bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular basis of the unusual catalytic preference for GDP/GTP in Entamoeba histolytica 3‐phosphoglycerate kinase
Authors: Encalada Rusely , Rojo‐Domínguez Arturo , Rodríguez‐Zavala José S. , Pardo Juan P. , Quezada Héctor , Moreno‐Sánchez Rafael , Saavedra Emma
Published: March 2009

Phosphoglycerate kinase (EC 2.7.2.3) catalyzes reversible phosphoryl transfer from 1,3‐bisphosphoglycerate to ADP to synthesize 3‐phosphoglycerate and ATP during glycolysis. Phosphoglycerate kinases from several sources can use GDP/GTP as alternative substrates to ADP/ATP; however, the maximal velocities (Vm) reached with the guanine nucleotides are ∼ 50% of those displayed with the adenine nucleotides. By contrast, Entamoeba histolytica phosphoglycerate kinase (EC 2.7.2.10) is the only reported phosphoglycerate kinase displaying higher activity with GDP/GTP and lower affinities for the adenine nucleotides. To elucidate the molecular basis of the Entamoeba histolytica phosphoglycerate kinase selectivity for GDP/GTP, a conformational analysis was carried out on a homology model based on crystallographic structures of yeast and pig phosphoglycerate kinases. Some amino acid residues involved in the purine ring binding site not previously described were detected. Accordingly, Y239, E309 and V311 were replaced by site‐directed mutagenesis in the Entamoeba histolytica phosphoglycerate kinase gene for the corresponding amino acid residues present in the adenine nucleotide‐dependent phosphoglycerate kinases and the recombinant proteins were purified. Kinetic analysis of the enzymes showed that the single mutants Y239F, E309Q, E309M and V311L increased their catalytic efficiencies (Vm/Km) with ADP/ATP as a result of both, increased Vm and decreased Km values. Furthermore, a higher catalytic efficiency in the double mutant Y239F/E309M was achieved, which was mainly due to an increased affinity for ADP/ATP with a concomitant diminished affinity for GDP/GTP. The main Entamoeba histolytica phosphoglycerate kinase amino acid residues involved in the selectivity for guanine nucleotides were thus identified.
Acute intermittent porphyria – impact of mutations found in the hydroxymethylbilane synthase gene on biochemical and enzymatic protein properties
Authors: Ulbrichova Dana , Hrdinka Matous , Saudek Vladimir , Martasek Pavel
Published: March 2009

Acute intermittent porphyria is an autosomal dominantly inherited disorder, classified as acute hepatic porphyria, caused by a deficiency of hydroxymethylbilane synthase (EC 2.5.1.61, EC 4.3.1.8, also known as porphobilinogen deaminase, uroporphyrinogen I synthase), the third enzyme in heme biosynthesis. Clinical features include autonomous, central, motor or sensory symptoms, but the most common clinical presentation is abdominal pain caused by neurovisceral crises. A diagnosis of acute intermittent porphyria is crucial to prevent life‐threatening acute attacks. Detection of DNA variations by molecular techniques allows a diagnosis of acute intermittent porphyria in situations where the measurement of porphyrins and precursors in urine and faeces and erythrocyte hydroxymethylbilane synthase activity is inconclusive. In the present study, we identified gene defects in six Czech patients with acute intermittent porphyria, as diagnosed based on biochemical findings, and members of their families to confirm the diagnosis at the molecular level and/or to provide genetic counselling. Molecular analyses of the hydroxymethylbilane synthase gene revealed seven mutations. Four were previously reported: c.76C>T, c.77G>A, c.518G>A, c.771 + 1G>T (p.Arg26Cys, p.Arg26His, p.Arg173Gln). Three were novel mutations: c.610C>A, c.675delA, c.750A>T (p.Gln204Lys, p.Ala226ProfsX28, p.Glu250Asp). Of particular interest, one patient had two mutations (c.518G>A; c.610C>A), both located in exon 10 of the same allele. To establish the effects of the mutations on enzyme function, biochemical characterization of the expressed normal recombinant and mutated proteins was performed. Prokaryotic expression of the mutant alleles of the hydroxymethylbilane synthase gene revealed that, with the exception of the p.Gln204Lys mutation, all mutations resulted in little, if any, enzymatic activity. Moreover, the 3D structure of the Escherichia coli and human protein was used to interpret structure–function relationships for the mutations in the human isoform.
X‐ray crystallography and structural stability of digestive lysozyme from cow stomach
Authors: Nonaka Yasuhiro , Akieda Daisuke , Aizawa Tomoyasu , Watanabe Nobuhisa , Kamiya Masakatsu , Kumaki Yasuhiro , Mizuguchi Mineyuki , Kikukawa Takashi , Demura Makoto , Kawano Keiichi
Published: March 2009

In ruminants, some leaf‐eating animals, and some insects, defensive lysozymes have been adapted to become digestive enzymes, in order to digest bacteria in the stomach. Digestive lysozyme has been reported to be resistant to protease and to have optimal activity at acidic pH. The structural basis of the adaptation providing persistence of lytic activity under severe gastric conditions remains unclear. In this investigation, we obtained the crystallographic structure of recombinant bovine stomach lysozyme 2 (BSL2). Our denaturant and thermal unfolding experiments revealed that BSL2 has high conformational stability at acidic pH. The high stability in acidic solution could be related to pepsin resistance, which has been previously reported for BSL2. The crystal structure of BSL2 suggested that negatively charged surfaces, a shortened loop and salt bridges could provide structural stability, and thus resistance to pepsin. It is likely that BSL2 loses lytic activity at neutral pH because of adaptations to resist pepsin.
Conservative management of ulcerated haemangioma – twenty years experience
Authors: Pandey Anand , Gangopadhyay Ajay Narayan , Sharma Shiv Prasad , Kumar Vijayendra , Gopal Saroj Chooramani , Gupta Dinesh Kumar
Published: March 2009

Ulceration is the most common complication of haemangioma and occurs in 5–15% of cases. The present study was carried out to evaluate the clinical features of ulcerated haemangioma and efficacy of the management protocol adopted by us over a period of 20 years. All patients with ulcerated haemangioma were evaluated on the basis of age at presentation, sex, rural or urban distribution and site of haemangioma. Treatment included application of topical antibiotic and systemic antibiotic and analgesic for pain. The total number of patients was 608. The male to female ratio was 1: 2·28. The rural:urban distribution was 2·43:1. The most common site of involvement was head and neck. Mean age of patients was 5·60 ± 2·44 months. Mean size of haemangioma and ulceration was 47·30 ± 20·67 cm2 and 7·49 ± 4·52 cm2, respectively. The mean time for ulcer healing was 40·06 ± 19·41 days. Ulcer size of more than 10 cm2 took more time to heal. Response to treatment was satisfactory. Ulcerated haemangioma usually occurs before completion of 1 year of age; hence, every patient with haemangioma needs careful attention. Adequate treatment and regular follow up brings satisfactory response in the patients.
Silver resistance in MRSA isolated from wound and nasal sources in humans and animals
Authors: Loh Jia V , Percival Steven L , Woods Emma J , Williams Nicola J , Cochrane Christine A
Published: March 2009

Methicillin‐resistant Staphylococcusaureus (MRSA) colonises skin, nasal passages and dermal wounds. Methods used to manage wounds infected and colonised with MRSA often include the use of topical antiseptics such as ionic silver and iodine. The objectives of this study were to determine the prevalence of silver‐resistance (sil) genes in MRSA and methicillin‐resistant coagulase‐negative staphylococci (MR‐CNS) isolated from wounds and nasal cavities of humans and animals, and also to determine the susceptibility of sil‐positive and sil‐negative MRSA isolates to a silver‐containing Hydrofiber® (SCH) wound dressing, on planktonic silE‐positive and silE‐negative MRSA. Polymerase chain reaction was used to determine the presence of three silver‐resistance (sil) genes, silE, silP and silS in 33 MRSA and 8 methicillin‐resistant staphylococci (MR‐CNS). SilP and silS genes were absent in all isolates tested; however, two MRSA strains were found to contain the silE gene, together with one isolate of MR‐CNS. Phenotypic resistance of the silE‐positive strains and their susceptibility to the SCH dressing was evaluated using the zone of inhibition test on Mueller Hinton agar, and confocal laser microscopy using a live/dead fluorescent stain. Results confirmed that the SCH dressing was effective in killing all MRSA strains with and without the silE gene. First, this study showed that the prevalence of sil genes was low in the isolates investigated; and secondly, that the presence of a silver‐resistance gene (silE) in MRSA and MR‐CNS did not afford protection to the organism in the presence of a SCH wound dressing. The use of topical antiseptics in chronic wound care should be considered before the use of antibiotics that can result in their overuse and the risk of further resistance.
Maggot debridement therapy with Lucilia cuprina: a comparison with conventional debridement in diabetic foot ulcers
Authors: Paul Aaron G , Ahmad Nazni W , Lee HL , Ariff Ashraff M , Saranum Masri , Naicker Amara S , Osman Zulkiflee
Published: March 2009

This is prospective case–control study of more than 18 months performed to assess the effectiveness of maggot debridement therapy (MDT) with the sterile larvae of Lucilia cuprina (a tropical blowfly maggot) for the treatment of diabetic foot ulcers. Literature thus far has only reported results with the temperate maggot, Lucilia sericata. This study documents outcome in diabetic foot wounds treated with maggot debridement versus those treated by conventional debridement alone. In this series of 29 patients treated with MDT, 14 wounds were healed, 11 were unhealed and 4 were classified under others. The control group treated by conventional debridement had 30 patients of which 18 wounds were healed, 11 unhealed and 1 classified under others. There was no significant difference in outcome between the two groups. The conclusion that can be made from this study is that MDT with L. cuprina is as effective as conventional debridement in the treatment of diabetic foot ulcers. It would be a feasible alternative to those at high risk for surgery or for those who refuse surgery.
Bacteriotherapy with Lactobacillus plantarum in burns
Authors: Peral Maria C , Huaman Martinez Miguel A , Valdez Juan C
Published: March 2009

Bacterial colonisation and infection remain the major causes of delayed healing and graft rejection following burns. Topical treatment is necessary to reduce the incidence of burn wound infection. Silver sulphadiazine (SD‐Ag) is an often used microbicidal agent. However, this treatment produces adverse reactions and side‐effects. On the basis of experimental data and clinical application of lactobacilli as probiotics, we performed this exploratory study to establish the effectiveness of bacteriotherapy with topical application of the innocuous bacteria Lactobacillus plantarum cultured in De Man, Rogosa and Sharpe medium to provide an alternative method for burn treatment using SD‐Ag as a reference. These innocuous bacteria would compete with other bacteria that are wound pathogens and would modify the wound environment and promote tissue repair. Eighty burned patients from the Plastic Surgery and Burns Unit were grouped into infected (delayed) second‐ and third‐degree and non infected (early) third‐degree burns and treated with L. plantarum or SD‐Ag. The proportion of patients with delayed second‐degree burns was 0·71 for L. plantarum and 0·73 for SD‐Ag (relative rate: −2·72%) with respect to the decrease in bacterial load (<105 bacteria/g of tissue), promotion of granulating tissue wound bed and healing. In early third‐degree burns, the values were 0·75 for L. plantarum and 0·84 for SD‐Ag (relative rate: −1·07%) in preventing wound infection and promotion of granulation tissue, 0·90 in graft taking for both treatments (relative rate: 0%) and 0·75 for L. plantarum and 0·77 for SD‐Ag (relative rate: −2·60%) in healing. In delayed third‐degree burns, values were 0·83 for L. plantarum and 0·71 for SD‐Ag (relative rate: +16·90%) with respect to the decrease in the bacterial load (<105 bacteria/g of tissue) and providing a granulating tissue wound bed, 0·90 in graft taking for both treatments (relative rate: 0%) and 0·75 for L. plantarum and 0·64 for SD‐Ag (relative rate: + 17·19%) in healing. Although the number of patients (between 12 and 15 per group) did not enable the application of a power statistical test, these results suggest that the L. plantarum treatment should be studied in greater depth and could be used as a valid alternative for the topical treatment of burns.
Synthesis, Biological Evaluation, Mechanism of Action and Quantitative Structure–Activity Relationship Studies of Chalcones as Antibacterial Agents
Authors: Sivakumar Ponnurengam Malliappan , Priya Sobana , Doble Mukesh
Published: March 2009

Forty‐eight chalcone analogs were synthesized and their in vitro antibacterial activity against Staphylococcus aureus NCIM 5021, Bacillus subtilis NCIM 2718, Phaseolus vulgaris NCIM 2813, Escherichia coli NCIM 2931, Salmonella typhi 2501 and Enterobacter aerogenes NCIM 5139 were evaluated by microdilution broth assay. Quantitative structure–activity relationships were developed for all the cases (r 2 = 0.68–0.79;  = 0.58–0.78; q 2 = 0.51–0.68; F = 13.02–61.51). Size, polarizability, electron‐donating/withdrawing and hydrophilic nature of the molecule determine the activity against these Gram‐positive and Gram‐negative bacteria. Staphylococcus aureus was the most and S. typhi was the least hydrophobic of these organisms. These chalcones act better against more hydrophobic organisms. The more active chalcones have log P between 1.5 and 3. Compound 24, one of the most active compounds, was found to act by damaging the cell wall of S. aureus. Slimicidal activity of five of the most active compounds (24, 31, 32, 34 and 37) was found to be in the range of 48–60% against S. aureus and 40–54% against E. coli. A correlation was observed among the hydrophobicity of the compounds, hydrophobicity of the bacterial cell surface and the antibacterial activity of the compound.
Discovery and Binding Studies on a Series of Novel Pin1 Ligands
Authors: Wu Bainan , Rega Michele F. , Wei Jun , Yuan Hongbin , Dahl Russell , Zhang Ziming , Pellecchia Maurizio
Published: March 2009

Pin1 plays a key role in various biological cellular processes via the recognition of phosphorylated Ser/Thr‐Proline motifs. Moreover, high expression levels of Pin1 are correlated to tumorgenesis in some cancer types. Here, we identify a novel series of small molecular weight compounds with a core structure mimicking the phoshorylated serine. The binding affinity and binding mode of the compounds for Pin1 are analyzed via NMR spectroscopy and computational studies. The reported chemical probes and relative binding data to Pin1 represent valuable stepping stones for the validation of Pin1 as target for drug discovery and for eventually the development of possible lead compounds.
Design of Conformationally Constrained Azole Antifungals: Efficient Synthesis and Antifungal Activity of trans‐3‐Imidazolylflavanones
Authors: Emami Saeed , Behdad Mehran , Foroumadi Alireza , Falahati Mehraban , Lotfali Ensieh , Sharifynia Somaye
Published: March 2009

An efficient protocol has been developed for the preparation of a series of trans‐3‐imidazolylflavanones and (Z)‐trans‐3‐imidazolylflavanone oximes, as potential antifungal agents, by the reaction of 2‐imidazolyl‐2′‐hydroxyacetophenone with different benzaldehyde derivatives and subsequence oximation reaction. The stereochemical and conformational aspects of compounds were also assigned by 1H NMR spectroscopy. The aryl group present at C‐2 and the imidazole ring present at C‐3 were assigned to occupy equatorial position in trans‐3‐imidazolylflavanones 3 and axial position in (Z)‐trans‐3‐imidazolylflavanone oximes 4. Most of the 3‐imidazolylflavanone derivatives containing a substituent on phenyl ring showed significant antifungal activity and modification of the 4‐oxo group to oxime or substituent on phenyl ring produced changes in the antifungal activity profile of these compounds. Generally, 3′‐chloro‐ analog of 3‐imidazolylflavanone 3c exhibited better profile of antifungal activity against strains of fungi tested, superior than reference drug fluconazole.
Adenosine infusion attenuates soluble RAGE in endotoxin‐induced inflammation in human volunteers
Authors: Soop A. , Sundén‐Cullberg J. , Albert J. , Hållström L. , Treutiger C.‐J. , Sollevi A.
Published: March 2009

Aim: To evaluate possible anti‐inflammatory effects of pre‐treatment with adenosine in a human experimental inflammatory model.

Methods: The study design was double‐blind, crossover, placebo‐controlled and randomized. In the Intensive Care Unit of a university hospital, 16 healthy male volunteers were treated for 5.5 h with infusions of adenosine 40 μg kg−1 min−1or placebo. Thirty minutes after the start of adenosine or placebo, 2 ng kg−1E‐Coli endotoxin was administered. Heart rate, body temperature, blood pressure, plasma cytokines (TNF‐α, IL‐6 and IL‐10), soluble RAGE and resistin, exhaled nitric oxide and nitrite/nitrate in urine were determined.

Results: Endotoxin elicited the expected clinical signs of an inflammatory reaction (tachycardia, fever) and led to prominent release of the cytokines studied (P < 0.001). Resistin in plasma increased after endotoxin (P < 0.001). After placebo treatment, soluble RAGE (sRAGE) in plasma increased 5 h after the endotoxin challenge (P < 0.001) but not after adenosine. After placebo, orally exhaled NO increased with a peak at 4 h (P < 0.001), although there was no statistically significant difference between the two treatments. Nitrite/nitrate in urine (n = 11) did not differ between adenosine and placebo treatments.

Conclusion: In conclusion, adenosine infusion starting before endotoxin challenge in humans attenuated sRAGE significantly but otherwise had no clear anti‐inflammatory effect. Adenosine as a potential anti‐inflammatory treatment in humans needs further study, including use of higher doses. The mechanism underlying the effect of adenosines on sRAGE remains unknown.
New Models to Investigate Mechanisms of Disease Genesis from Primitive BCR‐ABL+ Hematopoietic Cells
Authors: EAVES CONNIE , JIANG XIAOYAN , EISTERER WOLFGANG , CHALANDON YVES , PORADA G. , ZANJANI ESMAIL , EAVES ALLEN
Published: March 2009

Abstract: Three years ago we described a novel autocrine IL‐3/G‐CSF mechanism active in the leukemic CD34+ cells from chronic myeloid leukemia (CML) patients in chronic phase (PNAS 96: 12804–12809, [1999]). We also showed that exposure of the most primitive CD34+ cells from normal human bone marrow to excess IL‐3 stimulates not only the division of these cells but also their differentiation. In contrast, both IL‐3 and G‐CSF cause an expansion of the more mature types of normal CD34+ progenitors. These findings suggested that the autocrine IL‐3/G‐CSF mechanism active in CML stem cells can compromise their self‐renewal in spite of increasing their proliferative activity, which, in turn, might explain the paradoxically slow rate of expansion of this compartment over time in patients with latent disease. To investigate this hypothesis, we have begun to characterize the numbers and types of cells generated from chronic phase CML patients' cells transplanted into adult immunodeficient mice or fetal sheep, and also from transplants of primitive murine and human hematopoietic cells transduced with a retroviral BCR‐ABL vector. Our findings to date using these models reinforce the importance of the autocrine IL‐3/G‐CSF mechanism in the development of CML. BCR‐ABL appears to directly activate IL‐3 and G‐CSF production in primitive hematopoietic cells and this is important to their transplantable leukemogenic activity. However, the development in vivo of an overt leukemia from primitive BCR‐ABL+ hematopoietic cells can be very delayed. These models thus offer new opportunities for analyzing the molecular events that underlie the pathogenesis of human CML and the future testing of new therapeutic approaches.
Thrombopoietin
Authors: KAUSHANSKY KENNETH
Published: March 2009

Abstract: Although it is clear that thrombopoietin is the primary regulator of thrombopoiesis, several lines of evidence indicate that the hormone affects multiple aspects of hematopoiesis: the in vivo administration of TPO increases marrow levels of erythroid, myeloid, and megakaryocytic progenitor cells and its genetic elimination or that of its receptor (c‐mpl) reduces the numbers of these cells; all hematopoietic stem cells (HSCs) are c‐mpl+; genetic elimination of c‐mpl reduces the numbers of murine HSCs by 7–8‐fold; and its null mutation in humans leads to congenital amegakaryocytic thrombocytopenia, a disorder that almost invariably leads to aplastic anemia. Recently, we have begun to explore the role of TPO in the HSC self‐renewal and expansion that characterizes the post‐stem‐cell‐transplantation period. Using limiting dilution cell transplantation analyses, we found that HSC self‐renewal and expansion is reduced 10–20‐fold after transplantation of normal stem cells into tpo null mice compared to their wild‐type counterparts. Although the molecular mechanisms responsible for these findings are only now being explored, it is expected that a greater understanding of the roles played by TPO in HSC physiology will lead to novel therapeutic opportunities.
Nonpeptide Mediators in the Hematopoietic Microenvironment
Authors: MÖHLE ROBERT , BOEHMLER ANDREAS M. , DENZLINGER CLAUDIO , KANZ LOTHAR
Published: March 2009

Abstract: Migration of hematopoietic stem and progenitor cells (HPCs) is controlled by chemotactic factors released in the hematopoietic microenvironment. In particular, the chemokine SDF‐1, which activates the G protein‐coupled receptor (GPR) CXCR4, plays an important role in progenitor cell mobilization and homing. However, we provide evidence that ligands of other GPRs similarly act on CD34+ hematopoietic progenitors. These ligands comprise non‐peptide mediators, including the cysteinyl‐leukotriene receptor CysLT1, and stimulate migration and integrin‐dependent adhesion of HPCs. Moreover, continuous activation of a GPR by a specific ligand upregulates the responsiveness of other GPRs to their corresponding ligands. These findings suggest that HPC migration may not depend on a single chemokine receptor (e.g., CXCR4). Rather, mobilization and homing of HPCs involve several GPRs, which interact with each other as well as with adhesion molecules. Pharmacological activation and inhibition of the GPR may allow HPC mobilization and homing to be modulated.
Hematopoietic Stem Cell Transplantation across Major Genetic Barriers
Authors: REISNER YAIR , GUR HILIT , REICH‐ZELIGER SHLOMIT , MARTELLI MASSIMO F. , BACHAR‐LUSTIG ESTHER
Published: March 2009

Abstract: Studies in mice and humans demonstrate that transplantation of hematopoietic progenitors in numbers larger than commonly used (“megadose” transplants) overcomes major genetic barriers. In vitro studies suggest that veto cells, within the population of hematopoietic progenitors, facilitate this favorable outcome. Thus, when purified CD34+ cells were added to bulk mixed‐lymphocyte reactions (MLRs) they suppressed CTLs against the donor's stimulators, but not against stimulators from a third party. This tolerizing activity depends on cell contact and can be blocked by the caspase inhibitor BD‐FMK, suggesting that the effector host T cells are deleted by apoptosis upon interaction with the CD34+ cells. Early myeloid CD33+ cells generated by short‐term ex vivo expansion of CD34+ cells also exhibit veto activity, and these cells can be grown in large numbers. Tolerance induction can be further enhanced by other veto cells. Perhaps the most potent veto cell is the CD8+ CTL. However, this cell is also associated with marked GVHD (graft‐versus‐host disease. GVHD can be separated from the veto activity by generating anti‐third party CTLs under IL2 deprivation. Under such selective pressure only the stimulated clones which make IL2 can survive, while anti‐host clones die. In vivo studies show that such anti‐third party veto CTLs can be used safely for tolerance induction without GVHD.
Differential Gene Expression Underlying the Functional Distinctions of Primary Human CD34+ Hematopoietic Stem and Progenitor Cells from Peripheral Blood and Bone Marrow
Authors: STEIDL ULRICH , KRONENWETT RALF , HAAS RAINER
Published: March 2009

Abstract: The restorative capacity of human CD34+ hematopoietic cells is clinically used in the autologous and allogeneic transplant setting to support cytotoxic therapy. We examined gene expression patterns of highly enriched bone marrow CD34+ (BM‐CD34+) or G‐CSF‐mobilized peripheral blood CD34+ (PB‐CD34+) cells by cDNA array technology, quantitative real‐time RT‐PCR, and flow cytometry, to identify molecular causes underlying the functional differences between circulating and sedentary hematopoietic stem and progenitor cells. The greater cell cycle and DNA synthesis activity of BM‐CD34+ compared to PB‐CD34+ cells was reflected by the 2‐ to 5‐fold higher expression of 9 genes involved in cell cycle, 11 genes regulating DNA synthesis, and the cell cycle‐initiating transcription factor E2F‐1. The 2‐ to 3‐fold greater expression of 5 pro‐apoptotic genes in PB‐CD34+ cells indicated a higher apoptotic activity, which could functionally be corroborated by apoptosis assays. Thrombin receptor (PAR1), known to play a role in trafficking of malignant cells, was 3.6‐fold higher expressed in circulating CD34+ cells than in BM‐CD34+ cells. Guidance via thrombin receptor might molecularly mediate stem cell migration. In summary, our study provides gene expression profiles of primary human CD34+ hematopoietic cells of blood and marrow. Our data molecularly confirm and explain the finding that CD34+ cells residing in the bone marrow are cycling more rapidly, whereas circulating CD34+ cells consist of a higher number of quiescent stem and progenitor cells. Moreover, our data give novel molecular insights into stem cell migration and differentiation.
Hematopoietic Stem Cell Emergence in the Human Embryo and Fetus
Authors: PÉAULT BRUNO , TAVIAN MANUELA
Published: March 2009

Abstract: Two waves of hematopoietic stem cell generation take place in the first month of human gestation. The first one has long been known to occur in the yolk sac; only recently was another one identified that results in the development, from the 27th day, of clusters of hematopoietic cells on the ventral endothelium of the aorta and vitelline artery. This latter, intra‐embryonic phase of blood cell progenitor production is undoubtedly local since its presumptive territory of occurrence, the para‐aortic splanchnopleura, is endowed with blood‐forming potential in vitro from, at least, day 19 of development. The first multipotent, lympho‐myeloid stem cells emerge within that intra‐embryonic territory, whereas the yolk sac only produces myeloid precursor cells devoid of lymphoid potential. The forerunners of the hematopoietic stem cells born to intra‐embryonic arteries appear to migrate from the splanchnopleura as KDR+ angio‐hematopoietic cells that colonize the ventral wall of the aorta to give rise to hematogenous endothelium. All these results indicate that stem cells for human definitive hematopoiesis emerge within the embryo, in the walls of truncal arteries, from splanchnopleural ancestors and through a vascular endothelial intermediate stage. Less expectedly, blood‐forming endothelial cells were also encoutered in the embryonic and fetal liver and in the fetal and, even, adult bone marrow, suggesting that a filiation between vascular and hematopoietic cells may persist during the whole life.
Pluripotent Stem Cells Identified in Multiple Murine Tissues
Authors: HOWELL JONATHAN C. , LEE WEI‐HUA , MORRISON PAUL , ZHONG JIN , YODER MERVIN C. , SROUR EDWARD F.
Published: March 2009

Abstract: Pluripotential stem cells (PSCs) have been recently described in many tissues including skeletal muscle, brain, and bone marrow. However, the true nature of these cells is still unclear, and their precise definition has yet to be determined. We hypothesized that a common, rare population of PSCs with a broad tissue differentiation potential can be identified in multiple murine tissues and that these cells are capable of transdifferentiation into cells of different primordial germ layer origins in response to diverse microenvironmental cues. To examine this hypothesis, we isolated phenotypically defined cells from murine skeletal muscle and cultured these cells under different conditions tailored to promote differentiation into several cell types including myocytes. We report here that in conditions permissive for hematopoietic differentiation, muscle‐derived CD45−Sca‐1+c‐kit− cells differentiated into cells expressing hematopoietic‐specific mRNA; while in conditions promoting myogenic, neuronal, and adipocytic differentiation, cells morphologically typical of these cell types expressing tissue‐specific markers were identified 9–14 days in culture. When CD45−Sca‐1+c‐kit− cells from muscle or bone marrow were transplanted intracerebellarly into Purkinje cell degenerative (pcd) mice, the behavior of these mice improved 28 days after transplantation relative to mice injected with vehicle alone, suggesting that these cells contributed to the appearance of functional neuronal cells that may have improved the ataxic condition characteristic of these mice. Phenotypic analysis of single cell suspensions prepared from brain, blood, and intestinal epithelium revealed the presence of CD45−Sca‐1+c‐kit− cells in varying degrees. These studies suggest that a phenotypically common, multipotent cell can be identified in different tissues and that this cell may represent a universal pluripotent stem cell residing at different levels in multiple murine tissues.
Early Stages of Hematopoietic Differentiation
Authors: Spangrude Gerald J. , Perry S. Scott , Slayton William B.
Published: March 2009

Abstract: Mouse bone marrow contains hematopoietic stem cells as well as progenitor cells, which are partially differentiated offspring of stem cells. We have utilized several approaches to separate progenitors from stem cells in order to characterize essential differences between these two stages of development. As a first approach, we utilized the supravital fluorescent dye rhodamine‐123 (Rh‐123) to distinguish quiescent stem cells (Rh‐123low) from metabolically active progenitor cells (Rh‐123hi). Analysis of megakaryocyte potential in a tissue culture assay demonstrated that Rh‐123hi progenitor cells were capable of robust megakaryocyte differentiation, while Rh‐123low stem cells produced fewer colonies containing megakaryocytes. Transplantation of the two cell populations into irradiated recipients revealed the opposite outcome, suggesting that the tissue culture assay failed to predict behavior in a transplant setting. We also evaluated functional potential of lymphoid progenitors isolated by selecting for differential expression of Thy‐1.1 and c‐kit. The potential of defined cell populations to differentiate as T or B lymphocytes in vivo was dependent upon the time post transplant at which animals were evaluated. These studies underscore the need for caution in the interpretation of lineage potentials evaluated by both in vitro and in vivo assays.
Marrow Stem Cell Potential within a Continuum
Authors: QUESENBERRY P. , COLVIN G. , LAMBERT J. F. , ABEDI M. , CERNY J. , DOONER M. , MOORE B. , McAULIFFE C. , DEMERS D. , GREER D. , PARENT A. , BADIAVAS E. , LUM L. , FALANGA V.
Published: March 2009

Abstract: On the basis of our studies of the fluctuation of the hematopoietic stem cell phenotype with cell cycle trnsit, we hypothesize that the ability of marrow stem cells to convert to nonhematopoietic cells will also vary at different points in the cell cycle. The new biology of stem cells has an impact on many fields including developmental biology and stem cell biology and the clinical potential is enormous.
Antibody W7C5 Defines a CD109 Epitope Expressed on CD34+ and CD34− Hematopoietic and Mesenchymal Stem Cell Subsets
Authors: GIESERT CHRISTINA , MARXER ANKE , SUTHERLAND D. ROBERT , SCHUH ANDRE C. , KANZ LOTHAR , BÜHRING HANS‐JÖRG
Published: March 2009

Abstract: The cell surface antigen recognized by monoclonal antibody W7C5 is expressed at low levels on human CD34+ and CD34− bone marrow stem cell populations but at high levels on fetal liver CD34+ cells. To identify the recognized antigen, we performed partial amino acid sequence and MALDI analysis of purified W7C5 antigens. The analyses revealed that MoAb W7C5 detects CD109, a recently cloned glycophosphatidylinositol‐linked glycoprotein. The specificity of MoAb W7C5 was further confirmed by the selective recognition of CHO cells transfected with human CD109 cDNA.
The Mesenchymal Stroma Negatively Regulates B Cell Lymphopoiesis through the Expression of Activin A
Authors: SHOHAM TSIPI , PARAMESWARAN RESHMI , SHAV‐TAL YARON , BARDA‐SAAD MIRA , ZIPORI DOV
Published: March 2009

Abstract: The negative control of B cell generation is only partially resolved. We assessed the role of activin A in regulation of B lymphopoiesis in view of its specific inhibitory effects on tumor B lineage cells. Activin A is constitutively expressed in mouse hemopoietic organs and in cultured mesenchymal cell lines. We observed an inverse relationship between activin A titer and B lineage cell production. In the spleen, the red pulp exhibited a relatively higher abundance of the protein as compared with the lymphoid follicles, wherein B cell accumulation occurs. Furthermore, a specific shut off in activin A expression was observed in bone marrow and spleen following in vivo induction of B lymphocyte polyclonal activation. We further substantiated these in vivo observations by in vitro studies of primary bone marrow cultures, in which the expression of functional activin A was found to be diminished prior to the onset of B lymphopoiesis. The reduction in functional activin A is shown to concomitantly occur with spontaneous induction of the expression of activin A specific inhibitors. We therefore propose that the mesenchymal organ stroma expresses activin A that negatively controls B cell lymphopoiesis.
Segment Inventories
Authors: Mielke Jeff
Published: March 2009

The nature of speech sound inventories has been a focus of study by phonologists and phoneticians, facilitated in 1984 with the publication by Ian Maddieson of the UCLA Phonological Segment Inventory Database. This article gives an overview of the study of inventories and summarizes some of the major findings. Crosslinguistic comparison of inventories provides a window into the phonetic factors that shape languages, but also reflects non‐linguistic factors. Models of the phonetic and phonological factors argued to shape inventories are discussed, along with some challenges that need to be dealt with in order to isolate linguistically interesting facts.
Enhanced expression of CD71, transferrin receptor, on immature reticulocytes in patients with paroxysmal nocturnal hemoglobinuria
Authors: SATO S. , KOZUMA Y. , HASEGAWA Y. , KOJIMA H. , CHIBA S. , NINOMIYA H.
Published: March 2009

Erythroid cells in the marrow express CD71, transferrin receptor, and reticulocytes released from the marrow lose their expression during maturation. The immature reticulocyte fraction, the proportion of reticulocytes with the highest content of RNA, has been determined by hematology analysis. In the present study, we examined CD71 expression on immature reticulocytes by flow cytometry (FCM) in paroxysmal nocturnal hemoglobinuria (PNH) patients with reticulocytosis. We modified ‘reticulocyte‐gated FCM’ to multi‐color FCM, i.e. RNA/CD71, RNA‐CD59 or CD59/CD71/CD45. In PNH, in addition to the increased number of immature reticulocytes (%CD71‐positive), a more immature phenotype in regard to both CD71 intensity and RNA content levels was demonstrated. In seven PNH patients studied, %CD71‐positive reticulocytes were significantly increased at 32.2 ± 11.9% (n = 10, normal 10.4 ± 3.5%, P = 0.002). RNA content levels (assessed by mean fluorescence index, MFI) in CD71‐positive reticulocytes were significantly increased at 812.0 ± 215.2 MFI in PNH (n = 10, normal 508 ± 86.1 MFI, P = 0.002). These data indicate that stimulated erythropoietic conditions induce the release of more immature reticulocytes to the peripheral blood than ordinary erythropoietic conditions. CD71 intensity on immature reticulocytes was well correlated with their RNA content levels, indicating the usefulness of CD71 as an immature reticulocyte marker.
Modeling of Vertical Solute Dispersion in a Sediment Bed Enhanced by Wave‐Induced Interstitial Flow1
Authors: Qian Qin , Voller Vaughan R. , Stefan Heinz G.
Published: March 2009

Abstract: Mass (solute) transport in a stream or lake sediment bed has a significant effect on chemical mass balances and microbial activities in the sediment. A “1D vertical dispersion model” is a useful tool to analyze or model solute transfer between river or lake water and a sediment bed. Under a motionless water column, solute transfer into and within the sediment bed is by molecular diffusion. However, surface waves or bed forms create periodic pressure waves along the sediment/water interface, which in turn induce flows in the pores of the sediment bed. The enhancement of solute transport by these interstitial periodic flows in the pores has been incorporated in a 1D depth‐dependent “enhanced dispersion coefficient (DE).” Typically,DEdiminishes exponentially with depth in the sediment bed. Relationships have been developed to estimateDEas a function of the characteristics of sediment (particle size, hydraulic conductivity, and porosity) and pressure waves (wave length and height). In this paper, we outline and illustrate the calculation ofDEas well as the penetration depth (dp) of the flow effect. Sample applications to illustrate the computational procedure are provided for dissolved oxygen transfer into a stream gravel bed and release of phosphorus from a lake bed. The sensitivity of the results to input parameter values is illustrated, and compared with the errors obtained when interstitial flow is ignored. Maximum values ofDEnear the sediment surface can be on the order of 1 cm2/s in a stream gravel bed with standing waves, and 0.001 cm2/s in a fine sand lake bed under progressive surface waves, much larger than molecular diffusion coefficients.
Evaluating high‐throughput sequencing as a method for metagenomic analysis of nematode diversity
Authors: PORAZINSKA DOROTA L. , GIBLIN‐DAVIS ROBIN M. , FALLER LINA , FARMERIE WILLIAM , KANZAKI NATSUMI , MORRIS KRYSTALYNNE , POWERS THOMAS O. , TUCKER ABRAHAM E. , SUNG WAY , THOMAS W. KELLEY
Published: March 2009

Nematodes play an important role in ecosystem processes, yet the relevance of nematode species diversity to ecology is unknown. Because nematode identification of all individuals at the species level using standard techniques is difficult and time‐consuming, nematode communities are not resolved down to the species level, leaving ecological analysis ambiguous. We assessed the suitability of massively parallel sequencing for analysis of nematode diversity from metagenomic samples. We set up four artificial metagenomic samples involving 41 diverse reference nematodes in known abundances. Two samples came from pooling polymerase chain reaction products amplified from single nematode species. Two additional metagenomic samples consisted of amplified products of DNA extracted from pooled nematode species. Amplified products involved two rapidly evolving ~400‐bp sections coding for the small and large subunit of rRNA. The total number of reads ranged from 4159 to 14771 per metagenomic sample. Of these, 82% were > 199 bp in length. Among the reads > 199 bp, 86% matched the referenced species with less than three nucleotide differences from a reference sequence. Although neither rDNA section recovered all nematode species, the use of both loci improved the detection level of nematode species from 90 to 97%. Overall, results support the suitability of massively parallel sequencing for identification of nematodes. In contrast, the frequency of reads representing individual species did not correlate with the number of individuals in the metagenomic samples, suggesting that further methodological work is necessary before it will be justified for inferring the relative abundances of species within a nematode community.
Characterization of 10 microsatellite markers for the understorey Amazonian herb Heliconia acuminata
Authors: CÔRTES M. C. , GOWDA V. , KRESS W. J. , BRUNA E. M. , URIARTE M.
Published: March 2009

We characterized 10 microsatellite loci for the plant Heliconia acuminata from the Biological Dynamics of Forest Fragments Project (Manaus, Brazil). Markers were screened in 61 individuals from one population and were found to be polymorphic with an average of eight alleles per locus. We found moderate to high levels of polymorphic information content, and observed and expected heterozygosities. All 10 markers are suitable for spatial genetic structure and parentage analyses and will be used for understanding H. acuminata dynamics across a fragmented landscape.
Centaureaaksoyi sp. nov. (Asteraceae: Cardueae) from Turkey and a contribution to the sectional taxonomy
Authors: Hamzaoğlu Ergin , Budak Ümit
Published: March 2009

Centaurea
aksoyi Hamzaoğlu & Budak (Asteraceae: Cardueae) is described and illustrated as a new species from Turkey (central Anatolia, Yozgat province). Diagnostic characters, descriptions and taxonomic comments are given. The characteristics of the new species are compared to the related species Centaureapseudokotschyi Wagenitz (sect. Pseudoseridia Wagenitz) and Centaureapaphlagonica (Bornm.) Wagenitz (sect. Plumosipappus Wagenitz). The geographical distribution of the new and related species are given, and it is considered as Critically Endangered based on the IUCN criteria.
A novel method for floristic quality assessment using the vegetation of the Jiuding Mountain, Sichuan, China as an example
Authors: Tu Wei‐Guo , Gao Xin‐Fen , Wu Ning , Liu Shi‐Hua
Published: March 2009

Floristic Quality Assessment (FQA) is proposed as a new method to assess floristic integrity. In the present study, we use both the conventional method broadly adopted in the USA and a new one proposed herein to assess the floristic quality of the Jiuding Mountain. For the application of the FQA, each plant species was assigned an integer from 0 to 10 termed coefficient of conservatism (C) based on its global distribution area. The coefficient shows the degree of fidelity of each plant species to the flora of the west slope of the Jiuding Mountain and represents a rank based on its distribution area. Those species receiving a C value of 0–1 are distributed all over the world, those receiving a C value of 2–3 mostly in tropical areas, and those receiving a C value of 4–6 broadly in temperate areas. Species with C values of 7–8 are endemic to China and those with coefficients 9–10 are restricted to the Hengduan Mountains or even endemic to the Jiuding Mountain. Based on the fact that different plant species show different relative importance values (IV), the new method estimates the weighted averages for the coefficient of conservatism called the weighted mean coefficient of conservatism ( w). The floristic quality index (FQI) was then derived from
or
w, native species richness (N) and non‐native species (A). As expected when using the new method, the dominant species of the communities obtained different Cs, which resulted in different
ws and FQIws as compared to the conventional method.
Plastid DNA haplotype variation in Dactylorhiza incarnata (Orchidaceae): evidence for multiple independent colonization events into Scandinavia
Authors: Hedrén Mikael
Published: March 2009

The early marsh orchid, Dactylorhiza incarnata (L.) Soó s. l., grows in medium‐rich to rich fens and marshes over much of Europe and parts of Asia. The species is highly polymorphic and different forms may grow together at the same site. In the present study, I tested the hypothesis that these forms represent different migrant populations that have colonized Scandinavia independently of each other, possibly from different source areas. Accessions from Scandinavia and elsewhere were screened for variation at three size‐variable plastid marker loci, one polyA repeat, one polyA‐polyTA‐polyT repeat and one 9 bp indel. Ten haplotypes were defined on basis on the combined variation pattern. The common occurrence of several haplotypes in southern Scandinavia and adjacent areas to the south and the east of the Baltic Sea suggests that D. incarnata has been dispersed on repeated occasions across the Baltic. Also, there was some correlation between haplotype composition and morphological form on the island of Gotland, in agreement with the independent colonization hypothesis. Material from northernmost Sweden, Finland and northwest Russia was fixed for a single widespread haplotype, indicating that populations in this area are located farther away from the Pleistocene refugia. Dactylorhiza incarnata ssp. lobelii from southwest Norway was characterized by a haplotype that was not encountered elsewhere in Scandinavia. Given its proximity to British populations dominated by the same haplotype, it is suggested that D. incarnata ssp. lobelii was established independently of the other Scandinavian populations, from coastal refugia located in western Europe.
Magnolia bawangensis sp. nov. (Magnoliaceae) from Hainan, China
Authors: Liu Dong‐Ming , Zhou Ren‐Zhang , Zeng Qing‐Wen , Xing Fu‐Wu
Published: March 2009

Magnolia bawangensis Law, R. Z. Zhou & D. M. Liu (Magnoliaceae), a new species from the Mount Bawangling, Hainan Province, China, is described and illustrated. It is closely related to M. paenetalauma Dandy, but differs by being a smaller tree with fewer spathe scars, absence of pedicels, pubescent and more numerous tepals, white pubescent and more numerous stamens.
The German Clinical Trials Register: challenges and chances of implementing a bilingual registry
Authors: Hasselblatt Hanna , Dreier Gabriele , Antes Gerd , Schumacher Martin
Published: March 2009

Objective It is a moral responsibility towards patients, funding organizations, the scientific community and the general public to publish not only the results of clinical trials but also to inform prospectively about planned trials. This is especially important for scientists, funding organizations, and ethics committees, since they are involved in the planning of new trials, which should rely on the knowledge of existing trials. Moreover, patients can only participate in clinical trials if they or their physicians are aware of ongoing trials for which they are eligible. In order to provide a central portal for information on clinical research in Germany and to facilitate the search for specific trials, the German Clinical Trials Register was implemented in 2008.

Method An internet‐based prototype of the German Clinical Trials Register was developed in close co‐operation with the World Health Organization's registries network on the basis of the internationally harmonized data specification. This was further adapted to a bilingual registry's needs. Co‐operation with medical ethics committees in Germany has been initiated with the aim of a joint synergistic data acquisition procedure.

Results The German Clinical Trials Register was launched in August 2008. It was approved as the World Health Organization's primary registry for Germany in October 2008, and fulfils the requirements of the International Committee of Journal Editors.

Conclusion The German Clinical Trials Register provides a central bilingual portal for all users (including scientists, patients, physicians, funding organizations, and ethics committees) for information on planned, ongoing, and completed clinical trials conducted in Germany. The German Clinical Trials Register allows International Committee of Journal Editors and World Health Organization‐compliant registration of clinical trials in Germany.