Generation of induced pluripotent stem cell lines IRMBi003-A and IRMBi003-B from a healthy donor to model Alzheimer's disease.

Induced Pluripotent Stem Cell (iPSC) lines derived from healthy individuals are helpful and essential tools for disease modelling. Here, we described the reprogramming of skin fibroblasts obtained from a healthy 59-year-old individual without Alzheimer's disease. The generated iPSC lines have a normal karyotype, expressed pluripotency markers, and demonstrated the ability to differentiate into the three germ layers. The iPSC lines will be used as controls to study Alzheimer's disease mechanisms.

Influence of organic amendments on diuron leaching through an acidic and a calcareous vineyard soil using undisturbed lysimeters.

The influence of different organic amendments on diuron leaching was studied through undisturbed vineyard soil columns. Two composts (A and D), the second at two stages of maturity, and two soils (VR and Bj) were sampled. After 1 year, the amount of residues (diuron+metabolites) in the leachates of the VR soil (0.19-0.71%) was lower than in the Bj soil (4.27-8.23%), which could be explained by stronger diuron adsorption on VR. An increase in the amount of diuron leached through the amended soil columns, compared to the blank, was observed for the Bj soil only. This result may be explained by the formation of mobile complexes between diuron and water-extractable organic matter (WEOM) through the Bj soil, or by competition between diuron and WEOM for the adsorption sites in the soil. For both soils, the nature of the composts and their degree of maturity did not significantly influence diuron leaching.

Evaluating equilibrium and non-equilibrium transport of bromide and isoproturon in disturbed and undisturbed soil columns.

In this study, displacement experiments of isoproturon were conducted in disturbed and undisturbed columns of a silty clay loam soil under similar rainfall intensities. Solute transport occurred under saturated conditions in the undisturbed soil and under unsaturated conditions in the sieved soil because of a greater bulk density of the compacted undisturbed soil compared to the sieved soil. The objective of this work was to determine transport characteristics of isoproturon relative to bromide tracer. Triplicate column experiments were performed with sieved (structure partially destroyed to simulate conventional tillage) and undisturbed (structure preserved) soils. Bromide experimental breakthrough curves were analyzed using convective-dispersive and dual-permeability (DP) models (HYDRUS-1D). Isoproturon breakthrough curves (BTCs) were analyzed using the DP model that considered either chemical equilibrium or non-equilibrium transport. The DP model described the bromide elution curves of the sieved soil columns well, whereas it overestimated the tailing of the bromide BTCs of the undisturbed soil columns. A higher degree of physical non-equilibrium was found in the undisturbed soil, where 56% of total water was contained in the slow-flow matrix, compared to 26% in the sieved soil. Isoproturon BTCs were best described in both sieved and undisturbed soil columns using the DP model combined with the chemical non-equilibrium. Higher degradation rates were obtained in the transport experiments than in batch studies, for both soils. This was likely caused by hysteresis in sorption of isoproturon. However, it cannot be ruled out that higher degradation rates were due, at least in part, to the adopted first-order model. Results showed that for similar rainfall intensity, physical and chemical non-equilibrium were greater in the saturated undisturbed soil than in the unsaturated sieved soil. Results also suggested faster transport of isoproturon in the undisturbed soil due to higher preferential flow and lower fraction of equilibrium sorption sites.

The organic carbon derived from sewage sludge as a key parameter determining the fate of trace metals.

In a sandy agricultural soil of south-west of France, continuously cultivated with maize and amended with sewage-sludge over 20 years, the behavior of three trace metals (Cu, Pb, and Zn) was studied during the sludge applications (1974-1993) and after its cessation (1993-1998). Using the delta13C analysis, the dynamics of different sources of organic matter were followed in order to elucidate the influence of the sludge-derived organic matter on the fate of trace metals in the soil and its particle size fractions. This study revealed that sludge-derived organic matter contributed to the formation of macroaggregates through the binding of preexisting microaggregates. These macroagreggates were thus responsible for the accumulation of trace metals in the coarsest fraction as well as for the protection of maize-derived organic matter against biodegradation. After sludge application ceased, the disaggregation of macroaggregates occurred simultaneously with high losses in Cu and Pb. On the contrary, Zn appeared less affected by the cessation of sludge application, with only a location change from coarse to fine fractions.

Reversible encephalopathy associated with cholesterol embolism syndrome: magnetic resonance imaging and pathological findings.

We describe a patient found to have acute diffuse and reversible encephalopathy on magnetic resonance imaging (MRI) associated with cholesterol emboli syndrome (CES). The initial MRI showed extensive white matter, basal ganglia and cortical damage without evidence of brain infarction. Dramatic clinical and MRI improvement was observed with corticosteroids. Pathologically, cholesterol crystal emboli were found in the lumen of skin and brain arteries and were associated with varying degrees of inflammation of the arteriole wall. This case suggests that CES may be responsible for extensive, acute and reversible encephalopathy underlined by an inflammation of brain arteries.

Quantifying the contribution of nitrification and denitrification to the nitrous oxide flux using 15N tracers.

Microbial transformations of nitrification and denitrification are the main sources of nitrous oxide (N2O) from soils. Relative contributions of both processes to N2O emissions were estimated on an agricultural soil using 15N isotope tracers (15NH4+ or 15NO3-), for a 10-day batch experiment. Under unsaturated and saturated conditions, both processes were significantly involved in N2O production. Under unsaturated conditions, 60% of N-N2O came from nitrification, while denitrification contributed around 85-90% under saturated conditions. Estimated nitrification rates were not significantly different whatever the soil moisture content, whereas the proportion of nitrified N emitted as N2O changed from 0.13 to 2.32%. In coherence with previous studies, we interpreted this high value as resulting from the decrease in O2 availability through the increase in soil moisture content. It thus appears that, under limiting aeration conditions, some values for N2O emissions through nitrification could be underestimated.

[Congenital myasthenic syndromes: phenotypic expression and pathophysiological characterisation]. / Syndromes myasthéniques congénitaux.

Congenital Myasthenic Syndromes (CMS) are a heterogeneous group of diseases caused by genetic defects affecting neuromuscular transmission. The twenty five past Years saw major advances in identifying different types of CMS due to abnormal presynaptic, synaptic, and postsynaptic proteins. CMS diagnosis requires two steps: 1) positive diagnosis supported by myasthenic signs beginning in neonatal period, efficacy of anticholinesterase medications, positive family history, negative tests for anti-acetylcholine receptor (AChR) antibodies, electromyographic studies (decremental response at low frequency, repetitive CMAP after one single stimulation); 2) pathophysiological characterisation of CMS implying specific studies: light and electron microscopic analysis of endplate (EP) morphology, estimation of the number of AChR per EP, acetylcholinesterase (AChE) expression, molecular genetic analysis. Most CMS are postsynaptic due to mutations in the AChR subunits genes that alter the kinetic properties or decrease the expression of AChR. The kinetic mutations increase or decrease the synaptic response to ACh resulting respectively in Slow Channel Syndrome (characterized by a autosomal dominant transmission, repetitive CMAP, refractoriness to anticholinesterase medication) and fast channel, recessively transmitted. AChR deficiency without kinetic abnormalities is caused by recessive mutations in AChR genes (mostly epsilon subunit) or by primary rapsyn deficiency, a post synaptic protein involved in AChR concentration. Recently, mutations in SCN4A sodium channel have been reported in one patient. AChE deficiency is identified on the following data: recessive transmission, presence of repetitive CMAP, refractoriness to cholinesterase inhibitors, slow pupillary response to light and absent expression of the enzyme at EP. This synaptic CMS is caused by mutations in the collagenic tail subunit (ColQ) that anchors the catalytic subunits in the synaptic basal lamina. The most frequent presynaptic CMS is caused by mutations of choline acetyltransferase. Several CMS are still not characterized. Many EP molecules are potential etiological candidates. In these unidentified cases, other methods of investigations are required: linkage analysis, when sufficient number of informative relatives are available, microelectrophysiological studies performed in intercostal or anconeus muscles. Prognosis of CMS, depending on severity and evolution of symptoms, is difficult to assess, and it cannot not be simply derived from mutation identification. Most patients respond favourably to anticholinesterase medications or to 3,4 DAP which is effective not only in presynaptic but also in postsynaptic CMS. Specific therapies for slow channel CMS are quinidine and fluoxetine that normalize the prolonged opening episodes. Clinical benefits derived from the full characterisation of each case include genetic counselling and specific therapy.

Alachlor and bentazone losses from subsurface drainage of two soils.

Atrazine (6-chloro-N2-ethyl-N4-isopropyl-1,3,5-triazine-2,4-diamine) is frequently detected at high concentrations in ground water. Bentazone [3-isopropyl-1H-2,1,3-benzothiadiazin-4(3H)-one 2,2-dioxide] plus alachlor (2-chloro-2',6'-diethyl-N-methoxymethylacetanilide) is a potential herbicide combination used as a substitute for atrazine. Thus, the objective of this study was to assess the environmental risk of this blend. Drainage water contamination by bentazone and alachlor was assessed in silty clay (Vertic Eutrochrept) and silt loam (Aquic Hapludalf) soils under the same management and climatic conditions. Drainage volumes and concentrations of alachlor and bentazone were monitored after application. Herbicides first arrived at the drains after less than 1 cm of net drainage. This is consistent with preferential flow and suggests that about 3% of the pore volume was active in rapid transport. During the monitoring periods, bentazone losses were higher (0.11-2.40% of the applied amount) than alachlor losses (0.00-0.28%) in the drains of the silty clay and silt loam. The rank order of herbicide mass losses corresponded with the rank order of herbicide adsorption coefficients. More herbicide residues were detected in drainage from the silty clay, probably due to preferential flow and more intensive drainage in this soil than the silt loam. Surprisingly, herbicide losses were higher in the drains of both soils in the drier of the two study years. This could be explained by the time intervals between the treatments and first drainage events, which were longer in the wetter year. Results suggest that the drainage phases occurred by preferential flow in the spring-summer period, with correspondingly fast leaching of herbicides, and by matrix flow during the fall-winter period, with slower herbicide migration.

Inoculation of an atrazine-degrading strain, Chelatobacter heintzii Cit1, in four different soils: effects of different inoculum densities.

The possibility to improve atrazine degradation in soils by bioaugmentation was studied. The atrazine-mineralizing strain, Chelatobacter heintzii Cit1, was inoculated in four sterile and four non-sterile soils, at varying inoculum densities. Two soils, which had shown enhanced atrazine mineralization, were used to determine which inoculum density was capable of restoring their original mineralizing capacity after sterilization. The two other soils, with intermediate and low capacity to mineralize atrazine, were used in order to demonstrate that atrazine mineralization in such soils could be improved by inoculation. Mineralization kinetics were fitted using the Gompertz model. In the case of soils adapted to atrazine mineralization, inoculation of C. heintzii did not accelerate the rate of atrazine mineralization, which was essentially performed by the indigenous microflora. However, with soils that did not mineralize atrazine, the introduction of 10(4) cfug(-1) resulted in a 3-fold increase of atrazine mineralization capacity.

Photoinductive efficiency of soil extracted humic and fulvic acids.

Humic and fulvic acids extracted from soils of different genesis were investigated for their ability to photoinduce the transformation of fenuron (2 x 10(-4) mol(-1)) at 365 nm. The ratio of the initial rate of fenuron consumption over the rate of light absorption by humic substances was found to be higher for fulvic acids (range 2.0 x 10(-3) to 9.0 x 10(-5)) than for humic acids (range 1.7 x 10(-4) to - 3.6 x 10(-5)). Within the FAs population, this ratio decreased as the specific absorption coefficient at 365 nm increased. It seems therefore that most of 365-nm absorbing components have no photoinductive activity and even reduce that of photoinductive chromophores.

Assessing the transfer of pentachlorophenol through soil columns using 13[C]isotope.

The transfer of organic pollutants was studied through soil columns using 13[C]-labelled pentachlorophenol (PCP) as a model compound. The organic carbon content and the 13[C]/12[C] ratio were measured in two soil sections, 0-3 cm and 3-6 cm, and in percolated water using an Elemental Analyser coupled with a Magnetic Mass Sector. The mass balance of carbon was evaluated and the amount of PCP was calculated in each compartment of the soil-water systems. The results show that more than 80% of the PCP-derived 13[C] remained in the upper layer of the soil column. Approximately 20% was transferred to the lower soil layer, and less than 1% was found in the water leachates. The 13[C]-labelled tracers may thus be used as an alternative to radioactive compounds to follow the fate of organic pollutants in soil and water under field conditions.

Skin biopsy immunostaining with a Notch3 monoclonal antibody for CADASIL diagnosis.

CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy) is a small-artery disease of the brain caused by NOTCH3 mutations that lead to an abnormal accumulation of NOTCH3 within the vasculature. We aimed to establish whether immunostaining skin biopsy samples with a monoclonal antibody specific for NOTCH3 could form the basis of a reliable and easy diagnostic test. We compared the sensitivity and specificity of this method in two groups of patients suspected of having CADASIL with complete scanning of mutation-causing exons of NOTCH3 (in a retrospective series of 39 patients) and with limited scanning of four exons that are mutation hotspots (prospective series of 42 patients). In the retrospective series skin biopsy was positive in 21 (96%) of the 22 CADASIL patients examined and negative in all others; in the prospective series, seven of the 42 patients had a positive skin biopsy whereas only four had a mutation detected by limited NOTCH3 scanning. Our immunostaining technique is highly sensitive (96%) and specific (100%) for diagnosis of CADASIL.

The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.

Mutations in Notch3 cause CADASIL (cerebral autosomal dominant adult onset arteriopathy), which leads to stroke and dementia in humans. CADASIL arteriopathy is characterized by major alterations of vascular smooth muscle cells and the presence of specific granular osmiophilic deposits. Patients carry highly stereotyped mutations that lead to an odd number of cysteine residues within EGF-like repeats of the Notch3 receptor extracellular domain. Such mutations may alter the processing or the trafficking of this receptor, or may favor its oligomerization. In this study, we examined the Notch3 expression pattern in normal tissues and investigated the consequences of mutations on Notch3 expression in transfected cells and CADASIL brains. In normal tissues, Notch3 expression is restricted to vascular smooth muscle cells. Notch3 undergoes a proteolytic cleavage leading to a 210-kDa extracellular fragment and a 97-kDa intracellular fragment. In CADASIL brains, we found evidence of a dramatic and selective accumulation of the 210-kDa Notch3 cleavage product. Notch3 accumulates at the cytoplasmic membrane of vascular smooth muscle cells, in close vicinity to but not within the granular osmiophilic material. These results strongly suggest that CADASIL mutations specifically impair the clearance of the Notch3 ectodomain, but not the cytosolic domain, from the cell surface.

[CADASIL: genetics and physiopathology]. / CADASIL: génétique et physiopathologie.

CADASIL, an autosomal dominant adult onset arteriopathy causing stroke and dementia in humans, is underlaid by a non atherosclerotic non amyloid angiopathy involving mainly the media of small cerebral arteries; it is characterized by major lesions of vascular smooth muscle cells. Using a positional cloning approach, we mapped CADASIL locus on chromosome 19 and identified the mutated gene as being Notch3. This gene, previously unknown in humans, encodes for a large transmembrane receptor belonging to the Notch/lin12 gene family which are known to be involved in cell fate specification during development. Genetic analysis of more than 120 CADASIL unrelated families allowed us to show that these mutations are highly stereotyped and affect only the extra cellular domain of the protein. On the basis of these data, a molecular diagnostic test has been set up and is now widely required by clinicians involved in the diagnosis of vascular leukoencephalopathies. Using this test, we recently showed that CADASIL can also occur in patients who do not have any affected relative due to the existence of notch3 de novo mutations. As a first step to investigate the molecular and cellular mechanisms leading from Notch3 mutations to CADASIL phenotype, we analyzed by in-situ hybridization and immunohistochemistry the pattern of expression of this gene. Notch3 expression is highly restricted to the vascular smooth muscle cell in normal human adults. In CADASIL tissues there is a dramatic accumulation of the extracellular domain of the protein which suggests that one of the main mechanisms of CADASIL involves anomalies in the proteolytical cleavage and clearance of this protein. These data provide important clues to the mechanisms of this condition and current work should lead in the next future to a complete understanding of CADASIL and set up the basis of a rational therapeutical approach of this condition.

A laboratory study of the mineralization and binding of 14C-labeled herbicide rimsulfuron in a rendzina soil.

The fate of pyrimidine-2-14C-rimsulfuron in a rendzina soil was investigated using a laboratory microcosm approach. Measurement of CO2 evolution suggested that rimsulfuron applied at 5 times the recommended dose did not affect soil respiration. Under abiotic conditions, no mineralization of 14C-rimsulfuron into 14C-CO2 occurred and under biotic ones it was very low reaching 0.75% of the applied 14C-rimsulfuron after 246 days of incubation. The analysis of data showed that a three-half order model provided the best fit for the mineralization curve. Extractable 14C-residues decreased over time to 70-80% of the applied 14C-rimsulfuron at the end of the incubation. After 246 days of incubation, non extractable residues (NER) accounted for up to 24.7% of the applied 14C-rimsulfuron and were distributed according to organic carbon in soil size fractions, suggesting a progressive incorporation process of NER to soil humus.

A model for the formation and degradation of bound residues of the herbicide 14C-isoproturon in soil.

The humic monomer catechol was reacted with 14C-isoproturon and some of its metabolites, including 14C-4-isopropylaniline, in aqueous solution under a stream of oxygen. Only in the case of 14C-4-isopropylaniline, incorporation in oligomers, in fulvic acid-like polymers, and in humic acid-like polymers was observed. The main oligomer was identified by mass spectrometry as 4,5-bis-(4-isopropylphenylamino)-3,5-cyclohexadiene-1,2-dione. Oligomers and polymers containing bound 14C-4-isopropylaniline were subjected to biodegradation studies in a loamy agricultural soil during 55 days by quantifying 14CO2 evolved. In all cases, significant mineralization rates could be determined, which, however, were much smaller than those of free 14C-4-isoproturon and free 14C-4-isopropylaniline in the same soil.

Consequências do cultivo e do pousio sobre a matéria orgânica do solo sob floresta natural na Amazônia Oriental

In an area of yellow podzolic latossol, five differents conditions of soil use, were compared: natural forest, recently, one and five years of annual crops, and three years of fallow following two years of annual crops. The organic matter was characterized by grain-size fractionation method. In the soils after burring and one year of cropping, plant residues in the litter and 0-15 em layer werw much less abundant than under the natural vegetation. Such evolution continued up to five years of cultivation, but was cultivation, but was reserved when the soil was abondoned and fallow developed. Carbon of the very fine fractiones (less than 50 m), which concentrates the clay-humic complex material, remained almost unchanged. Chemical fractionation of this material shewed that under annual crops the humus composition changed, and the amount of fulvic acids, principally those soluble in sodium hydroxide, increased, contrary to the humic acid. This results suggest that hamus incorporated non-polymeric material from the crops, and that a de-polymerisation process of the pre-existing clay-humic complex material ocuured. This process stopped under fallow and the humus then returned to its initial equilibrium.

São comparadas num mesmo latossolo podzólico amarelo de textura média, situações sob vegetação natural, depois da queimada, após um e cinco anos de cultivo e após três anos de pousio consecutivos a dois anos de cultivo. Depois da queimada e um ano de cultivo, há uma forte diminuição de todos os resíduos vegetais. Esta diminuição foi quantidade na liteira e na camada 0-15 cm do solo através do fracionamento granulométrico da matéria orgânica. Esta evolução continua durante os cinco anos de cultivo e se inverte quando o solo é abandonado à capoeira após dois anos de cultivo. O carbono da fração 0-50 m, que concentra o complexo argilo-húmico, não sofre grandes variações quantitativas. O fracionamento químico desta fração mostra que, sob cultivo, o húmus é caracterizado por um equilíbrio, comportando mais ácidos fúlvicos, principalmente os extraídos pelo hidróxido de sódio, e menos ácidos húmicos do que no ecossistema natural. Isto indica que o húmus incorpora frações jovens pouco polimerizadas e que há, ao mesmo tempo, despolimerização das frações ligadas às argilas. O equilíbrio inicial se restabelece quando, depois de um curto período de cultivo, se deixa o solo em pousio.