Long-chain omega-3 fatty acids regulate bovine whole-body protein metabolism by promoting muscle insulin signalling to the Akt–mTOR–S6K1 pathway and insulin sensitivity

The ability of the skeletal musculature to use amino acids to build or renew constitutive proteins is gradually lost with age and this is partly due to a decline in skeletal muscle insulin sensitivity. Since long-chain omega-3 polyunsaturated fatty acids (LC n –3PUFA) from fish oil are known to improve insulin-mediated glucose metabolism in insulin-resistant states, their potential role in regulating insulin-mediated protein metabolism was investigated in this study. Experimental data are based on a switchback design composed of three 5 week experimental periods using six growing steers to compare the effect of a continuous abomasal infusion of LC n –3PUFA-rich menhaden oil with an iso-energetic control oil mixture. Clamp and insulin signalling observations were combined with additional data from a second cohort of six steers. We found that enteral LC n –3PUFA potentiate insulin action by increasing the insulin-stimulated whole-body disposal of amino acids from 152 to 308 μmol kg⁻¹ h⁻¹ ( P = 0.006). The study further showed that in the fed steady-state, chronic adaptation to LC n –3PUFA induces greater activation ( P < 0.05) of the Akt–mTOR–S6K1 signalling pathway. Simultaneously, whole-body total flux of phenylalanine was reduced from 87 to 67 μmol kg⁻¹ h⁻¹ ( P = 0.04) and oxidative metabolism was decreased ( P = 0.05). We conclude that chronic feeding of menhaden oil provides a novel nutritional mean to enhance insulin-sensitive aspects of protein metabolism.     

Functional proteomics mapping of a human signaling pathway

Access to the human genome facilitates extensive functional proteomics studies. Here, we present an integrated approach combining large-scale protein interaction mapping, exploration of the interaction network, and cellular functional assays performed on newly identified proteins involved in a human signaling pathway. As a proof of principle, we studied the Smad signaling system, which is regulated by members of the transforming growth factor beta (TGFbeta) superfamily. We used two-hybrid screening to map Smad signaling protein-protein interactions and to establish a network of 755 interactions, involving 591 proteins, 179 of which were poorly or not annotated. The exploration of such complex interaction databases is improved by the use of PIMRider, a dedicated navigation tool accessible through the Web. The biological meaning of this network is illustrated by the presence of 18 known Smad-associated proteins. Functional assays performed in mammalian cells including siRNA knock-down experiments identified eight novel proteins involved in Smad signaling, thus validating this integrated functional proteomics approach.     

Deposition and passage of transthyretin through the blood-nerve barrier in recipients of familial amyloid polyneuropathy livers

Familial amyloid polyneuropathy (FAP) is characterized by deposition of mutated transthyretin (TTR) in the peripheral nervous system. Prior to amyloid fibrils, nonfibrillar TTR aggregates are deposited inducing oxidative stress with increased nitration (3-NT). As the major source of TTR is the liver, liver transplantation (LT) is used to halt FAP. Given the shortage of liver donors, domino LT (DLT) using FAP livers is performed. The correlation between TTR deposition in the skin and nerve was tested in biopsies from normal individuals, asymptomatic carriers (FAP 0) and FAP patients; in FAP 0, nonfibrillar TTR was observed both in the skin and nerve in the same individuals; in patients, amyloid was detected in both tissues. The occurrence of amyloidosis in recipients of FAP livers was evaluated 1-7 years after DLT: TTR deposition occurred in the skin 3 years after transplantation either as amyloid or aggregates; in one of the recipients, fibrillar TTR was present in the epineurium 6 years after DLT. Deposits were scarce and 3-NT immunostaining was irrelevant. Nerve biopsies from DLT recipients had no FAP-related neuropathy. Our findings suggest that TTR amyloid formation occurs faster than predicted and that TTR of liver origin can cross the blood-nerve barrier. Recipients of FAP livers should be under surveillance for TTR deposition and tissue damage.     

Escape efficiency of diploid and polyploid frogs: A comparison ofOdontophrynus cultripes (2n=22) andO. americanus (4n=44)

The escape efficiency of two closely related species of frogs,Odontophrynus cultripes(2n=22) and the tetraploidO. americanus (4n=44), were compared in a shuttle box and under simulated naturalistic conditions.O. americanus was generally superior toO. cultripes, and females tended to outperform males within both species. The relative inefficiency ofO. cultripes escape behavior was examined in light of the animals' having an elaborate, passive defense mechanism in the form of well-marked venom glands. Escape efficiency was highly variable in both species. Possessing twice the amount of DNA, the tetraploid behavioral variation was paradoxically less than that of the diploid, but compatible with what has been found for morphological characters in other organisms.This research was carried out at the Instituto Butantan with the support of ongoing grants from the Brazilian CNPq, FAPESP, FEDIB, and PNUD while the first author was a visiting professor in the Departamento de Genética Humana, Instituto de Biociências, Universidade de São Paulo, under the auspices of the Programa Multinacional de Genética, Organization of American States.     

In vivo autofluorescence imaging of early cancers in the human tracheobronchial tree with a spectrally optimized system

The changes in the autofluorescence characteristics of the bronchial tissue is of crucial interest as a cancer diagnostic tool. Evidence exists that this native fluorescence or autofluorescence of bronchial tissues changes when they turn dysplastic and to carcinoma in situ. There is good agreement that the lesions display a decrease of autofluorescence in the green region of the spectrum under illumination with violet-light, and a relative increase in the red region of the spectrum is often reported. Imaging devices rely on this principle to detect early cancerous lesions in the bronchi. Based on a spectroscopic study, an industrial imaging prototype is developed to detect early cancerous lesions in collaboration with the firm Richard Wolf Endoskope GmbH, Germany. A preliminary clinical trial involving 20 patients with this spectrally optimized system shows that the autofluorescence can help to detect most lesions that would otherwise have remained invisible to an experienced endoscopist under white light illumination. A systematic off line analysis of the autofluorescence images pointed out that real-time decisional functions can be defined to reduce the number of false positive results. Using this method, a positive predictive value (PPV) of 75% is reached using autofluorescence only. Moreover, a PPV of 100% is obtained, when combining the white light (WL) mode and the autofluorescence (AF) mode, at the applied conditions. Furthermore, the sensitivity is estimated to be twice higher in the AF mode than in WL mode.     

Characterization of wild-type recombinant Bet v 1a as a candidate vaccine against birch pollen allergy

BACKGROUND: We describe the production in Escherichia coli as a recombinant protein of clinical grade wild-type Bet v 1a (rBet v 1a), to be used as a candidate vaccine against birch pollen allergy. METHODS: This recombinant protein was purified by hydrophobic interaction and ion exchange chromatography and characterized by SDS-PAGE, immunoprint and circular dichroism in parallel with natural Bet v 1 (nBet v 1) purified from a birch pollen extract. We also compared rBet v 1 and nBet v 1 for their capacity to induce histamine release from basophils and to stimulate T lymphocyte proliferation. RESULTS: rBet v 1a appears in SDS-PAGE as an 18-kDa monomeric protein, whereas purified nBet v 1 comprises a mixture of isoforms (resolving as three distinct bands and six spots after 1-dimensional and 2-dimensional electrophoresis, respectively). Both recombinant and natural purified Bet v 1 molecules are recognized by IgE from birch pollen-allergic patients as well as anti-Bet v 1 murine monoclonal antibodies, suggesting that the recombinant protein is correctly folded in a native configuration. Circular dichroism analysis confirmed that the two Bet v 1 molecules exhibit similar 3-dimensional structures, even if rBet v 1a appears more compact and stable in thermodenaturation/renaturation experiments. Both rBet v 1 and nBet v 1 induce the degranulation of sensitized basophils and proliferation of Bet v 1-specific T lymphocytes in a similar manner. CONCLUSIONS: On the basis of these structural and biological properties, rBet v 1a is a valid candidate vaccine against birch pollen allergy, currently evaluated in humans.     

<Emphasis Type="Italic">Blastocystis hominis</Emphasis>: neutral red supravital staining and its application to in vitro drug sensitivity testing

A simple in vitro drug sensitivity testing system for Blastocystis hominis clinical isolates was developed. Application of supravital staining by neutral red allowed quantitative viability assessment. Four xenic cultures, isolated from human sources, were grown in modified monophasic Robinson's medium and tested for sensitivities to nine available drugs. Assessment was done using the cell-count method from air-dried preparations after supravital staining with neutral red. Also, the light absorbence method was evaluated. Trimethoprim, metronidazole, quinacrine, tetracycline, paromomycin, and two new antiprotozoal drugs, nitazoxanide and deacetyl-nitazoxanide, showed cytostatic or cytocidal effects on all or some Blastocystis isolates. Chloroquine and sulphamethoxazole did not demonstrate any effect at the concentrations studied. Received: 20 November 1998 / Received last revision: 27 December 1999 / Accepted: 29 December 1999     

Intégration de connaissances et modélisation en imagerie médicale

Image processing has considerably evolved since the first works, forty years ago. The currently observable tendency is that the methods are more and more mathematically-based with an objective of generalization of the concepts for high-level image interpretation of images. Nevertheless, these methods, despite their generic formulation, are always specialized on image modality and research context in order to obtain better and more reliable results. This specialization is carried out by integrating knowledge. In medical imaging, the studied objects are anatomical and functional structures associated with a corpus of medical knowledge which can be advantageously exploited in the image processing techniques.     

Early steps of a thymic tumor in SV40 transgenic mice: hyperplasia of medullary epithelial cells and increased mature thymocyte numbers disturb thymic export

Bone marrow progenitors migrate to the thymus, where they proliferate and differentiate into immunologically competent T cells. In this report we show that mice transgenic for SV40 T and t antigens under the control of the L-pyruvate kinase promoter develop, in a first step, thymic hyperplasia of both thymocytes and epithelial cells. Morphological studies (histology, immunohistolabeling and electron microscopy) revealed modifications of the thymic microenvironment and gradual expansion of medullary epithelial cells in 1 month-old mice, taking over the cortical region. Then, a thymic carcinoma develops. Two-color labeling of frozen sections identified the transgene in medullary epithelial cells. Flow cytometry analysis demonstrated a marked increase in mature CD4+ and CD8+ thymocytes in adult mice (39 +/- 10 x 10(6) in transgenic mice and 12 +/- 5 x 10(6) in age-matched controls). Furthermore, thymocyte export was disturbed.