bluuebunny/crossref_metadata_2025 huggingface dataset:
Chemokine receptor expression in EBV-associated lymphoproliferation in hu/SCID mice: implications for CXCL12/CXCR4 axis in lymphoma generation
Authors: Piovan Erich , Tosello Valeria , Indraccolo Stefano , Cabrelle Anna , Baesso Ilenia , Trentin Livio , Zamarchi Rita , Tamamura Hirokazu , Fujii Nobutaka , Semenzato Gianpietro , Chieco-Bianchi Luigi , Amadori Alberto
Published: September 2004

The mechanisms by which intraperitoneal injection of peripheral blood mono-nuclear cells (PBMCs) from Epstein-Barr virus (EBV)–seropositive donors into severe combined immunodeficient (SCID) mice gives rise to lymphomas (hu/SCID tumors) are far from clear. This study addressed whether chemokine receptors and their ligands could be implicated in this experimental model. CXCR4 was found to be highly expressed in hu/SCID tumors; surface expression of CXCR4 was prevalently limited to a tumor cell subset poorly expressing CD23, whereas the CXCR4 ligand, CXCL12, was predominantly expressed by the tumor subpopulation expressing CD23. In vitro inhibition of this autocrine/paracrine CXCL12/CXCR4 axis significantly inhibited lymphoma proliferation and survival. Furthermore, CXCL12 was expressed in cells recovered from the mouse peritoneal cavity early after PBMC transfer as well as by EBV-transformed B cells but not by resting or activated B lymphocytes; also, lymphoma development was associated with a dramatic increase in the levels of murine CXCL12 present in the peritoneal cavity. Finally, antagonizing the CXCL12/CXCR4 axis in vivo strongly counteracted lymphoma development. These studies demonstrate that CXCL12 expression may be associated with EBV infection and suggest that the CXCR4/CXCL12 axis may participate in the EBV-associated lymphomagenesis process in immunodeficient hosts.
Quantitative analysis of nucleoside transporter and metabolism gene expression in chronic lymphocytic leukemia (CLL): identification of fludarabine-sensitive and -insensitive populations
Authors: Mackey John R. , Galmarini Carlos M. , Graham Kathryn A. , Joy Anil A. , Delmer Alain , Dabbagh Laith , Glubrecht Darryl , Jewell Lawrence D. , Lai Raymond , Lang Thack , Hanson John , Young James D. , Merle-Béral Helene , Binet Jacques L. , Cass Carol E. , Dumontet Charles
Published: September 2004

Resistance to fludarabine is observed in the clinic, and molecular predictive assays for benefit from chemotherapy are required. Our objective was to determine if expression of nucleoside transport and metabolism genes was associated with response to fludarabine therapy in patients with chronic lymphocytic leukemia (CLL). CLL cells from 56 patients were collected prior to treatment with fludarabine. Quantitative reverse transcriptase–polymerase chain reaction (RT-PCR) was performed on sample RNA to determine the relative levels of mRNA of 3 nucleoside transporters that mediate fludarabine uptake (human equilibrative nucleoside transporter 1 [hENT1], human equilibrative nucleoside transporter 2 [hENT2], and human concentrative nucleoside transporter 3 [hCNT3]), deoxycytidine kinase (dCK), and 3 5′-nucleotidases (ecto-5′nucleotidase [CD73], deoxynucleotidase-1 [dNT-1], and cytoplasmic high-Km 5-nucleotidase [CN-II]). Two-dimensional hierarchical cluster analysis of gene expression identified 2 distinct populations of CLL. Cluster 2 patients experienced a 3.4-fold higher risk of disease progression than cluster 1 patients (P = .0058, log-rank analysis). Furthermore, independent analysis of the individual genes of interest revealed statistically significant differences for risk of disease progression (adjusted hazard ratios [HRs]) with underexpression of dNT-1 (HR = 0.45; P = .042), CD73 (HR = 0.40; P = .022), and dCK (HR = 0.0.48; P = .035), and overexpression of hCNT3 (HR = 4.7; P = .0007) genes. Subjects with elevated hCNT3 expression experienced a lower complete response rate to fludarabine therapy (11% vs 69%; P = .002). No hCNT3-mediated plasma membrane nucleoside transport was detected in CLL samples expressing hCNT3 message, and hCNT3 protein was localized to the cytoplasm with immunohistochemical and confocal microscopy.
Cannabis-induced cytotoxicity in leukemic cell lines: the role of the cannabinoid receptors and the MAPK pathway
Authors: Powles Thomas , Poele Robert te , Shamash Jonathan , Chaplin Tracy , Propper David , Joel Simon , Oliver Tim , Liu Wai Man
Published: September 2004

Δ9-Tetrahydrocannabinol (THC) is the active metabolite of cannabis. THC causes cell death in vitro through the activation of complex signal transduction pathways. However, the role that the cannabinoid 1 and 2 receptors (CB1-R and CB2-R) play in this process is less clear. We therefore investigated the role of the CB-Rs in mediating apoptosis in 3 leukemic cell lines and performed microarray and immunoblot analyses to establish further the mechanism of cell death. We developed a novel flow cytometric technique of measuring the expression of functional receptors and used combinations of selective CB1-R and CB2-R antagonists and agonists to determine their individual roles in this process. We have shown that THC is a potent inducer of apoptosis, even at 1 × IC50 (inhibitory concentration 50%) concentrations and as early as 6 hours after exposure to the drug. These effects were seen in leukemic cell lines (CEM, HEL-92, and HL60) as well as in peripheral blood mononuclear cells. Additionally, THC did not appear to act synergistically with cytotoxic agents such as cisplatin. One of the most intriguing findings was that THC-induced cell death was preceded by significant changes in the expression of genes involved in the mitogen-activated protein kinase (MAPK) signal transduction pathways. Both apoptosis and gene expression changes were altered independent of p53 and the CB-Rs.
Final adult height of patients who received hematopoietic cell transplantation in childhood
Authors: Sanders Jean E. , Guthrie Katherine A. , Hoffmeister Paul A. , Woolfrey Ann E. , Carpenter Paul A. , Appelbaum Frederick R.
Published: September 2004

Growth impairment and growth hormone (GH) deficiency are complications after total body irradiation (TBI) and hematopoietic cell transplantation (HCT). To determine the impact of GH therapy on growth, the final heights of 90 GH-deficient children who underwent fractionated TBI and HCT for malignancy were evaluated. Changes in height standard deviation (SD) from the diagnosis of GH deficiency to the achievement of final height were compared among 42 who did and 48 who did not receive GH therapy. At HCT, GH-treated patients were younger (P = .001), more likely to have undergone central nervous system irradiation (P = .007), and shorter (P = .005) than patients who did not receive GH therapy. After HCT, GH deficiency was diagnosed at 1.5 years (range, 0.8-9.5 years) for GH-treated and 1.2 years (range, 0.9-8.8 years) for nontreated patients. GH therapy was associated with significantly improved final height in children younger than 10 years at HCT (P = .0001), but GH therapy did not impact the growth of older children. Girls (P = .0001) and children diagnosed with acute myelogenous leukemia (AML), chronic myelogenous leukemia (CML), or myelodysplastic syndromes (MDS) (compared with acute lymphoblastic leukemia [ALL] or non-Hodgkin lymphoma [NHL]; P = .02) also showed more rapid growth than their counterparts. These data demonstrate that GH therapy improves the final height of young children after fractionated TBI.
The pattern of methacholine responsiveness in mice is dependent on antigen challenge dose
Authors: Zosky Graeme R , von Garnier Christophe , Stumbles Philip A , Holt Patrick G , Sly Peter D , Turner Debra J
Published: September 2004

Background

Considerable variation exists in the protocols used to induce hyperresponsiveness in murine models of allergic sensitisation. We examined the effect of varying the number of antigen exposures at challenge on the development of methacholine responsiveness in systemically sensitised mice.

Methods

BALB/c mice were sensitised with ovalbumin (OVA), challenged with 1, 3 or 6 OVA aerosols. Lung function was measured using low frequency forced oscillations and partitioned into components representing the airways (Raw) and lung parenchyma (tissue damping (G) and tissue elastance (H)). Responsiveness to inhaled methacholine (MCh), inflammatory cell profile and circulating IgE were assessed 24 and 48 hours after challenge. The threshold dose of MCh required to elicit a detectable response (sensitivity) and response to 30 mg.mL-1 (maximal response) were determined for each compartment.

Results

Sensitivity; All three OVA protocols resulted in an increased sensitivity to MCh in Raw but not in G or H. These responses where present at 24 and 48 hrs, except 1 OVA aerosol in which changes had resolved by 48 hrs. Maximal response; 1 OVA aerosol increased maximal responses in Raw, G and H at 24 hrs, which was gone by 48 hrs. Three OVA aerosols increased responses in H at 48 hrs only. Six OVA challenges caused increases in Raw, G and H at both 24 and 48 hrs. Eosinophils increased with increasing antigen challenges. IgE was elevated by OVA sensitisation but not boosted by OVA aerosol challenge.

Conclusions

The pattern of eosinophilia, IgE and MCh responsiveness in mice was determined by antigen dose at challenge. In this study, increased sensitivity to MCh was confined to the airways whereas increases in maximal responses occurred in both the airway and parenchymal compartments. The presence of eosinophilia and IgE did not always coincide with increased responsiveness to inhaled MCh. These findings require further systematic study to determine whether different mechanisms underlie airway and parenchymal hyperresponsiveness post antigen challenge.
Depressed and Well Mothers' Emotion Interpretation Accuracy and the Quality of Mother—Infant Interaction
Authors: Broth Michelle Robbins , Goodman Sherryl H. , Hall Christine , Raynor Lynne Catherine
Published: September 2004

The inadequate parenting associated with mothers' depression may be related to mothers' problems in interpreting infants' emotional expressions. The relations between depressed and well mothers' accuracy at interpreting babies' facial expressions and the quality of the mothers' interactions with their infants were examined. In partial support of our hypotheses, depressed mothers' level of depressive symptoms was associated with less accuracy, especially regarding positive emotions. Contrary to expectations, depressed mothers did not differ from well mothers in terms of their emotion accuracy. Furthermore, depressed mothers' accuracy at interpreting infants' emotions was not significantly related to the quality of their interaction with their infants; in contrast, well mothers' accuracy for infants' negative emotions was associated with better interaction quality. These findings provide new information about depressed mothers' emotional interpretations and their parenting. The different pattern of findings for depressed and well mothers suggests that other mediating factors are important, which are discussed.
When Infants Lose Exclusive Maternal Attention: Is It Jealousy?
Authors: Hart Sybil L. , Carrington Heather A. , Tronick E. Z. , Carroll Sebrina R.
Published: September 2004

To characterize infant reactions to jealousy evocation, 94 6‐month‐olds and their mothers were videotaped in an episode where the mothers directed positive attention toward a lifelike doll, and in 2 contrasting interactions: face‐to‐face play and a still‐face perturbation. Cross‐context comparisons of affects and behaviors revealed that jealousy evocation responses were distinguished by diminished joy and heightened anger and intensity of negative emotionality, comparable to levels displayed during the still‐face episode; heightened sadness, with durations exceeding those displayed during still‐face exposure; and an approach response consisting of interest, looks at mother, and diminished distancing, which was more pronounced than that demonstrated during play. Infants' heightened anger and sadness during jealousy evocation correlated with heightened maternal sensitivity and dyadic vocal turn‐taking, respectively, during play; and infants' diminished joy and interest during jealousy evocation were associated with heightened maternal withdrawal and intrusiveness, respectively, during play. Both fear and mother‐directed gaze were greater in girls. The discussion argues for interpreting the infant's mixed and agitated reaction to jealousy evocation as evidence of jealousy.
Young Infants' Behavioral Reactivity to Mild Perturbation: Developmental Continuity, Stability, and Organization
Authors: Hsu Hui‐Chin , Porter Christin L.
Published: September 2004

This study investigated the developmental continuity, stability, and organization of infants' behavioral response to mild stress from 1 to 3 months of age. Changes in infant stress reactivity were expected to coincide with a suspected neurobiological shift around the 2nd month. A total of 53 infants and their mothers participated in this study. At 1 and 3 months, infants' stress reactivity to mild perturbation was assessed by withdrawing a pacifier during nonnutritive sucking. Infant baseline heart rate activity and maternal reports of infant temperament were also obtained. Focusing on facial and vocal negativity, behavioral measures of infant reactivity to pacifier withdrawal were coded from videos. Results showed that developmental continuity and individual instability characterized changes in infant stress reactivity over the 2nd‐month transition. Developmental reorganization was also evidenced by changes in the direction and magnitude of infant stress reactivity in relation to baseline heart period and maternal temperament ratings, respectively. These findings provide direct supporting evidence for the contention that changes in infant stress reactivity to mild perturbation may be a function of developmental shifts in underlying neurobiological mechanisms in early infancy.
The Onset of Working Memory in Infants
Authors: Reznick J. Steven , Morrow Judy D. , Goldman Barbara Davis , Snyder Jessica
Published: September 2004

We used an optimized configuration of the delayed‐response task to explore the ability of young infants to remember which of 2 locations was correct across 12 trials after a 1‐ to 2‐sec delay. Performance improved with age, particularly after 5.5 months. These findings suggest an onset of appreciable working memory for many infants in the middle of their 6th month.
A hypophosphorylated form of RPA34 is a specific component of pre-replication centers
Authors: Françon Patricia , Lemaître Jean-Marc , Dreyer Christine , Maiorano Domenico , Cuvier Olivier , Méchali Marcel
Published: September 2004

Replication protein A (RPA) is a three subunit single-stranded DNA-binding protein required for DNA replication. In Xenopus, RPA assembles in nuclear foci that form before DNA synthesis, but their significance in the assembly of replication initiation complexes has been questioned. Here we show that the RPA34 regulatory subunit is dephosphorylated at the exit of mitosis and binds to chromatin at detergent-resistant replication foci that co-localize with the catalytic RPA70 subunit, at both the initiation and elongation stages of DNA replication. By contrast, the RPA34 phosphorylated form present at mitosis is not chromatin bound. We further demonstrate that RPA foci assemble on chromatin before initiation of DNA replication at sites functionally defined as initiation replication sites. Association of RPA with these sites does not require nuclear membrane formation, and is sensitive to the S-CDK inhibitor p21. We also provide evidence that RPA34 is present at initiation complexes formed in the absence of MCM3, but which contain MCM4. In such conditions, replication foci can form, and short RNA-primed nascent DNAs of discrete size are synthesized. These data show that in Xenopus, the hypophosphorylated form of RPA34 is a component of the pre-initiation complex.
Cell cycle arrest at the initiation step of human chromosomal DNA replication causes DNA damage
Authors: Szüts Dávid , Krude Torsten
Published: September 2004

Cell cycle arrest in response to environmental effects can lead to DNA breaks. We investigated whether inhibition of DNA replication during the initiation step can lead to DNA damage and characterised a cell-cycle-arrest point at the replication initiation step before the establishment of active replication forks. This arrest can be elicited by the iron chelators mimosine, ciclopirox olamine or 2,2′-bipyridyl, and can be reversed by the removal of the drugs or the addition of excess iron. Iron depletion induces DNA double-strand breaks in treated cells, and activates a DNA damage response that results in focal phosphorylation of histone H2AX, focal accumulation of replication protein A (RPA) and ATR (ATM and Rad3-related kinase), and activation of CHK1 kinase. Abrogation of the checkpoint response does not abolish the cell cycle arrest before the establishment of active DNA replication forks. DNA breaks appear concomitantly with the arrival of cells at the arrest point and persist upon release from the cell cycle block. We conclude that DNA double-strand breaks are the consequence, and not the cause, of cell cycle arrest during the initiation step of DNA replication by iron chelation.
Tetraspanin CD82 controls the association of cholesterol-dependent microdomains with the actin cytoskeleton in T lymphocytes: relevance to co-stimulation
Authors: Delaguillaumie Alix , Harriague Julie , Kohanna Sylvie , Bismuth Georges , Rubinstein Eric , Seigneuret Michel , Conjeaud Hélène
Published: September 2004

T-cell activation is initiated by the concerted engagement of the T-cell receptor and different co-stimulatory molecules, and requires cytoskeleton-dependent membrane dynamics. Here, we have studied the relationships between tetraspanins, cytoskeleton and raft microdomains, and their relevance in T-cell signaling. Localization studies and density-gradient flotation experiments indicate that part of tetraspanins localizes in raft microdomains linked to the actin cytoskeleton. First, partial coalescence of lipid raft is triggered by tetraspanin cross-linking and results in large caps in which F-actin also concentrates. Second, the amount of tetraspanins, which are recovered in the cholesterol-dependent insoluble fractions of low and intermediate density, and which appears to be membrane vesicles by electron microscopy, is under cytoskeletal influence. Disruption of actin filaments enhances the amount of tetraspanins recovered in typical raft fractions, whereas F-actin-stabilizing agents induce the opposite effect. Our data also reveal that CD82 constitutes a link between raft domains and the actin cytoskeleton, which is functionally relevant. First, tetraspanin signaling induces a selective translocation of CD82 from detergent-resistant membrane fractions to the cytoskeleton-associated pellet. Second, all functional effects linked to CD82 engagement, such as adhesion to culture plates, formation of actin bundles and early events of tyrosine phosphorylation, are abolished, or strongly reduced, by cholesterol depletion. We also show that dynamic relocalization of CD82 and F-actin at the periphery of the immune synapse is induced upon contact of T cells with antigen-presenting cells. This suggests that the tetraspanin web might participate in the membrane dynamics required for proper T-cell signaling. More generally, the interaction of tetraspanins with raft domains and with the actin cytoskeleton might relate with their role in many cellular functions as membrane organizers.
p73 competes with co-activators and recruits histone deacetylase to NF-Y in the repression of PDGF β-receptor
Authors: Uramoto Hidetaka , Wetterskog Daniel , Hackzell Anders , Matsumoto Yoshiki , Funa Keiko
Published: September 2004

We investigated mechanisms of the p73α-mediated repression of the platelet-derived growth factor β-receptor (PDGFRB) promoter caused by its interaction with NF-Y. Treatment of cells with the histone deacetylase (HDAC) inhibitor, Trichostatin A, increases PDGFRB promoter activity through the CCAAT motif and counteracts the repression caused by p73α. Activation of the PDGFRB promoter by the co-activator p300 also occurs through the CCAAT motif. Expression of p73α counteracts both p300- and P/CAF-mediated activation of the PDGFRB promoter, and expression of p300 or P/CAF attenuates the p73α-mediated repression of the promoter activity. In concordance, p73α decreases the p300-mediated acetylation of NF-YC, p300 competes with p73α for binding NF-YB, and P/CAF competes with p73α for binding NF-YB and NF-YC. Furthermore, p73α, but not the oncogenic ΔNp73α, binds directly to HDAC1. We performed chromatin immunoprecipitation with antibodies against p73, ΔNp73, NFYB, p300 and HDAC1 at different periods after serum stimulation in serum-starved NIH3T3 cells. A marked decrease of ΔNp73, NF-YB and p300 was detected 6 hours after serum stimulation when the expression of PDGFRB decreased. Conversely, HDAC1 was found bound at its maximum and the anti-p73 detecting both TAp73 and ΔNp73 was found at all time points, indicating that p73, but not ΔNp73, remains bound at this time. Double immunofluorescence staining of TAp73 and HDAC1 revealed that both of these molecules exist in the nucleus at this time point, supporting the presence of endogenous interaction. These results suggest that p73 and ΔNp73 behave as physiological regulators for the transcription of the PDGFRB promoter.
Compartmentalization established by claudin-11-based tight junctions in stria vascularis is required for hearing through generation of endocochlear potential
Authors: Kitajiri Shin-ichiro , Miyamoto Tatsuo , Mineharu Akihito , Sonoda Noriyuki , Furuse Kyoko , Hata Masaki , Sasaki Hiroyuki , Mori Yoshiaki , Kubota Takahiro , Ito Juichi , Furuse Mikio , Tsukita Shoichiro
Published: September 2004

Claudins are cell adhesion molecules working at tight junctions (TJs) that are directly involved in compartmentalization in multicellular organisms. The cochlea includes a rather peculiar compartment filled with endolymph. This compartment is characterized by high K+ concentration (∼150 mM) and a positive endocochlear potential (∼90 mV; EP), both indispensable conditions for cochlear hair cells to transduce acoustic stimuli to electrical signals. These conditions are thought to be generated by the stria vascularis, which is adjacent to the endolymph compartment. The stria vascularis itself constitutes an isolated compartment delineated by two epithelial barriers, marginal and basal cell layers. Because TJs of basal cells are primarily composed of claudin-11, claudin-11-deficient (Cld11-/-) mice were generated with an expectation that the compartmentalization in stria vascularis in these mice would be affected. Auditory brainstem response measurements revealed that Cld11-/- mice suffered from deafness; although no obvious gross morphological malformations were detected in Cld11-/- cochlea, freeze-fracture replica electron microscopy showed that TJs disappeared from basal cells of the stria vascularis. In good agreement with this, tracer experiments showed that the basal cell barrier was destroyed without affecting the marginal cell barrier. Importantly, in the endolymph compartment of Cld11-/- cochlea, the K+ concentration was maintained around the normal level (∼150 mM), whereas the EP was suppressed down to ∼30 mV. These findings indicated that the establishment of the stria vascularis compartment, especially the basal cell barrier, is indispensable for hearing ability through the generation/maintenance of EP but not of a high K+ concentration in the endolymph.
Caenorhabditis eleganssyndecan (SDN-1) is required for normal egg laying and associates with the nervous system and the vulva
Authors: Minniti Alicia N. , Labarca Mariana , Hurtado Claudia , Brandan Enrique
Published: September 2004

In Caenorhabditis elegans, the identification of many enzymes involved in the synthesis and modification of glycosaminoglycans (GAGs), essential components of proteoglycans, has attained special attention in recent years. Mutations in all the genes that encode for GAG biosynthetic enzymes show defects in the development of the vulva, specifically in the invagination of the vulval epithelium. Mutants for certain heparan sulfate modifying enzymes present axonal and cellular guidance defects in specific neuronal classes. Although most of the enzymes involved in the biosynthesis and modification of heparan sulfate have been characterized in C. elegans, little is known regarding the core proteins to which these GAGs covalently bind in proteoglycans. A single syndecan homologue (sdn-1) has been identified in the C. elegans genome through sequence analysis. In the present study, we show that C. elegans synthesizes sulfated proteoglycans, seen as three distinct species in western blot analysis. In the sdn-1 (ok449) deletion mutant allele we observed the lack of one species, which corresponds to a 50 kDa product after heparitinase treatment. The expression of sdn-1 mRNA and sequencing revealed that sdn-1 (ok449) deletion mutants lack two glycosylation sites. Hence, the missing protein in the western blot analysis probably corresponds to SDN-1. In addition, we show that SDN-1 localizes to the C. elegans nerve ring, nerve cords and to the vulva. SDN-1 is found specifically phosphorylated in nerve ring neurons and in the vulva, in both wild-type worms and sdn-1 (ok449) deletion mutants. These mutants show a defective egg-laying phenotype. Our results show for the first time, the identification, localization and some functional aspects of syndecan in the nematode C. elegans.
Visualization of SHP-1-target interaction
Authors: Biskup Christoph , Böhmer Annette , Pusch Rico , Kelbauskas Laimonas , Gorshokov Alexander , Majoul Irina , Lindenau Jörg , Benndorf Klaus , Böhmer Frank-D.
Published: September 2004

Signaling of receptor tyrosine kinases (RTKs) is regulated by protein-tyrosine phosphatases (PTPs). We previously discovered the efficient downregulation of Ros RTK signaling by the SH2 domain PTP SHP-1, which involves a direct interaction of both molecules. Here, we studied the mechanism of this interaction in detail. Phosphopeptides representing the SHP-1 candidate binding sites in the Ros cytoplasmic domain, pY2267 and pY2327, display high affinity binding to the SHP-1 N-terminal SH2 domain (Kd=217 nM and 171 nM, respectively). Y2327 is, however, a poor substrate of Ros kinase and, therefore, contributes little to SHP-1 binding in vitro. To explore the mechanism of association in intact cells, functional fluorescent fusion proteins of Ros and SHP-1 were generated. Complexes of both molecules could be detected by Förster resonance energy transfer (FRET) in intact HEK293 and COS7 cells. As expected, the association required the functional SHP-1 N-terminal SH2 domain. Unexpectedly, pY2267 and pY2327 both contributed to the association. Mutation of Y2327 reduced constitutive association in COS7 cells. Ligand-dependent association was abrogated upon mutation of Y2267 but remained intact when Y2327 was mutated. A phosphopeptide representing the binding site pY2267 was a poor substrate for SHP-1, whereas Ros activation loop phosphotyrosines were effectively dephosphorylated. We propose a model for SHP-1-Ros interaction in which ligand-stimulated phosphorylation of Ros Y2267 by Ros, phosphorylation of Y2327 by a heterologous kinase, and inactivation of Ros by SHP-1-mediated dephosphorylation play a role in the regulation of complex stability.
Tetraspanin protein (TSP-15) is required for epidermal integrity inCaenorhabditis elegans
Authors: Moribe Hiroki , Yochem John , Yamada Hiromi , Tabuse Yo , Fujimoto Toyoshi , Mekada Eisuke
Published: September 2004

Epidermal integrity is essential for animal development and survival. Here, we demonstrate that TSP-15, a member of the tetraspanin protein family, is required for epithelial membrane integrity in Caenorhabditis elegans. Reduction of tsp-15 function by mutation or by RNA interference elicits abnormalities of the hypodermis, including dissociation of the cuticle and degeneration of the hypodermis. Lethality during molting often results. Examination of GFP transgenic animals, genetic mosaic analysis and rescue assays revealed that TSP-15 functions in hyp7, a large syncytium that composes most of the hypodermis. Assays with a membrane-impermeable dye or leakage analysis of a hypodermal-specific marker indicate that the barrier function of the hypodermal membrane is impaired owing to the loss or reduction of TSP-15. These results indicate that TSP-15 functions in the maintenance of epithelial cell integrity.
Mechanical stress induces profound remodelling of keratin filaments and cell junctions inepidermolysis bullosa simplexkeratinocytes
Authors: Russell David , Andrews Paul D. , James John , Lane E. Birgitte
Published: September 2004

The outer epidermal layer of the skin is an epithelium with remarkable protective barrier functions, which is subject to pronounced physical stress in its day-to-day function. A major candidate component for absorbing this stress is the K5/K14 keratin intermediate filament network. To investigate the part played by keratins in stress resilience, keratinocyte cell lines were subjected to mechanical stress. Repeated stretch and relaxation cycles over increasing time produced reproducible changes in the configuration of the keratin network. When wild-type cells were compared with cells carrying a keratin mutation associated with severe epidermolysis bullosa simplex-type skin fragility, the mutant keratin filaments were unable to withstand the mechanical stress and progressively fragmented yielding aggregates and novel ring structures. The cell junctions into which the keratin filaments are normally anchored also progressively disassembled, with all components tested of the cytoplasmic plaques becoming relocated away from the membrane and onto the keratin rings, while integral membrane receptors integrins and cadherins remained at the plasma membrane. The results suggest that maintenance of desmosomes and hemidesmosomes may require some tension, normally mediated by keratin attachments.
Macroscopic folding and replication of the homogeneously staining region in late S phase leads to the appearance of replication bands in mitotic chromosomes
Authors: Shimizu Noriaki , Shingaki Kenta
Published: September 2004

The chromosomal G/R bands are alternating domains differing in their nucleotide sequence biases. The bands are also related to the time of replication: pulse-labeling during S phase makes the replication sites as visible as replication bands that are close to the G/R bands in mitotic chromosomes. We previously showed that a plasmid bearing a mammalian replication origin efficiently generated a chromosomal homogeneously staining region (HSR). Here, we analyze the replication of this artificial HSR and show that it was replicated at the last stage of S phase. The HSR was composed of plasmid repeats only; nonetheless, we found that replication sites pulse-labeled during late S phase appeared as bands in the mitotic HSR and their number was dependent on the length of the HSR. Therefore, replication bands might not arise from sequence information per se. To understand the chronological order of appearance of replication sites, we performed a double pulse-chase experiment using IdU and CldU. Replication of the entire HSR required 100-120 minutes. During this period, the replicated sites appeared as bands at the first and last stages, but in between were apparently scattered along the entire HSR. An analysis of S-phase nuclei revealed that the replication started at the periphery of the globular HSR domain, followed by initiation in the internal domain. The replicated HSR appeared as a ring or a pair of extended spirals in late G2-phase nuclei. To account for these findings, we present a model in which the HSR is folded as a coiled-coil structure that is replicated from the outside to the inside in S phase nuclei.
RNA-directed DNA methylation
Authors: Mathieu Olivier , Bender Judith
Published: September 2004

Double-stranded RNAs (dsRNAs) and their `diced' small RNA products can guide key developmental and defense mechanisms in eukaryotes. Some RNA-directed mechanisms act at a post-transcriptional level to degrade target messenger RNAs. However, dsRNA-derived species can also direct changes in the chromatin structure of DNA regions with which they share sequence identity. For example, plants use such RNA species to lay down cytosine methylation imprints on identical DNA sequences, providing a fundamental mark for the formation of transcriptionally silent heterochromatin. Thus, RNA can feed backwards to modulate the accessibility of information stored in the DNA of cognate genes. RNA triggers for DNA methylation can come from different sources, including invasive viral, transgene or transposon sequences, and in some cases are derived from single-stranded RNA precursors by RNA-dependent RNA polymerases. The mechanism by which RNA signals are translated into DNA methylation imprints is currently unknown, but two plant-specific types of cytosine methyltransferase have been implicated in this process. RNA can also direct heterochromatin formation in fission yeast and Drosophila, but in these organisms the process occurs in the absence of DNA methylation.
Multiplex Sum-frequency Spectroscopy with Electronic Resonance Enhancement
Authors: Ishibashi Taka-aki , Onishi Hiroshi
Published: September 2004

Multiplex detection of broad bandwidth light brings remarkable improvements in the experimental design of sum-frequency spectroscopy. Highly sensitive, interface-specific observations of molecular vibrations are demonstrated with examples of n-alkyl chains covalently anchored on Si(111) and an organic compound chemisorbed on TiO2(110).
Lithium Alkoxide-promoted Michael Reaction between Silyl Enolates and α,β-Unsaturated Carbonyl Compounds
Authors: Mukaiyama Teruaki , Tozawa Takashi , Fujisawa Hidehiko
Published: September 2004

Michael reaction between silyl enolates and α,β-unsaturated carbonyl compounds by using a catalytic amount of Lewis base such as lithium alkoxide in DMF proceeds smoothly to afford the corresponding Michael-adducts in good yields with moderate to high diastereoselectivities. This reaction can be reasonably explained by considering an alkoxide anion-initiated autocatalytic process.
Automatic fall detectors and the fear of falling
Authors: Brownsell Simon , Hawley Mark S.
Published: September 2004

We studied the effect of automatic fall detection units on the fear of falling. Participants were community alarm users living in the community aged over 75 years or those aged 60–74 years who had experienced a fall in the previous six months. Of those approached, 31% consented to take part; the main reason given for potential participants declining involvement was that they were happy with the technology they already had. Subjects were assigned to a control group ( n = 21) or intervention group ( n = 34) based on age, the number of self-reported falls in the previous six months and their score on the self-administered Falls Efficacy Scale (FES), which measures fear of falling on a scale of 0–100, with higher scores indicating less fear. The monitoring period lasted a mean of 17 weeks (SD 3.1). There was no significant difference between the intervention and control groups in their mean ratings of fear of falls (40.3 vs 37.5, difference 2.8, 95% CI 6.2 to 11.8), health-related quality of life or morale. Differences in fear of falling between an intervention subgroup who wore their detector at least occasionally (62%) and those who did not (38%) suggested that some people may benefit from a fall detector while others may lose confidence if they are provided with one. Most users who wore their detectors at least occasionally felt more confident and independent and considered that the detector improved their safety.
Web-based diabetes control
Authors: Ferrer-Roca O. , Franco Burbano K. , Cárdenas A. , Pulido P. , Diaz-Cardama A.
Published: September 2004

We developed a diabetes management system with a Web interface that allowed patients to transmit their physiological data using either a PC Web browser or a mobile phone capable of working with the WAP protocol. The system could generate automatic responses to input values in accordance with a set of ranges previously defined for each user. User analysis was carried out with personal and online questionnaires. During a nine-month study period on the island of Tenerife, 12 patients were recruited. On average, they used the system every 2.0 days (SD 2.1) and the doctors reviewed their patient data every 4.0 days (SD 3.9). The average number of visits to the Website was 477 per month. Users were satisfied with the continuity and self-efficacy of care, but lack of time was a drawback for 38% of them and 75% expressed a preference for sending their data via the mobile phone short message service (SMS).
Nine years' experience of telecardiology in primary care
Authors: Molinari G. , Valbusa A. , Terrizzano M. , Bazzano M. , Torelli L. , Girardi N. , Barsotti A.
Published: September 2004

The ITMS telecardiology network started in April 1995. Over nine years, about 7000 peripheral medical users (mainly general practitioners), who were experienced in using transtelephonic electrocardiography and who were spread throughout Italy, conducted teleconsultations with the Telecardiology Centre. A total of 106,942 patients were evaluated. Teleconsultation was carried out for non-cardiac symptoms or routine control for 30,444 patients (28%) and for symptoms suggestive of heart disease for 76,498 (72%). After teleconsultation, 61,908 patients (58%) were reported to have no heart disease, 27,947 patients (26%) had their drug dose adjusted and remained at home, 11,503 patients (11%) were sent to their cardiologist for further investigations and 5584 patients (5%) were urgently hospitalized. Among the hospitalized patients, the echocardiogram demonstrated an ST-elevation coronary syndrome in 1785 patients (32%), a non-ST-elevation coronary syndrome in 2236 (40%) and a life-threatening arrhythmia in 1354 (24%). Telecardiology improves the decision making of general practitioners, avoids unnecessary hospitalizations, reduces the time before treatment in cardiac emergencies, rationalizes health-care costs and promotes home care.
Operators' experiences of emergency calls
Authors: Forslund Kerstin , Kihlgren Annica , Kihlgren Mona
Published: September 2004

In Sweden, the operators at emergency dispatch centres are responsible for allocating resources (e.g. ambulances, fire brigade, police) in response to calls. We analysed situations that the emergency operators experienced as difficult and their reflections on how they managed them. Interviews were conducted with all 16 emergency operators at a centre that serves a population of 275,000 and receives about 700,000 emergency calls annually. A phenomenological–hermeneutic approach was used for the analysis. Situations that operators experienced as difficult were characterized by uncertainty, communication difficulties and insufficient resources. Skills, knowledge and experience were regarded as important in the management of these situations, as were personal qualities such as sensitivity, insight, empathy and intuition. The emergency operators stated that they needed more guidance, feedback and education in their work. This would lead to an increased sense of certainty, which would lead to decreased stress and a better outcome for those in need.
A multi-functional, portable device with wireless transmission for home monitoring of children with a learning disability
Authors: Tura A. , Badanai M. , Longo D. , Quareni L.
Published: September 2004

A portable monitoring device was developed to assist in the management of children with a learning disability. The device was designed for continuous home monitoring of blood oxygen saturation, heart and respiration rates, and patient activity. It could be worn on a belt, while the patient continued normal activities. Data were stored on a multimedia card and automatically transmitted to a PC at prescribed intervals via a Bluetooth wireless link. From the PC the data were transmitted to a Web server, where the information was made available to the staff involved in the patient's care. Preliminary clinical studies were performed with nine patients (four with Down's syndrome, three with cerebral palsy and two with mental retardation). Patients and families considered the device easy to use and to wear. The monitoring device identified events of possible clinical interest. Although it was designed for monitoring children with a learning disability, it may also be useful with other groups, such as elderly people.
Telemedicine and coping skills groups for Pacific Island veterans with post-traumatic stress disorder: a pilot study
Authors: Morland Leslie A. , Pierce Kathleen , Wong Matthew Y.
Published: September 2004

Patients with post-traumatic stress disorder (PTSD) were randomly assigned to either an eight-week videoconferencing PTSD coping skills group or a traditional face-to-face PTSD coping skills group. Levels of attrition and compliance, patient satisfaction, clinician satisfaction and patients' retention of information were compared between the two conditions. Of the 41 referred veterans, 20 were eligible and agreed to participate in the study. Three of these participants withdrew from the study before randomization. By the end of the study, 89% of the patients remained in the videoconferencing group, whereas only 50% remained in the face-to-face group. Patients in the face-to-face group attended an average of 4.9 sessions and patients in the videoconferencing group attended 6.3 sessions (this difference was not significant). There was no difference between levels of patient satisfaction or clinician satisfaction at weeks 4 or 8. Patients' retention of information was similar in the two groups. The results show that videoconferencing can be used to provide coping skills groups for veteran patients with PTSD who reside in remote rural locations.
A Community-Based Educational Intervention to Improve Antithrombotic Drug Use in Atrial Fibrillation
Authors: Jackson Shane L , Peterson Gregory M , Vial Janet H
Published: September 2004

BACKGROUND

Despite evidence that antithrombotics are effective in reducing the risk of stroke in atrial fibrillation (AF), they remain underused.

OBJECTIVE

To perform a controlled trial of a comprehensive educational program promoting the rational prescribing of antithrombotics for stroke prevention in AF.

METHODS

The intervention was conducted in Southern Tasmania, Australia, using Northern Tasmania as a control area. General practitioners were sent locally produced guidelines on stroke risk stratification and antithrombotic drug use in AF, which were followed by academic detailing visits. Outcomes were measured using evaluation feedback from the general practitioners, and drug utilization data were provided by a series of patients presenting to the hospital with an admission diagnosis of AF and dispensing of antithrombotic therapy under the Australian Pharmaceutical Benefits Scheme.

RESULTS

During the educational intervention, 272 guidelines were mailed and, subsequently, 162 general practitioners were visited and the guidelines discussed. Hospital admission data before and after the intervention revealed a significant increase in the use of warfarin in patients at high risk of stroke (33% vs 46% of eligible patients; p < 0.05). Analysis of prescription data for warfarin also indicated that the increase in use of warfarin within the intervention region was significantly greater than for the control region (p < 0.001).

CONCLUSIONS

The educational program described here led to a significant increase in the prescribing of warfarin for stroke prevention in patients with AF.
The Human H2A and H2B Histone Gene Complement
Authors: Albig W. , Trappe R. , Kardalinou E. , Eick S. , Doenecke D.
Published: September 2004

Sequences and expression patterns of newly isolated human histone H2A and H2B genes and the respective proteins were compared with previously isolated human H2A and H2B genes and proteins. Altogether, 15 human H2A genes and 17 human H2B genes have been identified. 14 of these are organized as H2A/H2B gene pairs, while one H2A gene and three H2B genes are solitary genes. Two H2A genes and two H2B genes turned out to be pseudogenes. The 13 H2A genes code for at least 6 different amino acid sequences, and the 15 H2B genes encode 11 different H2B isoforms. Each H2A/H2B gene pair is controlled by a divergent promoter spanning 300 to 330 nucleotides between the coding regions of the two genes. The highly conserved divergent H2A/H2B promoters can be classified in two groups based on the patterns of consensus sequence elements. Group I promoters contain a TATA box for each gene, two Oct-1 factor binding sites, and three CCAAT boxes. Group II promoters contain the same elements as group I promoters and an additional CCAAT box, a binding motif for E2F and adjacent a highly conserved octanucleotide (CACAGCTT) that has not been described so far. Five of the 6 gene pairs and 4 solitary genes with group I promoters are localized in the large histone gene cluster at 6p21.3–6p22, and one gene pair is located at 1q21. All group II promoter associated genes are contained within the histone gene subcluster at D6S105, which is located at a distance of about 2 Mb from the major subcluster at 6p21.3–6p22 containing histone genes with group I promoters. Almost all group II H2A genes encode identical amino acid sequences, whereas group I H2A gene products vary at several positions. Using human cell lines, we have analyzed the expression patterns of functional human H2A/H2B gene pairs organized within the two histone gene clusters on the short arm of chromosome 6. The genes show varying expression patterns in different tumor cell lines.
Efficient Control of raf Gene Expression by CAP and Two Raf Repressors that Bend DNA in Opposite Directions
Authors: Muiznieks I. , Rostoks N. , Schmitt R.
Published: September 2004

The plasmid-borne
Studies with Lysine N6-Hydroxylase. Effect of a Mutation in the Assumed FAD Binding Site on Coenzyme Affinities and on Lysine Hydroxylating Activity
Authors: Stehr M. , Smau L. , Singh M. , Seth O. , Macheroux P. , Ghisla S. , Diekmann H.
Published: September 2004

The proposed FAD binding site of L-lysine N
The Recombinant Thermosome from the Hyperthermophilic Archaeon Methanopyrus kandleri: In Vitro Analysis of Its Chaperone Activity
Authors: Minuth T. , Henn M. , Rutkat K. , Andrä S. , Frey G. , Rachel R. , Stetter K.O. , Jaenicke R.
Published: September 2004

The archaeon

In order to obtain the chaperonin in amounts sufficient for functional investigations, the gene encoding the 60 kDa subunit was expressed in

The results demonstrate that the recombinant
Not More than Three Tissue Kallikreins Identified from Organs of the Guinea Pig
Authors: Fiedler F. , Betz G. , Hinz H. , Lottspeich F. , Raidoo D.M. , Bhoola K.D.
Published: September 2004

The large and varied multigene families of tissue kallikreins of rat and mouse are considered to selectively release as many bioactive peptides. In order to determine whether a similar family of enzymes is expressed in the organs of the guinea pig purification studies were performed. Tissue kallikreins from the submandibular gland, coagulating gland/prostate complex and the pancreas were separated by affinity chromatography on benzamidine-Sepharose. Amino-terminal sequences, the patterns of hydrolysis rates of a number of peptide

Guinea pig tissue kallikreins 1 and 2 have been reported previously. In the present study we have identified a third type, designated tissue kallikrein 1a because of its sequence similarity to kallikrein 1, which differs from the latter in the catalytic properties. The inferred occurrence of not more than two or three independent tissue kallikrein genes in the guinea pig contrasts with the varied family of enzymes expressed by the large number of such genes present in rats and mice. Expression in the guinea pig (and also in humans) of only a small number of tissue kallikreins makes specific processing of a multitude of biologically active peptides by such enzymes unlikely.