bluuebunny/crossref_metadata_2025 huggingface dataset:
The role of BMP‐7 in chondrogenic and osteogenic differentiation of human bone marrow multipotent mesenchymal stromal cells in vitro
Authors: Shen Bojiang , Wei Aiqun , Whittaker Shane , Williams Lisa A. , Tao Helen , Ma David D.F. , Diwan Ashish D.
Published: November 2009

This study addresses the role of bone morphogenetic protein‐7 (BMP‐7) in chondrogenic and osteogenic differentiation of human bone marrow multipotent mesenchymal stromal cells (BM MSCs) in vitro. BM MSCs were expanded and differentiated in the presence or absence of BMP‐7 in monolayer and three‐dimensional cultures. After 3 days of stimulation, BMP‐7 significantly inhibited MSC growth in expansion cultures. When supplemented in commonly used induction media for 7–21 days, BMP‐7 facilitated both chondrogenic and osteogenic differentiation of MSCs. This was evident by specific gene and protein expression analyses using real‐time PCR, Western blot, histological, and immunohistochemical staining. BMP‐7 supplementation appeared to enhance upregulation of lineage‐specific markers, such as type II and type IX collagens (COL2A1, COL9A1) in chondrogenic and secreted phosphoprotein 1 (SPP1), osteocalcin (BGLAP), and osterix (SP7) in osteogenic differentiation. BMP‐7 in the presence of TGF‐β3 induced superior chondrocytic proteoglycan accumulation, type II collagen, and SOX9 protein expression in alginate and pellet cultures compared to either factor alone. BMP‐7 increased alkaline phosphatase activity and dose‐dependently accelerated calcium mineralization of osteogenic differentiated MSCs. The potential of BMP‐7 to promote adipogenesis of MSCs was restricted under osteogenic conditions, despite upregulation of adipocyte gene expression. These data suggest that BMP‐7 is not a singular lineage determinant, rather it promotes both chondrogenic and osteogenic differentiation of MSCs by co‐ordinating with initial lineage‐specific signals to accelerate cell fate determination. BMP‐7 may be a useful enhancer of in vitro differentiation of BM MSCs for cell‐based tissue repair. J. Cell. Biochem. 109: 406–416, 2010. © 2009 Wiley‐Liss, Inc.
Glycolytic pyruvate regulates P‐Glycoprotein expression in multicellular tumor spheroids via modulation of the intracellular redox state
Authors: Wartenberg Maria , Richter Madeleine , Datchev André , Günther Sebastian , Milosevic Nada , Bekhite Mohamed M. , Figulla Hans‐Reiner , Aran Josep M. , Pétriz Jordi , Sauer Heinrich
Published: November 2009

ABC transporters like P‐glycoprotein (P‐gp/ABCB1) are membrane proteins responsible for the transport of toxic compounds out of non‐malignant cells and tumor tissue. Aim: To investigate the effect of glycolysis and the tissue redox state on P‐gp expression in multicellular tumor spheroids derived from prostate adenocarcinoma cells (DU‐145), glioma cells (Gli36), and the human cervix carcinoma cell line KB‐3‐1 transfected with a P‐gp–EGFP fusion gene that allows monitoring of P‐gp expression in living cells. During cell culture of DU‐145, Gli36, and KB‐3‐1 tumor spheroids P‐gp expression was observed as well as increased lactate and decreased pyruvate levels and expression of glycolytic enzymes. Inhibition of glycolysis for 24 h by either iodoacetate (IA) or 2‐deoxy‐D‐glucose (2‐DDG) downregulated P‐gp expression which was reversed upon coincubation with the radical scavenger ebselen as shown by semi‐quantitative immunohistochemisty in DU‐145 and Gli36 tumor spheroids, and by EGFP fluorescence in KB‐3‐1 tumor spheroids. Consequently endogenous ROS generation in DU‐145 tumor spheroids was increased in the presence of either IA or 2‐DDG, which was abolished upon coincubation with ebselen. Exogenous addition of pyruvate significantly reduced ROS generation, increased P‐gp expression as well as efflux of the P‐gp substrate doxorubicin. Doxorubicin transport was significantly blunted by 2‐DDG and IA, indicating that inhibition of glycolysis reversed the multidrug resistance phenotype. In summary our data demonstrate that P‐gp expression in tumor spheroids is closely related to the glycolytic metabolism of tumor cells and can be downregulated by glycolysis inhibitors via mechanisms that involve changes in the cellular redox state. J. Cell. Biochem. 109: 434–446, 2010. © 2009 Wiley‐Liss, Inc.
Effect of arachidonic acid on hypoxia‐induced IL‐6 production in mouse ES cells: Involvement of MAPKs, NF‐κB, and HIF‐1α
Authors: Lee Sang Hun , Lee Yu Jin , Han Ho Jae
Published: November 2009

This study examined the role of arachidonic acid (AA) in hypoxia‐induced production of interleukin (IL)‐6 and its related signaling pathways in mouse embryonic stem (ES) cells. Hypoxia with AA induced IL‐6 production, which was mediated by reactive oxygen species (ROS). In addition, hypoxia increased the levels of p38 mitogen‐activated protein kinases (MAPKs) and stress‐activated protein kinase/c‐jun NH2‐terminal kinase (SAPK/JNK) phosphorylation, which were blocked by antioxidant (vitamin C). Inhibition of p38 MAPK and SAPK/JNK blocked hypoxia‐ or hypoxia with AA‐induced nuclear factor‐kappa B (NF‐κB) activation. Furthermore, hypoxia‐induced increase in hypoxia‐inducible factor‐1α (HIF‐1α) expression was regulated by NF‐κB activation. Consequently, the increased HIF‐1α expression induced activation of matrix metalloproteinase (MMP)‐2 and MMP‐9. The expression of each signaling molecule stimulated an increase in IL‐6 production that was greater in hypoxic conditions with AA than with hypoxia alone. Finally, inhibition of IL‐6 production using IL‐6 antibody or soluble IL‐6 receptor attenuated the hypoxia‐induced increases in DNA synthesis of mouse ES cells. In conclusion, AA potentiates hypoxia‐induced IL‐6 production through the MAPKs, NF‐κB, and HIF‐1α pathways in mouse ES cells. J. Cell. Physiol. 222: 574–585, 2010. © 2009 Wiley‐Liss, Inc.
PKB/Akt activation inhibits p53‐mediated HIF1A degradation that is independent of MDM2
Authors: Choy Mun‐Kit , Movassagh Mehregan , Bennett Martin R. , Foo Roger S.‐Y.
Published: November 2009

Cross‐talk between the two transcription factors, p53 and hypoxia inducible factor 1α (HIF1A), is important in different pathophysiological conditions (Hammond and Giaccia, 2006, Clin Cancer Res 12:5007–5009) such as in the transition from myocardial hypertrophy to cardiac dilatation and heart failure. In that context, p53 induces HIF1A degradation which in turn provokes the transition from compensatory hypertrophy to myocardial thinning and chamber dilatation (Sano et al., 2007, Nature 446:444–448). In order to investigate the mechanism of p53‐induced HIF1A degradation, we used the established in vitro model of deferroxamine (DFX)‐induced HIF1A accumulation in H9c2 cardiac cells (Sano et al., 2007, Nature 446:444–448). Here, we report that opposite to HIF1A accumulation following exposure to DFX, prolonged DFX‐induced p53 activation and HIF1A protein decrease, without any change in Hif1a mRNA. HIF1A protein decrease accompanied upregulated HIF1A ubiquitination. MDM2, an ubiquitin E3 ligase target gene of p53, was upregulated following prolonged DFX, but using p53/Mdm2 double‐null mouse embryonic fibroblasts, we found that p53 upregulated HIF1A ubiquitination and degradation independently of MDM2. Moreover, with prolonged DFX treatment, an enhanced interaction between MDM2 and HIF1A was lacking. Instead, phospho‐Aktser473 was decreased during the phase coinciding with HIF1A degradation, and inhibition of PKB/Akt phosphorylation using PI3K inhibitor (LY294002) upregulated HIF1A ubiquitination. In summary, we propose that p53‐induced HIF1A degradation is not exclusively MDM2‐mediated, but reversible by PKB/Akt phosphorylation. J. Cell. Physiol. 222: 635–639, 2010. © 2009 Wiley‐Liss, Inc.
High glucose increases expression of cyclooxygenase‐2, increases oxidative stress and decreases the generation of nitric oxide in mouse microvessel endothelial cells
Authors: Aljofan Mohamad , Ding Hong
Published: November 2009

Hyperglycaemia is a key factor that contributes to the development of diabetes‐related microvascular disease. Both cyclooxygenase I and cyclooxygenase II are expressed in endothelial cells and play key roles in the regulation of cardiovascular function. In the current study we tested the hypothesis that hyperglycaemia‐induced increased expression of cyclooxygenase II is a contributing factor both to the increased oxidative stress and to the reduction in the generation of nitric oxide in microvessel endothelial cells following their exposure to high glucose. We demonstrated that the exposure of mouse microvascular endothelial cells to high glucose for 3 days decreased the generation of nitric oxide and enhanced production of superoxide. Western blots illustrated that exposure to high glucose also increased endothelial nitric oxide synthase and cyclooxygenase II protein expression levels and decreased the dimer/monomer ratio of endothelial nitric oxide synthase protein. All the changes induced by the high glucose culture media could be reversed by either the cyclooxygenase II inhibitor CAY10404, the non‐selective cyclooxygenase inhibitor indomethacin or the protein kinase C inhibitor chelerythrine, but not solely by preincubation with the antioxidant and putative NADPH oxidase inhibitor, apocynin. Our data indicate that high glucose induced oxidative stress is linked to an increase in the expression of cyclooxygenase II and a reduced generation of nitric oxide that is associated with an uncoupled endothelial nitric oxide synthase, possibly due to decreased dimer/monomer ratio. J. Cell. Physiol. 222: 669–675, 2010. © 2009 Wiley‐Liss, Inc.
Natural selection coupled with intragenic recombination shapes diversity patterns in the major histocompatibility complex class II genes of the giant panda
Authors: Chen Yi‐Yan , Zhang Ying‐Ying , Zhang He‐Min , Ge Yun‐Fa , Wan Qiu‐Hong , Fang Sheng‐Guo
Published: November 2009

Ample variations of the major histocompatibility complex (MHC) genes are essential for vertebrates to adapt to various environmental conditions. In this study, we investigated the genetic variations and evolutionary patterns of seven functional MHC class II genes (one DRA, two DRB, two DQA, and two DQB) of the giant panda. The results showed the presence of two monomorphic loci (DRA and DQB2) and five polymorphic loci with different numbers of alleles (seven at DRB1, six at DRB3, seven at DQA1, four at DQA2, six at DQB1). The presence of balancing selection in the giant panda was supported by the following pieces of evidence: (1) The observed heterozygosity was higher than expected. (2) Amino acid heterozygosity was significantly higher at antigen‐binding sites (ABS) compared with non‐ABS sequences. (3) The selection parameter ω (dN/dS) was significantly higher at ABS compared with non‐ABS sequences. (4) Approximately 95.45% of the positively selected codons (P>0.95) were located at or adjacent to an ABS. Furthermore, this study showed that (1) The Qinling subspecies exhibited high ω values across each locus (all >1), supporting its extensive positive selection. (2) The Sichuan subspecies displayed small ω at DRB1 (ω<0.72) and DQA2 (ω<0.48), suggesting that these sites underwent strong purifying selection. (3) Intragenic recombination was detected in DRB1, DQA1, and DQB1. The molecular diversity in classic Aime‐MHC class II genes implies that the giant panda had evolved relatively abundant variations in its adaptive immunity along the history of host–pathogen co‐evolution. Collectively, these findings indicate that natural selection accompanied by recombination drives the contrasting diversity patterns of the MHC class II genes between the two studied subspecies of giant panda. J. Exp. Zool. (Mol. Dev. Evol.) 314B:208–223, 2010. © 2009 Wiley‐Liss, Inc.
Mystery unsolved: missing limbs in deformed amphibians
Authors: Skelly David K. , Benard Michael F.
Published: November 2009

Ballengee and Sessions (2009) claim that predatory attacks by small predators such asSympetrumdragonfly larvae are sufficient to explain amphibian limb deformities in which the limb is partly or completely missing. This deformity type, the most common in nature, is not well explained byRibeiroiainfection which has also been nominated as a mechanism for limb deformities. We argue that the conclusions of the Ballanegee and Sessions study are not well founded. In part this is because the authors have provided no quantitative analysis of the association between limb deformities and predator densities. Our own data on frequencies of limb deformities suggest that missing hind limbs are often extremely rare even whenSympetrumand other small predators are common. While predatory attacks may contribute to observations of limb deformities, further study will be required to elucidate their role; other potential mechanisms deserve study as well. It is premature, and counterproductive, to draw any conclusions regarding the mechanisms behind the most common limb deformities recorded in natural populations.J. Exp. Zool. (Mol. Dev. Evol.) 314B:179–181, 2010. © 2009 Wiley‐Liss, Inc.
Radiosynthesis of 4‐[18F]fluoromethyl‐L‐phenylalanine and [18F]FET via a same strategy and automated synthesis module
Authors: Kersemans Ken , Mertens John , Caveliers Vicky
Published: November 2009

Currently there is still a need for more potent amino acid analogues as tumour imaging agents for peripheral tumour imaging with PET as it was recently reported that the success of O‐(2′‐[18F]fluoroethyl)‐L‐tyrosine ([18F]FET) is limited to brain, head and neck tumours. As the earlier described 2‐Amino‐3‐(2‐[18F]fluoromethyl‐phenyl)‐propionic acid (2‐[18F]FMP) suffered from intramolecular‐catalysed defluorination, we synthesized 2‐Amino‐3‐(4‐[18F]fluoromethyl‐phenyl)‐propionic acid (4‐[18F]FMP) as an alternative for tumour imaging with PET. Radiosynthesis of 4‐[18F]FMP, based on Br for [18F] aliphatic nucleophilic exchange, was performed with a customized modular Scintomics automatic synthesis hotboxthree system in a high overall yield of 30% and with a radiochemical purity of \gt 99%. 4‐[18F]FMP was found to be stable in its radiopharmaceutical formulation, even at high radioactivity concentrations. Additionally, for a comparative study, [18F]FET was synthesized using the same setup in 40% overall yield, with a radiochemical purity \gt 99%. The described automated radiosynthesis allows the production of two different amino acid analogues with minor alternations to the parameter settings of the automated system, rendering this unit versatile for both research and clinical practice. Copyright © 2009 John Wiley & Sons, Ltd.
Protein–surface interactions: challenging experiments and computations
Authors: Cohavi Ori , Corni Stefano , De Rienzo Francesca , Di Felice Rosa , Gottschalk Kay E. , Hoefling Martin , Kokh Daria , Molinari Elisa , Schreiber Gideon , Vaskevich Alexander , Wade Rebecca C.
Published: November 2009

Protein–surface interactions are fundamental in natural processes, and have great potential for applications ranging from nanotechnology to medicine. A recent workshop highlighted the current achievements and the main challenges in the field. Copyright © 2009 John Wiley & Sons, Ltd.
Viral infection in bone marrow transplants: Is JC virus involved?
Authors: Mischitelli Monica , Fioriti Daniela , Anzivino Elena , Bellizzi Anna , Barucca Valentina , Boldorini Renzo , Miglio Umberto , Sica Simona , Sorà Federica , De Matteis Silvia , Chiarini Fernanda , Pietropaolo Valeria
Published: November 2009

Hemorrhagic cystitis is characterized by hematuria due to inflammation of the bladder. In bone marrow transplants, this disease is linked to the infection by human polyomavirus BK, whereas the role of the human polyomavirus JC is unclear. The transcriptional control regions of both viruses contain important cellular transcription factor binding sites that undergo rearrangement process generating suitable variants that could be more active for viral replication and for the onset of hemorrhagic cystitis. In this study urine obtained from seven patients with bone marrow transplant were examined. Polyomavirus genomes were quantified by PCR and viral loads were compared. The transcriptional regions of both viruses were amplified and sequenced to determine the presence of variants. Subtypes of polyomaviruses were determined by amplification and sequencing of the viral protein 1 region. The results showed that four of seven patients were positive for BK DNA, two of seven patients had BK and JC DNA and one of seven had JC DNA. Positive samples were amplified and sequenced successively for transcriptional regions. The viral archetype was always found in both viruses. Finally, typing showed that BK virus subtype I infected patients with BK, whereas JC virus genotype IA and genotype 1B were found in patients infected with JC. The data suggest that new and different approaches are required to improve the morbidity and mortality caused by polyoma‐associated hemorrhagic cystitis, since it known that BK virus is involved in the onset of hemorrhagic cystitis, whereas the role of JC virus should be investigated further. J. Med. Virol. 82:138–145, 2010. © 2009 Wiley‐Liss, Inc.
Detection of a new yellow fever virus lineage within the South American genotype I in Brazil
Authors: de Souza Renato P. , Foster Peter G. , Sallum Maria Anice M. , Coimbra Terezinha L.M. , Maeda Adriana Y. , Silveira Vivian R. , Moreno Eduardo S. , da Silva Fernanda G. , Rocco Iray M. , Ferreira Ivani B. , Suzuki Akemi , Oshiro Fabíola M. , Petrella Selma M.C.N. , Pereira Luiz E. , Katz Giselda , Tengan Cilea H. , Siciliano Melissa M. , dos Santos Cecília L.S.
Published: November 2009

Nucleotide sequences of two regions of the genomes of 11 yellow fever virus (YFV) samples isolated from monkeys or humans with symptomatic yellow fever (YF) in Brazil in 2000, 2004, and 2008 were determined with the objective of establishing the genotypes and studying the genetic variation. Results of the Bayesian phylogenetic analysis showed that sequences generated from strains from 2004 and 2008 formed a new subclade within the clade 1 of the South American genotype I. The new subgroup is here designated as 1E. Sequences of YFV strains recovered in 2000 belong to the subclade 1D, which comprises previously characterized YFV strains from Brazil. Molecular dating analyses suggested that the new subclade 1E started diversifying from 1D about 1975 and that the most recent 2004–2008 isolates arose about 1985. J. Med. Virol. 82:175–185, 2010. © 2009 Wiley‐Liss, Inc.
Influence of amino‐acid polymorphism in the core protein on progression of liver disease in patients infected with hepatitis C virus genotype 1b
Authors: Kobayashi Mariko , Akuta Norio , Suzuki Fumitaka , Hosaka Tetsuya , Sezaki Hitomi , Kobayashi Masahiro , Suzuki Yoshiyuki , Arase Yasuji , Ikeda Kenji , Watahiki Sachiyo , Mineta Rie , Iwasaki Satomi , Miyakawa Yuzo , Kumada Hiromitsu
Published: November 2009

The substitution of amino acid (aa) 70 of arginine for glutamine and/or that of aa91 of leucine for methionine in the core protein in patients infected with hepatitis C virus (HCV) genotype 1b is associated with a poor response to pegylated interferon and ribavirin. Factors influencing these substitutions were sought in 1,097 patients infected with HCV‐1b who had not received antiviral treatment. HCV variants with Arg70 and Leu91 (wild‐type) decreased, while those with Gln70 and/or Met91 (mutant types) increased with age (P < 0.001). Of the 1,097 patients, 464 (42.3%) were infected with the Gln70 variant and the remaining 633 patients with the Arg70 variant. The proportion of patients with the Gln70 variant increased with the severity of liver disease (P < 0.001), elevated γ‐glutamyl transpeptidase (γ‐GTP) levels (P < 0.001) and a decrease in platelet count (P = 0.008). In univariate analysis patients with hepatocellular carcinoma, elevated aspartate aminotransferase (AST ≥ 58 IU/L) and γ‐GTP (≥61 IU/L), and decreased albumin levels (<3.9 g/dl) were more frequent in the patients with the Gln70 variant than the Arg70 variant (P = 0.003, 0.005, <0.001, and 0.031, respectively). In multivariate analysis HCC (odds ratio 1.829 [95% confidence interval 1.147–2.917]) and γ‐GTP ≥61 IU/L (1.647 [1.268–2.139]) increased the risk for the Gln70 variant. In conclusion, the substitution of amino aa70 of Arg for Gln in patients infected with HCV‐1b increases with age, and it is associated with severe liver disease accompanied by elevated AST and γ‐GTP levels, as well as the development of hepatocellular carinoma. J. Med. Virol. 82:41–48, 2010. © 2009 Wiley‐Liss, Inc.
Long‐term shedding of hepatitis E virus in the feces of pigs infected naturally, born to sows with and without maternal antibodies
Authors: Kanai Yuta , Tsujikawa Muneo , Yunoki Mikihiro , Nishiyama Shoko , Ikuta Kazuyoshi , Hagiwara Katsuro
Published: November 2009

Pigs are presumed reservoirs for hepatitis E virus (HEV) transmission to humans. To examine infection kinetics, two litters of domestic pigs (A and B, each containing 10 piglets) infected naturally with HEV were studied until pigs were 6 months old. Maternal IgG and IgA antibodies were detected in litter A piglets, but not in litter B ones. All pigs shed HEV in feces when they were 30–110 days old, and 17 developed viremia at 40–100 days of age. Phylogenetic analysis revealed a highly close sequence of HEV genotype 3 in all pigs. The serum levels of specific IgG and IgA were similar in all pigs, although IgA was not detected in the feces. Interestingly, the onset of both viremia and seroconversion was delayed significantly in litter A pigs. The kinetics of fecal virus shedding was similar in both litters; shedding was not detected after the pigs were 120 days old. The differences in the infection kinetics between litters A and B suggested that maternal antibodies delayed the onset of viremia and seroconversion. Quantitative real‐time reverse transcriptase‐polymerase chain reaction revealed that HEV RNA in feces peaked 10 days after initial shedding of approximately 106.0 copies/g. The viral load was much lower in the serum than in the feces. At 200 days of age, HEV RNA was found in the internal organs of 3 out of 13 pigs. These study findings improve the understanding of the dynamics of natural HEV transmission in pigs, which could help in controlling virus transmission from pigs to humans. J. Med. Virol. 82:69–76, 2010. © 2009 Wiley‐Liss, Inc.
Passage in egg culture is a major cause of apparent positive selection in influenza B hemagglutinin
Authors: Gatherer Derek
Published: November 2009

Several studies have identified residues apparently under positive selection in influenza B virus hemagglutinin. Host immune evasion is the main mechanism proposed to exert this selection pressure. However, these reports have not considered the culture history of the strains used to calculate positive selection. This article shows that passage of influenza B virus through egg culture is a strong contributory factor to the strength and statistical significance of positive selection on hemagglutinin. Non‐synonymous mutations resulting in the loss of the N‐glycosylation site at positions 197–199 of hemagglutinin have been positively selected to a far greater degree in egg‐cultured strains than in other strains. Once egg‐cultured strains are removed from the analysis, positive selection is found to be far weaker, less statistically significant, and more diffusely localized along the protein. Caution should therefore be exercised both in claims for the existence of positive selection in influenza B hemagglutinin, and in attribution of host immune evasion as its cause. The major cause of molecular adaptation in influenza B hemagglutinin proteins may well be laboratory eggs rather than natural hosts. J. Med. Virol. 82:123–127, 2010. © 2009 Wiley‐Liss, Inc.
Detection of GII‐4/2006b variant and recombinant noroviruses in children with acute gastroenteritis, South Korea
Authors: Chung Ju‐Young , Han Tae‐Hee , Park Sang‐Hun , Kim Sang Woo , Hwang Eung‐Soo
Published: November 2009

Norovirus (NoV), a single‐stranded, positive RNA virus, is an important etiologic agent of acute gastroenteritis in children worldwide. In this study, a total of 434 fecal samples collected from 434 children with acute gastroenteritis in Seoul, between September 2007 and July 2008 were tested to determine the molecular epidemiology of NoVs and characterize recombinant strains by using RT‐PCR followed by sequencing. Of the 434 specimens, NoV, rotavirus, and adenovirus were detected in 155 (35.8%), 72 (16.6%), and 19 specimens (4.3%), respectively. NoV GI was detected in 7 specimens (1.6%) and GII in 148 (34.1%) specimens. Phylogenetic analysis of capsid sequences in the GII‐positive specimens revealed the presence of the following strains: GII‐4, 111 (75.0%); GII‐3, 35 cases (23.6%); GII‐6b, 1 case; and GII‐16, 1 case. Most of the GII‐4 strains were grouped with the GII‐4/2006b variant with 98–100% nucleotide identity. Eleven strains were identified as recombinant (GII‐4/GII‐3 in 10 cases and GII‐b polymerase/GII‐16 capsid in 1 case) by sequencing based on the RdRP and capsid genes. The putative recombination point in the recombinant strains was the ORF‐1/ORF2 overlap, located at nucleotide 5,046 with reference to Lordsdale. In conclusion, GII‐4/2006b variants were detected predominantly and a new recombinant strain (GII‐4/GII‐3) was found in the Korean children with gastroenteritis. Continuous monitoring of the genetic diversity of NoVs is important to determine the trend of the predominant genotype and new recombinant strain. J. Med. Virol. 82:146–152, 2010. © 2009 Wiley‐Liss, Inc.
Proteomic analysis of cell lines expressing small hepatitis B surface antigen revealed decreased glucose‐regulated protein 78 kDa expression in association with higher susceptibility to apoptosis
Authors: Zhao Chao , Zhang Wei , Tian Xiaochen , Fang Caiyun , Lu Haojie , Yuan Zhenghong , Yang Pengyuan , Wen Yumei
Published: November 2009

Accumulating evidence suggests a key role of hepatocyte apoptosis in the pathogenesis of viral hepatitis B. It was found in this study that stable expression of small hepatitis B surface antigen (SHBs) in HepG2 and Huh7 cells increased susceptibility to apoptosis. Proteomic analysis of SHBs expressing HepG2 cells revealed 43 down‐regulated and 38 up‐regulated proteins. Some have been implicated in apoptosis, including glucose‐regulated protein 78 kDa (GRP78), heterogeneous nuclear ribonucleoprotein H3 (hnRNP H), Rho GDP dissociation inhibitor (GDI), cystatin B, far upstream element‐binding protein (FUSEbp), and TNF receptor‐associated protein 1 (TRAP1). Differential expression of GRP78 and several other proteins was confirmed by Western blot analysis. Replenishing GRP78 improved cellular resistance to apoptosis, whereas reduction of GRP78 by siRNA increased susceptibility even in the absence of SHBs. Taken together, these results suggest that HBsAg plays a pro‐apoptotic role through down‐regulation of GRP78. J. Med. Virol. 82:14–22, 2010. © 2009 Wiley‐Liss, Inc.
Molecular epidemiology of HCV genotypes among injection drug users in Taiwan: Full‐length sequences of two new subtype 6w strains and a recombinant form_2b6w
Authors: Lee Yuan‐Ming , Lin Huang‐Jie , Chen Yen‐Ju , Lee Cheng‐Ming , Wang Sheng‐Fan , Chang Kai‐Yuan , Chen Tzu‐Lang , Liu Hsin‐Fu , Chen Yi‐Ming Arthur
Published: November 2009

Human immunodeficiency virus type 1 (HIV‐1) circulating recombinant form (CRF) 07_BC strain has caused serious outbreaks among injection drug users in Taiwan since 2004. The objective of this study was to conduct a molecular epidemiological study of HCV genotypes in intravenous drug users in Taiwan. Blood samples and questionnaires from 591 intravenous drug users infected with HIV‐1 were collected nationwide. In total, 180 samples were selected for HCV genotyping using multiplex PCR and phylogenetic analysis of the core, E1 and NS5B regions. The Inno‐Lipa assay was used to confirm multiple infections with different genotypes. Eighty percent had a single infection with subtype 1b being the most common subtype (24%), 12% had double infections and two had triple infections. In addition, three recombinant forms (RFs)‐2a1a, 3a1b, and 2b6w were identified. Phylogenetic analyses showed that the 3a, 6a, and 6n strains were clustered with strains present in Thailand and mainland China. Full‐length sequence analysis showed that two 6w strains shared 89.4–90.2% sequence homology with the 6(r) strain from the Guangdong Province, China. Bootscan analysis revealed that the recombination breakpoint of RF_2b6w was located at the NS2‐NS3 junction. In summary, the distribution of HCV genotypes among Taiwanese intravenous drug users was complex and more than 12% of the drug users were infected with more than one genotype of HCV. J. Med. Virol. 82:57–68, 2010. © 2009 Wiley‐Liss, Inc.
Molecular epidemiology of KI and WU polyomaviruses in infants with acute respiratory disease and in adult hematopoietic stem cell transplant recipients
Authors: Debiaggi Maurizia , Canducci Filippo , Brerra Roberto , Sampaolo Michela , Marinozzi Maria Chiara , Parea Maurizio , Arghittu Milena , Alessandrino Emilio Paolo , Nava Stefano , Nucleo Elisabetta , Romero Egidio , Clementi Massimo
Published: November 2009

Polyomaviruses KI (KIPyV) and WU (WUPyV) were described recently in children with acute respiratory disease. The pathogenic potential of these human viruses has not been determined completely, but a correlation between immunosuppression and virus reactivation has been suggested. In the present study, the association between KI/WUPyV infection and immunosuppression was investigated using sequential nasopharyngeal aspirates from asymptomatic adult hematopoietic stem cell transplant recipients. In parallel, an investigation on the WU/KIPyV prevalence in children with acute respiratory disease was also carried out. Two of the 126 samples obtained from the 31 hematopoietic transplant recipients were positive for KIPyV (1 sample, 0.79%) and WUPyV (1 sample, 0.79%). Both samples were obtained 15 days after allogeneic transplantation and virus persistence was not observed in subsequent samples. In symptomatic children, 7 of the 486 nasopharyngeal aspirates were positive for WUPyV (1.4%) and 1 for KIPyV (0.2%). Single polyomavirus infection was detected in four patients, whereas the remaining patients were co‐infected with respiratory syncityal virus (three patients) or adenovirus (one patient). The results suggest that WU/KIPyVs have a limited circulation in Italy and a low pathogenic potential in young children. Brief and asymptomatic infection can occur in hematopoietic transplant recipients. J. Med. Virol. 82:153–156, 2010. © 2009 Wiley‐Liss, Inc.
Role of the HBx oncoprotein in carbonic anhydrase 9 induction
Authors: Holotnakova Tereza , Tylkova Lucia , Takacova Martina , Kopacek Juraj , Petrik Juraj , Pastorekova Silvia , Pastorek Jaromir
Published: November 2009

Carbonic anhydrase 9 (CA9), as one of the most hypoxia‐responsive genes, has been associated almost exclusively with hypoxic tumors. Its principal role is in pH regulation which helps tumor cells overcome intracellular acidosis and survive extended periods of time with low oxygen. Hypoxia‐inducible factor 1 (HIF‐1) is the main transcriptional activator of CA9. Hepatitis B virus X protein (HBx) has been shown to increase the transcriptional activity of HIF‐1. HBx is often expressed from the gene integrated in the hepatocytes infected persistently and contributes significantly to alterations in host gene expression that can lead to the development of hepatocellular carcinoma (HCC) associated with Hepatitis B virus (HBV). The aim of this study was to determine the effect of HBx on expression of CA9. Transient transfection of HBx led to an increase in the expression of CA9 as assessed by RT‐PCR and Western blotting. HBx was able to increase CA9 promoter activity significantly in several cell lines. The effect was mediated via HIF‐1 and a functional HRE element located −10/−3 bp upstream of the CA9 transcription initiation site. These data suggest that CA9 may be involved in the development of HCC by contributing to the survival of hepatocytes infected with HBV in liver tissue with fibrosis. J. Med. Virol. 82:32–40, 2010. © 2009 Wiley‐Liss, Inc.
Do human polyoma viruses and human immunodeficiency virus share common co‐receptors?
Authors: Borissov Kalin , Tsekov Iliya , Gavazova Rayna , Kalvatchev Zlatko , Argirova Radka
Published: November 2009

Host and/or viral factors involved in human polyomavirus (HPoV) infection in persons living with HIV remain unknown. A hypothesis is outlined suggesting the importance of the co‐receptors CCR5, CCR2, and CXCR4 not only for HIV, but also for HPoV. Functionally capable receptors coded by wild‐type (wt) genotypes could facilitate internalization of HPoV in the cell resulting in brain and/or kidney infection/s in HIV infected individuals. Forty‐nine Bulgarians with HIV, all treated by HAART, without neurological and/or kidney disorders, were tested for JCV and BKV and genotyped for CCR5 (CCR5del32), CCR2 (CCR2‐64I), and CXCR4 (SDF1‐3′A). In 27/49 (55.1%) individuals a co‐infection with HPoV was identified—BKV in 12/49 (24.5%), JCV—in another 12/49 (24.5%), and both viruses—in 3/49 (6.1%). A high frequency of wt CCR5 was found in patients with HPoV (91.7% for BKV and JCV and in 100% with both viruses). V/V of CCR2 was presented in 75% for BKV and JCV and in 66.7% for BKV plus JCV. SDF1‐3′G/G predominated in JCV infected patients (75%), while G/A and A/A genotypes were more frequent in patients with BKV (41.7%). Also, 21/22 (95.4%) persons without HPoV infection were heterozygous for SDF1 and CCR2. The number of individuals bearing wt of all co‐receptors in the group of persons not infected with HPoV was lower (P = 0.03) than that with polymorphism/s in one or two genes (SDF1 and CCR2) in the same group. The results suggest a probable role of co‐receptors used by HIV to facilitate infection with HPoV. J. Med. Virol. 82:8–13, 2010. © 2009 Wiley‐Liss, Inc.
Elevated levels of macrophage colony‐stimulating factor in human fracture healing
Authors: Sarahrudi Kambiz , Mousavi Mehdi , Thomas Anita , Eipeldauer Stefan , Vécsei Vilmos , Pietschmann Peter , Aharinejad Seyedhossein
Published: November 2009

Macrophage colony‐stimulating factor (M‐CSF) plays a unique role in bone remodeling. However, to our knowledge, no data on the role of M‐CSF in fracture healing in humans have been published so far. This study addressed this issue. One hundred and thirteen patients with long‐bone fractures were included in the study and divided into two groups, according to their course of fracture healing. The first group contained 103 patients with normal fracture healing. Ten patients with impaired fracture healing formed the second group of the study. Volunteers donated blood samples as control. Serum samples were collected over a period of 6 months, following a standardized time schedule. In addition, M‐CSF levels were measured in fracture hematoma and serum of 11 patients with bone fractures. M‐CSF concentrations were measured by enzyme‐linked immunosorbent assay (ELISA). Fracture hematoma contained significantly higher M‐CSF concentrations compared to M‐CSF concentrations in patient's serum. M‐CSF levels in fracture hematoma and in patient's serum were both significantly higher than M‐CSF concentrations measured in serum of healthy controls. Highly elevated M‐CSF serum concentrations were found in patients with physiological fracture healing over the entire observation period. Significant differences in the M‐CSF serum concentration between patients with normal fracture healing and patients with impaired fracture healing were not observed. This study indicates, for the first time, to our knowledge, a possible local and systemic involvement of M‐CSF in humans during fracture healing. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:671–676, 2010
Amino acid geochronology: its impact on our understanding of the Quaternary stratigraphy of the British Isles
Authors: Penkman Kirsty
Published: November 2009

This review aims to act as a brief primer to amino acid racemization and protein breakdown, as well as to document the major applications and results of amino acid dating on British Quaternary sediments. Amino acid results from fossil shells have had far‐reaching effects in developing our current understanding of British Quaternary stratigraphy and its links to continental Europe and the marine record. To appreciate the potential for amino acid geochronology, the technique's limitations and recent developments are discussed, as well as the future of the field. Copyright © 2009 John Wiley & Sons, Ltd.
Site‐specific deposition of Ag nanoparticles on ZnO nanorod arrays via galvanic reduction and their SERS applications
Authors: Song Wei , Han Xiaoxia , Chen Lei , Yang Youming , Tang Bin , Ji Wei , Ruan Weidong , Xu Weiqing , Zhao Bing , Ozaki Yukihiro
Published: November 2009

A controllable heterostructure consisting of ZnO nanorod arrays with attached Ag nanoparticles at only one end has been synthesized via a facile and convenient galvanic reduction method. Scanning electron microscopic images of these nanostructures showed good selectivity of Ag deposition on the tip of ZnO nanorod arrays. The formation of these regular AgZnO heterogeneous nanorod arrays can be explained by a localization of the electrons at the ends of the ZnO nanorods after the electron transfer step. By tuning the reaction time and the concentration of silver nitrate, the density of Ag nanoparticles on the tip of ZnO nanorods can be well controlled. Owing to the introduction of Ag nanoparticles with different densities, the resulting AgZnO heterogeneous nanorod arrays have been proved to be a versatile substrate for surface‐enhanced Raman scattering not only for common organic molecules but also for label‐free protein detection. Copyright © 2009 John Wiley & Sons, Ltd.
Protein adsorption drastically reduces surface‐enhanced Raman signal of dye molecules
Authors: Zhang Dongmao , Ansar Siyam M. , Vangala Karthikkeshwar , Jiang Dongping
Published: November 2009

There is an increasing interest in developing surface enhancement Raman spectroscopy methods for intracellular biomolecule and for in vitro protein detection that involve dye or protein–dye conjugates. In this work, we have demonstrated that protein adsorption on silver nanoparticle (AgNP) can significantly attenuate the surface‐enhanced Raman spectroscopy (SERS) signal of dye molecules in both protein/dye mixtures and protein/dye conjugates. SERS spectra of 12 protein/dye mixtures were acquired using 4 proteins [bovine serum albumin (BSA), lysozyme, trypsin, and concanavalin A] and three dyes [Rhodamine 6G, adenine, and fluorescein isothiocyanate (FITC)]. Besides the protein/dye mixtures, spectra were also obtained for the free dyes and four FITC‐conjugated proteins. While no SERS signal was observed in protein/FITC mixtures or conjugates, a significantly reduced SERS intensity (up to 3 orders of magnitude) was observed for both R6G and adenine in their respective protein mixtures. Quantitative estimation of the number of dye molecules absorbed onto AgNP implied that the degree of R6G SERS signal reduction in the R6G/BSA sample is 2 to 3 orders of magnitude higher than what could be accounted for by the difference in the amount of the absorbed dyes. This finding has significant implications for both intracellular SERS analyses and in vitro protein detection using SERS tagging strategies that rely on Raman dyes as reporter molecules. Copyright © 2009 John Wiley & Sons, Ltd.
Experimental and theoretical structure and vibrational analysis of ethyl trifluoroacetate, CF3CO2CH2CH3
Authors: Defonsi Lestard María Eliana , Tuttolomondo María Eugenia , Wann Derek A. , Robertson Heather E. , Rankin David W. H. , Altabef Aida Ben
Published: November 2009

The molecular structure and conformational properties of ethyl trifluoroacetate, CF3CO2CH2CH3, were determined in the gas phase by electron diffraction, and vibrational spectroscopy (IR and Raman). The experimental investigations were supplemented by ab initio (MP2) and DFT quantum chemical calculations at different levels of theory. Experimental and theoretical methods result in two structures with Cs (anti–anti) and C1 (anti–gauche) symmetries, the former being slightly more stable than the latter. The electron‐diffraction data are best fitted with a mixture of 56% anti–gauche and 44% anti–anti conformers. The conformational preference was also studied using the total energy scheme, and the natural bond orbital scheme. Also, the infrared spectra of CF3CO2CH2CH3 are reported for the gas, liquid and solid states, as is the Raman spectrum of the liquid. The comparison of experimental averaged IR spectra of Cs and C1 conformers provides evidence for the predicted conformations in the IR spectra. Harmonic vibrational wavenumbers and scaled force fields have been calculated for both conformers. Copyright © 2009 John Wiley & Sons, Ltd.
Recent advances in linear and non‐linear Raman spectroscopy. Part III
Authors: Kiefer W.
Published: November 2009

Following the first two reviews on recent advances in linear and non‐linear Raman spectroscopy, the present review summarises papers mainly published in the Journal of Raman Spectroscopy during 2008. This again serves to give a brief overview of recent advances in this research field and to provide readers of this journal a quick introduction to the various sub‐fields of Raman spectroscopy. It also reflects the current research interests and trends in the Raman community. Copyright © 2009 John Wiley & Sons, Ltd.
Transformation of pleomorphic adenoma to carcinoma ex pleomorphic adenoma of the parotid gland is independent of p53 mutations
Authors: Gedlicka Claudia , Item Chike B. , Wögerbauer Markus , Martinek Helga , Heiduschka Gregor , Erovic Boban M. , Ch.Grasl Matthaeus , Thurnher Dietmar
Published: November 2009

Background and Objectives

This retrospective study was performed to evaluate the status of p53 in pleomorphic adenomas and carcinomas ex pleomorphic adenoma in the parotid gland. As loss or mutation of p53 can cause malignant transformation, the possible degeneration of pleomorphic adenomas to carcinomas ex pleomorhic adenoma was investigated by mutational analysis.

Methods

Twenty‐five Patients including 14 patients with pleomorphic adenomas and 11 patients with carcinoma ex pleomorphic adenoma of the parotid gland were examined for p53 status. DNA was extracted out of paraffin‐embedded tissue and PCR was performed for the coding exons 2–11. Denaturing gradient gel electrophoresis (DGGE) was carried out for mutational analysis and DNA sequencing was performed in case of a suspected mutation.

Results

Fourteen pleomorphic adenomas and 11 carcinomas ex pleomorphic adenoma were screened for p53 status and potent mutations. Subsequent sequencing of the distinct exons showed no mutation.

Conclusion

We could not detect mutations of p53 neither in benign nor malignant parotid tumors and we therefore assume that p53 plays no role in the transformation from pleomorphic adenoma to carcinoma ex pleomorphic adenoma. J. Surg. Oncol. 2010;101:127–130. © 2009 Wiley‐Liss, Inc.
Lippia origanoides chemotype differentiation based on essential oil GC‐MS and principal component analysis
Authors: Stashenko Elena E. , Martínez Jairo R. , Ruíz Carlos A. , Arias Ginna , Durán Camilo , Salgar William , Cala Mónica
Published: November 2009

Chromatographic (GC/flame ionization detection, GC/MS) and statistical analyses were applied to the study of essential oils and extracts obtained from flowers, leaves, and stems of Lippia origanoides plants, growing wild in different Colombian regions. Retention indices, mass spectra, and standard substances were used in the identification of 139 substances detected in these essential oils and extracts. Principal component analysis allowed L. origanoides classification into three chemotypes, characterized according to their essential oil major components. α‐ and β‐phellandrenes, p‐cymene, and limonene distinguished chemotype A; carvacrol and thymol were the distinctive major components of chemotypes B and C, respectively. Pinocembrin (5,7‐dihydroxyflavanone) was found in L. origanoides chemotype A supercritical fluid (CO2) extract at a concentration of 0.83±0.03 mg/g of dry plant material, which makes this plant an interesting source of an important bioactive flavanone with diverse potential applications in cosmetic, food, and pharmaceutical products.
A sol‐gel based solid phase microextraction fiber for the analysis of aliphatic alcohols in apple juices
Authors: Farhadi Khalil , Maleki Ramin , Tahmasebi Raheleh
Published: November 2009

A new fiber based on titania‐chitin sol‐gel coated on a silver wire for the headspace solid phase microextraction of aliphatic alcohols from apple juice samples was developed. The influences of fiber coating composition and microextraction conditions (extraction temperature, extraction time, and ionic strength of the sample matrix) on the fiber performance were investigated. Also, the influence of temperature and time on desorption of analytes from fiber were studied. Under the optimized conditions, a porous fiber with a high extraction capacity and good thermal stability (up to 250°C) was obtained. The proposed headspace solid‐phase microextraction‐GC method was successfully used for the analysis of aliphatic alcohols in apple juice and concentrate samples. The recovery values were from 92.8 to 98.6%. The RSD (n=5) for all analytes were below 7.8%.
Adsorption of a single protein interacting with multiple ligands: Inner radial humps in the concentration profiles induced by non‐uniform ligand density distributions
Authors: Riccardi E. , Liapis A. I.
Published: November 2009

The dynamic behavior of the concentration profiles of a single protein in the pore solution and the adsorbed phase is studied in different adsorbent media when the spatial density distribution of the immobilized ligands is either uniform or non‐uniform and at the same time the single protein is forming one‐site and two‐site adsorbate–ligand complexes with the immobilized monovalent ligands. The competition for the formation of one‐site and two‐site interaction complexes leads to the formation of inner radial humps in the concentration profiles of the two‐site adsorbate–ligand complex in adsorbent particles having either uniform or non‐uniform spatial ligand density distributions. The results show that inner radial humps in the concentration profiles of the adsorbed protein (total concentration of adsorbed protein by one‐site and two‐site interactions) occur only in adsorbent media whose spatial ligand density distributions are non‐uniform and have maxima or minima occurring in radial positions located between the center and the outer surface of the particles. The non‐uniform spatial ligand density distributions satisfying this property provide the cause for the occurrence of inner radial humps in the concentration profiles of a single adsorbed protein, while the multi‐site adsorption interactions affect the magnitude and the rate of propagation of the inner radial humps in the concentration profiles of the single adsorbed protein. It is also demonstrated that adsorbent media having certain non‐uniform functional forms of spatial distribution in the density of immobilized ligands could provide more efficient adsorption of a protein than an adsorbent medium whose spatial distribution of the density of immobilized ligands is uniform. Furthermore, the results in this study suggest the type of information that could be obtained from finite bath experiments and could be used to (i) determine whether multi‐site adsorbate–ligand complexes are formed during the adsorption of a single adsorbate and (ii) select between alternative adsorbent media the adsorbent particles that could provide the highest overall adsorption rate for a given adsorbate of interest. The results clearly indicate that it is very important to study the dynamic behavior of an adsorption system having a non‐uniform spatial ligand density distribution and where the values of the pH and ionic strength are such that the electrophoretic effects are active.