Nuclear scaffold proteins are differently sensitive to stabilizing treatment by heat or Cu++.

The distribution of three nuclear scaffold proteins (of which one is a component of a particular class of nuclear bodies) has been studied in intact K562 human erythroleukemia cells, isolated nuclei, and nuclear scaffolds. Nuclear scaffolds were obtained by extraction with the ionic detergent lithium diidosalicylate (LIS), using nuclei prepared in the absence of divalent cations (metal-depleted nuclei) and stabilized either by a brief heat exposure (20 min at 37C or 42C) or by Cu++ ions at 0C. Proteins were visualized by in situ immunocytochemistry and confocal microscopy. Only a 160-kD nuclear scaffold protein was unaffected by all the stabilization procedures performed on isolated nuclei. However, LIS extraction and scaffold preparation procedures markedly modified the distribution of the polypeptide seen in intact cells, unless stabilization had been performed by Cu++. In isolated nuclei, only Cu++ treatment preserved the original distribution of the two other antigens (M(r), 125 and 126 kD), whereas in heat-stabilized nuclei we detected dramatic changes. In nuclear scaffolds reacted with antibodies to 125 and 126-kD proteins, the fluorescent pattern was always disarranged regardless of the stabilization procedure. These results, obtained with nuclei prepared in the absence of Mg+2 ions, indicate that heat treatment per se can induce changes in the distribution of nuclear proteins, at variance with previous suggestions. Nevertheless, each of the proteins we have studied behaves in a different way, possibly because of its specific association with the nuclear scaffold.

Control of introduced species using Trojan sex chromosomes.

To control introduced exotic species that have predominantly genetic, but environmentally reversible, sex determination (e.g. many species of fish), Gutierrez and Teem recently modeled the use of carriers of Trojan Y chromosomes--individuals who are phenotypically sex reversed from their genotype. Repeated introduction of YY females into wild populations should produce extreme male-biased sex ratios and eventual elimination of XX females, thus leading to population extinction. Analogous dynamics are expected in systems in which sex determination is influenced by one or a few major genes on autosomes.

Efforts to standardise Ki-67 counting in breast cancer. A pilot study of the Swiss Working Group of Gyneco- and Breast Pathologists

Goals: Adjuvant chemotherapy decisions in breast cancer are increasing based on the pathologist's assessment of the proliferation fraction in the tumor. Yet, how good and how reproducible are we pathologists at providing reliable Ki-67 readings on breast carcinomas. Exactly how to count and in which areas to count within a tumor remains inadequately standardized. The Swiss Working Group of Gyneco- and Breast Pathologists has tried to appreciate this dilemma and to propose ways to obtain more reproducible results.Methods: In a first phase, 5 pathologists evaluated Ki67 counts in 10 breast cancers by exact counting (500 cells) and by eyeballing. Pathologists were free to select the region in which Ki67 was evaluated. In a second phase 16 pathologists evaluated Ki-67 counts in 3 breast cancers also by exact counting and eyeballing, but in predefined fields of interest. In both phases, Ki67 was assessed in centrally immunostained slides (ZH) and on slides immunostained in the 11 participating laboratories. In a third phase, these same 16 pathologists were once again asked to read the 3 cases from phase 2, plus three new cases, and this time exact guidelines were provided as to what exactly is considered a Ki-67 positive nucleus.Results: Discordance of Ki67 assessment was due to each of the following 4 factors: (i) pathologists' divergent definitions of what counts as a positive nucleus (ii) the mode of assessment (counting vs. eyeballing), (iii) immunostaining technique/protocol/antibody, and (iv) the selection of the area in which to count.Conclusion: Providing guidelines as to where to count (representative field in the tumor periphery and omitting hot spots) and what nuclei to count (even faintly immunostained nuclei count as positive) reduces the discordance rates of Ki67 readings between pathologists. Laboratory technique is only of minor importance (even over a large antibody dilution range), and counting nuclei does not improve accuracy, but rather aggravates deviations from the group mean values.Disclosure of Interest: None Declared

Ultrastructural and morphometrical analyses of the brown adipocyte nucleus in a hibernating dormouse.

The fine morphology, size, and perichromatin granule frequency were analysed in brown adipocyte nuclei from hibernating, arousing, and euthermic dormice, Muscardinus avellanarius. Unusual nuclear structural constituents such as nuclear amorphous bodies, coiled body-like constituents and bundles of nucleoplasmic filaments were described as typical of hibernating nuclei. Morphometrical findings showed significant difference in total nuclear and nucleolar size in the three physiological conditions investigated as well as decreasing frequency of perichromatin granules in nuclei of hibernating to arousing to euthermic animals. A possible involvement of these granules in the intranuclear transport or storage of pre-mRNA is discussed in the context of other experimental evidence.

Method for retinal gene repair in neonatal mouse.

Gene correction at the site of the mutation in the chromosome is the absolute way to really cure a genetic disease. The oligonucleotide (ODN)-mediated gene repair technology uses an ODN perfectly complementary to the genomic sequence except for a mismatch at the base that is mutated. The endogenous repair machinery of the targeted cell then mediates substitution of the desired base in the gene, resulting in a completely normal sequence. Theoretically, it avoids potential gene silencing or random integration associated with common viral gene augmentation approaches and allows an intact regulation of expression of the therapeutic protein. The eye is a particularly attractive target for gene repair because of its unique features (small organ, easily accessible, low diffusion into systemic circulation). Moreover therapeutic effects on visual impairment could be obtained with modest levels of repair. This chapter describes in details the optimized method to target active ODNs to the nuclei of photoreceptors in neonatal mouse using (1) an electric current application at the eye surface (saline transpalpebral iontophoresis), (2) combined with an intravitreous injection of ODNs, as well as the experimental methods for (3) the dissection of adult neural retinas, (4) their immuno-labelling, and (5) flat-mounting for direct observation of photoreceptor survival, a relevant criteria of treatment outcomes for retinal degeneration.

Genetic exchange in an arbuscular mycorrhizal fungus results in increased rice growth and altered mycorrhiza-specific gene transcription.

Arbuscular mycorrhizal fungi (AMF) are obligate symbionts with most terrestrial plants. They improve plant nutrition, particularly phosphate acquisition, and thus are able to improve plant growth. In exchange, the fungi obtain photosynthetically fixed carbon. AMF are coenocytic, meaning that many nuclei coexist in a common cytoplasm. Genetic exchange recently has been demonstrated in the AMF Glomus intraradices, allowing nuclei of different Glomus intraradices strains to mix. Such genetic exchange was shown previously to have negative effects on plant growth and to alter fungal colonization. However, no attempt was made to detect whether genetic exchange in AMF can alter plant gene expression and if this effect was time dependent. Here, we show that genetic exchange in AMF also can be beneficial for rice growth, and that symbiosis-specific gene transcription is altered by genetic exchange. Moreover, our results show that genetic exchange can change the dynamics of the colonization of the fungus in the plant. Our results demonstrate that the simple manipulation of the genetics of AMF can have important consequences for their symbiotic effects on plants such as rice, which is considered the most important crop in the world. Exploiting natural AMF genetic variation by generating novel AMF genotypes through genetic exchange is a potentially useful tool in the development of AMF inocula that are more beneficial for crop growth.

Novel nuclear ribonucleoprotein structural components in the dormouse adrenal cortex during hibernation.

Adrenocortical cell nuclei of the dormouse Muscardinus avellanarius were investigated by electron microscopic immunocytochemistry in hibernating, arousing and euthermic individuals. While the basic structural constituents of the cell nucleus did not significantly modify in the three groups, novel structural components were found in nuclei of hibernating dormice. Lattice-like bodies (LBs), clustered granules (CGs), fibrogranular material (FGM) and granules associated with bundles of nucleoplasmic fibrils (NF) all contained ribonucleoproteins (RNPs), as shown by labeling with anti-snRNP (small nuclear RNP), anti-m3G-capped RNA and anti-hnRNP (heterogeneous nuclear RNP) antibodies. Moreover, the FGM also showed immunoreactivity for the proliferation associated nuclear antigen (PANA) and the non-snRNP splicing factor SC-35. All these nuclear structural components disappeared early during arousal and were not found in euthermic animals. These novel RNP-containing structures, which have not been observed in other tissues investigated so far in the same animal model, could represent storage and/or processing sites for pre-mRNA during the extreme metabolic condition of hibernation, to be quickly released upon arousal. NFs, which had been sometimes found devoid of associated granules in nuclei of brown adipose tissue from hi-bernating dormice, were present in much higher amounts in adrenocortical cell nuclei; they do not contain RNPs and their role remains to be elucidated. The possible roles of these structures are discussed in the frame of current knowledge of morpho-functional relationships in the cell nucleus.

Distribution and anatomical localization of the glucose transporter 2 (GLUT2) in the adult rat brain--an immunohistochemical study.

The aim of this work was to study the distribution and cellular localization of GLUT2 in the rat brain by light and electron microscopic immunohistochemistry, whereas our ultrastructural observations will be reported in a second paper. Confirming previous results, we show that GLUT2-immunoreactive profiles are present throughout the brain, especially in the limbic areas and related nuclei, whereas they appear most concentrated in the ventral and medial regions close to the midline. Using cresyl violet counterstaining and double immunohistochemical staining for glial or neuronal markers (GFAp, CAII and NeuN), we show that two limited populations of oligodendrocytes and astrocytes cell bodies and processes are immunoreactive for GLUT2, whereas a cross-reaction with GLUT1 cannot be ruled out. In addition, we report that the nerve cell bodies clearly immunostained for GLUT2 were scarce (although numerous in the dentate gyrus granular layer in particular), whereas the periphery of numerous nerve cells appeared labeled for this transporter. The latter were clustered in the dorsal endopiriform nucleus and neighboring temporal and perirhinal cortex, in the dorsal amygdaloid region, and in the paraventricular and reuniens thalamic nuclei, whereas they were only a few in the hypothalamus. Moreover, a group of GLUT2-immunoreactive nerve cell bodies was localized in the dorsal medulla oblongata while some large multipolar nerve cell bodies peripherally labeled for GLUT2 were scattered in the caudal ventral reticular formation. This anatomical localization of GLUT2 appears characteristic and different from that reported for the neuronal transporter GLUT3 and GLUT4. Indeed, the possibility that GLUT2 may be localized in the sub-plasmalemnal region of neurones and/or in afferent nerve fibres remains to be confirmed by ultrastructural observations. Because of the neuronal localization of GLUT2, and of its distribution relatively similar to glucokinase, it may be hypothesized that this transporter is, at least partially, involved in cerebral glucose sensing.

Generic finite-size enhancement of pairing in mesoscopic Fermi systems

The finite-size-dependent enhancement of pairing in mesoscopic Fermi systems is studied under the assumption that the BCS approach is valid and that the two-body force is size independent. Different systems are investigated such as superconducting metallic grains and films as well as atomic nuclei. It is shown that the finite size enhancement of pairing in these systems is in part due to the presence of a surface which accounts quite well for the data of nuclei and explains a good fraction of the enhancement in Al grains.

Hadronic vacuum polarization and the Lamb shift

Recent improvements in the determination of the running of the fine-structure constant also allow an update of the hadronic vacuum-polarization contribution to the Lamb shift. We find a shift of -3.40(7) kHz to the 1S level of hydrogen. We also comment on the contribution of this effect to the determination by elastic electron scattering of the rms radii of nuclei.

Two and three nucleon induced photon absorption

We have investigated the mechanisms leading to two and three body photon absorption in nuclei. At photon energies around the pion production threshold we obtain a fraction of three body absorption of less than 10% of the total, contradicting previous theoretical claims that it dominates the absorption process. The strength of the three body channel grows smoothly with the photon energy reaching a maximum of about 60% of the total direct absorption at energies of the photon around 400 MeV.

Variational Wigner-Kirkwood approach to relativistic mean field theory

The recently developed variational Wigner-Kirkwood approach is extended to the relativistic mean field theory for finite nuclei. A numerical application to the calculation of the surface energy coefficient in semi-infinite nuclear matter is presented. The new method is contrasted with the standard density functional theory and the fully quantal approach.

Allergies alimentaires rares [Rare food allergies]

A limited number of foods explain the majority of food allergies. These allergies can be due to a weak allergenicity (garlic, onion, potato), or a weak (or increasing) exposure to emergent food allergens which can be imported (exotic fruits), or recently introduced (lupin, buckwheat, sesame, inulin) or modified by the industry (lysats, lecithins, traces of antibiotics, caseinates, molds, dust mite). Others are in relation with rarer cross-reactivity food allergy syndrome (Apiaceae-Compositae-mugwort syndrome, egg-bird syndrome, cat epithelium-pork meat syndrome). Others are rarely identified, because the food is masked (pepper, basilic). We illustrate rare cases of food allergy and discuss the diagnostic management which is based on a meticulous patient history. Un nombre restreint d'aliments explique la majorité des allergies alimentaires. Les allergies alimentaires rares sont dues à une faible allergénicité (ail, oignon, pomme de terre) ou à une exposition faible ou croissante à des aliments émergents, importés (fruits exotiques), introduits (lupin, sarrasin, sésame, inuline), ou modifiés par l'industrie (lysats, lécithines, traces d'antibiotiques, caséinates, moisissures, acariens). D'autres sont en relation avec des croisements d'allergènes rares (syndrome croisé ombellifères-composées-armoise, syndrome oeuf-oiseau, syndrome épithélium de chat-viande de porc). D'autres enfin sont rarement identifiées, car l'allergène est masqué (poivre, basilic). Nous décrivons des cas rares illustratifs et rappelons la démarche diagnostique qui s'appuie sur une anamnèse minutieuse.

Effects of new nonlinear couplings in relativistic effective field theory

We extend the relativistic mean field theory model of Sugahara and Toki by adding new couplings suggested by modern effective field theories. An improved set of parameters is developed with the goal to test the ability of the models based on effective field theory to describe the properties of finite nuclei and, at the same time, to be consistent with the trends of Dirac-Brueckner-Hartree-Fock calculations at densities away from the saturation region. We compare our calculations with other relativistic nuclear force parameters for various nuclear phenomena.