Search

By means of its antiangiogenic activity, thrombospondin-1 (TSP-1) exerts indirect antitumoral action on solid tumors. Here, we investigated potential antitumor action in an in vitro cell model for promyelocytic leukemia (NB4-LR1), resistant to retinoid maturation. Purified soluble TSP-1 added to cultures induced a strong dose-dependent growth inhibition and a slowly developing maturation-independent cell death. Recombinant fragments of TSP-1 allowed mapping of these activities to its type 3 repeat/C-terminal domain, features that are distinct from those of TSP-1 action on solid tumors, previously ascribed to the type 1 repeat domain. Cell death in leukemia was characterized as a caspase-independent mechanism, without DNA fragmentation, but phosphatidylserine externalization followed by membrane permeabilization. Mitochondria membrane depolarization was inherent to TSP-1 action but did not produce release of death-promoting proteins (eg, noncaspase apoptosis regulators, apoptosis-induced factor [AIF], endonuclease G, or Omi/HtrA2 or the caspase regulators, cytochrome c or second mitochondrial activator of caspase/direct inhibitor of apoptosis protein-binding protein with low isoelectric point [Smac/DIABLO]). Although detected, reactive oxygen species (ROS) production was likely not involved in the death process. Finally, receptor agonist RFYVVM and RGD peptides indicated that TSP-1 death effects are mediated by membrane receptors CD47 and αvβ3. These results demonstrated a new domain-specific antitumoral activity of TSP-1 on a leukemia cell line, which extends TSP-1 therapeutic potential outside the area of vascularized solid tumors. (Blood. 2005;106:658-667)

Protein acetyltransferases and deacetylases have been implicated in oncogenesis, apoptosis and cell cycle regulation. Most of the protein acetyltransferases described acetylate ε-amino groups of lysine residues within proteins. Mouse ARD1 (homologue of yeast Ard1p, where Ard1p stands for arrest defective 1 protein) is the only known protein acetyltransferase catalysing acetylation of proteins at both α- (N-terminus) and ε-amino groups. Yeast Ard1p interacts with Nat1p (N-acetyltransferase 1 protein) to form a functional NAT (N-acetyltransferase). We now describe the human homologue of Nat1p, NATH (NAT human), as the partner of the hARD1 (human ARD1) protein. Included in the characterization of the NATH and hARD1 proteins is the following: (i) endogenous NATH and hARD1 proteins are expressed in human epithelial, glioma and promyelocytic cell lines; (ii) NATH and hARD1 form a stable complex, as investigated by reciprocal immunoprecipitations followed by MS analysis; (iii) NATH–hARD1 complex expresses N-terminal acetylation activity; (iv) NATH and hARD1 interact with ribosomal subunits, indicating a co-translational acetyltransferase function; (v) NATH is localized in the cytoplasm, whereas hARD1 localizes both to the cytoplasm and nucleus; (vi) hARD1 partially co-localizes in nuclear spots with the transcription factor HIF-1α (hypoxia-inducible factor 1α), a known ε-amino substrate of ARD1; (vii) NATH and hARD1 are cleaved during apoptosis, resulting in a decreased NAT activity. This study identifies the human homologues of the yeast Ard1p and Nat1p proteins and presents new aspects of the NATH and hARD1 proteins relative to their yeast homologues.

Abstract: We define a probabilistic model for the analysis of a Non-Repudiation protocol that guarantees fairness, without resorting to a trusted third party, by means of a probabilistic algorithm. By using the PRISM model checker, we estimate the probability for a malicious user to break the non-repudiation property, depending on various parameters of the protocol. [Copyright &y& Elsevier]

Our aim is to study the information flow problem in hybrid systems, namely systems consisting of a discrete program with an analog environment. Information flows compromise the security of a system because they cause leakage of protected information. In order to tackle information flow in real-life systems, we introduce new classes of hybrid systems that extend the known ones while preserving their properties. Then, we define a logic to specify information flow. By means of some examples, we show that, by this logic, we are able to express information flows in hybrid systems and to certify that some suspect behaviors of these systems do not give rise to any information flow. We give a model checking procedure for our logic, and we prove that it gives a correct answer whenever it terminates. Moreover, for a particular class of hybrid systems, we give a version of the procedure that always terminates.

ABSTRACTIn the south of France, leishmaniasis due to Leishmania infantumoccurs in the following five foci of endemicity (from west to east): Pyre´ne´es-Orientales, Ce´vennes, Provence, Co^te d'Azur, and Corsica. Between 1981 and 2002, 712 Leishmaniastrains obtained from humans, dogs, cats, and sand flies were studied by isoenzyme analysis. In total, seven zymodemes were identified: MON-1, MON-11, MON-24, MON-29, MON-33, MON-34, and MON-108. The Pyre´ne´es-Orientales focus is characterized by a predominance of human cutaneous leishmaniasis and a high enzymatic polymorphism (five zymodemes). In the other foci, where human visceral leishmaniasis is predominant, only two zymodemes are present. L. infantumMON-1 is the parasite most frequently found, in patients both with and without concomitant human immunodeficiency virus infection. MON-1 is the only zymodeme present in dogs, which act as the reservoir host in all of the foci. In Ce´vennes, where the complete life cycle of zymodeme MON-1 has been identified, Phlebotomus perniciosusand Phlebotomus ariasiare vectors. The enzymatic polymorphism is compared to that of neighboring countries (Spain and Italy). In Pyre´ne´es-Orientales, small variant zymodemes with electromorphs of heterozygote-like and homozygotic patterns can be explained by different genetic hypotheses.

A hybrid system consists of a finite number of locations, variables and transitions. Different classes are considered in the literature. In this paper we study the different expressive power of these classes. [Copyright &y& Elsevier]

To determine the extent to which the vagina, endocervix, and amniotic fluid screen the Escherichia coli strains responsible for neonatal infections, we studied the genetic relationships among 105 E. coli strains isolated from all of the ecosystems involved in this infectious process. Twenty-four strains were isolated from the intestinal flora, and 25 strains were isolated from the vaginas of pregnant women. Twenty-seven strains were isolated from the amniotic fluid, blood, and cerebrospinal fluid (CSF) of infected neonates. The intraspecies genetic characteristics of all of the isolates were determined by random amplified polymorphic DNA (RAPD) analysis, PCR ECOR (E. coli reference) grouping, and PCR virulence genotyping. A correlation was found between the intraspecies distributions of the strains in the A, B1, B2, and D ECOR groups and in the two major RAPD groups (I and II). Nevertheless, the distribution of the E. coli strains in the RAPD groups according to their anatomical origins was more significant than their distribution in the ECOR groups. This may be explained by the existence of an E. coli subpopulation, defined by the RAPD I group, within the ECOR B2 group. This RAPD I group presents a major risk for neonates: 75% of the strains isolated from patients with meningitis and 100% of the strains isolated from patients with bacteremia were in this group. The vagina and the amniotic fluid are two barriers that favor colonization by highly infectious strains. Indeed, only 17% of fecal strains belonged to the RAPD I group, whereas 52% of vaginal strains and 67% of amniotic fluid strains belonged to this subpopulation. The ibeA and iucC genes were significantly associated with CSF strains, whereas the hly and sfa/foc genes were more frequent in blood strains. These findings could serve as a basis for developing tools to recognize vaginal strains, which present a high risk for neonates, for use in prophylaxis programs.

ABSTRACTTo determine the extent to which the vagina, endocervix, and amniotic fluid screen the Escherichia colistrains responsible for neonatal infections, we studied the genetic relationships among 105 E. colistrains isolated from all of the ecosystems involved in this infectious process. Twenty-four strains were isolated from the intestinal flora, and 25 strains were isolated from the vaginas of pregnant women. Twenty-seven strains were isolated from the amniotic fluid, blood, and cerebrospinal fluid (CSF) of infected neonates. The intraspecies genetic characteristics of all of the isolates were determined by random amplified polymorphic DNA (RAPD) analysis, PCR ECOR (E. coli reference) grouping, and PCR virulence genotyping. A correlation was found between the intraspecies distributions of the strains in the A, B1, B2, and D ECOR groups and in the two major RAPD groups (I and II). Nevertheless, the distribution of the E. colistrains in the RAPD groups according to their anatomical origins was more significant than their distribution in the ECOR groups. This may be explained by the existence of an E. colisubpopulation, defined by the RAPD I group, within the ECOR B2 group. This RAPD I group presents a major risk for neonates: 75% of the strains isolated from patients with meningitis and 100% of the strains isolated from patients with bacteremia were in this group. The vagina and the amniotic fluid are two barriers that favor colonization by highly infectious strains. Indeed, only 17% of fecal strains belonged to the RAPD I group, whereas 52% of vaginal strains and 67% of amniotic fluid strains belonged to this subpopulation. The ibeAand iucCgenes were significantly associated with CSF strains, whereas the hlyand sfa/focgenes were more frequent in blood strains. These findings could serve as a basis for developing tools to recognize vaginal strains, which present a high risk for neonates, for use in prophylaxis programs.

Differentiation therapy for acute myeloid leukemia uses transcriptional modulators to reprogram cancer cells. The most relevant clinical example is acute promyelocytic leukemia (APL), which responds dramatically to either retinoic acid (RA) or arsenic trioxide (As2O3). In many myeloid leukemia cell lines, cyclic adenosine monophosphate (cAMP) triggers growth arrest, cell death, or differentiation, often in synergy with RA. Nevertheless, the toxicity of cAMP derivatives and lack of suitable models has hampered trials designed to assess the in vivo relevance of theses observations. We show that, in an APL cell line, cAMP analogs blocked cell growth and unraveled As2O3-triggered differentiation. Similarly, in RA-sensitive or RA-resistant mouse models of APL, continuous infusions of 8-chloro-cyclic adenosine monophosphate (8-Cl-cAMP) triggered major growth arrest, greatly enhanced both spontaneous and RA- or As2O3-induced differentiation and accelerated the restoration of normal hematopoiesis. Theophylline, a well-tolerated phosphodiesterase inhibitor which stabilizes endogenous cAMP, also impaired APL growth and enhanced spontaneous or As2O3-triggered cell differentiation in vivo. Accordingly, in an APL patient resistant to combined RA–As2O3 therapy, theophylline induced blast clearance and restored normal hematopoiesis. Taken together, these results demonstrate that in vivo activation of cAMP signaling contributes to APL clearance, independently of its RA-sensitivity, thus raising hopes that other myeloid leukemias may benefit from this therapeutic approach.

The exact origin of strains of Escherichia coliresponsible for infectious diseases in intensive care units (ICUs) remains partly unknown. Our aim was to determine the nature of the link between strains from the intestinal flora of hospital staff, strains from the intestinal flora of patients hospitalized in ICUs and strains isolated from ICU patients with invasive diseases. For this purpose, 77 strains of E. coliwere genetically characterized by exploring their entire genomes by random amplified polymorphism of DNA (RAPD), and by determining their phylogenetic position in ECOR (E. colireference) groups, the virulence factors harboured (pap, sfa, afa, hly, aerand cnf) and their ability to mutate. The strains isolated from the intestinal flora of hospital staff were found to constitute a genetically heterogeneous population compared with the strains isolated from ICU carriers, which were highly clustered. The latter strains harboured numerous virulence factors, and 80% belonged to the group ECOR B2. The strains isolated from infected patients harboured fewer virulence factors than those from the ICU carriers, and only half belonged to ECOR B2. Moreover, these strains were more genetically related to strains from hospital staff than to strains from ICU carriers. Thus, the exogenous origin of the E. colistrains is probably almost as important as translocation from intestinal flora in ICUs. Moreover, a strong mutator phenotype had a minor, or no, role in the rapid adaptation to modifications in the ecological environment.

Expectations about future events are known to trigger neural mechanisms that affect both perception and action. Here we report that different and opposite expectations of bad and good motor performance modulate the therapeutic effects of subthalamic nucleus stimulation in Parkinsonian patients who had undergone chronic implantation of electrodes for deep brain stimulation. By analyzing the effects of subthalamic stimulation on the velocity of movement of the right hand, we found hand movement to be faster when the patients expected a good motor performance. The expectation of good performance was induced through a placebo-like procedure, thus indicating that placebo-induced expectations have influence on the treatment outcome. All these effects occurred within minutes, suggesting that expectations induce neural changes very quickly.

We model a timing attack on web privacy proposed by Felten and Schneider by using three different approaches: HL-Timed Automata, SMV model checker, and tSPA Process Algebra. Some comparative analysis on the three approaches is derived. [Copyright &y& Elsevier]

We study the problem of privacy in the framework of Timed Automata. By distinguishing between secret and observable actions we formulate a property of no-privacy in terms of a property of the language accepted by a Timed Automaton, and we give an algorithm checking such property. [Copyright &y& Elsevier]

Co/Fe multilayers were electron beam evaporated in ultra-high vacuum and analyzed by Alternating Gradient Force Magnetometry, Magnetic Force Microscopy, Conversion Electron Mössbauer Spectroscopy, and Transmission Electron Microscopy. The multilayer of 10 nm Co and 30 nm Fe layer thickness showed a single-phase magnetic behavior because of a strong exchange coupling established between the layers. The system exhibits stripe domains which were correlated to the presence of a perpendicular magnetic anisotropy. The study performed on multilayers where Co was intercalated by very thin 57Fe layers showed that the interfaces were very clean and sharp.

Acute promyelocytic leukemia (APL) is characterized by the specific chromosome translocation t(15;17) with promyelocytic leukemia-retinoic acid receptor-a (PML-RARA) fusion gene and the ability to undergo terminal differentiation as an effect of all-trans retinoic acid (ATRA). Recently, arsenic trioxide (As2O3) has been identified as an alternative therapy in patients with both ATRA-sensitive and ATRA-resistant APL. At the cellular level, As2O3 triggers apoptosis and a partial differentiation of APL cells in a dose-dependent manner; both effects are observed in vivo among patients with APL and APL animal models. To further explore the mechanism of As2O3-induced differentiation, the combined effects of arsenic and a number of other differentiation inducers on APL cell lines (NB4 and NB4-R1) and some fresh APL cells were examined. The data show that a strong synergy exists between a low concentration of As2O3 (0.25 µM) and the cyclic adenosine monophosphate (cAMP) analogue, 8-CPT-cAMP, in fully inducing differentiation of NB4, NB4-R1, and fresh APL cells. Furthermore, cAMP facilitated the degradation of As2O3-mediated fusion protein PML-RARa, a process considered to play a key role in overcoming the differentiation arrest of APL cells. On the other hand, cAMP could significantly inhibit cell growth by modulating several major players in G1/S transition regulation. Interestingly, H89, an antagonist of protein kinase A, could block the differentiation-inducing effect of As2O3potentiated by cAMP. These results thus support the existence of a novel signaling cross-talk for APL maturation, which may deepen understanding of As2O3-induced differentiation in vivo, and thus furnish insights for new therapeutic strategies.

Objectives: Bilateral chronic high frequency stimulation of the subthalamic nucleus (STN), through the stereotactical placement of stimulating electrodes, effectively improves the motor symptoms of severe Parkinson's disease. Intraoperative neurophysiological and clinical monitoring techniques (neuronal electrical activity recording and intraoperative stimulation) may improve and refine the localisation of the nucleus. The objective of this work was to compare the preoperative CT and MRI localisation with the intraoperative neurophysiological identification of STN. The relation between the localisation of the STN and the position of the most effective contact of the permanent quadripolar electrode at a 3 month and 1 year follow up was also studied. Methods: Fourteen consecutive parkinsonian patients were submitted to bilateral implant for STN stimulation. All the patients underwent a standard MRI and stereotactic CT to obtain, by image fusion and localisation software, the stereotactical coordinates of STN. The STN extension and boundaries were identified by a semimicrorecording of the neuronal electrical activity. The definitive quadripolar electrode was positioned to locate at least two contacts within the STN recording area. Intraoperative macrostimulation was performed to confirm the correct position of the electrode. Postoperative clinical evaluation of the effects of stimulation was checked for each contact of the quadripolar electrode testing the improvement on contralateral rigidity to select the best contact. This evaluation was repeated at 3 months and 1 year after surgery. Results: In 35.7% of the procedures it was necessary to perform more than one track to get a recording of neuronal activity consistent with STN. The mean position of the central point of all the 28 STN recording areas in respect of the AC-PC line midpoint was 2.7 mm posterior (SD 0.7), 3.8 mm inferior (SD 1.1), and 11.6 mm lateral (SD 0.9), and the mean distance between the anatomical target and the central point of the STN as defined by intraoperative recording was 0.5 mm (SD 0.5) on the anteroposterior plane, 0.7 mm (SD 0.7) on the lateral plane, and 0.9 mm (SD 0.6) on the vertical plane. At 1 year the mean position of the central point of the most effective contact of the electrode in respect of the AC-PC line midpoint was 1.7 mm posterior (SD 0.9), 1.7 mm inferior (SD 1.5), and 12.3 mm lateral (SD 0.9). Conclusion: The results highlight the role of the intraoperative recording to get a more accurate localisation of the STN in surgery for Parkinson's disease, allowing the identification of the boundaries and of the extension of the nucleus. The most effective contact of the quadripolar electrode was always in the upper part of the STN recording area or immediately above it, suggesting a role of this region in the clinical effectiveness of the STN electrical stimulation.

The magnetic, magnetoelastic, and structural properties of the Fe62.5Co6Ni7.5Zr6Cu1Nb2B15 at.- alloy, recently produced by rapid quenching from the melt, have been observed and explained with regards to the proper choice of components and their percentages. In particular, it was shown that a giant magnetoelastic wave amplitude, obtained with a very low magnetising field, is a consequence of simultaneous very high values of permeability, magnetomechanical coupling factor, and Youngs modulus. The importance of the new amorphous alloy, both for applications and basic research, is founded on its structural development by means of isothermal heat treatments. In particular, the tendency to maintain structural disorder at temperatures above where the first crystallisation occurrs is fundamental. X-ray analysis demonstrates that this property determines the presence of nanocrystalline phases after heating at a temperature well above that of the first crystallisation.

To better understand the role of retinoids in myelopoiesis, expression of the retinoid receptor genes (retinoic acid receptors [RARs] and retinoid X receptors [RXRs]) were examined during differentiation of factor-dependent cell-Paterson (FDCP)–mixA4 murine progenitor cells. The major receptor expressed in undifferentiated A4 cells was RARα (primarily the RARα1 isoform). Following induction of myelomonocytic differentiation with granulocyte and granulocyte-macrophage colony-stimulating factors, a dramatic increase in RARα expression (particularly the RARα2 isoform) was seen. In contrast, expression of both RARα isoforms was rapidly extinguished upon induction of erythroid differentiation with erythropoeitin (EPO). A modest induction of RXRα expression was seen, particularly during differentiation in the myelomonocytic lineage. Low expression levels of RARγ2 and RXRβ remained unchanged, irrespective of differentiation pathway. Consistent with the gene expression patterns, RARα agonists and antagonists stimulated myelomonocytic and erythroid differentiation of FDCP-mixA4 cells, respectively. Taken together, these results suggest that erythropoiesis and granulopoiesis require diminished and enhanced RARα activities, respectively, which at physiological all-trans-retinoic acid (RA) concentrations may be accomplished by reciprocal effects of EPO and myelomonocytic growth factors on its expression. This hypothesis is corroborated by data showing that RA, which positively regulates RARα2 expression, can exert inhibitory effects on erythroid differentiation.

To better understand the role of retinoids in myelopoiesis, expression of the retinoid receptor genes (retinoic acid receptors [RARs] and retinoid X receptors [RXRs]) were examined during differentiation of factor-dependent cell-Paterson (FDCP)–mixA4 murine progenitor cells. The major receptor expressed in undifferentiated A4 cells was RARα (primarily the RARα1 isoform). Following induction of myelomonocytic differentiation with granulocyte and granulocyte-macrophage colony-stimulating factors, a dramatic increase in RARα expression (particularly the RARα2 isoform) was seen. In contrast, expression of both RARα isoforms was rapidly extinguished upon induction of erythroid differentiation with erythropoeitin (EPO). A modest induction of RXRα expression was seen, particularly during differentiation in the myelomonocytic lineage. Low expression levels of RARγ2 and RXRβ remained unchanged, irrespective of differentiation pathway. Consistent with the gene expression patterns, RARα agonists and antagonists stimulated myelomonocytic and erythroid differentiation of FDCP-mixA4 cells, respectively. Taken together, these results suggest that erythropoiesis and granulopoiesis require diminished and enhanced RARα activities, respectively, which at physiological all-trans-retinoic acid (RA) concentrations may be accomplished by reciprocal effects of EPO and myelomonocytic growth factors on its expression. This hypothesis is corroborated by data showing that RA, which positively regulates RARα2 expression, can exert inhibitory effects on erythroid differentiation.

OBJECTIVE: To investigate the relation between the variation of the parameters of stimulation and the clinical effectiveness in parkinsonian patients treated with deep brain stimulation of the subthalamic nucleus (STN), to provide information on the electrical parameter setting and the mechanism of action of deep brain stimulation. METHODS: Ten patients with Parkinson's disease bilaterally implanted in the STN were studied. For every patient the intensity of the stimulus necessary to obtain the disappearance of contralateral wrist rigidity (required clinical effect, RCE) and the side effect threshold in 20 different conditions of stimulation, coupling four pulse width values (60, 120, 210, 450 μs) with five rate values (10, 50, 90, 130, 170 Hz) were determined. All the patients were tested after a 12 hour withdrawal of antiparkinsonian drugs, and the clinical evaluation was double blind. RESULTS: In all the patients it was impossible to obtain the RCE using 10 and 50 Hz stimulus rates. For all the other stimulus rate values, the intensity-pulse width curves (IPWCs) for the RCE and for the side effect threshold showed a hyperbolic trend. For every pulse width value, increasing the rate from 90 to 130 and to 170 Hz progressively decreased the intensity of the stimulus necessary to reach the RCE, but the differences were not significant. Within the same rate value, the progressive reduction of the stimulus intensity necessary to obtain the RCE, obtained with the lengthening of the pulse width was significant (p<0.05) only comparing 60 with 210 μs and 60 with 450 μs. CONCLUSIONS: The findings give some useful indications for the electrical parameter setting in deep brain stimulation of the STN, and some information about the mechanism of action of deep brain stimulation.

Fifty-two Leishmania strains, obtained from human patients and dogs in a visceral leishmaniasis focus in Sudan, were characterized by isoenzyme electrophoresis (15 enzymes). The phylogenetic analysis showed that the 7 Leishmania zymodemes obtained hold ancestral positions on the phylogenetic tree, supporting the hypothesis of an East African origin of visceral leishmaniasis.

Abstract:: A composite material of a polyethylene matrix filled by a fine silver powder was prepared with different Ag contents and physical behaviours ranging from insulator to conductor. Ac differential magnetic susceptibility χ measurements show the samples are paramagnetic up to an Ag concentration of ∼65%. At low temperatures the composite is spin-glass type, whereas the transition from insulator to conductor corresponds to an abatement of χ at zero magnetizing field. Magneto-conductivity effects have been observed in resistivity measurements at low temperatures. They can be explained in terms of an effective exchange electronic scattering mechanism between the conduction electrons and the diluted magnetic moments arising from unpaired electron spins of boundary silver particles. Moreover, the presence of a broad minimum in the resistivity curve at T = ∼ 20 K, observed in samples with an Ag concentration just above the percolation threshold, addresses to possible interference effects similar to those reported in disordered materials.

Not Available

Two cases of skin lesions similar to those caused by Leishmania parasites have been reported from Martinique. Parasites isolated from these lesions were unlike Leishmania reference strains by isoenzyme analysis and electron microscopy and were assumed to be monoxenous trypanosomatids which normally only infect invertebrates. Both strains have now been retyped by isoenzyme analysis and found to be identical to each other and distantly related to all other Leishmania species. The sequence of the 18S ribosomal RNA gene and partial sequences of the DNA polymerase alpha and RNA polymerase II largest subunit genes were obtained. These sequences indicated that the Martinique parasites clustered with L. enriettii and were basal to all other euleishmania. However, support for both the position basal to all euleishmania and the clustering with L. enriettii was low. The Martinique parasites may cluster with L. (Leishmania) or L. (Viannia) or form a novel clade within the euleishmania either with or without L. enriettii.

Cytochrome P450 (P450)-dependent metabolism of all-trans-retinoic acid (atRA) is important for the expression of its biological activity. Because the human P450s involved in the formation of the principal atRA metabolites have been only partially identified, the purpose of this study was to identify the human P450s involved in atRA metabolism. The use of phenotyped human liver microsomes (n= 16) allowed the identification of the following P450s: 2B6, 2C8, 3A4/5, and 2A6 were involved in the formation of 4-OH-RA and 4-oxo-RA; 2B6, 2C8, and 2A6 correlated with the formation of 18-OH-RA; and 2A6, 2B6, and 3A4/5 activities correlated with 5,6-epoxy-RA formation (30-min incubation, 10 μM atRA, HPLC separation, UV detection 340 nm). The use of 15 cDNA-expressed human P450s from lymphoblast microsomes, showed the formation of 4-OH-RA by CYP3A7 > CYP3A5 > CYP2C18 > CYP2C8 > CYP3A4 > CYP2C9, whereas the 18-OH-RA formation involved CYPs 4A11 > 3A7 > 1A1 > 2C9 > 2C8 > 3A5 > 3A4 >2C18. Kinetic studies identified 3A7 as the most active P450 in the formation of three of the metabolites: for 4-OH-retinoic acid, 3A7 showed a Vmax/Kmof 127.7, followed by 3A5 (Vmax/Km= 25.6), 2C8 (Vmax/Km= 24.5), 2C18 (Vmax/Km= 15.8), 3A4 (Vmax/Km= 5.7), 1A1 (Vmax/Km= 5.0), and 4A11 (Vmax/Km= 1.9); for 4-oxo-RA, 3A7 showed a Vmax/Kmof 13.4, followed by a 10-fold lower activity for both 2C18 and 4A11 (Vmax/Km= 1.2); and for 18-OH-RA, 3A7 showed a Vmax/Kmof 10.5 compared with a Vmax/Kmof 2.1 for 4A11 and 2.0 for 2C8. 5,6-Epoxy-RA was only detected at high substrate concentrations in this system (>10 μM), and P450s 2C8, 2C9, and 1A1 were the most active in its formation. The use of embryonic kidney cells (293) stably transfected with human P450 cDNA confirmed the major involvement of P450s 3A7, 1A1, and 2C8 in the oxidation of atRA, and to a lesser extent, 1A2, 2C9, and 3A4. In conclusion, several human P450s involved in atRA metabolism have been identified, the expression of which was shown to direct atRA metabolism toward the formation of specific metabolites. The role of these human P450s in the biological and anticancer effects of atRA remains to be elucidated.

Cytochrome P450 (P450)-dependent metabolism of all-trans-retinoic acid (atRA) is important for the expression of its biological activity. Because the human P450s involved in the formation of the principal atRA metabolites have been only partially identified, the purpose of this study was to identify the human P450s involved in atRA metabolism. The use of phenotyped human liver microsomes (n = 16) allowed the identification of the following P450s: 2B6, 2C8, 3A4/5, and 2A6 were involved in the formation of 4-OH-RA and 4-oxo-RA; 2B6, 2C8, and 2A6 correlated with the formation of 18-OH-RA; and 2A6, 2B6, and 3A4/5 activities correlated with 5, 6-epoxy-RA formation (30-min incubation, 10 microM atRA, HPLC separation, UV detection 340 nm). The use of 15 cDNA-expressed human P450s from lymphoblast microsomes, showed the formation of 4-OH-RA by CYP3A7 > CYP3A5 > CYP2C18 > CYP2C8 > CYP3A4 > CYP2C9, whereas the 18-OH-RA formation involved CYPs 4A11 > 3A7 > 1A1 > 2C9 > 2C8 > 3A5 > 3A4 >2C18. Kinetic studies identified 3A7 as the most active P450 in the formation of three of the metabolites: for 4-OH-retinoic acid, 3A7 showed a V(max)/K(m) of 127.7, followed by 3A5 (V(max)/K(m) = 25.6), 2C8 (V(max)/K(m) = 24.5), 2C18 (V(max)/K(m) = 15.8), 3A4 (V(max)/K(m) = 5.7), 1A1 (V(max)/K(m) = 5.0), and 4A11 (V(max)/K(m) = 1.9); for 4-oxo-RA, 3A7 showed a V(max)/K(m) of 13.4, followed by a 10-fold lower activity for both 2C18 and 4A11 (V(max)/K(m) = 1.2); and for 18-OH-RA, 3A7 showed a V(max)/K(m) of 10.5 compared with a V(max)/K(m) of 2.1 for 4A11 and 2.0 for 2C8. 5,6-Epoxy-RA was only detected at high substrate concentrations in this system (>10 microM), and P450s 2C8, 2C9, and 1A1 were the most active in its formation. The use of embryonic kidney cells (293) stably transfected with human P450 cDNA confirmed the major involvement of P450s 3A7, 1A1, and 2C8 in the oxidation of atRA, and to a lesser extent, 1A2, 2C9, and 3A4. In conclusion, several human P450s involved in atRA metabolism have been identified, the expression of which was shown to direct atRA metabolism toward the formation of specific metabolites. The role of these human P450s in the biological and anticancer effects of atRA remains to be elucidated.

The Jem-1(JEM-1,HGMW-approved symbol BLZF1) gene mapping to human chromosome 1q24 codes for a ubiquitously expressed 3-kb mRNA, translated in a 45-kDa nuclear protein. Recent studies have shown a deficient expression of this gene in acute promyelocytic leukemia (APL). However, treatment with retinoids was able to upregulate JEM-1mRNA in maturing NB4 leukemia cells. Here, we report the characterization of the structural organization of JEM-1.By hybridization screening of a human genomic library derived from blood mononuclear cells, five overlapping genomic DNA clones were isolated. These clones extend over 34 kb of the human genome and comprise the complete JEM-1gene and a 4-kb 5′flanking region. Determination of the exon–intron structure of Jem-1revealed seven exons whose junctions with introns exhibited typical splice sequences. A shorter transcript (Jem-1s, 1.3 kb) generated by exon 3 extension and polyadenylation was identified. Its translation generated a 23-kDa protein that exhibited a cytoplasmic localization. 5′RACE–PCR identified a major transcription start site (TSS) located at 403 nt upstream of the ATG. Computer analysis of the 1.8-kb 5′flanking region showed that it lacks a TATA box, Inr motifs or DPE motifs, but it contains a typical CCAAT box located 95 bp upstream of the TSS. Sequencing also revealed potential cis-acting elements for multiple transcription regulators including Sp1, GATA, C/EBP, AP-1, and Pu1. No retinoic acid receptor elements or retinoic X receptor elements were detected. This 1.8-kb DNA sequence showed a strong constitutive promoter activity determined by a luciferase-reporter gene assay in transiently transfected HeLa cells. Retinoids further increased luciferase expression 2.7-fold. We demonstrated that the 1-kb distal sequence contains yet unidentified elements reducing constitutive transcription. Thus, the maximal constitutive promoter activity was assigned to a −432 + 101 region overlapping the TSS. These data support the idea of a constitutive expression of JEM-1,but a negative regulation in APL released by retinoids.

To elucidate the molecular mechanism of all-trans-retinoic acid (ATRA)–induced differentiation of acute promyelocytic leukemia (APL) cells, the gene expression patterns in the APL cell line NB4 before and after ATRA treatment were analyzed using complementary DNA array, suppression-subtractive hybridization, and differential-display–polymerase chain reaction. A total of 169 genes, including 8 novel ones, were modulated by ATRA. The ATRA-induced gene expression profiles were in high accord with the differentiation and proliferation status of the NB4 cells. The time courses of their modulation were interesting. Among the 100 up-regulated genes, the induction of expression occurred most frequently 12-48 hours after ATRA treatment, while 59 of 69 down-regulated genes found their expression suppressed within 8 hours. The transcriptional regulation of 8 induced and 24 repressed genes was not blocked by cycloheximide, which suggests that these genes may be direct targets of the ATRA signaling pathway. A balanced functional network seemed to emerge, and it formed the foundation of decreased cellular proliferation, maintenance of cell viability, increased protein modulation, and promotion of granulocytic maturation. Several cytosolic signaling pathways, including JAKs/STAT and MAPK, may also be implicated in the symphony of differentiation.

Strips of Fe62.5Co6Ni7.5Zr6Cu1Nb2B15, prepared by rapid quenching from the melt, were annealed at different temperatures and studied performing Mössbauer and magnetoelastic measurements. By increasing annealing temperature, the long range order increases strengthening the exchange interactions. The maximum of magnetoelastic response corresponds to the maximum of the out of plane magnetization. A possible explanation for the onset of the perpendicular magnetic anisotropy is proposed.

Abstract: Leishmaniasis is widespread in Colombia and is found in 30 of 32 Departments. More than 200 infection zones have been reported from different regions, which vary from sea-level to an altitude of 2,300 m along the Atlantic Coast, Pacific coast, Amazon basin, Cauca and Magdalena valleys. We report 76 Leishmania stocks isolated from humans, dogs and phlebotomine hosts. Isoenzyme electrophoresis revealed 16 zymodemes, which could be divided into four phylogenetic complexes, i.e., L. braziliensis, L. amazonensis, L. guyanensis/panamensis and L. infantum. Three zymodemes became integrated into the subgenus Leishmania and the other zymodemes into the subgenus Viannia. Cutaneous infections were due to the L. braziliensis (9.2%) and L. guyanensis/panamensis (85.54%) complexes. Mucous secondary involvement was due to the L. braziliensis and L. guyanensis/panamensis complexes. In this work the specific status of L. (V.) guyanensis and L. (V.) panamensis is discussed.

•Streptococcus pneumoniaeinfections are frequent and mostly community-based.•In France, the pneumococcal surveillance is monitored by the network of ORP, in collaboration with the National Reference Center•The proportion of PDSP isolated in meningitis increased both in children (29.2, 2009 vs 51.4% in 2021) and in adults (26.1 in 2009 vs 46.1% in 2021)•Most of the serotypes included in the PCV13 have disappeared after vaccine introduction, while those not included in the PCV13 vaccine have emerged

The existence of incompressible flow producing negative magnetic eddy diffusivities is demonstrated. This provides for a dynamo mechanism, alternative to α-type effects, requiring neither the presence of mean heliciiy nor the breaking of parity invariance. In the kinematic dynamo phase, the magnetic field grows exponentially with a growth rate proportional to the square of the wavenumber. The concrete example, analyzed by means of multiscale techniques, is a parity-invariant flow of the Taylor-Green type.

Medulloblastoma is one of the most common primary tumors of the central nervous system seen in children; in contrast, it is somewhat uncommon in adult age. Due to the infrequent occurrence, data on incidence rates are sparse. The present study was aimed at ascertaining the epidemiological characteristics of medulloblastoma in adult age in Piedmont during the period 1976–1995. Piedmont is a region in northwest Italy, which had a total population of 4.30 millions/year for the period mentioned. From the files of clinical records of patients hospitalized in neurologic and neurosurgical departments, 45 cases (32 males, 13 females) of histologically verified medulloblastoma were recorded. The incidence rate (annual per million) in the whole period studied was 0.5 (95% confidence interval, 0.36–0.67). The incidence rate was high in the age group 15 to 19 years (2.33/million/year) and decreased up to age 40, consistent with the embryonal origin of the tumor. No time-trend of incidence rate was found. Male excess was evident in all age groups and in each time period. Median survival time was 17.6 years; the 5-year survival rate was 69.9%. Survival rate in the present group of adult medulloblastoma is slightly better than that reported in clinical series. A comparison was made with incidence data concerning pediatric medulloblastoma reported in the Registry of Childhood Cancer of Piedmont: from 1980 to 1989, adult medulloblastomas represented 34% of medulloblastomas. The figure is higher than that generally assumed, and indicates that the occurrence of this embryonal tumor in adult age is relevant. Our epidemiological data are consistent with an embryonal origin of medulloblastoma. Int. J. Cancer 80:689–692, 1999. © 1999 Wiley-Liss, Inc.

Magnetic field dependences of the piezomagnetic dynamics Zdand moduli of elasticity in the Fe73.5Cu1Ta3Si15.5B7alloy strips in as-quenched state and after annealing for 1 h in vacuum in the temperature range from 400°C to 600°C were studied. The elasticity moduli were changed from 75 to 210 GPa depending on magnetic bias field and annealing temperature. A crystalline structure having nanometer sized crystals was observed after annealing at 500°C.

Treatment of IPC‐81 cells led to inhibition of protein synthesis, which was accompanied by an increase in the average size of polysomes and a decreased rate of elongation, indicating that it involved inhibition of peptide chain elongation. This inhibition was also associated with increased phosphorylation of elongation factor eEF2 (which inhibits its activity) and enhanced Ca2+/calmodulin‐independent activity of eEF2 kinase. Previous work has shown that phosphorylation of eEF2 kinase by cAMP‐dependent protein kinase (cAPK) in vitro induces such activator‐independent activity, and the present data show that such a mechanism can occur in intact cells to link physiological levels of cAPK activation with inhibition of protein synthesis.

The membrane monosialoganglioside GM1, the high affinity receptor for cholera toxin, is generally considered ubiquitous on normal cells. It was found to be abundant both on normal mature hemopoietic cells and on leukemic cells. By contrast, the normal factor‐dependent cell lines, which achieve indefinite proliferation in the presence of the multilineage hemopoietic growth factor apparently displayed the unique character of having low or undetectable levels of surface membrane and cytoplasmic cholera toxin receptors. These results were obtained by the Scatchard analysis of 125iodinated toxin binding, immunofluorescence studies and gel electrophoresis autoradiography. This corroborated the fact that these cells were highly resistant to growth inhibition by cholera toxin (microM to fM) while normal mature cells and leukemic cells of similar phenotype were sensitive.

Acute promyelocytic leukemia (APL) is due to a chromosomal t(15;17) translocation which involves a novel human gene, Myl, (also named PML) and the retinoic acid (RA) receptor alpha (RAR‐alpha) gene. We report here the characterization of Myl and of the reciprocal MylRAR (PMLRAR) and RARMyl (RARPML) fusion transcripts which are found in two classes of APL patients. Myl displays similarities with a new family of proteins of which some members are fused to protooncogenes in the transforming proteins RFP‐ret and T18. The speckled nuclear localization of Myl, as well as its sequence homology with the 52 kDa component of the RO/SSA ribonucleoprotein particle, suggest that Myl may be present in a ribonucleoprotein complex. In contrast to both Myl and RAR‐alpha whose localization is essentially nuclear in the presence or absence of RA, MylRAR which is largely cytoplasmic in the absence of RA appears to be translocated to the nucleus in the presence of RA. Myl and MylRAR can associate in vitro and this association is mediated by a coiled coil in the Myl sequence. In vivo this association results in a colocalization of Myl and MylRAR which is identical to that of MylRAR alone. Studies of activation of transcription from the promoters of several RA target genes indicate that MylRARs have altered transcription activation properties when compared with RAR‐alpha. Most notably, MylRAR represses markedly the activity of some RA target promoters in the absence of RA. Western blot analyses of patient samples show that MylRAR is expressed to a much higher level than wild type RAR‐alpha originating from the normal allele. Taken together, these results suggest that MylRAR may interfere in a dominant manner with both Myl and RAR functions.

Acute promyelocytic leukemia (APL) is a well-defined entity among acute leukemia, cytogenetically characterized by a t(15;17) (q22;q11-12) translocation. In vitro and in vivo studies suggest that all-transretinoic acid (RA) treatment restores cell maturation. We have isolated the first permanent cell line with t(15;17), derived from the marrow of a patient with APL in relapse. The establishment of the cell line, its morphologic, karyotypic, and immunohistochemical features are reported. RA induced cell line maturation. Cells strongly expressed myeloid markers, but also some T-cell markers. Additional karyotypic abnormalities, a 12p rearrangement and the possible presence of a homogenous staining region (HSR) on 19q+ are discussed both in relation to T-cell (CD2, CD4) and monocyte (CD9) markers, and to the acquired cell growth autonomy. The cell line represents a remarkable tool for biomolecular studies.

In the acute promyelocytic leukemia (APL) cell line NB4, as well as in APL patients' cells, arsenic trioxide (As2O3) leads to incomplete cell maturation, induction of apoptosis, as well as to the degradation of the oncogenic PML/RARa fusion protein. We have isolated an arsenic-resistant NB4 subline (NB4-AsR), which fails to undergo apoptosis, but maintains the partial differentiation response to this drug. When grown in the presence of As2O3, NB4-AsRcells degrade PML/RARa, slightly differentiate, and become more sensitive to serum deprivation-induced apoptosis. Similarly, in RA-resistant NB4-R1 cells, RA induced a significant PML/RARa degradation and yet failed to induce cell maturation. Thus, As2O3- or retinoic acid (RA)-induced PML/RARa degradation may be a prerequisite, but is not sufficient for the full differentiative/apoptotic response to these drugs. Strikingly, RA-triggered differentiation and apoptosis were greatly accelerated in As2O3-treated NB4-AsR cells. The synergism between these two agents in this setting could provide an experimental basis for combined or sequential RA/As2O3 therapies.