Effect of pps disruption and constitutive expression of srfA on surfactin productivity, spreading and antagonistic properties of Bacillus subtilis 168 derivatives.

AIMS: To analyse the effects of plipastatin operon disruption and constitutive expression of surfactin operon in Bacillus subtilis 168 on surfactin productivity, in vitro invasive growth and antagonism against fungi. METHODS AND RESULTS: The srfA native promoter was replaced by the constitutive promoter P(repU) in B. subtilis 168 after integration of a functional sfp gene. Moreover, the plipastatin synthesis was further disrupted in the B. subtilis 168 derivatives. In liquid media, an earlier and higher expression of P(repU), than that found with P(srfA), led to a specific surfactin production fivefold higher after 6 h of culture. On solid media, not only the invasive growth and the haemolytic activity but also the antifungal activity of the constitutive strains were improved when compared to the parental strain BBG111. As expected, the disruption of the plipastatin operon strongly reduced in vitro antifungal properties but, interestingly, enhanced specific surfactin production (1.47 g g(-1) of biomass), spreading behaviour and haemolytic activity of the strains. CONCLUSIONS: This work demonstrates for the first time the interdependency of surfactin and plipastatin regarding their biosynthesis as well as their influence on the biological activities of the producing strain. SIGNIFICANCE AND IMPACT OF THE STUDY: The constitutive overproduction of surfactin enhances the invasive growth and the in vitro antagonistic activity of the mutant strain. Both properties are known to play an important role in the biocontrol of plant diseases. Plipastatin operon disruption increases the surfactin productivity of mutant strains. These mutants are interesting for use in continuous bioprocesses for surfactin production or in bioremediation.

Hyperdiploid karyotypes in acute myeloid leukemia define a novel entity: a study of 38 patients from the Groupe Francophone de Cytogenetique Hematologique (GFCH).

A series of 38 patients with acute myeloblastic leukemia (AML) with 49 or more chromosomes and without structural abnormalities was selected within the Groupe Francophone de Cytogénétique Hématologique (GFCH) to better define their characteristics. The median age of the patients was 65 years, and all FAB subtypes were represented. Although all chromosomes were gained, some seems to prevail: chromosome 8 (68%), 21 (47%), 19 (37%), and 13 and 14 (34% each). Since MLL rearrangement leads patients in a group with an unfavorable prognosis, search for cryptic rearrangements of MLL was performed in 34 patients and showed abnormalities in 5 (15%). When we applied the most frequent definition of complex karyotypes (three or more abnormalities), all patients with high hyperdiploid AML fall in the unfavorable category. Among the 18 patients without MLL rearrangement receiving an induction therapy, 16 (89%) reached CR and 6 (33%) were still alive after a 31-month median follow-up (14-61 months). Although this study was retrospective, these results suggest that high hyperdiploid AML without chromosome rearrangement seems to be a subgroup of uncommon AML (less than 1%), and may be better classified in the intermediate prognostic group.

Report of 34 patients with clonal chromosomal abnormalities in Philadelphia-negative cells during imatinib treatment of Philadelphia-positive chronic myeloid leukemia.

Imatinib mesylate (Gleevec), an inhibitor of the BCR-ABL tyrosine kinase, was introduced recently into the therapy of chronic myeloid leukemia (CML). Several cases of emergence of clonal chromosomal abnormalities after therapy with imatinib have been reported, but their incidence, etiology and prognosis remain to be clarified. We report here a large series of 34 CML patients treated with imatinib who developed Philadelphia (Ph)-negative clones. Among 1001 patients with Ph-positive CML treated with imatinib, 34 (3.4%) developed clonal chromosomal abnormalities in Ph-negative cells. Three patients were treated with imatinib up-front. The most common cytogenetic abnormalities were trisomy 8 and monosomy 7 in twelve and seven patients, respectively. In 15 patients, fluorescent in situ hybridization with specific probes was performed in materials archived before the initiation of imatinib. The Ph-negative clone was related to previous therapy in three patients, and represented a minor pre-existing clone that expanded after the eradication of Ph-positive cells with imatinib in two others. However, in 11 patients, the new clonal chromosomal abnormalities were not detected and imatinib may have had a direct effect. No myelodysplasia was found in our cohort. With a median follow-up of 24 months, one patient showed CML acceleration and two relapsed.

New mechanisms of AML1 gene alteration in hematological malignancies.

The human AML1 gene (also named CBFA2 or RUNX1), located in the 21q22 chromosomal band, encodes for one of the two subunits forming a heterodimeric transcription factor, the human core binding factor (CBF). AML1 protein contains a highly evolutionary conserved domain of 128 amino acids called runt domain, responsible for both heterodimerization with the beta subunit of CBF and for DNA binding. AML1 is normally expressed in all hematopoietic lineages and acts to regulate the expression of various genes specific to hematopoiesis playing a pivotal role in myeloid differentiation. AML1 is one of the genes most frequently deregulated in leukemia through different mechanisms including translocation, mutation and amplification. Translocations lead to the formation of fusion genes encoding for chimerical proteins such as AML1-ETO which induces leukemogenesis. Recently, new mechanisms of AML1 deregulation by point mutations or amplification have been reported. To our knowledge, 51 patients (among 805 studied) with AML1 point mutations have been described. Forty of them have acute myeloid leukemia (AML) most often M0 AML. In this subtype of AML, the frequency of AML1 mutation is significantly higher; 21.5% of patients mutated (34/158). Mutations have also been found with lower frequency in other FAB subtype AML (6 cases), in myeloproliferative disorders (6 cases), in myelodysplastic syndrome (3 cases) and rarely in acute lymphoblastic leukemia (1 case). AML1 gene amplification has been found essentially in childhood ALL (12 cases) and more rarely in myeloid malignancies (4 cases). Here, we reviewed all these cases of AML1 point mutations and amplification and focused on the mechanisms of AML1 deregulation induced by these alterations.

The effect of hypertonic saline dextran solutions on hypoxic pulmonary vasoconstriction in anaesthetised piglets.

Hypoxic pulmonary vasoconstriction (HPV) is a regulatory mechanism by which blood is diverted from poorly ventilated to better ventilated areas of the lung. The aim of the present study was to assess the extent to which hypertonic saline dextran and dextran solutions modify the magnitude of HPV during isovolumic haemodilution in intact acutely instrumented piglets. Eighteen large white piglets were anesthetised and assigned to two groups. Mean pulmonary arterial pressure (PAP) and cardiac output (Q), systemic arterial pressure and left arterial pressure (LAP) were measured. A decrease in Q was obtained by reducing venous return. This enabled measurement of transpulmonary pressures (mean PAP minus LAP) at four levels of Q in hyperoxia (inspiratory oxygen fraction (FiO2)=0.4) then in hypoxia (Fi,O2=0.1) in the two groups before blood soustraction (10 mL x kg(-1)) and after loading with sodium chloride (NaCl) 7.5% and dextran 6% or with dextran 6% alone. Dextran alone led to a decrease in mean PAP-LAP/Q values, and NaCl with dextran was associated with a significant shift of mean PAP-LAP/Q plots to higher pressures in hypoxia. Hypertonic saline dextran solution, as replacement fluid in isovolaemic haemodilution increased the magnitude of hypoxic pulmonary vasoconstriction, whereas dextran solution reduced it.

Abnormal activation in the visual cortex after corneal refractive surgery for myopia: demonstration by functional magnetic resonance imaging.

OBJECTIVE: To try to correlate subjective photophobic symptoms with visual pathway modifications (from the retinal image to the visual cortex) after refractive surgery by exploring brain activation on photic stimulation. DESIGN: Noncomparative case series. PARTICIPANTS: Four subjects reporting discomfort produced by luminance (glare, halos, starbursts, or a combination thereof) in one eye after laser in situ keratomileusis (LASIK) were enrolled. The contralateral myopic eye (control) had no visual impairment and had undergone LASIK without complications or had not had previous surgery. METHODS: Functional magnetic resonance imaging was performed during photic stimulation, delivered by an optical fiber, of the affected and unaffected eyes. RESULTS: Functional magnetic resonance imaging provided evidence that most subjective visual symptoms correlated with anatomic flap abnormalities are associated with a higher signal increase in the visual association cortices compared with a nonsymptomatic eye. CONCLUSIONS: Functional magnetic resonance imaging of the visual cortex may help in exploring the mechanisms involved in glare effects after refractive surgery.

Occurrence of two superoxide dismutases in Aeromonas hydrophila: molecular cloning and differential expression of the sodA and sodB genes.

Aeromonas spp., considered as emerging opportunistic pathogens, belong to the family Vibrionaceae. Among the criteria currently used for their classification is the presence of a single FeSOD (iron-containing superoxide dismutase), which distinguishes them from Enterobacteriacea. In this paper the cloning of the sodA and sodB genes encoding two different SODs in Aeromonas hydrophila ATCC 7966 is reported. The sodB gene encoded an FeSOD (196 amino acids, 21.5 kDa), was constitutively expressed and showed 75% homology with the E. coli FeSOD. The sodA gene encoded a protein of 206 amino acids (22.5 kDa) with MnSOD (manganese-containing SOD) activity and showed 55% homology with the Escherichia coli MnSOD. The MnSOD of A. hydrophila was detected only during the stationary phase of growth under high aeration or when induced by lack of iron. Nevertheless, paraquat had no detectable effect on its production. The amino-terminal part of the Mn-containing protein contained a putative signal sequence which could permit a periplasmic localization.

Ocular dominance columns in the adult New World Monkey Callithrix jacchus.

In the marmoset Callithrix jacchus, ocular dominance columns (ODC) have been reported to be present in young animals, but absent in adults (Spatz, 1989). We have studied in juvenile and adult animals the postnatal organization of the retino-geniculo-cortical afferents by means of transneuronal labeling. We show in the present work that ODC are present in the primary visual cortex of Callithrix jacchus, both in the adult and in the juvenile animal. The present work confirms the presence of ODC in the visual cortex of juvenile marmoset before the end of the first postnatal month. In 2-month-old animals, ODC are well demarcated in IVcalpha and IVcbeta. In the adult marmosets, the present data clearly show that the primary visual cortex is also organized with ODC. In horizontal sections, they form a mosaic through the ventral and dorsal calcarine cortex and through the dorso-lateral occipital part of the striate cortex. In frontal sections, their presence is manifest in IVcbeta within the calcarine cortex and they only faintly appear in IVcalpha. These new findings are important since they underline the usefulness of the adult New World Monkeys as a model in visual research.

Effects of sevoflurane on hypoxic pulmonary vasoconstriction in anaesthetized piglets.

In vitro, halogenated agents reduce the pulmonary vasoconstrictor response to alveolar hypoxia in isolated perfused lungs. However, studies in intact animals have been less convincing. The aim of the present study was to assess the effect of sevoflurane on hypoxic pulmonary vasoconstriction (HPV) in anaesthetized piglets using the pressure/cardiac index relationship (P/Q). Ten large white piglets were anaesthetized and mechanically ventilated, alternately in hyperoxia (FIO2 = 0.4) and hypoxia (FIO2 = 0.12). Multipoint plots of pulmonary arterial pressure (PAP) or differences between PAP and left atrial pressure (LAP) against Q were generated by gradual inflation of a balloon introduced into the inferior vena cava. P/Q relationships were established in hyperoxia and hypoxia at baseline, and then with sevoflurane. In hypoxia, pressure gradients (PAP-LAP) increased at every level of Q, thus demonstrating active pulmonary vasoconstriction. Sevoflurane at 1 MAC did not affect these P/Q relationships in hyperoxia or hypoxia as compared with baseline. Sevoflurane at a clinically relevant concentration (1 MAC) has no significant effect on HPV in anaesthetized piglets.

Effect of monocular deprivation on NMDAR1 immunostaining in ocular dominance columns of the marmoset Callithrix jacchus.

We previously showed that immunoreactivity to N-Methyl-D-aspartate (NMDA) receptors in primary visual cortex of Callithrix jacchus is regulated by visual activity during the second and third postnatal months (Fonta et al., 1997). The purpose of the present study was to show that the columnar pattern of high and low NMDAR1 immunoreactivity observed in monocularly deprived animals corresponds to ocular dominance columns linked to the nondeprived and deprived eye, respectively. We compared cortical distribution of NMDAR1 receptors and the projection zones of thalamic afferents, revealed by transneuronal transport of tritiated proline, in 2-month-old, either monocularly deprived or control, marmosets. The data show that ocular dominance columns exist in 2-month-old marmosets and that a 2-week monocular deprivation by means of eyelid suture leads to a modification of the thalamo-cortical afferents organization. Experiments of neuronal tracing and immunohistochemistry performed on the same animals demonstrated that cortical domains with decreased NMDAR1 level correspond to the deprived eye columns. These investigations, coupled to the previous results, strongly suggest that the NMDA receptors, regulated by visual activity, are involved in the refining of ocular dominance columns in the primary visual cortex of juvenile marmoset.

Effect of light on germinating spores of Streptomyces viridosporus.

Our study showed that the effect of light on germinating spores of Streptomyces was variable: some species were indifferent, whereas others, such as Streptomyces viridosporus, displayed a marked inhibition of CFU numbers on growth medium. A special study with S. viridosporus showed that light only had an impact during the first few hours of spore incubation. The effects of scavengers of toxic forms of oxygen and of photosensitizers, along with the oxidative stress of illuminated spores evidenced by the superoxide dismutase levels, suggested that light and oxygen had a combined action.

'LABNOTE', a laboratory notebook system designed for academic genomics groups.

We have developed a relational laboratory database system, adapted to the daily book-keeping needs of laboratories that must keep track of information acquired on hundreds or thousands of clones in an effective and user-friendly fashion. Data, whether final or related to experiments in progress, can be accessed in many different ways, e.g. by clone name, by gene, by experiment or through DNA sequence. Updating, import and export of results is made easier by specially developed tools. This system, in network version, serves several groups in our Institute and (over the Internet) elsewhere, and is instrumental in collaborative studies based on expression profiling. It can be used in many similar situations involving progressiveaccumulation of information on sets of clones or related objects.

Scanning direction and line bisection: a study of normal subjects and unilateral neglect patients with opposite reading habits.

Sixty normal dextrals (30 left-to-right and 30 right-to-left readers) and two left unilateral neglect patient with opposite reading habits performed a passive line bisection task. In order to study the effect of scanning direction on performance, subjects had to stop a mark moving on the to-be-bisected line either from the left to the right or in the opposite direction. Results showed that the position of the subjective middle was dependent upon the scanning direction of the line for all the subjects. A leftward deviation appeared for left to right scanning, whereas a rightward shift occurred when the mark moved from the right to the left. These results emphasize the role of scanning direction in space organization and are discussed with respect to the explanatory hypotheses of unilateral neglect.

On the iron requirement of lactobacilli grown in chemically defined medium.

The iron requirement of four strains of lactobacilli (L. acidophilus, L. delbrueckii subsp. bulgaricus, L. plantarum, and L. pentosus) was studied in a synthetic medium under aerobic or anaerobic conditions. Effects of iron salt and iron-chelated compounds were tested on bacterial growth in manganese-free or -supplemented media. No significant growth stimulation was observed in any condition. These results support the absolute manganese requirement for optimum growth of lactobacilli and the needless incorporation of iron in growth media.

The cell cycle gene SKP1 is regulated by light in postnatal rat brain.

During the early postnatal phase of high neuronal plasticity, an altered visual input leads to great modifications of visual cortex organization [Y. Frégnac, M. Imbert, Development of neuronal selectivity in primary visual cortex of cat, Physiol. Rev., 64 (1984) 375-434; D.H. Hubel, T.N. Wiesel, S. LeVay, Plasticity of ocular dominance columns in monkey striate cortex, Philos. Trans. R. Soc. London, Ser. B, 278 (1977) 377-409.]. We used refined differential screening of an organized cDNA library to identify the genes that may participate in this plasticity. We isolated a candidate plasticity gene encoding for a 163 aa protein that is closely related to the human and yeast Skp1p, a key factor in cell cycle progression [C. Baï, K. Hofman, L. Ma, M. Goebl, J.W. Harper, S.J. Elledge, SKP1 connects cell cycle regulators to the ubiquitin proteolysis machinery through a novel motif, the F-box, Cell, 86 (1996) 263-274; C. Connelly, P. Hieter, Budding yeast SKP1 encodes an evolutionary conserved kinetochore protein required for cell cycle progression, Cell, 86 (1996) 275-285; H. Zhang, R. Kobayashi, K. Galaktionov, D. Beach, p19Skp1 and p45Skp2 are essential elements of the cyclin A-CDK2 S phase kinase, Cell, 82 (1995) 915-925.]. Northern blot analysis showed that the expression of SKP1 (Skp1p gene) dramatically decreased after 2 h of light stimulation in the visual cortex of young dark-reared rats. This down regulation lasted at least 72 h. It was specific for the critical period as we did not observe any significant regulation of SKP1 mRNA by light in adult dark-reared rat brain. The down regulation was observed in the superior colliculus but also in the frontal cortex and in the hippocampus. The fact that this down regulation was not restricted to the visual system, suggested that it could be produced by dark rearing-induced hormonal changes. The significance of SKP1 expression in the brain and its regulation are discussed.

Diagnosis, classification, and cytogenetics of myelodysplastic syndromes.

BACKGROUND AND OBJECTIVE: The diagnosis of myelodysplastic syndromes (MDS) is essentially morphological and based on the presence of dysplastic features in the peripheral blood and bone marrow. The French-American-British (FAB) Cooperative Group proposed a classification based on easily obtainable laboratory information. In spite of some limitations, the FAB criteria have been useful for a long time. Currently, the recognition of other distinct morphological MDS subgroups such as hypocellular MDS and MDS with myelofibrosis, the increasing incidence of MDS in children as well as that of therapy-related MDS, and the finding of specific chromosomal alterations associated with different morphological features, reveal the insufficiency of this classification. The aim of the present review is to examine some new aspects of the diagnosis, classification, and cytogenetics of MDS. EVIDENCE AND INFORMATION SOURCES: The authors of this review have been actively working and contributing original papers on MDS for the last 15 years. They also organized or participated in the Fourth International Symposium on MDS (Barcelona, April 24-27, 1997). In addition, the present review critically examines relevant articles and abstracts published in journals covered by the Science Citation Index and Medline. STATE OF THE ART AND PERSPECTIVES: Most of investigators working on MDS tend to integrate morphology and cytogenetics in the diagnosis and classification of these disorders. FAB criteria remain useful particularly for patients with not available cytogenetic study. Refractory cytopenia with multilineage dysplasia should be considered as a new MDS subtype. Some authors propose considering all patients with more than 20% of blast cells in peripheral blood or bone marrow as having acute leukemia. Chronic myelomonocytic leukemia with myeloproliferative features may be included among chronic myeloproliferative disorders. MDS with myelofibrosis is recognized as a new MDS subtype. Therapy-related MDS (t-MDS) should be classified according to the involved agents. Finally, besides including chromosomal abnormalities in the diagnosis (e.g., RAEB with trisomy 8), several cytogenetic abnormalities such as deletion 5q and deletion 17q, associated to specific clinical-morphological features, should be of help to identify new MDS syndromes.

A mathematical approach to the connectivity between the cortical visual areas of the macaque monkey.

The visual cortex of the macaque monkey is divided into many distinct visual information processing areas. In many cases, anatomical and physiological results allow one to determine the presence or the absence of neuronal connections from one area to another. We have approached the topology of this neuronal network within the mathematical framework of graph theory. At first, we studied the unknown part of the network, i.e. the part where anatomical and physiological results are lacking. Relying on a specific topological property of the network established on the known part, we developed an interpolation algorithm for reducing the level of uncertainty concerning the unknown part. From these results, we then constructed a connectional model of the neuronal network for the entire cortical visual system. Subsequently, a topological analysis of this model, with the help of factorial analysis and clustering technics, shows its structural properties and singular vertices. This analysis suggests the existence of two distinct classes of areas, one in the parietal part of the cortex and the other in the temporal part, which are connected to each other via relay areas, especially involving the frontal eye field. These results may help to understand the functional role of particular cortical areas in vision and, more generally, to explore how visual information flows within the visual cortex.