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The design of enzymatic routes for the production of biosourced copolymers represents an attractive alternative to chemical synthesis from fossil carbon. In this paper, we explore the potential of glycosynthesizing enzymes to produce novel block copolymers composed of various covalently-linked α-glucans with contrasting structures and physicochemical properties. To this end, various glucansucrases able to synthesize α-glucans with different types of α-osidic bonds from sucrose were tested for their ability to elongate oligosaccharide and polysaccharide acceptors with different structures from the native polymer synthesized by each enzyme. We showed that two enzymes – namely, the alternansucrase from Leuconostoc mesenteroides NRRL B-1355 (specific for α(1 → 6)/α(1 → 3)-linked alternan synthesis) and the dextransucrase DSR-MΔ1 from Leuconostoc citreum NRRL B-1299 (specific for α(1 → 6)-linked dextran formation) – were able to elongate α(1 → 4)-linked amylose and α(1 → 6)/α(1 → 3)-linked alternan respectively. Carrying out stepwise acceptor reactions, and after optimization of the acceptor size and donor/acceptor ratio, two types of diblock copolymers were synthesized – a dextran-b-alternan and an alternan-b-amylose – as well as the triblock copolymer dextran-b-alternan-b-amylose. Their structural characterization, performed by combining chromatographic, NMR and permethylation analyses, showed that the copolymer polymerization degree ranged from 29 to 170, which is the highest degree of polymerization ever reported for an enzymatically synthesized polysaccharide-based copolymer. The addition of dextran and alternan blocks to amylose resulted in conformational modifications and related flexibility changes, as demonstrated by small angle X-ray scattering.

Resume Introduction Les instillations endovesicales de BCG constituent le traitement de reference des TVNIM de risque intermediaire et eleve. Un traitement d’entretien est fortement recommande, neanmoins la frequence des effets secondaire responsable de l’arret du traitement d’entretien chez plus de quatre patients sur cinq avant la troisieme annee pourrait justifier une diminution voire un espacement des instillations. L’objectif de l’etude URO-BCG-4 etait l’evaluation d’un nouveau schema d’entretien par instillations endovesicales de BCG associant une diminution de dose (un tiers de dose) mais aussi une diminution du nombre d’instillation par cycle (deux ou trois). Patients et methodes Etude multicentrique du CCAFU (12 centres hospitalo-universitaires), randomisee, prospective, comparant le schema de reference de BCG-therapie d’entretien au un tiers de dose usuelle (groupe I) a un schema associant un tiers de dose et diminution du nombre d’instillations par cycle (deux au lieu de trois) (groupe II). Nous presentons les resultats preliminaires a un an de ce programme hospitalier de recherche clinique (2003-081 CHU de Rouen Promoteur). Resultats Le taux de recidive tumorale etait respectivement de 9 et 7 % ( p = 0,678) dans les groupes I et II. Le taux de progression tumorale etait de 3 et 2,8 % dans les groupes I et II ( p = 1). La tolerance des instillations endovesicales de BCG evaluee selon la classification OMS (Geneve 1979) etait comparable dans les deux groupes. Conclusion La diminution de la dose de BCG (un tiers de dose) et la modification du nombre et du rythme des instillations n’a pas modifie le taux de survie sans recidive tumorale. La toxicite des instillations endovesicales de BCG etait identique dans les deux groupes. L’utilisation de la classification OMS a montre ses limites dans l’etude des effets secondaires du BCG car trop complexe et souvent non exhaustive. Le taux de progression musculaire etait comparable dans les deux groupes neanmoins un recul clinique plus important est necessaire.

Féral C, Le Gall S, Leymarie P. Angiotensin II modulates steroidogenesis in granulosa and theca in the rabbit ovary: its possible involvement in atresia. Eur J Endocrinol 1995;133:747–53. ISSN 0804–4643 Accumulating evidence has shown the ovary of mammals to contain an intrinsic renin–angiotensin system that has been ascribed an autocrine-paracrine role. The present study in the female rabbit ovary investigated the putative in vitro action of angiotensin II (A II) on basal and gonadotropin-induced steroidogenesis. Ovarian follicles from immature female rabbits treated with pregnant mare's serum gonadotropin (PMSG) were dissected out and a complete separation of the theca interna from the granulosa layer was performed, to demonstrate that A II affects separately the two individual cellular components of the follicular wall. We could show that theca is a source of estradiol whose production under human chorionic gonadotropin (hCG) stimulation was reduced by A II. At the same time, A II increased the in vitro hCG-stimulated secretion of testosterone by theca. In granulosa, A II decreased hCG-stimulated aromatization of androstenedione to estradiol but did not alter the release of hCG-stimulated progesterone production. These results suggest that A II could induce locally an increase in follicular fluid androgen/estrogen ratio and possibly participate in causing atresia. C. Féral, EP CNRS 009, Laboratoire de Biochimie, CHU Caen, F-14032 Caen Cedex, France

Background HIV-specific CD8 T cells responses rely on the recognition of peptide-MHC-I complexes by cognate T cell receptors. HIV-derived MHC-I epitopes result from the degradation of viral proteins by the cellular processing machinery including proteasomes and aminopeptidases. Interferon gamma changes proteasome composition and peptidase activities. We hypothesize that HIV infection might affect the expression or activities of the antigen processing machinery, either through a direct effect of the virus or indirectly through cellular activation or from the release of cytokines by surrounding infected cells.

Background Whether HIV-infectable subsets, such as CD4 T cells, monocytes, macrophages and dendritic cells (DCs), have equivalent capacity to produce and present MHC-I restricted epitopes to HIV-specific CD8 T cells is unknown. MHC-I epitopes are processed by an intracellular degradation pathway involving multiple proteases. In this study we analyzed the effect of toll-like receptor (TLR) agonist-mediated maturation on the processing and presentation of HIV antigens in monocyte-derived DCs and macrophages.

The circulating reninangiotensin system (RAS) participates in the regulation of blood pressure and electrolyte metabolism. Renin, a proteolytic enzyme, synthesized in the kidney from its biological precursor, prorenin, cleaves its substrate angiotensinogen in the blood to form the active octapeptide, angiotensin II (AII). All the RAS components are present in the reproductive system of mammals. During pregnancy, the level of prorenin increases in the plasma. The ovary is the source of this prorenin during early pregnancy and maternal decidua later on. During the menstrual cycle, the thecal of preovulatory follicles synthesize prorenin, renin and AII. Thecal renin synthesis is controlled by LH/hCG as demonstrated in vivo and in vitro in the rabbit. Ovarian renin seems to be identical to kidney renin. Prorenin appears to be the major secretory product rather than renin, which remains intracellular. AT2-type angiotensin II-receptors are expressed in the rat on follicular granulosa cells and could be down-regulated by FSH. The bovine thecal cells also express AT2-receptors, up-regulated by LH. These data are consistent with an autrocrine or paracrine role for ovarian RAS. It has been implicated in neovascularization of the follicle and regulation of steroidogenesis by increasing the androgen/estrogen ratio, an index of follicular atresia.

Hypermutation is an important mechanism used by different Salmonella enterica subspecies enterica to regulate genetic stability in adaptation to changing environments, including antimicrobial treatments and industrial processes. Strong hypermutator strains generally contain a mutation in genes of the methyl mismatch repair (MMR) system and have mutation frequencies up to 1000-fold higher than wild type strains. The objectives of this study were to determine the distribution of mutation frequencies from a collection of 209 Salmonella strains, to genetically characterize a strong mutator, and to study MMR mutated protein-DNA binding interactions. Only one strain of S. Heidelberg was determined to have a hypermutator phenotype by virtue of its high mutation rate. Sequencing of genes of the MMR system showed a 12bp deletion in the mutS gene was present. The MMR mutated protein-DNA binding interactions were studied by bioanalysis, using the available crystal structure of a similar MutS protein from Escherichia coli. This analysis showed the small deletion in the Salmonella MutS was localized within the core domain. A retardation assay with MutS from hypermutable and wild type strains showed this mutation has no effect on MutS DNA binding. A better understanding of the genetic mechanisms of hypermutation will help to anticipate the behavior of hypermutator strains in various conditions.

The origin of ovarian renin and its regulation by hCG were investigated in rabbit periovulatory follicles and cultured preovulatory follicular cells. Intracellular content of renin in thecal cells was 8-fold greater than of granulosa cells. In vivo, administration of hCG increased intracellular content of renin in thecal but not granulosa cells. Similar results were obtained for cultured follicular cells, from which renin was partly released into the medium. In vitro, hCG increased intracellular renin content of thecal but not granulosa cells, without obvious effect on release. Approximately 95% of ovarian renin was inactive, but could be activated by trypsin. Thecal renin was antagonized in vitro by renin antiserum, indicating a specific renin activity. Our study establishes in the rabbit the thecal cell origin of ovarian inactive renin and demonstrates hCG regulation of its synthesis.

Background Epitopes displayed by MHC-I come from the multistep degradation of proteins by intracellular peptidases such as proteasome and aminopeptidases or cathepsins in the exogenous pathway. We hypothesize that due to structural homologies HIV protease inhibitors (PIs) used in antiretroviral therapies may affect activities of cellular peptidases involved in epitope processing and may affect epitope presentation to immune cells. Methods Using a fluorogenic assay the effect of 5 HIV-1 PIs (Ritonavir, Saquinavir, Nelfinavir, Indinavir, Atazanavir) on proteasome, aminopeptidase and cathepsin activities was tested in PBMCs from at least 6 healthy donors. Using PBMC cytosol as a source of peptidases and HPLC and mass spectrometry to define and quantify the degradation products, the effect of HIV PIs on HIV peptide processing kinetics and HIV epitope half-life was assessed. Finally we assessed the impact of PIs on the endogenous processing and presentation of epitopes by infected cells to CD8 T cells using a fluorescence-based cytotoxicity assay. Results HIV PIs variably altered proteasome, post-proteasomal aminopeptidases and cathepsin activities. Depending on the PI, some activities were inhibited (from 1.1 to 5 folds, p

Background HIV-1 Nef is a multifunctional protein frequently targeted by host CD8+ T cell responses in early and chronic phases of HIV-1 infection. In vivo reversions of CTL escape mutations within Nef have been reported, suggesting a possible impact of immune-selected mutations in Nef on viral fitness. The goal of this work was to determine whether CD8+ T cell selected mutations in regions outside of Gag, such as in Nef, also impair viral replication and may thus contribute to early immune control of HIV-1.

Les virus sont des ennemis des systemes immunitaires dans une guerre tres ancienne faite d'adaptations constantes aux armes et aux strategies de l'adversaire. Pour echapper a la detection et a la destruction par leur hote, les virus ont mis en place des mecanismes varies. Leur comprehension accroit nos connaissances sur les fonctionnements des systemes immunitaires et des virus. Les strategies mises en oeuvre par le virus de l'immunodeficience humaine (VIH) concernent notamment un processus-cle de la reponse immunitaire qui est la presentation antigenique par les molecules du complexe majeur d'histocompatibilite (CMH) aux lymphocytes T. Les cellules dendritiques sont des cellules specialisees dans la presentation antigenique possedant la propriete de mettre en route des reponses immunitaires. Dans l'infection par le VIH, les cellules dendritiques sont essentielles au virus pour sa penetration dans l'organisme, pour sa production et sa dissemination et, probablement, dans la pathogenie du SIDA.

In the Calyptraeidae, which are protandric hermaphrodite molluscs, the external male genital tract and the gonad develop synchronously. The penis appears in young animals but regresses at the time of sex inversion. Experiments carried out in Crepidula fornicata, using the organ culture technique, enabled us to prove that involution of the penis in vitro is not the result of an intrinsic cycle of this organ. Hormonal exposure which had occurred in vivo was sufficient to condition further development of the isolated penis in vitro. A pedal morphogenetic factor and a pleural regressive factor are responsible for the differentiation of the male genital tract. The programming depends upon a balance of these neuroendocrine factors in the animal that provided the explant. Notwithstanding its regression in vitro, the penis of Crepidula represents an organ unable to evolve by itself, inasmuch as its development is directly dependent on the neuroendocrine system.

The innervation of the endocrine optic gland of Sepia, which controls sexual maturation, was studied by immunocytochemistry. Anti-FMRFamide (Phe-Met-Arg-Phe-NH2) serum revealed immunoreactive neurons in the olfactory and basal-dorsal lobe of the supra-esophageal brain mass. The axons of these neurons form a network from which fibers run to the optic gland. The fibers form many varicosities on the glandular cells, indicating synaptic innervation. Apparently, the two brain lobes containing the immunopositive cells function as a unit where visual and olfactory cues are integrated to regulate the endocrine activity of the optic gland.