Incidental Encoding Strategies Did Not Improve Contextual Memory in Parkinson`s Disease Patients

Background. This study investigated the performance of patients with idiopathic Parkinson`s disease (PD) without dementia for incidental recognition memory and the effect of encoding strategies on contextual memory. Methods. The authors studied 21 patients with PD (ages 60-85, 12 women; Hoehn and Yahr I-III, Activities of Daily Living 70%-100%) and 22 healthy controls (ages 60-84, 18 women). Participants completed the vocabulary subtest of the Wechsler Adult Intelligence Scale and the Wisconsin Card Sorting Test (WCST). To assess the incidental recognition memory for item (object) and context (location of the object), participants of each group were assigned to 1 of 2 encoding conditions: (a) an incidental associative instruction to bind the object to its location or (b) a nonassociative, nonspecific instruction. Results. PD patients showed performance comparable to the control group`s on the vocabulary subtest and WCST. In contrast to controls, PD patients were unable to take advantage of the associative encoding instruction, which also had a deleterious effect on item recognition. Conclusion. This sample of participants with PD showed diminished item and context recognition memory and an impaired ability to use incidental memory encoding strategy, suggesting a compromised cognitive reserve. The fact that these alterations occurred in early stages of PD, and prior to more general cognitive alterations such as executive dysfunction, should be considered in the management of patients by using specific cognitive rehabilitation interventions.

Adaptive communication: languages with more non-native speakers tend to have fewer word forms

Explaining the diversity of languages across the world is one of the central aims of typological, historical, and evolutionary linguistics. We consider the effect of language contact-the number of non-native speakers a language has-on the way languages change and evolve. By analysing hundreds of languages within and across language families, regions, and text types, we show that languages with greater levels of contact typically employ fewer word forms to encode the same information content (a property we refer to as lexical diversity). Based on three types of statistical analyses, we demonstrate that this variance can in part be explained by the impact of non-native speakers on information encoding strategies. Finally, we argue that languages are information encoding systems shaped by the varying needs of their speakers. Language evolution and change should be modeled as the co-evolution of multiple intertwined adaptive systems: On one hand, the structure of human societies and human learning capabilities, and on the other, the structure of language.

Optical encoding of microbeads for gene screening: alternatives to microarrays

Rapid access to genetic information is central to the revolution presently occurring in the pharmaceutical industry, particularly In relation to novel drug target identification and drug development. Genetic variation, gene expression, gene function and gene structure are just some of the important research areas requiring efficient methods of DNA screening. Here, we highlight state-of-the-art techniques and devices for gene screening that promise cheaper and higher-throughput yields than currently achieved with DNA microarrays. We include an overview of existing and proposed bead-based strategies designed to dramatically increase the number of probes that can be interrogated in one assay. We focus, in particular, on the issue of encoding and/or decoding (bar-coding) large bead-based libraries for HTS.

Controlling gene expression in enterobacteriaceae: studies on sRNAs and strategies for synthetic biology

Dissertation presented to obtain the Master Degree in Molecular, Genetics and Biomedicine

Additives and Protein-DNA Combinations Modulate the Humoral Immune Response Elicited by a Hepatitis C Virus Core-encoding Plasmid in Mice

Humoral and cellular immune responses are currently induced against hepatitis C virus (HCV) core following vaccination with core-encoding plasmids. However, the anti-core antibody response is frequently weak or transient. In this paper, we evaluated the effect of different additives and DNA-protein combinations on the anti-core antibody response. BALB/c mice were intramuscularly injected with an expression plasmid (pIDKCo), encoding a C-terminal truncated variant of the HCV core protein, alone or combined with CaCl2, PEG 6000, Freund's adjuvant, sonicated calf thymus DNA and a recombinant core protein (Co.120). Mixture of pIDKCo with PEG 6000 and Freund's adjuvant accelerated the development of the anti-core Ab response. Combination with PEG 6000 also induced a bias to IgG2a subclass predominance among anti-core antibodies. The kinetics, IgG2a/IgG1 ratio and epitope specificity of the anti-core antibody response elicited by Co.120 alone or combined with pIDKCo was different regarding that induced by the pIDKCo alone. Our data indicate that the antibody response induced following DNA immunization can be modified by formulation strategies.

Cl gene cluster encoding several small nucleolar RNAs: a comparison amongst trypanosomatids

Small nucleolar RNAs (snoRNAs) are small non-coding RNAs that modify RNA molecules such as rRNA and snRNA by guiding 2'-O-ribose methylation (C/D box snoRNA family) and pseudouridylation reactions (H/ACA snoRNA family). H/ACA snoRNAs are also involved in trans-splicing in trypanosomatids. The aims of this work were to characterise the Cl gene cluster that encodes several snoRNAs in Trypanosoma rangeli and compare it with clusters from Trypanosoma cruzi, Trypanosoma brucei, Leishmania major, Leishmania infantum, Leishmania braziliensis and Leptomonas collosoma. The T. rangeli Cl gene cluster is an 801 base pair (bp) repeat sequence that encodes three C/D (Cl1, Cl2 and Cl4) and three H/ACA (Cl3, Cl5 and Cl6) snoRNAs. In contrast to T. brucei, the Cl3 and Cl5 homologues have not been annotated in the Leishmania or T. cruzi genome projects (http//:www.genedb.org). Of note, snoRNA transcribed regions have a high degree of sequence identity among all species and share gene synteny. Collectively, these findings suggest that the Cl cluster could constitute an interesting target for therapeutic (gene silencing) or diagnostic intervention strategies (PCR-derived tools).

Novel immunotherapeutic strategies against human papillomavirus type 16 (HPV 16) induced lesions and cervical cancer

RESUME Le cancer du col de l'utérus, deuxième cause de mort par cancer chez la femme, a pu être associé à une infection par plusieurs types de virus du Papillome Humain (HPV), et en particulier HPV 16. Les vaccins prophylactiques sont efficaces à prévenir le cancer du col utérin alors que les lésions de haut grade sont généralement traitées par ablation chirurgicale et par d'éventuels traitements additionnels. Les risques de récurrence liés aux ablations et le taux de mortalité (50%) lié au cancer, démontrent le besoin de développer des stratégies alternatives afin de cibler les lésions précancéreuses. A ce jour, les vaccins thérapeutiques ont démontré peu de résultats cliniques, contrastant avec les régressions de tumeurs ectopiques observées après vaccination dans des modèles murins avec tumeurs associées à HPV. L'induction de réponses immunitaires protectrices dans la muqueuse génitale semble être cruciale pour l'efficacité des vaccins thérapeutiques HPV et évaluer leur efficacité dans un modèle murin avec tumeurs-HPV génitales représente un pré-requis important avant de procéder à des études cliniques. Par conséquent, nous avons établi un modèle murin orthotopique où des tumeurs se développent dans (a muqueuse génitale après une instillation intra-vaginale (i.vag) de cellules tumorales exprimant les oncogènes E6/E7 d'HPV 16 et transduites par un vecteur lentiviral codant la luciferase afin de suivre le développement de ces tumeurs in vivo par imagerie. La caractérisation histologique a démontré que les tumeurs grandissaient dans l'épithélium vaginal et en accord avec leur localisation, des cellules Τ CD8 spécifiques à E7 induites par la tumeur n'étaient détectées que dans la muqueuse génitale et les ganglions drainants. Une infiltration de cellules Τ régulatrices a aussi été mise en évidence, empêchant la régression spontanée de ces tumeurs. Par conséquent, ce modèle devrait être plus adéquat pour tester des stratégies thérapeutiques, étant donné qu'il partage certaines similarités immunologiques avec les lésions génitales naturelles causées par HPV. Etant donné que les oncogènes E6 et E7 d'HPV sont nécessaires à la maintenance du phénotype cancéreux des cellules cervicales, elles représentent des antigènes cibles pour la vaccination thérapeutique. Nous avons démontré que des souris immunisées par voie sous-cutanée (s.c.) avec une dose d'un vaccin à base de polypeptide E7 d'HPV 16 et d'adjuvants, présentaient de nombreuses cellules Τ CD8 sécrétant de l'IFN-γ spécifiquement à E7 dans leurs organes lymphatiques mais également dans la muqueuse génitale. De plus, le manque de corrélation entre les réponses spécifiques mesurées dans la périphérie et dans la muqueuse génitale souligne la nécessité et l'importance de déterminer les réponses immunitaires localement là où les lésions dues à HPV se développent. Si une vaccination par voie muqueuse est plus propice à traiter/régresser des infections génitales/tumeurs que le voie parentérale est un sujet débattu. Nos données montrent que seule la voie s.c. était capable de régresser la quasi totalité des tumeurs génitales chez la souris bien que des réponses CD8 spécifiques à E7 similaires étaient mesurées dans la muqueuse génitale après des vaccinations intra-nasale et i.vag. Afin d'augmenter la réponse spécifique au vaccin dans la muqueuse génitale, des immunostimulants ont été administrés par voie i.vag après vaccination. Nous avons démontré qu'une application i.vag d'agonistes des Toll like receptors après une vaccination s.c. induisait de manière significative une augmentation des cellules Τ CD8 sécrétant de l'IFN-γ spécifiquement à E7 dans la muqueuse génitale. Plus précisément et concernant les CpG et Poly l:C, l'effet était probablement associé à une attraction locale des cellules Τ CD8 et deuxièmement dépendait respectivement des voies de signalisation TLR9 et TLR3/Mda5. Finalement, cette stratégie combinatoire a permis de régresser des grosses tumeurs génitales chez la souris, suggérant qu'une telle immunothérapie pourrait adéquatement traiter des lésions dues à HPV chez les femmes. SUMMARY Cervical cancer is the second leading cause of cancer mortality in women worldwide and results from an infection with a subset of Human Papillomavirus (HPV), HPV 16 representing the most prevalent type. The available prophylactic vaccines are an effective strategy to prevent cervical cancer while already established high grade lesions usually require surgical ablation of lesion with possible additional treatments. Recurrence risks linked to conventional ablations and the high mortality (50%) related to cervical cancer demonstrate the need for alternative strategies like immunotherapies to target pre¬cancerous lesions. Until now, therapeutic vaccines only showed limited clinical results, which strongly contrast with the regression of ectopic tumors observed in the available murine HPV tumor models after vaccination. Induction of protective immune responses in the genital mucosa (GM) may be crucial for efficacy of HPV therapeutic vaccines and evaluating their efficacy in a murine model with genital HPV- tumors represents an important prerequisite for clinical trials. Thus, we have here established an orthotopic mouse model where tumors in the GM develop after an intravaginal (i.vag) instillation of HPV 16 E6/E7 oncogenes-expressing tumor cells transduced with a luciferase encoding lentivirus vector for in vivo imaging of tumor growth. Histological characterization showed that tumor grew within the vaginal epithelium and according to their mucosal location tumor- induced E7-specific CD8 Τ cells were restricted to the GM and genital draining lymph nodes together with high Τ regulatory cells infiltrates preventing spontaneous regression. Consequently, sharing several immunological similarities with natural genital HPV lesions, this novel genital tumor model may be more adequate to test therapeutic strategies. As E6 and/or E7 HPV oncogenes expression is required for the maintenance of the cancerous phenotype of cervical cells, they represent target antigens for therapeutic vaccination. We reported that mice subcutaneously (s.c.) immunized once with an adjuvanted HPV 16 E7 polypeptide vaccine harbored high E7-specific IFN-γ secreting CD8 Τ cells in their lymphoid organs and more importantly in the GM. In addition, the lack of correlation between specific responses measured in the periphery with those measured in the GM highlighted the necessity and relevance to determine the immune responses locally where HPV 16-induced lesions develop. Whether a mucosal route of immunization is better to treat/regress genital infections/tumors than parenteral immunization is still debated. Our data shows that although similar E7-specific IFN-γ secreting CD8 Τ cells responses were measured in the GM upon mucosal routes of E7 vaccine delivery (nasal and vaginal immunizations), only the s.c immunization was able to regress at least all genital tumors in mice. To further increase the vaccine-specific responses in the GM, immunostimulatory agents were i.vag administrated after vaccination. We demonstrated that a single i.vag application of toll like receptor (TLR) agonists after a s.c. E7 vaccination induced a significant increase of E7-specific IFN-γ secreting CD8 Τ cells in the GM. More precisely, regarding CpG and Poly l:C, the effect is most probably associated with a local attraction of total CD8 Τ cells and secondly depends on TLR9 and TLR3/Mda5 signaling pathways, respectively. Finally, this combinatorial strategy induced tumor regression in mice harboring large genital tumors, suggesting that such an immunotherapy could be adequate to treat HPV-induced lesions in women.

Induction of potent antitumor CTL responses by recombinant vaccinia encoding a melan-A peptide analogue.

There is considerable interest in the development of vaccination strategies that would elicit strong tumor-specific CTL responses in cancer patients. One strategy consists of using recombinant viruses encoding amino acid sequences corresponding to natural CTL-defined peptide from tumor Ags as immunogens. However, studies with synthetic tumor antigenic peptides have demonstrated that introduction of single amino acid substitutions may dramatically increase their immunogenicity. In this study we have used a well-defined human melanoma tumor Ag system to test the possibility of translating the immunological potency of synthetic tumor antigenic peptide analogues into recombinant vaccinia viruses carrying constructs with the appropriate nucleotide substitutions. Our results indicate that the use of a mutated minigene construct directing the expression of a modified melanoma tumor Ag leads to improved Ag recognition and, more importantly, to enhanced immunogenicity. Thus, recombinant vaccinia viruses containing mutated minigene sequences may lead to new strategies for the induction of strong tumor-specific CTL responses in cancer patients.

Optimal strategies for sending information through a quantum channel

Quantum states can be used to encode the information contained in a direction, i.e., in a unit vector. We present the best encoding procedure when the quantum state is made up of N spins (qubits). We find that the quality of this optimal procedure, which we quantify in terms of the fidelity, depends solely on the dimension of the encoding space. We also investigate the use of spatial rotations on a quantum state, which provide a natural and less demanding encoding. In this case we prove that the fidelity is directly related to the largest zeros of the Legendre and Jacobi polynomials. We also discuss our results in terms of the information gain.

Strategies of "Pseudomonas aeruginosa" to overcome copper limitation

Summary Copper is an important trace element and micronutrient for living organisms as it is the cofactor of several enzymes involved in diverse biological redox processes such as aerobic respiration, denitrification and photosynthesis. Despite its importance, copper may be poorly bioavailable in soils and aquatic environments, as well as in the human body, especially at physiological or alkaline pH. In this work, we have investigated the strategies that the versatile bacterium and opportunistic pathogen Pseudomonas aeruginosa has evolved to face and overcome copper limitation. The global response of the P. aeruginosa to copper limitation was assessed under aerobic conditions. Numerous iron uptake functions (including the siderophores pyoverdine and pyochelin) were down-regulated whereas expression of cioAB (encoding an alternative, copper-independent, cyanide-resistant ubiquinol oxidase) was up-regulated. Wild type P. aeruginosa was able to grow aerobically in a defined glucose medium depleted of copper by a copper chelator, whereas a cioAB mutant did not grow. Thus, P. aeruginosa relies on the CioAB enzyme to cope with severe copper deprivation. A quadruple cyo cco1 cco2 cox mutant, which was deleted for all known heme-copper terminal oxidases of P. aeruginosa, grew aerobically, albeit more slowly than did the wild type, indicating that the CioAB enzyme is capable of energy conservation. However, the expression of a cioA'-'lacZ fusion was less dependent on the copper status in the quadruple mutant than in the wild type, suggesting that copper availability might affect cioAB expression indirectly, via the function of the heme-copper oxidases. These results suggest that the CioAB enzyme can be used as a by-pass strategy to overcome severe copper limitation and perform aerobic respiration even if virtually no copper is available. The PA0114 gene, which encodes a protein of the SCOT/SenC family, was found to be important for copper acquisition and aerobic respiration in low copper conditions. A PA0114 (sent) mutant grew poorly in low copper media and had low terminal oxidase activity with TMPD (N,N,N',N'-tetramethyl-p-phenylenediamine), but expressed the CioAB enzyme at elevated levels. Addition of copper reversed these phenotypes, suggesting that periplasmic copper capture by the SenC protein is another strategy that helps P. aeruginosa to adapt to copper deprivation. RESUME Le cuivre est un micronutriment important pour les organismes vivants. Il représente le cofacteur de plusieurs enzymes impliquées dans une multitude de processus biologiques tels que la respiration aérobie, la dénitrification et la photosynthèse. Malgré son importance, le cuivre peut être peu disponible dans les sols, les environnements aquatiques et le corps humain, spécialement à pH physiologique ou alcalin. Dans ce travail nous avons étudié les stratégies développées par la bactérie pathogène opportuniste Pseudomonas aeruginosa PAO1 afm de faire face et de surmonter le manque de cuivre. La réponse globale de P. aeruginosa à la carence de cuivre a été analysée dans des conditions aérobie. Les résultats obtenus ont montré que plusieurs gènes impliqués dans l'acquisition du fer, tels que les gènes codant pour les sidérophores (pyoverdine et pyochéline), étaient réprimés, tandis que l'expression de l'opéron cioAB, codant pour l'oxydase terminale insensible au cyanure (CIO), était augmentée. La souche sauvage P. aeruginosa est capable de croître dans un milieu où la concentration en cuivre est limitée, due à la présence d'un chélateur spéciftque de cuivre, tandis que le mutant cioAB ne croît pas dans ces conditions. Nous avons conclu que P. aeruginosa nécessite l'oxydase terminale CIO pour faire face à la carence en cuivre. Un quadruple mutant affecté dans toutes les oxydases dépendantes du cuivre (cyo ccol cco2 cox) et appartenant aux oxydases de type hème-cuivre, peut croître en aérobie, néanmoins plus lentement que la souche sauvage, ce qui montre que l'enzyme CIO est capable de conserver l'énergie. L'expression de la fusion rapportrice cioA'-'IacZ chez le quadruple mutant est moins dépendante de la disponibilité de cuivre que chez la souche sauvage. Ces résultats suggèrent que la disponibilité de cuivre influence l'expression de cioAB d'une façon indirecte, par le biais des oxydases terminales de type héme-cuivre. Il est donc possible qu'en cas de carence de cuivre, P. aeruginosa utilise l'enzyme CIO comme stratégie afin de surmonter ce manque et de réaliser la respiration aérobie. Nous avons démontré que le gène PA0114, codant pour une protéine appartenant à la famille SCO1/SenC, est important dans l'acquisition et dans la respiration aérobie dans des environnements où le cuivre est présent en faible concentration. En ces conditions, la croissance du mutant senC est faible; de plus, l'activité des oxydases terminales en présence du donneur d'électrons TMPD (N,N,N,N'-tetraméthyl-p-phénylenediamine) est basse. Toutefois, l'addition de cuivre au milieu de culture permet de restaurer le phénotype du type sauvage. Ces résultats montrent que la protéine SenC est capable d'acquérir le cuivre et représente donc une autre stratégie chez P. aeruginosa pour s'adapter à un manque de cuivre.

Deletion of the Vaccinia Virus Gene A46R, Encoding for an Inhibitor of TLR Signalling, Is an Effective Approach to Enhance the Immunogenicity in Mice of the HIV/AIDS Vaccine Candidate NYVAC-C.

Viruses have developed strategies to counteract signalling through Toll-like receptors (TLRs) that are involved in the detection of viruses and induction of proinflammatory cytokines and IFNs. Vaccinia virus (VACV) encodes A46 protein which disrupts TLR signalling by interfering with TLR: adaptor interactions. Since the innate immune response to viruses is critical to induce protective immunity, we studied whether deletion of A46R gene in a NYVAC vector expressing HIV-1 Env, Gag, Pol and Nef antigens (NYVAC-C) improves immune responses against HIV-1 antigens. This question was examined in human macrophages and in mice infected with a single A46R deletion mutant of the vaccine candidate NYVAC-C (NYVAC-C-ΔA46R). The viral gene A46R is not required for virus replication in primary chicken embryo fibroblast (CEF) cells and its deletion in NYVAC-C markedly increases TNF, IL-6 and IL-8 secretion by human macrophages. Analysis of the immune responses elicited in BALB/c mice after DNA prime/NYVAC boost immunization shows that deletion of A46R improves the magnitude of the HIV-1-specific CD4 and CD8 T cell immune responses during adaptive and memory phases, maintains the functional profile observed with the parental NYVAC-C and enhances anti-gp120 humoral response during the memory phase. These findings establish the immunological role of VACV A46R on innate immune responses of macrophages in vitro and antigen-specific T and B cell immune responses in vivo and suggest that deletion of viral inhibitors of TLR signalling is a useful approach for the improvement of poxvirus-based vaccine candidates.

Strategies and tools for preventing neurotoxicity: to test, to predict and how to do it

A change in paradigm is needed in the prevention of toxic effects on the nervous system, moving from its present reliance solely on data from animal testing to a prediction model mostly based on in vitro toxicity testing and in silico modeling. According to the report published by the National Research Council (NRC) of the US National Academies of Science, high-throughput in vitro tests will provide evidence for alterations in"toxicity pathways" as the best possible method of large scale toxicity prediction. The challenges to implement this proposal are enormous, and provide much room for debate. While many efforts address the technical aspects of implementing the vision, many questions around it need also to be addressed. Is the overall strategy the only one to be pursued? How can we move from current to future paradigms? Will we ever be able to reliably model for chronic and developmental neurotoxicity in vitro? This paper summarizes four presentations from a symposium held at the International Neurotoxicology Conference held in Xi"an, China, in June 2011. A. Li reviewed the current guidelines for neurotoxicity and developmental neurotoxicity testing, and discussed the major challenges existing to realize the NCR vision for toxicity testing. J. Llorens reviewed the biology of mammalian toxic avoidance in view of present knowledge on the physiology and molecular biology of the chemical senses, taste and smell. This background information supports the hypothesis that relating in vivo toxicity to chemical epitope descriptors that mimic the chemical encoding performed by the olfactory system may provide a way to the long term future of complete in silico toxicity prediction. S. Ceccatelli reviewed the implementation of rodent and human neural stem cells (NSCs) as models for in vitro toxicity testing that measures parameters such as cell proliferation, differentiation and migration. These appear to be sensitive endpoints that can identify substances with developmental neurotoxic potential. C. Sun ol reviewed the use of primary neuronal cultures in testing for neurotoxicity of environmental pollutants, including the study of the effects of persistent exposures and/or in differentiating cells, which allow recording of effects that can be extrapolated to human developmental neurotoxicity.