tumor-initiating capacity in serial transplantations, and direct cell tracking experiments. Integrative transcriptome analysis revealed common traits enriched for stemness-associated genes, although each individual CSC gene expression signature exhibited activation of different oncogenic pathways (e.g.,

EGFR

,

SRC,

and

MYC).

The common CSC signature was associated with malignant progression, which is enriched in poorly differentiated tumors, and was highly predictive of prognosis in liver and other cancers. Conclusion: Epigenetic modulation may provide a tool for prospective isolation and in-depth analysis of CSC. The liver CSC gene signatures are defined by a pernicious interaction of unique oncogene-specific and common stemness traits. These data should facilitate the identifications of therapeutic tools targeting both unique and common features of CSCs. (HEPATOLOGY 2011;)
NOX1/nicotinamide adenine dinucleotide phosphate, reduced form (NADPH) oxidase promotes proliferation of stellate cells and aggravates liver fibrosis induced by bile duct ligation
Authors: Cui Wenhao , Matsuno Kuniharu , Iwata Kazumi , Ibi Masakazu , Matsumoto Misaki , Zhang Jia , Zhu Kai , Katsuyama Masato , Torok Natalie J. , Yabe-Nishimura Chihiro
Published: May 2011

Among multiple isoforms of nicotinamide adenine dinucleotide phosphate, reduced form (NADPH) oxidase expressed in the liver, the phagocytic NOX2 isoform in hepatic stellate cells (HSCs) has been demonstrated to play a key role in liver fibrogenesis. The aim of this study was to clarify the role of NOX1, a nonphagocytic form of NADPH oxidase, in the development of fibrosis using

Nox1

-deficient mice (Nox1KO). Liver injury and fibrosis were induced by bile duct ligation (BDL) and carbon tetrachloride in Nox1KO and wildtype littermate mice (WT). Primary HSCs were isolated to characterize the NOX1-induced signaling cascade involved in liver fibrogenesis. Following BDL, a time-dependent increase in NOX1 messenger RNA (mRNA) was demonstrated in WT liver. Compared with those in WT, levels of collagen-1α mRNA and hydroxyproline were significantly suppressed in Nox1KO with a reduced number of activated HSCs and less severe fibrotic lesions. The expression levels of α-smooth muscle actin, a marker of HSCs activation, were similar in cultured HSCs isolated from both genotypes. However, cell proliferation was significantly attenuated in HSCs isolated from Nox1KO. In these cells, the expression of p27kip1, a cell cycle suppressor, was significantly up-regulated. Concomitantly, a significant reduction in phosphorylated forms of Akt and forkhead box O (FOXO) 4, a downstream effector of Akt that regulates the transcription of p27kip1 gene, was demonstrated in Nox1KO. Finally, the level of the oxidized inactivated form of phosphatase and tensin homolog (PTEN), a negative regulator of PI3K/Akt pathway, was significantly attenuated in HSCs of Nox1KO. These findings indicate that reactive oxygen species derived from NOX1/NADPH oxidase oxidize and inactivate PTEN to positively regulate the Akt/FOXO4/p27kip1 signaling pathway. NOX1 may thus promote proliferation of HSCs and accelerate the development of fibrosis following BDL-induced liver injury. (HEPATOLOGY 2011;)
Measurement of spectra of normal incidence absorptance of surfaces in life space
Authors: Wakabayashi Hidenobu , Makino Toshiro
Published: May 2011

The thermal and radiative environment of human life space is characterized strongly by solar radiation and by the radiation from surrounding walls as well as by air temperature and humidity. To consider the environmental radiation quantitatively, absorptance spectra and total emittance of the surfaces should be measured properly. In this work we measure spectra of normal incidence absorptance of such surfaces in a near‐ultraviolet through infrared region of wavelength of 0.30–11 μm. On the basis of measured spectra we evaluate solar absorptance of the surfaces to respond to the needs of radiation characteristics of the surfaces in the fields of architecture, life science, and solar engineering. Objective surfaces of the measurement are surfaces of indoor and outdoor environments: cloths, indoor wall materials, painted surfaces, ceramics, bricks, outdoor wall materials, road materials, leaves, etc. We also observe the transition of the absorptance spectra of a water–wet cement surface in a drying process. © 2011 Wiley Periodicals, Inc. Heat Trans Asian Res; Published online in Wiley Online Library (wileyonlinelibrary.com/journal/htj). DOI 10.1002/htj.20356
High‐risk human papillomavirus in oral squamous cell carcinoma of young patients
Authors: Kaminagakura Estela , Villa Luísa Lina , Andreoli Maria Antonieta , Sobrinho João Simão , Vartanian José Guilherme , Soares Fernando Augusto , Nishimoto Inês Nobuko , Rocha Rafael , Kowalski Luiz Paulo
Published: May 2011

The aim of this study was to investigate a possible relation between oral squamous cell carcinoma (SCC), the presence of high‐risk human papillomavirus (HR‐HPV) DNA and p16 expression in young patients. Paraffin‐embedded tumor blocks from 47 oral SCC of young (≤40‐year old) patients were evaluated. The presence of HPV DNA in tumor specimens was analyzed by polymerase chain reaction (PCR) using GP5+/GP6+ generic primers (L1 region) followed by dot blot hybridization for HPV typing. When necessary, the HPV16 positivity was confirmed by PCR HPV16 E7‐specific primers. Cases involving young patients were compared with 67 oral SCC from patients ≥50‐year old (controls). Demographic and clinical data were collected to analyze patient outcomes. p16ink4 expression was evaluated by immunostaining of tissue microarrays. HPV16 was detected in 22 (19.2%) cases; 15 (68.2%) young and 7 (31.8%) control patients, a statistically significant difference (p = 0.01). In 1 (1.7%) young group specimen, HPV DNA 16 and 18 was detected. p16 expression was observed in 11 (25.6%) cases from the young group and in 11 (19.6%) controls (p = 0.48). Association between HPV and p16 was verified, and it was statistically significant (p = 0.002). The higher prevalence of high‐risk HPV types, especially HPV16, may be a contributing factor to oral carcinogenesis in younger individuals.
Carcinogen metabolism genes, red meat and poultry intake, and colorectal cancer risk
Authors: Wang Jun , Joshi Amit D. , Corral Román , Siegmund Kimberly D. , Marchand Loïc Le , Martinez Maria Elena , Haile Robert W. , Ahnen Dennis J. , Sandler Robert S. , Lance Peter , Stern Mariana C.
Published: May 2011

Diets high in red meat are established risk factors for colorectal cancer (CRC). Carcinogenic compounds generated during meat cooking have been implicated as causal agents. We conducted a family‐based case‐control study to investigate the association between polymorphisms in carcinogen metabolism genes (CYP1A2 ‐154A>C, CYP1B1 Leu432Val, CYP2E1 ‐1054C>T, GSTP1 Ile105Val, PTGS2 5UTR ‐765, EPHX1 Tyr113His, NAT2 Ile114Thr, NAT2 Arg197Gln and NAT2 Gly286Glu) and CRC risk. We tested for gene‐environment interactions using case‐only analyses (N = 577) and compared statistically significant results to those obtained using case‐unaffected sibling comparisons (N = 307 sibships). Our results suggested that CYP1A2 ‐154A>C might modify the association between intake of red meat cooked using high temperature methods and well done on the inside and CRC risk (case‐only interaction OR = 1.53; 95% CI = 1.19–1.97; p = 0.0008) and the association between intake of red meat heavily browned on the outside and rectal cancer risk (case‐only interaction OR = 0.65; 95% CI = 0.48–0.86; p = 0.003). We also found that GSTP1 Ile105Val might modify the association between intake of poultry cooked with high temperature methods and CRC risk (p = 0.0035), a finding that was stronger among rectal cancer cases. Our results support a role for heterocyclic amines that form in red meat as a potential explanation for the observed association between diets high in red meat and CRC. Our findings also suggest a possible role for diets high in poultry cooked at high temperatures in CRC risk.
MAGE‐D4B is a novel marker of poor prognosis and potential therapeutic target involved in breast cancer tumorigenesis
Authors: Germano Serena , Kennedy Susan. , Rani Sweta , Gleeson Grainne , Clynes Martin , Doolan Padraig , McDonnell Susan , Hughes Linda , Crown John , O'Driscoll Lorraine
Published: May 2011

Melanoma‐associated antigen (MAGE) family members are generally described as tumor‐specific antigens. An association between MAGE‐D4B and breast cancer has yet to be reported and the functional role of the encoded protein has never been established. We performed microarray analysis of 104 invasive breast tumors and matched non‐cancerous breast biopsies. qPCR was used for validation in an independent biobank. To investigate the biological relevance of MAGE‐D4B in breast tumorigenesis, its phenotypic effects were assessed in vitro. Overall, MAGE‐D4B was detected in 43% of tumors while undetected in normal breast tissue. MAGE‐D4B was found to correlate with tumor progression and to be an independent prognostic marker for poor outcome in term of relapse‐free and overall survival, with potential predictive relevance in relation to response to chemotherapy. RNA interference‐mediated knockdown of MAGE‐D4B significantly hampered the invasive properties of Hs578T cells by affecting anchorage‐independent growth, adhesion, migration and invasion affecting anchorage‐independent growth, adhesion, migration and invasion and by modulating expression of invasion‐suppressor gene E‐cadherin.
Comparison of hematopoietic progenitor cell collections using the COBE Spectra version 7 and Amicus version 3.1 for patients with al amyloidosis
Authors: Burgstaler Edwin A. , Winters Jeffrey L.
Published: May 2011

To address concerns about infused fluid volume during HPC collections in patients with AL amyloidosis, our institution has used a 26:1 anticoagulant (AC) ratio on the COBE Spectra and on the Fenwal Amicus. In this study, in a cohort of AL amyloid patients, we compared the Amicus version 3.1 to the Spectra version 7 MNC collections with regard to infused fluid volume, CD34+ cell yield, lymphocyte yield, cross‐cellular content, and adverse reactions. Both instruments used a 26:1 AC ratio but the Amicus delivered significantly less AC per procedure (Amicus 678 mL vs. Spectra 753 mL). With comparable baseline CD34+ cell counts (Amicus 33 cells/μL vs. Spectra 27 cells/μL); Amicus collected significantly more CD34+ cells (3.1 vs. 1.5 × 106/kg) and equivalent lymphocytes (18.7 vs. 14.5 × 109). Amicus collected significantly fewer WBC (51.8 vs. 72.7 × 109), granulocytes (15.1 vs. 27.5 × 109), and PLT (2.3 vs. 3.9 × 1011) per procedure with equivalent RBC content (26 vs. 30 mL). CD34+ cell (5.0 vs. 4.4 × 106/kg) and lymphocyte doses (32.7 vs. 33.9 × 109) were equivalent in infused products collected on the Amicus and Spectra but the frequency of high volume products was lower for Amicus. Frequency and severity of adverse reactions during collection and infusion were similar for both. In this group of AL amyloid patients, Amicus was superior to Spectra with regard to fluid infused, CD34+ cell yield, and cross‐cellular contamination with equivalent lymphocyte yield and reaction incidence. © 2011 Wiley‐Liss, Inc.
Quantitative symmetry and chirality—A fast computational algorithm for large structures: Proteins, macromolecules, nanotubes, and unit cells
Authors: Dryzun Chaim , Zait Amir , Avnir David
Published: May 2011

Symmetry is one of the most fundamental properties of nature and is used to understand and investigate physical properties. Classically, symmetry is treated as a binary qualitative property, although other physical properties are quantitative. Using the continuous symmetry measure (CSM) methodology one can quantify symmetry and correlate it quantitatively to physical, chemical, and biological properties. The exact analytical procedures for calculating the CSM are computationally expensive and the calculation time grows rapidly as the structure contains more atoms. In this article, we present a new method for calculating the CSM and the related continuous chirality measure (CCM) for large systems. The new method is much faster than the full analytical procedures and it reduces the calculation time dependency fromN! toN2, whereNis the number of atoms in the structure. We evaluate the cost of the applied approximations, estimate the error of the method, and show that deviations from the analytical solutions are within an error of 2%, and in many cases even less. The method is applicable at the moment for the cyclic symmetry point groups—Ci,Cs,Cn, andSn, and therefore it can be used also for chirality measures, which are the minimal of theSnmeasures. We demonstrate the application of the method for large structures across chemistry: proteins, macromolecules, nanotubes, and large unit cells of crystals. © 2011 Wiley Periodicals, Inc. J Comput Chem, 2011
SDOCK: A global protein‐protein docking program using stepwise force‐field potentials
Authors: Zhang Changsheng , Lai Luhua
Published: May 2011

Fast Fourier transform (FFT) method limits the forms of scoring functions in global protein‐protein docking. On the other hand, force field potentials can effectively describe the energy hyper surface of biological macromolecules. In this study, we developed a new protein‐protein docking program, SDOCK, that incorporates van der Waals attractive potential, geometric collision, screened electrostatic potential, and Lazaridis‐Karplus desolvation energy into the scoring function in the global searching process. Stepwise potentials were generated from the corresponding continuous forms to treat the structure flexibility. After optimization of the atom solvation parameters and the weights of different potential terms based on a new docking test set that contains 142 cases with small or moderate conformational changes upon binding, SDOCK slightly outperformed the well‐known FFT based global docking program ZDOCK3.0. Among the 142 cases tested, 52.8% gave at least one near‐native solutions in the top 100 solutions. SDOCK was also tested on six blind testing cases in Critical Assessment of Predicted Interactions rounds 13 to 18. In all six cases, the near‐native solutions could be found within the top 350 solutions. Because the SDOCK approach performs global docking based on force‐field potentials, one of its advantages is that it provides global binding free energy surface profiles for further analysis. The efficiency of the program is also comparable with that of other FFT based protein‐protein docking programs. SDOCK is available for noncommercial applications at http://mdl.ipc.pku.edu.cn/cgi‐bin/down.cgi. © 2011 Wiley Periodicals, Inc. J Comput Chem, 2011
Induction of torpor: Mimicking natural metabolic suppression for biomedical applications
Authors: Bouma Hjalmar R. , Verhaag Esther M. , Otis Jessica P. , Heldmaier Gerhard , Swoap Steven J. , Strijkstra Arjen M. , Henning Robert H. , Carey Hannah V.
Published: May 2011

Mammalian hibernation consists of periods of depressed metabolism and reduced body temperature called “torpor” that are interspersed by normothermic arousal periods. Numerous cellular processes are halted during torpor, including transcription, translation, and ion homeostasis. Hibernators are able to survive long periods of low blood flow and body temperature followed by rewarming and reperfusion without overt signs of organ injury, which makes these animals excellent models for application of natural protective mechanisms to human medicine. This review examines efforts to induce torpor‐like states in non‐hibernating species using pharmacological compounds. Elucidating the underlying mechanisms of natural and pharmacologically induced torpor will speed the development of new clinical approaches to treat a variety of trauma and stress states in humans. J. Cell. Physiol. 227: 1285–1290, 2012. © 2011 Wiley Periodicals, Inc.
The nuclear form of glutathione peroxidase 4 is associated with sperm nuclear matrix and is required for proper paternal chromatin decondensation at fertilization
Authors: Puglisi Rossella , Maccari Irene , Pipolo Simona , Conrad Marcus , Mangia Franco , Boitani Carla
Published: May 2011

The nuclear isoform of the selenoprotein Phospholipid Hydroperoxide Glutathione Peroxidase (nGPx4) is expressed in haploid male germ cells, contains several cysteines and is able to oxidize protein thiols, besides glutathione. In this study we have investigated the subnuclear localization of this isoform in isolated mouse male germ cells at different steps of maturation. Immunoblotting and confocal microscopy analyses of subnuclear fractions showed that nGPx4 is localized to the nuclear matrix together with well known markers of this subnuclear compartment like lamin B and topoisomerase IIβ at all stages of germ cell differentiation. The peculiar nGPx4 distribution was confirmed by both biochemical and morphological analyses of COS‐1 cells overexpressing Flag‐tagged nGPx4. To test the functional role of nGPx4 in the process of chromatin assembly, sperm isolated from the caput and the cauda epididymides of wild‐type (WT) and genetically deficient in nGPx4 (nGPx4‐KO) mice were analyzed in an in vitro chromatin decondensation assay. Results showed that sperm from nGPx4‐KO mice were more prone to decondense than those from WT mice at all stages of epididymal maturation, providing conclusive evidence that nGPx4 is required for a correct sperm chromatin compaction. We next addressed the issue of whether the lack of nGPx4 impacts on early events occurring at fertilization. Indeed, in vitro fertilization experiments showed an acceleration of sperm chromatin dispersion in oocytes fertilized by nGpx4‐KO sperm compared with control. Overall these data indicate that the absence of nGPx4 leads to sperm nuclear matrix/chromatin instability that may negatively affect the embryo development. J. Cell. Physiol. 227: 1420–1427, 2012. © 2011 Wiley Periodicals, Inc.