Benefit of continuous positive airway pressure on work quality in patients with severe obstructive sleep apnea.

BACKGROUND: The objective of this prospective study was to assess the effect of CPAP therapy on job productivity and work quality for patients with severe obstructive sleep apnea (OSA). METHODS: A convenience sample of patients diagnosed with severe OSA using polysomnography or polygraphy and with a therapeutic indication for CPAP was enrolled in our study. Patients completed two self-administered questionnaires: the first before CPAP therapy and the second during the first 6 months after CPAP treatment. OSA symptoms were evaluated through self-administered questionnaires assessing potential effects on occupational activity: excessive daytime sleepiness was rated by the Epworth Sleepiness Scale (ESS), emotional status was rated by the Hospital Anxiety and Depression (HAD) scale, work quality was rated by the Work Role Functioning Questionnaire (WRFQ). RESULTS: Forty patients (30 men, mean age 47.3 ± 8.3, mean BMI 31.6 ± 7.4, mean apnea-hypopnea index 51.8 ± 16.3) showed a beneficial effect of CPAP therapy on ESS score (mean 11.6 to 8.2, p < 0.0001), the anxiety dimension (mean 57.5% to 20%, p = 0.0002), and the overall anxiety-depressive score (mean 50% to 22.5%, p = 0.0006). Mean WRFQ scores were significantly improved in the second questionnaire for the dimensions of timetable requirements (69.3% to 83.5%, p < 0.0001), productivity requirements (71.4% to 82.2%, p < 0.0001), mental requirements (72.0% to 84.3%, p < 0.0001), and social requirements (82.6% to 91.4%, p < 0.003). CONCLUSIONS: We observed that adherence to CPAP therapy for patients with severe OSA mitigates the impact of symptoms on work including excessive daytime sleepiness, impairment of work ability, and anxiety and depressive disorders.

Microstructure, surface characterization and long-term stability of new quaternary Ti-Zr-Ta-Ag alloy for implant use.

The novel Ti-20Zr-5Ta-2Ag alloy was characterised concerning its microstructure, morphology, mechanical properties, its passive film composition and thickness, its long-term electrochemical stability, corrosion resistance, ion release rate in Ringer solution of acid, neutral and alkaline pH values and antibacterial activity. The new alloy has a crystalline α microstructure (by XRD). Long-term XPS and SEM analyses show the thickening of the passive film and the deposition of hydroxyapatite in neutral and alkaline Ringer solution. The values of the electrochemical parameters confirm the over time stability of the new alloy passive film. All corrosion parameters have very favourable values in time which attest a high resistance to corrosion. Impedance spectra evinced a bi-layered passive film formed by the barrier, insulating layer and the porous layer. The monitoring of the open circuit potentials indicated the stability of the protective layers and their thickening in time. The new alloy releases (by ICP-MS measurements) very low quantities of Ti, Zr, Ag ions and no Ta ions. The new alloy exhibits a low antibacterial activity.

[Lung abscess due to community acquired Klebsiella pneumonia]. / Pneumopathie communautaire abcédée à Klebsiella pneumoniae.

INTRODUCTION: In western countries, community-acquired pneumonias due to Klebsiella pneumoniae (Kp) are rare and associated with a poor prognosis and a high mortality. The severity is in part linked to the virulence of Kp. Immuno-depression, sepsis and visceral abscesses are frequently found, constituting other classical risk factors for severity and contributing to the poor prognosis. The therapeutic strategy is based on third generation cephalosporins, aminoglycosides and quinolones. CASE REPORT: We report the case of a young adult, with undiagnosed diabetes, hospitalized as an emergency for septic shock complicating a community-acquired pneumonia due to Kp and associated with multiple brain and lung abscesses. After several weeks of treatment, initially with empirical then specific antibiotics, a favourable outcome was obtained. CONCLUSION: This case report underlines the particular severity of infections due to Kp and their main pathophysiological mechanisms. It is also an opportunity to highlight the potential responsibility of Kp in the presence of a pneumonia with lung abscesses and finally to update the principles of antibiotic therapy.

[Ewing's tumor]. / La tumeur d'Ewing.

Ewing's tumor (ET) is a malignant bone tumor occurring in children and young adults. ET affects mainly bones of the central axis, and almost always involves soft tissue infiltration. The discovery of a unique genetic alteration, which is a reciprocal translocation most frequently resulting in the fusion of the EWS gene situated on chromosome 22 with the FLI-1 gene on chromosome 11, currently places ET among neuroectodermal tumors. Moreover, this translocation is a tumor-specific genetic marker at the basis of defining ET today and is used as a diagnostic and potentially prognostic tool complementary to imaging and histopathological work-up. Since the 1970 s, important progress has been made in the clinical management of ET patients. Multiagent chemotherapy in association with local treatment (surgery and/or radiation) has clearly improved outcome. The introduction of systemic treatment was justified by the frequent sub-clinical diffusion of apparently localized ET. Intensified therapeutic strategies have for the first time cured some metastatic ET patients, but at the cost of major side effects. Treatment is currently adapted as a result of a better definition of prognostic factors as well as a better assessment of its adverse effects. Improvement in global patient care and increased management of specific acute complications associated with ET (often interwoven with iatrogeneous effects) represent an important step towards improving the quality of life for ET patients as well as preventing long term complications. In the light of present studies, the majority of surviving adults today describe their health and quality of life as good. ET is a fascinating example of the progress made not only in the diagnostic and therapeutic approach to cancer but also in the comprehension of the mechanisms behind carcinogenesis, and consequently reflects the revolution of medicine over the last century.

[Sequential approach in a case of tracheal hemangioma: segmental tracheal resection after Nd-YAG laser application]. / Abord secvential intr-un caz de hemangiom traheal: rezectie traheala segmentara precedata de Nd-YAG laser.

Tracheal tumors of malignant or benign origin are very rare. The symptoms may mimic asthmatic crisis, dyspnea at rest or light efforts appear only when the tumor obstructs 60% of the tracheal diameter. We present the case of a 50 year old patient, ex-smoker with symptoms present 5 years before admittance with dyspnea and small hemoptysis. Diagnosis was based on bronchoscopic examination, CT scan and histological examination of the resection sample revealing a rare benign tracheal tumor: a hemangioma. The sequential treatment of the disease is presented: interventional endoscopy and surgical resection. The excellent postoperative evolution emphasized the diagnostic and therapeutic value of bronchoscopy as well as surgery in benign tracheal tumors.

Caspase-dependent deubiquitination of monoubiquitinated nucleosomal histone H2A induced by diverse apoptogenic stimuli.

Enzymatic deubiquitination of mono-ubiquitinated nucleosomal histone H2A (uH2A) and H2B (uH2B) is closely associated with mitotic chromatin condensation, although the function of this histone modification in cell division remains ambiguous. Here we show that rapid and extensive deubiquitination of nucleosomal uH2A occurs in Jurkat cells undergoing apoptosis initiated by anti-Fas activating antibody, staurosporine, etoposide, doxorubicin and the proteasome inhibitor, N-acetyl-leucyl-leucyl-norlucinal. These diverse apoptosis inducers also promoted the accumulation of slowly migrating, high molecular weight ubiquitinated proteins and depleted the cellular pool of unconjugated ubiquitin. In apoptotic cells, ubiquitin was cleaved from uH2A subsequent to the appearance of plasma membrane blebbing, and deubiquitination of uH2A closely coincided with the onset of nuclear pyknosis and chromatin condensation. Nucleosomal uH2A deubiquitination, poly (ADP-ribose)polymerase (PARP) cleavage and chromatin condensation were prevented in cells challenged with apoptosis inducers by pretreatment with the pan-caspase inhibitor, zVAD-fmk, or by over-expressing anti-apoptotic Bcl-xL protein. These results implicate a connection between caspase cascade activation and nucleosomal uH2A deubiquitination. Transient transfection of 293 cells with the gene encoding Ubp-M, a human deubiquitinating enzyme, promoted uH2A deubiquitination, while an inactive mutated Ubp-M enzyme did not. However, Ubp-M-promoted deubiquitination of uH2A was insufficient to initiate apoptosis in these cells. We conclude that uH2A deubiquitination is a down-stream consequence of procaspase activation and that unscheduled cleavage of ubiquitin from uH2A is a consistent feature of the execution phase of apoptosis rather than a determining or initiating apoptogenic event. Nucleosomal uH2A deubiquitination may function as a cellular sensor of stress in situations like apoptosis through which cells attempt to preserve genomic integrity.

An antisense oligodeoxynucleotide targeted to chromaffin cell scinderin gene decreased scinderin levels and inhibited depolarization-induced cortical F-actin disassembly and exocytosis.

Chromaffin cell secretion requires cortical F-actin disassembly and it has been suggested that scinderin, a Ca2+ dependent F-actin severing protein, controls cortical actin dynamics. An antisense oligodeoxynucleotide targeting the scinderin gene was used to decrease the expression of the protein and access its role in secretion. Treatment with 2 microM scinderin antisense oligodeoxynucleotide for 4 days produced a significant decrease in scinderin expression and its mRNA levels. The expression of gelsolin, another F-actin severing protein, was not affected. Scinderin decrease was accompanied by concomitant and parallel decreases in depolarization-evoked cortical F-actin disassembly and exocytosis. Similar treatment with a mismatched oligodeoxynucleotide produced no effects. Scinderin antisense oligodeoxynucleotide treatment was also a very effective inhibitor of exocytosis in digitonin-permeabilized cells stimulated with increasing concentrations of Ca2+. This ruled out scinderin antisense interference with stimulation-induced depolarization or Ca2+ channel activation. Scinderin antisense treatment decreased the maximum (B(max)) secretory response to Ca2+ without modifying the affinity (K(m)) of the cation for the exocytotic machinery. Moreover, the antisense treatment did not affect norepinephrine uptake or the expression of dopamine ss-hydroxylase, suggesting that the number and function of chromaffin vesicles was not modified. In addition, scinderin antisense treatment did not alter the expression of proteins involved in vesicle-plasma membrane fusion, such as synaptophysin, synaptotagmin or syntaxin, indicating a lack of effects on the fusion machinery components. These observations strongly suggest that scinderin is a key player in the events involved in the secretory process.

Sensitivity of mature Erbb2 to geldanamycin is conferred by its kinase domain and is mediated by the chaperone protein Hsp90.

ErbB receptors are a family of ligand-activated tyrosine kinases that play a central role in proliferation, differentiation, and oncogenesis. ErbB2 is overexpressed in >25% of breast and ovarian cancers and is correlated with poor prognosis. Although ErbB2 and ErbB1 are highly homologous, they respond quite differently to geldanamycin (GA), an antibiotic that is a specific inhibitor of the chaperone protein Hsp90. Thus, although both mature and nascent ErbB2 proteins are down-regulated by GA, only nascent ErbB1 is sensitive to the drug. To reveal the underlying mechanism behind these divergent responses, we made a chimeric receptor (ErbB1/2) composed of the extracellular and transmembrane domains of ErbB1 and the intracellular domain of ErbB2. The ErbB1/2 protein is functional since its kinase activity was stimulated by epidermal growth factor. The sensitivity of ErbB1/2 to GA was similar to that of ErbB2 and unlike that of ErbB1, indicating that the intracellular domain of the chimera confers GA sensitivity. This finding also suggests that the GA sensitivity of mature ErbB2 depends on cytosolic Hsp90, rather than Grp94, a homolog of Hsp90 that is restricted to the lumen of the endoplasmic reticulum, although both chaperones bind to and are inhibited by GA. Lack of Grp94 involvement in mediating ErbB2 sensitivity to GA is further suggested by the fact that a GA derivative with low affinity for Grp94 efficiently depleted ErbB2 protein in treated cells. To localize the specific region of ErbB2 that confers GA sensitivity, we made truncated receptors with progressive deletions of the cytoplasmic domain and tested the GA sensitivity of these molecules. We found that ErbB2 constructs containing an intact kinase domain retained GA sensitivity, whereas those lacking the kinase domain (ErbB2/DK) lost responsiveness to GA completely. Hsp90 co-immunoprecipitated with all ErbB2 constructs that were sensitive to GA, but not with ErbB2/DK or ErbB1. Both tyrosine-phosphorylated and non-phosphorylated ErbB2 proteins were similarly sensitive to GA, as was a kinase-dead ErbB2 mutant. These data suggest that Hsp90 uniquely stabilizes ErbB2 via interaction with its kinase domain and that GA stimulates ErbB2 degradation secondary to disruption of ErbB2/Hsp90 association.

The heat shock protein 90 antagonist novobiocin interacts with a previously unrecognized ATP-binding domain in the carboxyl terminus of the chaperone.

Heat shock protein 90 (Hsp90), one of the most abundant chaperones in eukaryotes, participates in folding and stabilization of signal-transducing molecules including steroid hormone receptors and protein kinases. The amino terminus of Hsp90 contains a non-conventional nucleotide-binding site, related to the ATP-binding motif of bacterial DNA gyrase. The anti-tumor agents geldanamycin and radicicol bind specifically at this site and induce destabilization of Hsp90-dependent client proteins. We recently demonstrated that the gyrase inhibitor novobiocin also interacts with Hsp90, altering the affinity of the chaperone for geldanamycin and radicicol and causing in vitro and in vivo depletion of key regulatory Hsp90-dependent kinases including v-Src, Raf-1, and p185(ErbB2). In the present study we used deletion/mutation analysis to identify the site of interaction of novobiocin with Hsp90, and we demonstrate that the novobiocin-binding site resides in the carboxyl terminus of the chaperone. Surprisingly, this motif also recognizes ATP, and ATP and novobiocin efficiently compete with each other for binding to this region of Hsp90. Novobiocin interferes with association of the co-chaperones Hsc70 and p23 with Hsp90. These results identify a second site on Hsp90 where the binding of small molecule inhibitors can significantly impact the function of this chaperone, and they support the hypothesis that both amino- and carboxyl-terminal domains of Hsp90 interact to modulate chaperone activity.

Repetitive acute pancreatic injury in the mouse induces procollagen alpha1(I) expression colocalized to pancreatic stellate cells.

Pancreatic stellate cells may be a major source of extracellular matrix deposition during injury. This study was undertaken to establish whether pancreatic stellate cells are a source of Type I collagen in vivo and whether they continue to be a source of matrix production in the post-injury fibrotic pancreas. To induce pancreatic fibrogenesis, acute pancreatic injury was induced in mice three times weekly with supraphysiologic doses of cerulein. Animals were treated for 6 weeks and allowed to recover for an additional 6 weeks. Stellate cell activation and pancreatic collagen expression were measured by immunohistochemistry, whole tissue RNA analysis, and in situ hybridization. Histology and digital image analysis demonstrated the development of substantial pancreatic fibrosis after 6 weeks of treatment. During recovery, incomplete resolution of the fibrosis was found. Procollagen alpha1(I) mRNA increased more than 15-fold during treatment and continued to be 5-fold elevated during the post-injury phase. In situ hybridization studies demonstrated that collagen gene expression was colocalized to activated pancreatic stellate cells. Collagen expression and fibrosis persisted in focal areas during recovery. These findings show that pancreatic stellate cells are the major source of collagen during repetitive injury in vivo. Additionally, focal areas of sustained pancreatic fibrogenesis persist after cessation of cerulein treatment, and these areas may contribute to sustained total organ collagen expression in the absence of ongoing injury.

Scinderin, a Ca2+-dependent actin filament severing protein that controls cortical actin network dynamics during secretion.

Secretory vesicles are localized in specific compartments within neurosecretory cells. These are different pools in which vesicles are in various states of releasability. The transit of vesicles between compartments is controlled and regulated by Ca2+, scinderin and the cortical F-actin network. Cortical F-actin disassembly is produced by the filament severing activity of scinderin. This Ca2+-dependent activity of scinderin together with its Ca2+-independent actin nucleating activity, control cortical F-actin dynamics during the secretory cycle. A good understanding of the interaction of actin with scinderin and of the role of this protein in secretion has been provided by the analysis of the molecular structure of scinderin together with the use of recombinant proteins corresponding to its different domains.

Novobiocin and related coumarins and depletion of heat shock protein 90-dependent signaling proteins.

BACKGROUND: Heat shock protein 90 (Hsp90) interacts with and stabilizes several oncogenic protein kinases (e.g., p185(erbB2), p60(v-src), and Raf-1) and is required for the stability and dominant-negative function of mutated p53 protein. Two unrelated antibiotics, geldanamycin and radicicol, bind specifically to an atypical nucleotide-binding pocket of Hsp90, a site that shares homology with the adenosine triphosphate (ATP)-binding domain of bacterial DNA gyrase B. This interaction leads to destabilization of proteins that interact with Hsp90. Since the nucleotide-binding site of gyrase B is targeted by coumarin antibiotics (e.g., novobiocin), we investigated whether these drugs can also interact with Hsp90 and affect its activity. METHODS: We used immobilized novobiocin, geldanamycin, or radicicol to isolate either endogenous Hsp90 from cell lysates or Hsp90 deletion fragments translated in vitro. Effects of the coumarin antibiotics novobiocin, chlorobiocin, and coumermycin A1 on several proteins interacting with Hsp90 were assessed in vitro and in vivo. RESULTS: Hsp90 binding to immobilized novobiocin was competed by soluble coumarins and ATP but not by geldanamycin or radicicol. A carboxy-terminal Hsp90 fragment bound immobilized novobiocin but not immobilized geldanamycin, while a geldanamycin-binding amino-terminal fragment did not bind novobiocin. All three coumarins markedly reduced cellular levels of p185(erbB2), p60(v-src), Raf-1, and mutated p53. Furthermore, novobiocin reduced Raf-1 levels in the spleens of mice treated with the drug. CONCLUSIONS: These coumarin antibiotics, particularly novobiocin, represent a first-generation alternative to other Hsp90-targeting drugs that are not as well tolerated. Novobiocin's unique interaction with Hsp90 identifies an additional site on this protein amenable to pharmacologic interference with small molecules.

Localization by segmental deletion analysis and functional characterization of a third actin-binding site in domain 5 of scinderin.

Scinderin is a Ca2+-dependent actin filament severing protein present in a variety of secretory cells. Previous work suggests that scinderin-evoked cortical F-actin disassembly is required for secretion because local disassembly of cortical cytoskeleton allows secretory vesicle exocytosis (Vitale, M. L., Rodríguez Del Castillo, A., Tchakarov, L., and Trifaró, J.-M. (1991) J. Cell Biol. 113, 1057-1067). Scinderin has six domains each containing three internal sequence motifs, two actin, and two phosphatidylinositol disphosphate-binding sites in domains 1 and 2. In this paper we report the presence of another actin-binding site at the NH2-terminal of domain 5 (Sc511-518). This site binds actin in a Ca2+-independent manner and a recombinant fragment (Sc5-6 or Sc502-715) containing this site binds to actin-DNase-I-Sepharose 4B beads, co-sediments with actin and is able to nucleate actin assembly. Recombinant ScL5-6, a fusion protein devoid of the actin-binding site (Sc519-715), did not exhibit these properties. Moreover, Sc-ABP3, a peptide constructed with sequence (RLFQVRRNLASIT) identical to Sc511-523 blocked the binding of Sc5-6 to actin. Sc5-6 and Sc-ABP3 also prevented the actin severing activity of recombinant full-length scinderin (r-Sc) and inhibited the potentiation by r-Sc of Ca2+-evoked release of serotonin from permeabilized platelets. On the other hand, ScL5-6 failed to block the effect of r-Sc on platelet serotonin release. Sc1-4,6, a construct devoid of domain 5, was able to sever but unable to nucleate actin, indicating that an actin nucleation site of scinderin was in domain 5. The results suggest that scinderin, in addition to binding actin on sites present in domains 1 and 2, must bind actin on a third site in domain 5 to sever and nucleate actin effectively.

Aphid species--vectors of plum pox virus.

Plum pox virus (PPV) is widely spread by natural vectors present in plum orchards. The efficiency of transmission is dependent on the frequency of the occurrence of vectors and the cultivar susceptibility to this pathogen. Having in view that PPV has a wide range of annual and multiannual host plants and vectors, there is great concern for obtaining PPV-resistant cultivars. This report deals with the following vectors: Hyalopterus pruni, Brachycaudus cardui, Brachycaudus helichrysi. Myzus persicae and Phorodon humuli aphids, and Aculus fockeni mite. Seven different cultivars of Prunus domestica were utilized. To assess the virus transmission rate, 50-100 individual vectors per tree were used. The treatment of the trees was performed every four weeks and then the disease symptoms were observed. PPV was transmitted by all vectors studied, the rate ranging in dependence on the susceptibility of cultivars used. Thus, in cvs. Centenar, Pescarus, d'Agen, Stanley and Tuleu Gras, the transmission rate ranged from 20% to 60%, while in susceptible cvs. Vanat romanesc and Vanat de Italia--from 40% to 80%.

Recombinant scinderin enhances exocytosis, an effect blocked by two scinderin-derived actin-binding peptides and PIP2.

The cortical F-actin cytoskeleton represents a negative control for secretion, and it must be locally disassembled to allow chromaffin vesicle exocytosis. Recombinant scinderin (a Ca(2+)-dependent F-actin-severing protein) potentiated Ca(2+)-evoked F-actin disassembly and exocytosis in permeabilized chromaffin cells, an effect blocked by peptides Sc-ABP1 and Sc-ABP2 (with sequences corresponding to two actin-binding sites of scinderin), exogenous gamma-actin, or phosphatidylinositol 4,5-bisphosphate (PIP2). PIP2 effect was blocked by peptide Sc-PIP2BP (with sequence corresponding to a PIP2-binding site of scinderin). Truncated scinderin254-715 (lacking actin-severing domains) did not potentiate exocytosis. Sc-ABP1, Sc-ABP2, and gamma-actin also inhibited exocytosis in the absence of recombinant scinderin, suggesting an inhibition of endogenous scinderin. Results suggest that scinderin-evoked cortical F-actin disassembly is required for secretion and that scinderin is an important component of the exocytotic machinery.

Recombinant scinderin, an F-actin severing protein, increases calcium-induced release of serotonin from permeabilized platelets, an effect blocked by two scinderin-derived actin-binding peptides and phosphatidylinositol 4,5-bisphosphate.

In response to vessel injury or exposure to different substances, platelets undergo activation which consists of shape changes, formation of cellular pseudopodia, aggregation, and secretion. These dramatic changes are accompanied by cycles of actin depolymerization and polymerization. Previous work has shown the presence in platelets of gelsolin and scinderin, two Ca(2+)-dependent F-actin severing proteins. Recent published evidence suggests that scinderin is a component of the exocytotic machinery in chromaffin cells. The present work describes the preparation of recombinant scinderin and peptides Sc-ABP1 and Sc-ABP2 with sequences corresponding to two actin-binding sites of scinderin. Recombinant scinderin and peptides Sc-ABP1 and Sc-ABP2 were tested for their effects on Ca(2+)-induced serotonin release from digitonin permeabilized platelets. The results indicated that recombinant scinderin potentiates Ca(2+)-evoked serotonin release, an effect blocked in the presence of Sc-ABP1, Sc-ABP2, exogenous gamma-actin, or the addition of phosphatidylinositol 4,5-bisphosphate (PIP2). In the presence of a mismatched peptide (MMP) the potentiating effect of recombinant scinderin was not affected. Moreover, Sc-ABP1, Sc-ABP2, and gamma-actin inhibited Ca(2+)-induced release of serotonin in the absence of recombinant scinderin, suggesting an inhibition of platelet endogenous scinderin. MMP was ineffective under these conditions. The results suggest that F-actin disassembly, perhaps at a specific site, is required for platelet secretion and that scinderin might be an important component of the exocytotic machinery in platelets.

Molecular cloning and functional expression of chromaffin cell scinderin indicates that it belongs to the family of Ca(2+)-dependent F-actin severing proteins.

Scinderin is a Ca(2+)-dependent actin filament severing protein present in chromaffin cells, platelets and a variety of secretory cells. It has been suggested that scinderin is involved in chromaffin cell F-actin dynamics and that this actin network controls the delivery of secretory vesicles to plasma membrane exocytotic sites. Moreover, scinderin redistribution and activity may be regulated by pH and Ca2+ in resting and stimulated cells. Here we describe the molecular cloning, the nucleotide sequence and the expression of bovine chromaffin cell scinderin cDNA. The fusion protein obtained cross-reacts with native scinderin antibodies and binds phosphatidylserine (PS), phosphatidylinositol 4,5-bisphosphate (PIP2) and actin in a Ca(+)-dependent manner. Antibodies raised against the fusion protein produced the same cellular staining patterns for scinderin as anti-native scinderin. Nucleotide and amino acid sequence analysis indicate that scinderin has six domains each containing three internal sequence motifs, two actin and two PIP2 binding sites and has 63 and 53% homology with gelsolin and villin. These data indicate that scinderin is a novel member of the family of Ca(2+)-dependent F-actin severing proteins which includes gelsolin and villin.