Far-Field Raman Imaging of Short-Wavelength Particle Plasmons on Gold Nanorods

The surface plasmon fields of gold nanorods with a diameter of 100 nm and lengths of 1–5 m are imaged by using far-field Raman scattering of methylene blue adsorbed on the rods. When optically exciting the nanorods under total internal reflection with wave vector and electric field vector orientations along the rod axis, the plasmon field intensity along this axis is observed to be periodically modulated. This modulation is attributable to a beating of the exciting light wave and the nanorod plasmon mode. The plasmon wavelength deduced from the beat frequency is 379 nm, which is considerably smaller than the exciting laser wavelength of 647 nm. In general, Raman imaging is shown to be a powerful technique to probe local plasmon fields using far-field spectroscopy.     

Proteomic analysis of anaplastic lymphoma cell lines: identification of potential tumour markers

Anaplastic large-cell lymphomas (ALCL) are high grade lymphomas of T or null phenotype often associated with the t(2;5) translocation leading to the expression of a chimeric protein consisting of the N-terminal portion of nucleophosmin (NPM) and the intracellular domain of the anaplastic lymphoma kinase (ALK). Although ALCL are recognized as distinct clinical, biological and cytogenetic entities, heterogeneities persist in this group of tumours, which exhibit a broad spectrum of morphological features. Particularly, the common type tumour consisting in large cells contrast with the small cell variant that is sometimes associated with a leukemic phase. The ALK-negative ALCL is often associated with a poor prognosis. Here, we investigated the proteome of these subtypes of tumours using patient-derived cell lines. We compared the proteome of the cytosolic fraction of NPM-ALK-positive versus NPM-ALK-negative cells on one hand, and the proteome of common cell type versus small cell variant on the other hand. The identification of a set of proteins differentially expressed in the subtypes of ALCL points to new diagnosis/prognosis markers. This study also provides interesting information on the molecular mechanisms responsible for the different subtypes of ALCL.     

Acetylcholinesterase genes within the Diptera: takeover and loss in true flies

It has recently been reported that the synaptic acetylcholinesterase (AChE) in mosquitoes is encoded by the ace-1 gene, distinct and divergent from the ace-2 gene, which performs this function in Drosophila. This is an unprecedented situation within the Diptera order because both ace genes derive from an old duplication and are present in most insects and arthropods. Nevertheless, Drosophila possesses only the ace-2 gene. Thus, a secondary loss occurred during the evolution of Diptera, implying a vital function switch from one gene (ace-1) to the other (ace-2). We sampled 78 species, representing 50 families (27% of the Dipteran families) spread over all major subdivisions of the Diptera, and looked for ace-1 and ace-2 by systematic PCR screening to determine which taxonomic groups within the Diptera have this gene change. We show that this loss probably extends to all true flies (or Cyclorrhapha), a large monophyletic group of the Diptera. We also show that ace-2 plays a non-detectable role in the synaptic AChE in a lower Diptera species, suggesting that it has non-synaptic functions. A relative molecular evolution rate test showed that the intensity of purifying selection on ace-2 sequences is constant across the Diptera, irrespective of the presence or absence of ace-1, confirming the evolutionary importance of non-synaptic functions for this gene. We discuss the evolutionary scenarios for the takeover of ace-2 and the loss of ace-1, taking into account our limited knowledge of non-synaptic functions of ace genes and some specific adaptations of true flies.     

Structure-activity relationships in platelet-activating factor. Part 13: synthesis and biological evaluation of piperazine derivatives with dual anti-PAF and anti-HIV-1 or pure antiretroviral activity

HIV-1 infection of the brain and PAF neurotoxicity are implicated in AIDS dementia complex. We previously reported that a trisubstituted piperazine derivative is able to diminish both HIV-1 replication in monocyte-derived macrophages and PAF-induced platelet aggregation. We report in this work new compounds obtained by modifying its piperazine substituents. The structure-activity relationship study shows that a better dual activity or even pure antiretroviral compounds can be obtained in this series.     

The Th2 lymphoproliferation developing in LatY136F mutant mice triggers polyclonal B cell activation and systemic autoimmunity

Lat(Y136F) knock-in mice harbor a point mutation in Tyr(136) of the linker for activation of T cells and show accumulation of Th2 effector cells and IgG1 and IgE hypergammaglobulinemia. B cell activation is not a direct effect of the mutation on B cells since in the absence of T cells, mutant B cells do not show an activated phenotype. After adoptive transfer of linker for activation of T cell mutant T cells into wild-type, T cell-deficient recipients, recipient B cells become activated. We show in vivo and in vitro that the Lat(Y136F) mutation promotes T cell-dependent B cell activation leading to germinal center, memory, and plasma cell formation even in an MHC class II-independent manner. All the plasma and memory B cell populations found in physiological T cell-dependent B cell responses are found. Characterization of the abundant plasmablasts found in secondary lymphoid organs of Lat(Y136F) mice revealed the presence of a previously uncharacterized CD93-expressing subpopulation, whose presence was confirmed in wild-type mice after immunization. In Lat(Y136F) mice, B cell activation was polyclonal and not Ag-driven because the increase in serum IgG1 and IgE concentrations involved Abs and autoantibodies with different specificities equally. Although the noncomplement-fixing IgG1 and IgE are the only isotypes significantly increased in Lat(Y136F) serum, we observed early-onset systemic autoimmunity with nephritis showing IgE autoantibody deposits and severe proteinuria. These results show that Th2 cells developing in Lat(Y136F) mice can trigger polyclonal B cell activation and thereby lead to systemic autoimmune disease.     

Impairment of dendritic cell functionality and steady-state number in obese mice

There is a finely tuned interplay between immune and neuroendocrine systems. Metabolic disturbances like obesity will have serious consequences on immunity both at the cellular and at the cytokine expression levels. Our in vivo results confirm the immune deficiency of ob/ob mice, leptin deficient and massively obese, characterized by a reduced Ag-specific T cell proliferation after keyhole limpet hemocyanin immunization. In this report, we show that dendritic cells (DCs), major APCs involved in T lymphocyte priming, are affected in obese mice. Both their function and their steady-state number are disturbed. We demonstrate that DCs from ob/ob mice are less potent in stimulation of allogenic T cells in vitro. This impaired functionality is not associated with altered expression of phenotypic markers but with the secretion of immunosuppressive cytokines such as TGF-beta. Moreover, we show increased in vivo steady-state number of epidermal DCs in ob/ob mice, which is not due to a migratory defect. The ob/ob mice are characterized by the absence of functional leptin, a key adipokine linking nutrition, metabolism, and immune functions. Interestingly, intradermal injection of leptin is able to restore epidermal DC number in obese mice. Thus, DCs might be directly sensitive to metabolic disturbances, providing a partial explanation of the immunodeficiency associated with obesity.     

Genotoxic effects of bitumen fumes in Big Blue transgenic rat lung

Road paving workers are exposed to bitumen fumes (CAS No. 8052-42-4), a complex mixture of volatile compounds and particles containing carcinogenic and non-carcinogenic polycyclic aromatic hydrocarbons. However, epidemiological and experimental animal studies failed to draw unambiguous conclusions concerning their toxicity. In order to gain better insights on their genotoxic potential, we used an experimental design able to generate bitumen fumes at road paving temperature (temperature: 170 degrees C, total particulate matter: 100mg/m3) and perform a nose-only exposure of Big Blue transgenic rodents 6h/day for five consecutive days. The mutagenic properties of bitumen fumes were determined by analyzing the mutation frequency and spectrum of the neutral reporter gene cII inserted into the rodent genome. We previously observed in mouse lung, that bitumen fumes did not induce an increase of cII mutants, a modification of the mutation spectrum, nor the formation of DNA adducts. Since DNA adducts were found in the lungs of rats exposed to asphalt fumes in similar conditions, we decided to carry out an analogous experiment with Big Blue rats. A DNA adduct was detected 3 and 30 days after the end of treatment suggesting that these genetic alterations were quite steady. Thirty days after exposure, the cII mutant frequency was similar in control and exposed rats. In addition, a slight but not significant modification of the mutation spectrum associated with an increase of G:C to T:A and A:T to C:G transversions was noticeable in the treated animals. Then, these data failed to demonstrate a pulmonary mutagenic potential for bitumen fumes generated at road paving temperature in our experimental conditions despite the presence of a DNA adduct. These results may provide information concerning the pulmonary mechanism of action of this aerosol and may contribute to the occupational health hazard assessment.     

Isolation and development of a molecular sex marker for <Emphasis Type="Italic">Bassiana duperreyi</Emphasis>, a lizard with XX/XY sex chromosomes and temperature-induced sex reversal

Sex determination in the endemic Australian lizard Bassiana duperreyi (Scincidae) is influenced by sex chromosomes and incubation temperature, challenging the traditional dichotomy in reptilian sex determination. Analysis of those interactions requires sex chromosome markers to identify temperature-induced sex reversal. Here, we report the isolation of Y chromosome DNA sequence from B. duperreyi using amplified fragment length polymorphism PCR, the conversion of that sequence to a single-locus assay, and its combination with a single-copy nuclear gene (C-mos) to form a duplex PCR test for chromosomal sex. The accuracy of the assay was tested on an independent panel of individuals with known phenotypic sex. When used on offspring from field nests, our test identified the likely occurrence of a low rate of natural sex reversal in this species. This work represents the first report of Y chromosome sequence from a reptile and one of the few reptile sex tests.     

The role of HGF on invasive properties and repopulation potential of human fetal hepatic progenitor cells

The success of hepatocyte transplantation has been limited by the low efficiency of transplanted cell integration into liver parenchyma. Human fetal hepatic progenitor cells (hepatoblasts) engraft more effectively than adult hepatocytes in mouse livers. However, the signals required for their integration are not yet fully understood. We investigated the role of HGF on the migration and invasive ability of human hepatic progenitors in vitro and in vivo.Hepatoblasts were isolated from the livers of human fetuses between 10 and 12 weeks of gestation. Their invasive ability was assessed in the presence or absence of HGF. These cells were also transplanted into immunodeficient mice and analyzed by immunohistochemistry.In contrast to TNF-alpha, HGF increased the motogenesis and invasiveness of hepatoblasts, but not of human adult hepatocytes, via phosphorylation of extracellular signal-regulated kinase (ERK) 1/2. The invasive ability of human hepatoblasts correlated with the expression and secretion of matrix metalloproteinases (MMPs). Hepatoblasts stimulated with HGF prior transplantation into newborn mice migrated from the portal area into the hepatic parenchyma.Conclusions: In contrast to adult hepatocytes, hepatoblasts display invasive ability that can be modulated by HGF in vitro and in vivo.     

Fine Mapping and Marker Development for the Crossability Gene SKr on Chromosome 5BS of Hexaploid Wheat (Triticum aestivum L.)

Most elite wheat varieties cannot be crossed with related species thereby restricting greatly the germplasm that can be used for alien introgression in breeding programs. Inhibition to crossability is controlled genetically and a number of QTL have been identified to date, including the major gene Kr1 on 5BL and SKr, a strong QTL affecting crossability between wheat and rye on chromosome 5BS. In this study, we used a recombinant SSD population originating from a cross between the poorly crossable cultivar Courtot (Ct) and the crossable line MP98 to characterize the major dominant effect of SKr and map the gene at the distal end of the chromosome near the 5B homeologous GSP locus. Colinearity with barley and rice was used to saturate the SKr region with new markers and establish orthologous relationships with a 54-kb region on rice chromosome 12. In total, five markers were mapped within a genetic interval of 0.3 cM and 400 kb of BAC contigs were established on both sides of the gene to lay the foundation for map-based cloning of SKr. Two SSR markers completely linked to SKr were used to evaluate a collection of crossable wheat progenies originating from primary triticale breeding programs. The results confirm the major effect of SKr on crossability and the usefulness of the two markers for the efficient introgression of crossability in elite wheat varieties.DURING domestication and selection of a number of important crop species, diversity has eroded resulting in increased vulnerability to biotic and abiotic stresses while also jeopardizing the potential for sustained genetic improvement of elite cultivars over the long term (Tanksley and McCouch 1997; Fu and Somers 2009). The reintroduction of the remarkable diversity present in the different gene pools into elite varieties through intra- and interspecific crosses (primary and secondary gene pools) and intergeneric crosses (tertiary gene pools) has been practiced for decades in cereals (Feuillet et al. 2008). Despite some highly significant successes, including the incorporation of dwarfing and disease-resistance genes that fueled the Green Revolution, introgression remains laborious and, for complex characters, largely unfulfilled. Wheat (Triticum aestivum L.) has been crossed with a wide range of related species from the Triticeae tribe (Jiang et al. 1994), such as Aegilops, Agropyron, Haynaldia, Secale, and Hordeum, which represent a reservoir of interesting alleles for improving wheat resistance to biotic (diseases, insects) and abiotic stresses (cold, salinity, and drought) as well as for quality traits such as grain protein content (Fedak 1985). Intergeneric crosses have resulted in the transfer of desirable rye (Secale cereale L.) characteristics into wheat (Florell 1931) with one of the best examples being the 1BL/1RS chromosomal translocation that provided novel race-specific resistance to rust diseases, improved adaptation and stress tolerance, superior aerial biomass, and higher kernel weight to wheat varieties (Zarco-Hernandez et al. 2005). However, most of the adapted wheat germplasm is not crossable with alien species thereby restricting the panel of lines that can be used for alien introgression in wheat breeding (Krolow 1970) or for the production of primary triticale, a man-made wheat–rye hybrid.Beginning in the early 1900s, researchers were producing experimental crosses between bread wheat, T. aestivum L. (2n = 6x = 42) as a recipient, and rye, S. cereale L. (2n =14) as the pollen donor (Backhouse 1916). Genetic studies conducted by Lein (1943) showed that dominant alleles of two genes, named Kr1 and Kr2, are responsible for the poor crossability between bread wheat and rye. Kr1 and Kr2 genes were localized roughly on chromosome 5B and 5A, respectively (Riley and Chapman 1967) and subsequently located more precisely on the long arms of these two chromosomes (Lange and Riley 1973; Sitch et al. 1985). Further studies indicated that the dominant alleles driving incompatibility of crossing wheat with rye act by actively inhibiting the production of intergeneric hybrids (Riley and Chapman 1967; Lange and Wojciechowska 1976; Jalani and Moss 1980, 1981; Cameron and Reger 1991 ). Other crossability genes, such as Kr3 on chromosome 5D (Krolow 1970) and Kr4 on chromosome 1A (Zheng et al. 1992), were identified later. Finally, a study elucidated that chromosome 1A, derived from the Chinese (Sichuan) tetraploid wheat T. turgidum L. cv. Ailanmai, carries a recessive allele for high crossability with rye (Liu et al. 1999). Genetic studies also indicated that Kr genes have different effects on wheat–rye crossability. For example, by testing the cultivars Chinese Spring (CS), Hope, and the substitution lines CS/Hope 5B and CS/Hope 5A, Riley and Chapman (1967) demonstrated that Kr1 has a stronger effect than Kr2, whereas Kr3 seemed weaker than the two other genes (Krolow 1970).To further explore the mechanisms controlling crossability in wheat, Snape et al. (1979) performed crosses between the wild barley Hordeum bulbosum and the wheat cultivars Chinese Spring and Hope as well as 21 substitution lines carrying individual chromosomes of Hope in the background of Chinese Spring. The results revealed that the Kr1 and Kr2 genes on chromosomes 5B and 5A that govern crossability between wheat and rye also are involved in controlling crossability between wheat and barley, although the percentage of crossability observed was significantly lower than with rye. Using crosses between Chinese Spring, Hope, and the entire series of substitution lines with the cultivated barley (H. vulgare L.) cv. Betzes, Fedak and Jui (1982) also suggested that the homeologous alleles of the Kr genes on chromosome group 5 of Chinese Spring (5A, 5B, and 5D) favor crossability with additive effects.In 1998, a new locus, named SKr, controlling crossability between wheat and rye was detected using a mapping population of 187 double haploid (DH) lines produced by anther culture from F₁ hybrids of a cross between the noncrossable (NC) French wheat cv. Courtot (Ct) and the Chinese crossable (C) cv. Chinese Spring (Tixier et al. 1998). SKr was identified as a major QTL located on the distal end of the short arm of chromosome 5B within a confidence interval ranging from 8.7 to 20.9 cM (Lamoureux et al. 2002). In this population, the effect of SKr was stronger (22.1% of heritability) than the one of a QTL identified on 5BL (supposedly Kr1, 5.5% of heritability), whereas no significant effect was detected on 5AL (for Kr2). Moreover, the results indicated a 95% crossability rate for cv. Chinese Spring and ∼10% for Courtot, suggesting that the Courtot genotype is Kr1Kr1/kr2kr2, whereas Chinese Spring would be kr1kr1/kr2kr2.Here, we report the construction of a high resolution genetic map at the SKr locus using a single seed descent (SSD) population derived from a cross between Courtot and a crossable DH line (MP98) that allowed us to assess the crossability phenotype as a single “Mendelian factor” and to demonstrate the major dominant effect of SKr on crossability. Synteny between wheat, barley, and rice was used to increase the density of markers and reduce the genetic interval around the SKr gene to 0.3 cM. BAC contigs from Chinese Spring were established with closely linked and cosegregating markers to lay the foundation for positional cloning of the gene. Finally, a SSR marker cosegregating with SKr was developed and its value for the exploitation of SKr in breeding was assessed in a collection of crossable lines used to produce primary triticale.