bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transduction of Primary Hematopoietic Cells by Retroviral Vectors
Authors: Cornetta Kenneth , Pollok Karen E. , Miller A. Dusty 
Published: April 2008

INTRODUCTIONNoncycling cells are relatively resistant to transduction with retroviral vectors. Because most immortalized cell lines are actively proliferating, this is not an issue. However, for many primary cells, especially quiescent populations such as primitive progenitor and stem cells, the gene-transfer rate can be particularly low. This protocol describes transduction of primary hematopoietic cells. Two interventions are combined to maximize gene transfer in hematopoietic progenitor cells: (1) cytokines and other growth factors are used to stimulate cell cycling in hematopoietic cells, and (2) matrix proteins such as fibronectin are used to mediate colocalization of target cells and vector.
Scanning Electron Microscopy (SEM) of Plant Tissues
Authors: Bomblies Kirsten , Shukla Vipula , Graham Charles 
Published: April 2008

INTRODUCTIONScanning electron microscopy (SEM) is used to produce detailed images of surface structures. This article details the steps required to prepare a sample for SEM. In brief, the sample is fixed, dehydrated, subjected to critical-point drying, mounted, and coated with an electrically conducting material such as gold or palladium.
Mouse Mutagenesis Using N-Ethyl-N-Nitrosourea (ENU): Figure 1.
Authors: Salinger Andrew P. , Justice Monica J. 
Published: April 2008

INTRODUCTIONThis protocol describes chemical mutagenesis of male mice using N-ethyl-N-nitrosourea (ENU), which is the most efficient method for obtaining mouse mutations in phenotype-driven screens. A fractionated dose of ENU, an alkylating agent, can produce a mutation rate as high as 1.5 × 10−3 in male mouse spermatogonial stem cells. Treatment with ENU produces point mutations that provide a unique mutant resource: They reflect the consequences of single gene changes independent of position effects, provide a fine structure dissection of protein function, display a range of mutant effects from complete or partial loss of function to exaggerated function, and discover gene functions in an unbiased manner. After treatment with ENU, mice are mated in genetic screens designed to uncover mutations of interest. Screens for dominant, recessive, and modifying mutations can be performed.
Desorption Electrospray Ionization (DESI) Analysis of Intact Proteins/Oligopeptides
Authors: Takats Zoltan , Wiseman Justin M. , Ifa Demian R. , Cooks R. Graham 
Published: April 2008

INTRODUCTIONDesorption electrospray ionization (DESI) is amenable to the study of intact proteins in complex mixtures, including blood or other biological media. Intact proteins can be desorbed and ionized from the surface under gentle (soft) conditions to produce compact conformations of the protein. A procedure for DESI analysis of intact proteins and oligopeptides using mass spectrometry (MS) is described here. DESI-MS is an emerging technique with great promise, but its application range is still being investigated. Therefore, the protocol presented here provides general procedures used for the applications that have been investigated so far. Optimal ion source parameters and surface types may vary depending on the application.
In Situ Desorption Electrospray Ionization (DESI) Analysis of Tissue Sections
Authors: Takats Zoltan , Wiseman Justin M. , Ifa Demian R. , Cooks R. Graham 
Published: April 2008

INTRODUCTIONDesorption electrospray ionization (DESI) allows in situ analysis of biological tissues. The analysis of less abundant protein constituents within a tissue sample often requires the removal of lipid species prior to analysis, similar to the situation with matrix-assisted laser desorption/ionization (MALDI). After removal of lipid constituents, the tissue can be treated with protease to degrade proteins present in the tissue. The tryptic products can be investigated directly from the tissue using DESI. The spectra obtained feature ions of tryptic fragments from abundant proteins present in the tissue sample. The digestion is usually not complete; hence, the presence of missed cleavage sites is typical in the peptides detected. The signal is more stable for longer times than in the case of deposited samples, so the recording of mass spectrometry (MS)/MS data is simple in this case. DESI-MS is an emerging technique with great promise, but its application range is still being investigated. Therefore, the protocol for DESI-MS analysis of tissue sections presented here provides general procedures used for the applications that have been investigated so far. Optimal ion source parameters and surface types may vary depending on the application.
Retroviral Vectors for Gene Transfer
Authors: Cornetta Kenneth , Pollok Karen E. , Miller A. Dusty 
Published: April 2008

INTRODUCTIONRetroviral vectors from the γ-retrovirus genus were the first retroviral vectors to be developed. They have been called oncoretroviral vectors or simple retroviral vectors because of their derivation from oncogenic retroviruses having a simple gag-pol-env genome structure. Later additions to the retroviral vector family include the lentiviral and foamy viral vectors derived from more complex retroviruses that contain multiple accessory genes in addition to the standard gag-pol-env genes. This article describes the advantages and disadvantages of retroviral vectors for gene therapy. It also discusses the issues that must be considered in designing retroviral vectors and in choosing retroviral packaging cell lines.
The Maximum Likelihood Approach for Phylogenetic Prediction: Figure 1.
Authors: Mount David W. 
Published: April 2008

INTRODUCTIONMaximum likelihood (ML) methods are especially useful for phylogenetic prediction when there is considerable variation among the sequences in the multiple sequence alignment (msa) to be analyzed. ML methods start with a simple model, in this case a model of rates of evolutionary change in nucleic acid or protein sequences and tree models that represent a pattern of evolutionary change, and then adjust the model until there is a best fit to the observed data. For phylogenetic analysis, the observed data are the observed sequence variations found within the columns of an msa. The ML method is similar to the maximum parsimony method in that the analysis is performed on each column of an msa.
Identification of SMURF1 as a possible target for 7q21.3‐22.1 amplification detected in a pancreatic cancer cell line by in‐house array‐based comparative genomic hybridization
Authors: Suzuki Ayako , Shibata Tatsuhiro , Shimada Yutaka , Murakami Yoshinori , Horii Akira , Shiratori Keiko , Hirohashi Setsuo , Inazawa Johji , Imoto Issei 
Published: April 2008

Pancreatic cancer (PC) cell lines provide a useful starting point for the discovery and functional analysis of genes driving the genesis and progression of this lethal cancer. To increase our understanding of the gene copy number changes in pancreatic carcinomas and to identify key amplification and deletion targets, we applied genome‐wide array‐based comparative genomic hybridization using in‐house array (MCG Cancer Array‐800) to 24 PC cell lines. Overall, the analyses revealed high genomic complexity, with several copy number changes detected in each line. Homozygous deletions (log2ratio < –2) of eight genes (clones) were seen in 14 of the 24 cell lines, whereas high‐level amplifications (log2ratio > 2) of 10 genes (clones) were detected in seven lines. Among them, we focused on high‐level amplification at 7q22.1, because target genes for this alteration remain unknown. Through precise mapping of the altered region by fluorescence in situ hybridization, determination of the expression status of genes located within those regions, and functional analysis using knockdown of the gene expression or the ectopic overexpression approach in PC cell lines, as well as immunohistochemical analyses of candidates in primary tumors of PC, we successfully identified SMURF1 as having the greatest potential as a 7q21.3‐22.1 amplification target. SMURF1 may work as a growth‐promoting gene in PC through overexpression and might be a good candidate as a therapeutic target. Our results suggest that array‐based comparative genomic hybridization analysis combined with further genetic and functional examinations is a useful approach for identifying novel tumor‐associated genes involved in the pathogenesis of this lethal disease. (Cancer Sci 2008; 99: 986–994)
Induction of multinucleated cells and apoptosis in the PC‐3 prostate cancer cell line by low concentrations of polyethylene glycol 1000
Authors: Fukuta Katsuhiro , Kohri Kenjiro , Fukuda Hirokazu , Watanabe Masatoshi , Sugimura Takashi , Nakagama Hitoshi 
Published: April 2008

Polyethylene glycol (PEG) has been reported to inhibit the development of colonic lesions in carcinogen‐treated rats when administered orally. However, the precise mechanism for the chemopreventive activity of PEG remains largely elusive. Based on a characteristic feature of PEG as a ‘fusogen’, we investigated its potential as a chemotherapeutic agent through the induction of multinucleated cell formation and apoptosis induction in PC‐3 prostate cancer cells. When PC‐3 cells were treated with 0.5 and 1.0% PEG 1000, multinucleated cells were induced at a frequency of 8.4 and 13%, respectively, 36 h after PEG treatment under high cell density (1 × 106 cells in 100 µL PEG solution) in vitro. Although abnormality of cell cycle progression was not evident in PEG‐treated PC‐3 cells, multinucleated cells substantially disappeared at around 38 h due to apoptosis. In contrast, no apparent growth suppression was observed when PC‐3 cells were exposed to up to 1.0% PEG at a much lower cell density, namely under ordinary culture conditions. Furthermore, injection of 0.5% PEG solution in vivo into PC‐3 xenografts implanted in BALB/c‐nu/nu male mice significantly suppressed tumor growth compared to phosphate‐buffered saline injection. Multinucleated TdT‐mediated dUTP‐biotin nick end‐labeling (TUNEL)‐positive cells were observed inside the PEG‐injected tumors. PEG was here demonstrated to have anticell proliferation and antitumor effects via induction of apoptosis, possibly by cell fusion. PEG injection therapy could therefore be adopted as an alternative chemotherapeutic strategy for localized prostate cancers, including those that become refractory to androgen‐deprivation therapy. (Cancer Sci 2008; 99: 1055–1062)
Downregulation of Tie2 gene by a novel antitumor sulfolipid, 3′‐sulfoquinovosyl‐1′‐monoacylglycerol, targeting angiogenesis
Authors: Mori Yoko , Sahara Hiroeki , Matsumoto Kayo , Takahashi Nobuaki , Yamazaki Takayuki , Ohta Keisuke , Aoki Satoko , Miura Masahiko , Sugawara Fumio , Sakaguchi Kengo , Sato Noriyuki 
Published: April 2008

We previously reported that 3′‐sulfoquinovosyl‐1′‐monoacylglycerol (SQMG) was effective in suppressing the growth of solid tumors due to hemorrhagic necrosis in vivo. In the present study, we investigated the antiangiogenic effect of SQMG. In vivo assessment of antitumor assays showed that some tumor cell lines, but not others, were sensitive to SQMG. Microscopic study suggested that in SQMG‐sensitive tumors, but not SQMG‐resistant tumors, angiogenesis was reduced. We next investigated gene expression relating to angiogenesis in tumor tissues by quantitative real‐time polymerase chain reaction. Consequently, although vascular endothelial growth factor gene expression was not detected with significant differences among the cases, significant downregulation of Tie2 gene expression was observed in all SQMG‐sensitive tumors as compared with controls, but not in SQMG‐resistant tumors. These data suggested that the antitumor effects of SQMG could be attributed to antiangiogenic effects, possibly via the downregulation of Tie2 gene expression in SQMG‐sensitive tumors. (Cancer Sci 2008; 99: 1063–1070)
Effects of imatinib (Glivec) on the pharmacokinetics of metoprolol, a CYP2D6 substrate, in Chinese patients with chronic myelogenous leukaemia
Authors: Wang Yanfeng , Zhou Li , Dutreix Catherine , Leroy Elisabeth , Yin Qi , Sethuraman Venkat , Riviere Gilles‐Jacques , Yin Ophelia Q. P. , Schran Horst , Shen Zhi‐Xiang 
Published: April 2008

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Imatinib, a tyrosine kinase inhibitor, exhibits a competitive inhibition on the CYP450 2D6 isozyme with a Ki value of 7.5 μm. However, the clinical significance of the inhibition and its relevance to 2D6 polymorphisms have not been evaluated. The pharmacokinetics of imatinib have been well studied in Caucasians, but not in a Chinese population.• Metoprolol, a CYP2D6 substrate, has different clearances among patients with different CYP2D6 genotypes. It is often used as a CYP2D6 probe substrate for clinical drug–drug interaction studies. WHAT THIS STUDY ADDS • Co‐administration of imatinib at 400 mg twice daily increased the plasma AUC of metoprolol by approximately 23% in 20 Chinese patients with chronic myeloid leukaemia (CML), about 17% increase in CYP2D6 intermediate metabolizers (IMs) (n = 6), 24% in extensive metabolizers (EMs) (n = 13), and 28% for the subject with unknown 2D6 status (n = 1) suggesting that imatinib has a weak to moderate inhibition on CYP2D6 in vivo.• The clearance of imatinib in Chinese patients with CML showed no difference between CYP2D6 IMs and EMs, and no major difference from Caucasian patients with CML based on data reported in the literature.AIMSTo investigate the effect of imatinib on the pharmacokinetics of a CYP2D6 substrate, metoprolol, in patients with chronic myeloid leukaemia (CML). The pharmacokinetics of imatinib were also studied in these patients.METHODSPatients (n = 20) received a single oral dose of metoprolol 100 mg on day 1 after an overnight fast. On days 2–10, imatinib 400 mg was administered twice daily. On day 8, another 100 mg dose of metoprolol was administered 1 h after the morning dose of imatinib 400 mg. Blood samples for metoprolol and α‐hydroxymetoprolol measurement were taken on study days 1 and 8, and on day 8 for imatinib.RESULTSOf the 20 patients enrolled, six patients (30%) were CYP2D6 intermediate metabolizers (IMs), 13 (65%) extensive metabolizers (EMs), and the CYP2D6 status in one patient was unknown. In the presence of 400 mg twice daily imatinib, the mean metoprolol AUC was increased by 17% in IMs (from 1190 to 1390 ng ml−1 h), and 24% in EMs (from 660 to 818 ng ml−1 h). Patients classified as CYP2D6 IMs had an approximately 1.8‐fold higher plasma metoprolol exposure than those classified as EMs. The oral clearance of imatinib was 11.0 ± 2.0 l h−1 and 11.8 ± 4.1 l h−1 for CYP2D6 IMs and EMs, respectively.CONCLUSIONSCo‐administration of a high dose of imatinib resulted in a small or moderate increase in metoprolol plasma exposure in all patients regardless of CYP2D6 status. The clearance of imatinib showed no difference between CYP2D6 IMs and EMs.
Cow's milk allergy in adults is rare but severe: both casein and whey proteins are involved
Authors: Lam H‐Y. , Van Hoffen E. , Michelsen A. , Guikers K. , Van Der Tas C. H. W. , Bruijnzeel‐Koomen C. A. F. M. , Knulst A. C. 
Published: April 2008

SummaryBackground
Studies on cow's milk allergy (CMA) in adults are scarce. Little is known about the clinical symptoms, eliciting doses (ED), and allergens involved.Objective
The aim of this study was to analyse the clinical symptoms, ED and allergen recognition in adult CMA patients, compared with cow's milk (CM)‐sensitized, but tolerant controls.Methods
Adult CMA patients were evaluated by standardized questionnaires (n=30), skin prick tests (SPTs) and specific IgE for CM allergens (n=18), and a double‐blind placebo‐controlled food challenge (DBPCFC, n=10). A control group (n=25) of CM‐sensitized, but tolerant adults was included.Results
The majority of CMA patients (20/30, 67%) reported severe symptoms. In all patients participating in DBPCFC, CMA was confirmed. ED for subjective symptoms (0.3–300 mg CM protein) were significantly lower than that for objective symptoms (300–9000 mg CM protein). The severity of CMA by history and ED was not correlated with SPT or IgE. Patients had higher SPT reactivity than controls for CM, α‐lactalbumin and β‐lactoglobulin (P=0.002, P=0.014 and P=0.004) but not for casein. Specific IgE to CM tended to be higher (P=0.068) and IgE to casein was higher in patients than that in controls (P=0.016). No difference was observed for IgE to α‐lactalbumin and β‐lactoglobulin.Conclusion
Adult CMA is severe in nature. ED are low, starting from 0.3 mg CM protein. Patients with CMA recognize the same major allergens (casein and whey proteins) as controls, but display a stronger SPT and IgE reactivity.
IL‐13 induced increases in nitrite levels are primarily driven by increases in inducible nitric oxide synthase as compared with effects on arginases in human primary bronchial epithelial cells
Authors: Chibana K. , Trudeau J. B. , Mustovitch A. T. , Hu H. , Zhao J. , Balzar S. , Chu H. W. , Wenzel S. E. 
Published: April 2008

SummaryBackground
Exhaled nitric oxide is increased in asthma, but the mechanisms controlling its production, including the effects of T‐helper type 2 (Th2) cytokines, are poorly understood. In mouse and submerged human epithelial cells, Th2 cytokines inhibit expression of inducible nitric oxide synthase (iNOS). Arginases have been proposed to contribute to asthma pathogenesis by limiting the arginine substrate available to NOS enzymes, but expression of any of these enzymes has not been extensively studied in primary human cells.Objectives
We hypothesized that primary human airway epithelial cells in air–liquid interface (ALI) culture would increase iNOS expression and activity in response to IL‐13, while decreasing arginase expression.Methods
iNOS and arginase mRNA (real‐time PCR) and protein expression (Western blot and immunofluorescence) as well as iNOS activity (nitrite levels) were measured in ALI epithelial cells cultured from bronchial brushings of normal and asthmatic subjects following IL‐13 stimulation.Results
IL‐13 up‐regulated iNOS mRNA primarily at a transcriptional level in epithelial cells. iNOS protein and activity also increased, arginase1 protein expression decreased while arginase 2 expression did not change. The changes in iNOS protein correlated strongly with changes in nitrites, and inclusion of arginase (1 or 2) did not substantially change the relationship. Interestingly, iNOS mRNA and protein were not correlated.Conclusions
These results contrast with many previous results to confirm that Th2 stimuli enhance iNOS expression and activity. While arginase 1 protein decreases in response to IL‐13, neither arginase appears to substantially impact nitrite levels in this system.
Panbiogeographical analysis of the genus Bomarea (Alstroemeriaceae)
Authors: Alzate Fernando , Quijano‐Abril Mario Alberto , Morrone Juan J. 
Published: April 2008

AbstractAim  A panbiogeographical analysis of the genus Bomarea was undertaken in order to determine generalized tracks and biogeographical nodes, and to evaluate the current distribution of the genus based on the available tectonic information and the biogeographical regionalization of Latin America.Location  The Neotropical region from northern Mexico to northern Argentina, and the Nearctic and Andean regions.Method  A total of 2205 records of 101 species were analysed, representing 95% of the species assigned to Bomarea. Localities were represented on maps and their individual tracks were drawn. Based on their comparison, generalized tracks were detected and mapped. Nodes were identified in the areas where different generalized tracks were superimposed.Results  Five generalized tracks were recovered. One is located in the Eastern Central America and Western Panamanian Isthmus provinces (Caribbean subregion, Neotropical region), which was supported by three species of Central American distribution. The four remaining generalized tracks were located in South America, in the North Andean Paramo, Cauca and Puna biogeographical provinces. These tracks were supported by species of Bomarea with an Andean distribution. Biogeographical nodes were established in the Central Andean region of Colombia, central Ecuador and central Peru.Main conclusions  The nodes obtained for Bomarea support a hybrid origin for the Andean region, which presents diverse components from both northern and southern South America. Likewise, the track recovered between Colombia and Ecuador includes Andean and Neotropical areas, providing further support for this hypothesis. The nodes obtained are coherent with vicariant elements evident for Bomarea. Species of three clades proposed for Bomarea are distributed in specific generalized tracks.
Tertiary relict trees in a Mediterranean climate: abiotic constraints on the persistence of Prunus lusitanica at the eroding edge of its range
Authors: Pulido Fernando , Valladares Fernando , Calleja Juan Antonio , Moreno Gerardo , González‐Bornay Guillermo 
Published: April 2008

AbstractAim  To investigate the ecophysiological traits allowing persistence of a subtropical relict tree (Prunus lusitanica L.) under a dry Mediterranean climate at the eroding edge of its range.Location  A glasshouse for the study under controlled conditions and two marginal populations located in riparian forests of central Spain and exposed to summer drought, in contrast to subtropical populations that grow in mountain cloud forests.Methods  Two experiments were conducted to assess tolerance to the abiotic conditions found in riparian habitats. In a glasshouse experiment, gas‐exchange and light‐use parameters were measured to evaluate seedling responses to a factorial combination of irradiance (60%, 10%, 2% and 0.5% full sun) and moisture (well watered control and drought stress). In a parallel field experiment, irradiance and soil moisture were measured as predictors of seedling survival at two sites in which half the seedlings were subjected to a threefold increase in summer precipitation by adding water every 10 days.Results Soil moisture strongly determined survival both in the glasshouse and in the two field sites. In the field, periodic addition of water failed to increase survival. Water‐use efficiency (WUE) increased with drought. Seedlings did not tolerate deep shade (2–0.5%) and their performance and survival were better when exposed to moderate (10%) or high (60%) irradiance. The effect of water stress on seedling performance was stronger at moderate irradiance, moderate at high irradiance and negligible at very low light. Seedling size (height and number of leaves) attained after 1 month of experimental treatments had a positive effect on survival at the end of the summer, hence greater survival was not achieved at the expense of vertical growth.Main conclusions  While studies in Macaronesia have shown that P. lusitanica occupies a wide range of moisture conditions along mountain slopes, it behaves as an obligate riparian species in dry peripheral populations. Intolerance to deep shade and tolerance to moderate and high irradiance allow the species to grow in small and moist gaps, or in treeless river sections. Despite the species’ low range filling in marginal, drought‐prone regions, long‐term persistence might have been achieved through shifts to riparian habitats serving as local refuges.