bluuebunny/crossref_metadata_2025 huggingface dataset:
Adolescent musculoskeletal injuries in a football academy
Authors: Deehan D. J. , Bell K. , McCaskie A. W.
Published: January 2007

Interest in football continues to increase, with ever younger age groups participating at a competitive level. Football academies have sprung up under the umbrella of professional clubs in an attempt to nurture and develop such talent in a safe manner. However, increased participation predisposes the immature skeleton to injury. Over a five-year period we have prospectively collected data concerning all injuries presenting to the medical team at Newcastle United football academy. We identified 685 injuries in our cohort of 210 players with a mean age of 13.5 years (9 to 18). The majority of injuries (542;79%) were to the lower limb. A total of 20 surgical procedures were performed. Contact injuries accounted for 31% (210) of all injuries and non-contact for 69% (475).The peaks of injury occurred in early September and March. The 15- and 16-year-old age group appeared most at risk, independent of hours of participation. Strategies to minimise injury may be applicable in both the academy setting and the wider general community.
Amino-Terminal Pro–Brain Natriuretic Peptide, Brain Natriuretic Peptide, and Troponin T for Prediction of Mortality in Acute Heart Failure
Authors: Sakhuja Rahul , Green Sandy , Oestreicher Eveline M , Sluss Patrick M , Lee-Lewandrowski Elizabeth , Lewandrowski Kent B , Januzzi James L
Published: January 2007

Background: Combining testing for natriuretic peptides [amino-terminal pro–brain natriuretic peptide (NT-proBNP) and brain natriuretic peptide (BNP)] and cardiac troponin T (cTnT) may help predict mortality in patients with acute heart failure (HF).

Methods: We studied 209 patients with acute HF at an urban academic center and used ROC curves and multivariate analyses to examine the relationship of outcome to natriuretic peptide and cTnT concentrations at presentation.

Results: Higher concentrations of natriuretic peptides and cTnT at presentation were predictors of death at 60 days and 1 year (P <0.001 and P <0.01, respectively, at both time points). Optimal cutoff points for NT-proBNP, BNP, and cTnT for predicting death by 60 days or 1 year were 5562 and 3174 ng/L, 428 and 352 ng/L, and 0.01 and 0.01 μg/L, respectively. Most decedents demonstrated increased concentrations of both natriuretic peptides and cTnT and had a 25% mortality rate at the 60-day time point (P <0.001). Mortality rates were low (<4%) among patients with either no increase or an increase in only 1 marker. Decedents with increases in both a natriuretic peptide and cTnT at presentation had the highest death rate at 1 year (45%, P <0.001). This combination was strongly predictive of death [NT-proBNP plus cTnT: hazard ratio (HR), 7.66; 95% confidence interval (CI), 3.06–17.8; BNP plus cTnT: HR, 6.82; 95% CI, 2.99–16.5].

Conclusions: A dual-marker strategy incorporating a natriuretic peptide and cTnT is superior to either marker alone for estimating short- and longer-term risk in patients with acute HF.
Cystatin C and Estimates of Renal Function: Searching for a Better Measure of Kidney Function in Diabetic Patients
Authors: Pucci Laura , Triscornia Stefano , Lucchesi Daniela , Fotino Carmen , Pellegrini Giovanni , Pardini Ennia , Miccoli Roberto , Del Prato Stefano , Penno Giuseppe
Published: January 2007

Background: Early identification of impairment in renal function is crucial in diabetic patients. Serum cystatin C may be the most sensitive indicator of glomerular filtration rate (GFR) in the clinical setting.

Methods: We compared cystatin C with creatinine, the Cockcroft-Gault (C-G) formula, and the Modification of Diet in Renal Disease (MDRD) study equation for the assessment of early decreased renal function in 288 diabetic patients (125 type 1, 163 type 2) with renal impairment [GFR: 4–222 mL · min−1 · (1.73 m2)−1]. Relationships of cystatin C, creatinine, and iohexol clearance were linearized by plotting their reciprocals in a simple regression model. Diagnostic efficiency was calculated from ROC curves.

Results: In this study population, cystatin C (P = 0.0013) was better correlated with GFR (r = 0.857) than were creatinine (r = 0.772), C-G (r = 0.750), and MDRD (r = 0.806), a result replicated in patients with normal renal function (P = 0.023, type 1; P = 0.011, type 2), but not in those with decreased GFR. Mean cystatin C concentrations showed step-by-step statistically significant increases as GFR decreased, allowing very early detection of reduction in renal function. At 90 mL · min−1 · (1.73 m2)−1 and 75 mL · min−1 · (1.73 m2)−1 cut-points, diagnostic efficiencies of cystatin C (89% and 92%) were better than those of the other variables (79%–82% and 85%–86%, respectively; P = 0.01).

Conclusions: All data supported the value of serum cystatin C compared with conventional estimates based on serum creatinine measurement for detecting very early reduction of renal function. Use of cystatin C to measure renal function will optimize early detection, prevention, and treatment strategies for diabetic nephropathy.
Cell-Free Fetal DNA in Amniotic Fluid: Unique Fragmentation Signatures in Euploid and Aneuploid Fetuses
Authors: Lapaire Olav , Bianchi Diana W , Peter Inga , O’Brien Barbara , Stroh Helene , Cowan Janet M , Tantravahi Umadevi , Johnson Kirby L
Published: January 2007

Background: Circulating cell-free fetal deoxyribonucleic acids (cffDNA) are novel biomarkers with many clinical applications. Amniotic fluid (AF) is a rich source of cffDNA. We investigated the biophysical characteristics of cffDNA in AF, hypothesizing that they would differ from cffDNA in maternal plasma.

Methods: We obtained 10 mL of fresh AF supernatant from women carrying euploid fetuses (n = 39) and aneuploid fetuses (n = 4). To test the effects of storage and karyotype, samples from euploid fetuses (n = 19) and aneuploid fetuses with trisomies 21 (n = 16), 18 (n = 9), or 13 (n = 3); triploidy (n = 4); or monosomy X (n = 2) were frozen at −80 °C. AF cffDNA was characterized by real-time quantitative PCR amplification of glyceraldehyde-3-phosphate dehydrogenase, gel electrophoresis, and analysis of the DNA fragmentation signature.

Results: We observed a significant correlation of concentration with gestational age for fresh AF cffDNA from euploid fetuses (R2 = 0.77, P <0.0001) but not for frozen cffDNA (P = 0.63). The median amount of cffDNA in frozen euploid samples was significantly lower than in fresh samples (P <0.0001). After adjustment for gestational age, there was a statistically significant decrease in the median amount of cffDNA in frozen aneuploidy samples compared with frozen euploid samples (P = 0.0005). Analysis of the cffDNA size distribution showed different and qualitatively unique patterns for each karyotype.

Conclusions: Gestational age, karyotype, and sample storage time affect concentrations and fragment size of AF cff DNA. These effects may be attributable to fundamental differences in tissue sources, excretion modes, or kinetic pathways. Characteristic signature patterns for each common aneuploidy offer the possibility of using DNA fragmentation analysis as a means of triaging AF samples.
Influence of Seed Depth and Pathogens on Fatal Germination of Velvetleaf (Abutilon theophrasti) and Giant Foxtail (Setaria faberi)
Authors: Davis Adam S. , Renner Karen A.
Published: January 2007

Fatal germination of weed seeds occurs when a weed seed germinates, but the seedling dies before reaching the soil surface. Controlled-environment bioassays of velvetleaf and giant foxtail seed fate in Michigan field soil (Kalamazoo silt loam, 1.1% soil organic matter) were used to determine the role of pathogenic fungi and seed burial depth in fatal germination of these species. Fatal germination at 2 cm seed depth was nonexistent for giant foxtail, and rare (< 10% of seeds studied) for velvetleaf. At greater depths, fatal germination remained close to zero for giant foxtail, whereas it increased to as high as 40% for velvetleaf at a 10-cm burial depth. Cultures taken from fatally germinated velvetleaf seedlings were found to containPythium ultimum, a soilborne pathogen known as the causal agent for pea root rot. When samples of infected media taken from these cultures were used to inoculate field soil in pots, fatal germination of velvetleaf from depths of 4 to 6 cm increased, compared with field soil inoculated with sterile media. At seed burial depths of 8 and 10 cm, fatal germination of velvetleaf increased to 20 and 40%, respectively, and was the same for unsterilized soil andP. ultimum–inoculated soil. Given that maximal fatal germination of velvetleaf occurred in the unsterilized soil treatment at the 10 cm depth, burial of newly shed velvetleaf seeds to a 10 cm, or possibly greater, depth with tillage equipment may be a practical way of reducing velvetleaf seed banks through fatal germination.
Afterripening Requirements and Optimal Germination Temperatures for Nuttall's Alkaligrass (Puccinellia nuttalliana) and Weeping Alkaligrass (Puccinellia distans)
Authors: Tarasoff Catherine S. , Ball Daniel A. , Mallory-Smith Carol A.
Published: January 2007

In the Grande Ronde Valley of eastern Oregon, two perennial grass species in the genusPuccinellia, weeping alkaligrass and Nuttall's alkaligrass, are weeds of Kentucky bluegrass grass-seed production fields. Weeping alkaligrass is introduced from Eurasia, whereas Nuttall's alkaligrass is native to the region. These two species were studied to determine dormancy attributes and optimal temperature conditions for seed germination. Results from the current studies indicate that both species have a high level of embryonic dormancy immediately following seed harvest, which is primarily eliminated through dry storage (afterripening) and an incubation temperature of 20 C. Following adequate afterripening, a prechill treatment of 5 d at 5 C had an inconsistent effect on germination of weeping alkaligrass (P = 0.012 in 2002, 0.156 in 2003) and improved germination of Nuttall's alkaligrass over both years (P < 0.0001). The afterripening requirement for weeping alkaligrass was more than 90 d, whereas Nuttall's alkaligrass required more than 180 d. Following adequate afterripening, both species had rapid and well-synchronized germination at fluctuating day/night temperatures of 30/10 C given unlimited moisture conditions. Given these results, it is unlikely that seeds of either species would germinate in eastern Oregon during the summer months. The data predict a long viability period under dry storage for both species. Weeping alkaligrass and Nuttall's alkaligrass should exhibit a rapid, well-synchronized germination in the spring as observed in the field.
Influence of Winter Seed Position and Recovery Date on Hairy Nightshade (Solanum sarrachoides) Recruitment and Seed Germination, Dormancy, and Mortality
Authors: Peachey R. Edward , Mallory-Smith Carol
Published: January 2007

Understanding the factors regulating recruitment in diverse tillage systems will improve the efficacy of weed-management strategies. Experiments measured the effect of hairy nightshade winter seed position (burial depth) on seedling recruitment and seed germination, dormancy, and mortality. Hairy nightshade seeds were placed in soil tubes at 1 cm below the soil line and buried in the fall so that seeds were positioned in the soil at 1, 6, 13, and 25 cm below the soil surface. Tubes with undisturbed soil and seeds were removed from the field in March, April, and May of 2004 and 2005 and placed in wells in a heated, aluminum block with a linear temperature gradient from 22.7 to 36.0 C at 2.7 C intervals, and recruitment was measured. Seeds were also recovered from the soil tubes to determine effects of seed position on germination rate, mortality, and seed dormancy. Hairy nightshade seedling recruitment was greater for seeds positioned at 6, 13, and 25 cm during the winter than at 1 cm. Seed dormancy and mortality were greatest for seeds positioned at 1 cm, but that did not adequately account for the significant decrease in recruitment at 1 cm compared to seeds buried at 6, 13, or 25 cm. Protecting the seeds buried at 1 cm from rainfall during the winter increased seedling recruitment from 0 to 20% of buried seeds but had a negligible effect on seed mortality and dormancy. Soil density was negatively correlated with recruitment.
Shikimate Accumulation in Sunflower, Wheat, and Proso Millet after Glyphosate Application
Authors: Brien Henry W. , Shaner Dale L. , West Mark S.
Published: January 2007

Experiments were conducted to examine the utility of a spectrophometric leaf disc assay for detecting shikimate accumulation after glyphosate application in sunflower, proso millet, and wheat. The assay was conducted on both greenhouse- and field-grown plants. Glyphosate was applied at five rates ranging from 840 to 53 g ae ha−1. Shikimate accumulation data were generated at 1, 4, 7, and 14 d after application (DAA). Sunflower accumulated shikimate more rapidly and at lower glyphosate rates than the other two species. At 14 DAA, glyphosate at the two highest rates remained detectable in all three species. Plants receiving lower glyphosate doses (210, 105, and 53 g ae ha−1) had begun to grow out of the injury, or at least the shikimate levels in the plants were no longer significantly different than that present in the untreated controls. This spectrophotometric assay is both rapid and simple, with respect to other means of detecting shikimate, and it can be used to detect glyphosate drift. For it to be used by crop managers, samples from potentially drift-affected crops should be taken as soon as possible after the suspected drift event or immediately after the appearance of glyphosate injury.
Native and Exotic Distributions of Siamweed (Chromolaena odorata) Modeled Using the Genetic Algorithm for Rule-Set Production
Authors: Raimundo Rafael Luís Galdini , Fonseca Rafael Luís , Schachetti-Pereira Ricardo , Townsend Peterson A. , Lewinsohn Thomas Michael
Published: January 2007

Siamweed is an asteraceous shrub native to the Neotropics that ranks among the world's most widespread and troublesome invasive species. It was introduced in several regions of Africa, Southeast Asia, and the Pacific Islands, where it severely infests natural habitats and plantation crops. Although extensive data document the weed's abundance and distribution throughout the invaded continents, the details of its current range are not fully known, especially within its native region. In this study, we used point-occurrence data and digital maps summarizing relevant environmental parameters to generate predictions for the species' geographic distributional potential—specifically, we modeled the native range of siamweed in the Neotropics using the genetic algorithm for rule-set prediction, an evolutionary computing approach. The native range occurrence data set contained 239 published and herbarium records. Models were trained on a random subset of half the points and tested using the other half. The rule sets of the native-range models were projected onto the invaded continents to predict the weed's potential for invasion, blind to its known occurrences in such regions. Native-range models predicted a wide potential distribution of siamweed throughout tropical America, from southern United States to northern Argentina and southern Brazil. The weed's occurrence has been confirmed on the northern Pacific coast, in southeast Brazil, and in other South American areas, where it was supposed to be absent. Independent model projections to Africa, Asia, and Oceania are supported by known occurrence records. Four regions are predicted to be susceptible to siamweed spread: (1) Central Africa, currently being invaded from Western Africa; (2) Infestations spreading northward from South Africa, which have already reached Swaziland and Mozambique and may extend to East Africa and Madagascar; and (3) northern New Zealand and (4) Australia, which are at risk from uncontrolled infestations on several western Pacific islands.
Emergence Timing and Recruitment of Volunteer Spring Wheat
Authors: De Corby Kristi A. , Van Acker Rene C. , Brûlé-Babel Anita L. , Friesen Lyle F.
Published: January 2007

With the recent interest in genetically engineered (GE) wheat and the commercialization of novel-trait imidazolinone herbicide-resistant wheat in North America, volunteer wheat as a weed has also been the subject of renewed interest, specifically, its recruitment and persistence in annual cropping systems. The recruitment of seed from a wheat seedbank established the previous autumn was monitored in a flax crop at two field sites in southern Manitoba, Canada, in 2003 and 2004. Seeds of eight Canadian Western Hard Red spring wheat cultivars, which exhibit a range of preharvest sprouting-resistance characteristics, were broadcast and incorporated into the soil in the autumn at 500 seeds m−2. Tillage treatments consisted of autumn tillage only, and autumn and spring tillage. Recruitment the following spring occurred very early in terms of accumulated growing–degree days (base temperature of 0 C) but expressed as a proportion of total seeds broadcast was low and variable. Total cumulative emergence of wheat over all 4 site yr ranged from 0.9 to 13.1%, with an overall average of 4.3%. There was no relationship between preharvest sprouting-resistance characteristics and recruitment proportion, and no significant influence of tillage treatment on wheat recruitment. Wheat seed that did not recruit was rapidly degraded in the soil and did not persist for more than 12 mo. However, some emerged volunteer wheat plants escaped all control measures normally used in establishing and growing a typical flax crop, and these escaped volunteer wheat plants set viable seed. Therefore, results of this study indicate that efforts and attention should be directed toward achieving very high levels of volunteer wheat control in subsequent rotational crops and that reseeding by escaped volunteer wheat plants may be a more important persistence mechanism for spring wheat than multiyear soil seedbank persistence.
A Rationale for Atrazine Stewardship in Corn
Authors: Swanton Clarence J. , Gulden Robert H. , Chandler Kevin
Published: January 2007

In several European nations, including France and Germany, atrazine has been banned because of environmental concerns. However, in Canada, atrazine remains an important component of modern weed control in corn. The objectives of this study were to determine the value of atrazine to corn producers by examining weed control efficacy, yield of corn, adjusted gross return, and the variability associated with PRE and POST herbicides applied alone or in combination with atrazine. A randomized complete-block design experiment was conducted at two locations for 3 yr to evaluate the performance of selected PRE and POST herbicides with and without atrazine. The addition of atrazine to PRE herbicides increased weed control (25%), improved herbicide performance consistency, increased corn yields (8%), increased adjusted gross return (Can$59 ha−1), and reduced risk ($30 ha−1) over sites and years. Although improving weed control, the addition of atrazine to POST herbicides increased the risk of return compared with treatments without atrazine by about $20 ha−1because the increased cost of atrazine was not always offset by higher corn yields. Our results clearly demonstrate a value of atrazine for broadleaf weed control in corn, both in terms of efficacy and economic return. From our findings, we estimated that the economic benefit of atrazine to Ontario, Canada, corn producers to be at least $26.1 million in 2004. Under current economic pressures facing agricultural producers, our findings show that a balance between the environmental effects and the benefits of atrazine to corn producers must be found because no alternative herbicide with equal economic and agronomic attributes is available at this time. To meet this balance, research on further reducing atrazine use rates while maintaining effective weed control in corn and on developing a sustainable stewardship plan is warranted.
Seed Size and Burial Effects on Giant Ragweed (Ambrosia trifida) Emergence and Seed Demise
Authors: Harrison S. K. , Regnier E. E. , Schmoll J. T. , Harrison J. M.
Published: January 2007

Giant ragweed is a competitive, allergenic weed that persists in agricultural fields and early successional sites. Field experiments were conducted to determine the effects of seed size and seed burial depth on giant ragweed emergence and seed demise. In a seedling emergence experiment, small (< 4.8 mm in diameter) and large (> 6.6 mm in diameter) seeds were buried 0, 5, 10, and 20 cm in fall 1997, and weed emergence was monitored over the next seven growing seasons. A generalized linear mixed model fit to the cumulative emergence data showed that maximum emergence for both seed sizes occurred at the 5-cm burial depth, where probability of emergence was 19% for small seeds and 49% for large seeds. Emergence probability at the 10-cm burial depth was 9% for small seeds and 30% for large seeds, and no seedlings emerged from the 20-cm burial depth. The model predicted that ≥ 98% of total cumulative emergence was completed after four growing seasons for large seeds buried 5 cm, five growing seasons for small seeds buried 5 cm and large seeds buried 10 cm, and seven growing seasons for small seeds buried 10 cm. Seed size and burial treatment effects on seed demise were tested in a second experiment using seed packets. Rates of seed demise were inversely proportional to burial depth, and the percentage of viable seeds remaining after 4 yr ranged from 0% on the soil surface to 19% at the 20-cm burial depth. Some seeds recovered from the 20-cm burial depth were viable after 9 yr of burial. These results, coupled with previous research, suggest that seed size polymorphism facilitates giant ragweed adaptation across habitats and that a combination of no-tillage cropping practices, habitat modification, and timely weed control measures can reduce its active seed bank in agricultural fields by 90% or more after 4 yr.
Stem/Progenitor Cell-Like Properties of Desmoglein 3dim Cells in Primary and Immortalized Keratinocyte Lines
Authors: Wan Hong , Yuan Ming , Simpson Cathy , Allen Kirsty , Gavins Felicity N.E. , Ikram Mohammed S. , Basu Subham , Baksh Nuzhat , O'Toole Edel A. , Hart Ian R.
Published: January 2007

We showed previously that primary keratinocytes selected for low desmoglein 3 (Dsg3) expression levels exhibited increased colony-forming efficiency and heightened proliferative potential relative to cells with higher Dsg3 expression levels, characteristics consistent with a more “stem/progenitor cell-like” phenotype. Here, we have confirmed that Dsg3dim cells derived from cultured primary human adult keratinocytes have comparability with α6bri/CD71dim stem cells in terms of colony-forming efficiency. Moreover, these Dsg3dim cells exhibit increased reconstituting ability in in vitro organotypic culture on de-epidermalized dermis (DED); they are small, actively cycling cells, and they express elevated levels of various p63 isoforms. In parallel, using the two immortalized keratinocyte cell lines HaCaT and NTERT, we obtained essentially similar though occasionally different findings. Thus, reduced colony-forming efficiency by Dsg3bri cells consistently was observed in both cell lines even though the cell cycle profile and levels of p63 isoforms in the bri and dim populations differed between these two cell lines. Dsg3dim cells from both immortalized lines produced thicker and better ordered hierarchical structural organization of reconstituted epidermis relative to Dsg3bri and sorted control cells. Dsg3dim HaCaT cells also show sebocyte-like differentiation in the basal compartment of skin reconstituted after a 4-week organotypic culture. No differences in percentages of side population cells (also a putative marker of stem cells) were detected between Dsg3dim and Dsg3bri populations. Taken together our data indicate that Dsg3dim populations from primary human adult keratinocytes and long-term established keratinocyte lines possess certain stem/progenitor cell-like properties, although the side population characteristic is not one of these features.

Disclosure of potential conflicts of interest is found at the end of this article.
Human AB Serum and Thrombin-Activated Platelet-Rich Plasma Are Suitable Alternatives to Fetal Calf Serum for the Expansion of Mesenchymal Stem Cells from Adipose Tissue
Authors: Kocaoemer Asli , Kern Susanne , Klüter Harald , Bieback Karen
Published: January 2007

MSCs are currently in focus regarding their clinical potential in cell therapy and tissue engineering. However, most isolation and expansion protocols for clinical-scale production of MSCs use fetal calf serum (FCS) as a supplement, which poses a potential risk for infections as well as immunological reactions. To find a suitable FCS substitute, we investigated the effects of pooled human AB serum (AB-HS) and thrombin-activated platelet-rich plasma (tPRP) on adipose tissue MSCs (AT-MSCs) with FCS as the standard control medium. AT-MSCs of 10 donors were cultured under three different conditions: (a) 10% FCS, (b) 10% AB-HS, and (c) 10% tPRP. Colony-forming units, cumulative population doubling rates, and differentiation capacity toward the adipogenic and osteogenic lineages were assessed, along with immunophenotype. We demonstrated that AB-HS and tPRP provide a significantly higher proliferative effect on AT-MSCs than does FCS. In the first six passages, AB-HS and tPRP MSCs exhibited a fold expansion of 66.6 ± 15.7 and 68.1 ± 6.7, respectively, compared with 24.4 ± 0.7 for FCS. Differentiation capacity was preserved throughout long-term culture. Immunophenotype was characteristic for MSCs and comparable for all culture conditions with the exception of a distinct CD45-/CD14-positive side population for AB-HS and tPRP that tended to diminish with prolonged culture. We showed that pooled human AB serum and thrombin-activated platelet-rich plasma are alternatives to FCS for AT-MSCs. These human sources are better characterized regarding potential infectious threats, while providing a higher proliferation rate and retaining differentiation capacity and mesenchymal stem cell marker expression throughout long-term culture.

Disclosure of potential conflicts of interest is found at the end of this article.
Severe Hypoxia Defines Heterogeneity and Selects Highly Immature Progenitors Within Clonal Erythroleukemia Cells
Authors: Giuntoli Serena , Rovida Elisabetta , Gozzini Antonella , Barbetti Valentina , Cipolleschi Maria Grazia , Olivotto Massimo , Dello Sbarba Persio
Published: January 2007

We showed that resistance to severe hypoxia defines hierarchical levels within normal hematopoietic populations and that hypoxia modulates the balance between generation of progenitors and maintenance of hematopoietic stem cells (HSC) in favor of the latter. This study deals with the effects of hypoxia (0.1% oxygen) in vitro on Friend's murine erythroleukemia (MEL) cells, addressing the question of whether a clonal leukemia cell population comprise functionally different cell subsets characterized by different hypoxia resistance. To identify leukemia stem cells (LSC), we used the Culture Repopulating Ability (CRA) assay we developed to quantify in vitro stem cells capable of short-term reconstitution (STR). Hypoxia strongly inhibited the overall growth of MEL cell population, which, despite its clonality, comprised progenitors characterized by markedly different hypoxia-resistance. These included hypoxia-sensitive colony-forming cells and hypoxia-resistant STR-type LSC, capable of repopulating secondary liquid cultures of CRA assays, confirming what was previously shown for normal hematopoiesis. STR-type LSC were found capable not only of surviving in hypoxia but also of being mostly in cycle, in contrast with the fact that almost all hypoxia-surviving cells were growth-arrested and with what we previously found for HSC. However, quiescent LSC were also detected, capable of delayed culture repopulation with the same efficiency as STR-like LSC. The fact that even quiescent LSC, believed to sustain minimal residual disease in vivo, were found within the MEL cells indicates that all main components of leukemia cell populations may be present within clonal cell lines, which are therefore suitable to study the sensitivity of individual components to treatments.

Disclosure of potential conflicts of interest is found at the end of this article.
Differential Developmental Ability of Embryos Cloned from Tissue-Specific Stem Cells
Authors: Inoue Kimiko , Noda Shinichi , Ogonuki Narumi , Miki Hiromi , Inoue Shinichi , Katayama Kazufumi , Mekada Kazuyuki , Miyoshi Hiroyuki , Ogura Atsuo
Published: January 2007

Although cloning animals by somatic cell nuclear transfer is generally inefficient, the use of certain nuclear donor cell types may significantly improve or deteriorate outcomes. We evaluated whether two multipotent stem cell lines produced in vitro—neural stem cells (NSCs) and mesenchymal stem cells (MSCs)—could serve as nuclear donors for nuclear transfer cloning. Most (76%) NSC-derived embryos survived the two-cell–to–four-cell transition, the stage when the major zygotic gene activation occurs. Consistent with this observation, the expression patterns of zygotically active genes were better in NSC-derived embryos than in fibroblast clone embryos, which arrested at the two-cell stage more frequently. Embryo transfer experiments demonstrated that at least some of these NSC embryos had the ability to develop to term fetuses (1.6%, 3/189). In contrast, embryos reconstructed using MSCs showed a low rate of in vitro development and never underwent implantation in vivo. Chromosomal analysis of the donor MSCs revealed very frequent aneuploidy, which probably impaired the potential for development of their derived clones. This is the first demonstration that tissue-specific multipotent stem cells produced in vitro can serve as donors of nuclei for cloning mice; however, these cells may be prone to chromosomal aberrations, leading to high embryonic death rates. We found previously that hematopoietic stem cells (HSCs) are very inefficient donor cells because of their failure to activate the genes essential for embryonic development. Taken together, our data led us to conclude that tissue-specific stem cells in mice, namely NSCs, MSCs, and HSCs, exhibited marked variations in the ability to produce cloned offspring and that this ability varies according to both the epigenetic and genetic status of the original genomes.

Disclosure of potential conflicts of interest is found at the end of this article.
Radiotherapy-induced signal transduction
Authors: Yacoub Adly , Miller Anna , Caron Ruben W , Qiao Liang , Curiel David A , Fisher Paul B , Hagan Michael P , Grant Steven , Dent Paul
Published: January 2007

Exposure of tumor cells to ionizing radiation causes compensatory activation of multiple intracellular survival signaling pathways to maintain viability. In human carcinoma cells, radiation exposure caused an initial rapid inhibition of protein tyrosine phosphatase function and the activation of ERBB receptors and downstream signaling pathways. Radiation-induced activation of extracellular regulated kinase (ERK)1/2 promoted the cleavage and release of paracrine ligands in carcinoma cells which caused re-activation of ERBB family receptors and intracellular signaling pathways. Blocking ERBB receptor phosphorylation or ERK1/2 pathway activity using small-molecule inhibitors of kinases for a short period of time following exposure (3 h) surprisingly protected tumor cells from the toxic effects of ionizing radiation. Prolonged exposure (48–72 h) of tumor cells to inhibition of ERBB receptor/ERK1/2 function enhanced radiosensitivity. In addition to ERBB receptor signaling, expression of activated forms of RAS family members and alterations in p53 mutational status are known to regulate radiosensitivity apparently independent of ERBB receptor function; however, changes in RAS or p53 mutational status, in isogenic HCT116 cells, were also noted to modulate the expression of ERBB receptors and ERBB receptor paracrine ligands. These alterations in receptor and ligand expression correlated with changes in the ability of HCT116 cells to activate ERK1/2 and AKT after irradiation, and to survive radiation exposure. Collectively, our data in multiple human carcinoma cell lines argues that tumor cells are dynamic and rapidly adapt to any single therapeutic challenge, for example, radiation and/or genetic manipulation e.g. loss of activated RAS function, to maintain tumor cell growth and viability.
Chemotherapy: induction of stress responses
Authors: Tiligada E
Published: January 2007

Eukaryotic cells, from yeast to mammals, respond and adapt to environmental and microenvironmental stressors by evolutionary conserved multicomponent endogenous systems that utilise a network of signal transduction pathways to regulate the adaptive and protective phenotype. The balance between cell survival and cell death is decisive for sensitivity or resistance to DNA-damaging chemotherapeutic agents. Anticancer drugs may themselves act as stressors to induce adaptive signals that could limit their clinical value. Related research has been focused on the modulation of the expression and function of the heat shock proteins, the unfolded protein response, the mechanisms of subcellular translocation of signalling components, the genomic and non-genomic actions of drugs and endogenous functional components like hormonal pathways, the input of inflammation and alterations in the microenvironmental milieu and on the control of the cell cycle and proliferation. The outcome seems to be driven by the first-line responses that support cellular integrity and by specific mechanisms that depend on the type of cell and the nature, and duration and severity of the noxious stimulus. Data obtained from experimental organisms like the yeast have added valuable information on the basic conservation in cellular stress-related processes in eukaryotes and on the consequences that may accompany the adaptive and protective phenotype during the stress response to anticancer agents. Understanding the complex molecular pathways mediating these processes has started to contribute to the reevaluation of the current therapeutic regiments and to revolutionise the approaches for improved anticancer therapy.
Resistance to endocrine therapy in breast cancer: exploiting estrogen receptor/growth factor signaling crosstalk
Authors: Massarweh Suleiman , Schiff Rachel
Published: January 2007

Targeting the estrogen receptor (ER) is the oldest form of molecular targeted therapy, and the widespread use of the selective estrogen receptor modulator tamoxifen in breast cancer is responsible for major improvements in cure rates, quality of life, and disease prevention in the last 25 years. Newer forms of endocrine therapy now available for the management of endocrine responsive breast cancer include a new generation of aromatase inhibitors, which lower the estrogen ligand for ER, and pure ER antagonists which destroy the receptor. Despite these recent clinical advances, intrinsic and acquired resistance to these endocrine therapies is still a common feature that limits the success of this therapeutic strategy. Recent research into the molecular biology of ER signaling has revealed a remarkably complex interactive signaling with other growth factor signaling pathways in breast cancer cells, potentially explaining some of the reasons behind endocrine therapy action as well as resistance. This view of a more complex ER signaling system has uncovered new molecular targets which, if present in a cancer cell, might be additionally targeted using various signal transduction inhibitors to overcome or prevent resistance to endocrine therapy. In addition, the dynamic inverse relationship between the expression of ER and growth factor receptors brings more excitement to the potential of restoring ER expression in apparently ER-negative cells by inhibition of growth factor signaling. Ongoing clinical trials of endocrine therapy combined with growth factor pathway inhibitors or their downstream signaling elements promise to further improve the present care for breast cancer patients.
Deciphering antihormone-induced compensatory mechanisms in breast cancer and their therapeutic implications
Authors: Gee J M W , Shaw V E , Hiscox S E , McClelland R A , Rushmere N K , Nicholson R I
Published: January 2007

Breast cancer inhibition by antihormones is rarely complete, and our studies using responsive models reveal the remarkable flexibility of breast cancer cells in recruiting alternative signalling to limit maximal anti-tumour effects of oestrogen receptor α (ER) blockade. The recruited mechanism involves antihormone-induced expression of oestrogen-repressed signalling genes. For example, epidermal growth factor receptor gene (EGFR) is induced by antioestrogens and maintains residual kinase and ER phosphorylation, cell survival genes, and thereby allows incomplete antihormone response and emergence of resistance. Microarrays are revealing the breadth of antihormone-induced genes that may attenuate growth inhibition, including NFκB, Bag1, 14-3-3ζ and tyrosine kinases, such as HER2 and Lyn. Three concepts are emerging: first, some genes are induced exclusively by antioestrogens, while others extend to oestrogen deprivation; secondly, some are transiently induced, while others persist into resistance; finally, some confer additional adverse features when tumour cells are in an appropriate context. Among the latter is CD59 whose antioestrogen induction may permit evasion of immune surveillance in vivo. Also, induction of pro-invasive genes (including NFκB, RhoE and δ-catenin) may underlie our findings that antioestrogens can markedly stimulate migratory behaviour when tumour intercellular contacts are compromised. Based on our promising studies selectively inhibiting EGFR (gefitinib), NFκB (parthenolide) or CD59 (neutralising antibody) together with antioestrogens, we propose that co-targeting strategies could markedly improve anti-tumour activity (notably enhancing cell kill) during the antihormone-responsive phase. Furthermore, subverting those induced signalling genes that are retained into resistance (e.g. EGFR, NFκB, HER2) may prove valuable in this state. Alongside future deciphering and targeting of genes underlying antioestrogen-promoted invasiveness, embracing of intelligent combination strategies could significantly extend patient survival.
Growth factor receptor interplay and resistance in cancer
Authors: Jones Helen E , Gee Julia M W , Hutcheson Iain R , Knowlden Janice M , Barrow Denise , Nicholson Robert I
Published: January 2007

Aberrant signalling through the epidermal growth factor receptor (EGFR) plays a major role in the progression and maintenance of the malignant phenotype and the receptor is therefore a rational anti-cancer target. A variety of approaches have been developed to specifically target the EGFR which include monoclonal antibodies and small molecule tyrosine kinase inhibitors, such as gefitinib (Iressa). However, the recent clinical experience across a range of cancer types is revealing that despite the anti-EGFR agents demonstrating some anti-tumour activity, there is a high level of de novo and acquired resistance to such treatments and moreover, overexpression of the EGFR is clearly not the sole determinant of response to such therapies. Such adverse phenomena, which serve to limit the overall therapeutic impact of these new agents, implies the existence of a greater complexity involved in the regulation of EGFR signalling than was previously assumed. Indeed, evidence is accumulating which demonstrates that signalling interplay occurs between the EGFR, and the IGF-1 receptor (IGF-1R) and the review will focus on the emerging concept of growth factor pathway switching between these two receptors as a means of influencing the effectiveness of anti-EGFR agents such as gefitinib.
IGF1R signalling and its inhibition
Authors: Riedemann J , Macaulay V M
Published: January 2007

The type 1 IGF receptor (IGF1R) is a transmembrane tyrosine kinase that is frequently overexpressed by tumours, and mediates proliferation and apoptosis protection. IGF signalling also influences hypoxia signalling, protease secretion, tumour cell motility and adhesion, and thus can affect the propensity for invasion and metastasis. Therefore, the IGF1R is now an attractive anti-cancer treatment target. This review outlines the effects of IGF1R activation in tumour cells, and will describe the strategies that are available to block IGF signalling, both as investigational tools and as novel anti-cancer therapeutics. Design of specific IGF1R inhibitors has been problematic due to close homology with the insulin receptor, but recently it has proved possible to design selective IGF1R inhibitors. These compounds and IGF1R antibodies are showing promise in preclinical models of human cancer, and several agents are now in early phase clinical trials. Both classes of agents affect insulin receptor signalling, either by direct kinase inhibition or antibody-induced insulin receptor downregulation. This effect may lead to clinical toxicity, but could be therapeutically beneficial in blocking signalling via variant insulin receptors capable of a mitogenic response to IGF-II. Specificity for IGF1R targeting can be achieved by antisense and siRNA-mediated IGF1R downregulation; these approaches have undoubted utility as research tools, and may in future generate nucleic-acid-based therapeutics. It will be important to use data from preclinical and early clinical trials to establish the molecular correlates of sensitivity to IGF1R blockade, and the optimum means of combining this new approach with standard treatment modalities.
Role of hypoxia-inducible factor-1α as a cancer therapy target
Authors: Patiar Shalini , Harris Adrian L
Published: January 2007

Hypoxia occurs in solid tumours due to a mismatch between tumour growth and angiogenesis. Hypoxia in solid tumours is associated with an aggressive phenotype and resistance to radiation therapy and chemotherapy leading to poor patient prognosis. Hypoxia-inducible factor-1 (HIF-1) is a transcription factor, which is activated in response to intratumoural hypoxia and as a result of genetic alterations that activate oncogenes and inactivate tumour suppressor genes. It plays a key role in the adaptation of tumour cells to hypoxia by activating the transcription of genes, which regulate several biological processes including angiogenesis, cell proliferation and survival, glucose metabolism, pH regulation and migration. This makes HIF-1 an attractive target for the development of anticancer agents. The success of these agents depends on reliable methods to identify those patients most likely to benefit from HIF-1-targeted therapy. Several novel small molecule inhibitors of HIF-1 have been identified and are moving towards clinical trials, but none of these are specific for HIF-1. Further work is ongoing to identify more selective HIF-1 inhibitors.
Src as a therapeutic target in anti-hormone/anti-growth factor-resistant breast cancer
Authors: Hiscox Stephen , Morgan L , Green Tim , Nicholson Robert I
Published: January 2007

Endocrine therapy is the treatment of choice in hormone receptor-positive breast cancer. However, the effectiveness of anti-hormone drugs, such as tamoxifen, is limited because of the development of resistance, ultimately leading to disease progression and patient mortality. Using in vitro cell models of anti-hormone resistance, we have previously demonstrated that altered growth factor signalling contributes to an endocrine insensitive phenotype. Significantly, our recent studies have revealed that the acquisition of endocrine resistance in breast cancer is accompanied by a greatly enhanced migratory and invasive phenotype. Furthermore, therapeutic intervention using anti-growth factor monotherapies, despite an initial growth suppressive phase, again results in the development of a resistant state and a further augmentation of their invasive phenotype.

Using the dual specific Src/Abl kinase inhibitor, AZD0530, we have highlighted a central role for Src kinase in promoting the invasive phenotype that accompanies both anti-hormone and anti-growth factor resistance. Importantly, the use of Src inhibitors in combination with anti-growth factor therapies appears to be additive, producing a marked inhibitory effect on cell growth, migration and invasion and ultimately prevents the emergence of a resistant phenotype. These observations suggest that the inhibition of Src activity may present a novel therapeutic intervention strategy, particularly when used as an adjuvant in endocrine-resistant breast disease, with the potential to delay or prevent the acquisition of subsequent resistance to anti-growth factor therapies.
Role of receptor complexes in the extranuclear actions of estrogen receptor α in breast cancer
Authors: Song Robert X-D , Fan Ping , Yue Wei , Chen Yucai , Santen Richard J
Published: January 2007

Our recent studies have examined the role of various receptor complexes in the mediation of rapid, extranuclear effects of estradiol. This review describes 17β-estradiol (E2)-initiated extranuclear signaling pathways, which involve the insulin-like growth factor 1 receptor (IGF-1R) and epidermal growth factor receptor (EGFR) and result in the activation of several kinase cascades. The biologic results of these effects are the enhancement of cell proliferation and diminution of programmed cell death (apoptosis). Until recently, most studies assigned priority to the nuclear transcriptional actions of estrogen receptor α (ERα). Present investigative emphasis focuses on the additional importance of ERα residing in or near the plasma membrane. A small fraction of ERα is associated with the cell membrane and mediates the rapid effects of E2. Unlike classical growth factor receptors, such as IGF-1R and EGFR, ERα has no transmembrane and kinase domains and is known to initiate E2 rapid signals by forming protein/protein complexes with many signaling molecules. Our recent studies demonstrate that the IGF-1R is involved in tethering ERα to the plasma membrane, in activating the EGFR, and in the initiation of mitogen-activated protein kinase and phosphoinositide 3-kinase signaling. The formation of a multi-protein complex containing these receptors as well as adaptor proteins is a critical step in this process. A full understanding of the mechanisms underlying these relationships with the ultimate aim of abrogating specific steps, should lead to more targeted strategies for treatment of hormone-dependent breast cancer.