Doppler ultrasound in the measurement of pulse wave velocity: agreement with the Complior method

Aortic stiffness is an independent predictor factor for cardiovascular risk. Different methods for determining pulse wave velocity (PWV) are used, among which the most common are mechanical methods such as SphygmoCor or Complior, which require specific devices and are limited by technical difficulty in obtaining measurements. Doppler guided by 2D ultrasound is a good alternative to these methods. We studied 40 patients (29 male, aged 21 to 82 years) comparing the Complior method with Doppler. Agreement of both devices was high (R = 0.91, 0.84-0.95, 95% CI). The reproducibility analysis revealed no intra-nor interobserver differences. Based on these results, we conclude that Doppler ultrasound is a reliable and reproducible alternative to other established methods for themeasurement of aortic PWV

Coupled reproductive ecologies in a plant-pollinator-seed predator system

Résumé de la thèseBien que le mutualisme puisse être considéré comme une relation harmonieuse entre différentes espèces, son étude révèle plutôt une exploitation réciproque où chaque partenaire tente de maximiser ses bénéfices tout en réduisant ses coûts. Dans ce contexte, l'identification des facteurs qui favorisent ou contrarient, au cours de l'évolution, une issue mutualiste est une étape majeure pour pouvoir reconstruire les étapes clés menant à l'apparition et au maintien des interactions mutualistes. Le but de ce doctorat était l'identification des traits phénotypiques qui permettent à la plante Silene latofolia (Caryophyllacée)et à son pollinisateur - prédateur de graines, la phalène Hadena bicruris (Noctuidé), d'augmenter les bénéfices nets que chacun retire de l'interaction. Ce système d'étude est particulièrement bien approprié à l'étude de ces traits, car on peut assez facilement estimer la qualité et la quantité des descendants (fitness) des deux partenaires. En effet, la femelle papillon pond un oeuf dans la fleur qu'elle pollinise et sa larve se développe dans le fruit, consommant les graines de la plante. Ainsi, sur une même plante, il est possible d'estimer les succès respectifs de la plante et du papillon à obtenir une descendance. De plus, le conflit d'intérêt autour des graines qui sont indispensables, à la fois à la plante et au papillon, peut stimuler l'évolution de traits qui limitent la surexploitation réciproque des partenaires. Dans une première étude, j'ai montré que le papillon mâle était un pollinisateur efficace de S. latifolia et qu'ainsi, il permettait à la plante d'augmenter le nombre de graines produites (i.e.bénéfice) sans pour autant augmenter la quantité de larves sur la plante. Dans ce système, les papillons pondent un seul oeuf par fleur, déposé soit à l'intérieur de la fleur, dans le tube de corolle, soit sur le pétale. Ma seconde étude montre que les plantes répondent différemment à la présence des oeufs suivant leur position. Aussi, quand l'oeuf est placé dans la fleur, la plante a davantage tendance à ne pas développer le fruit de la fleur infesté ou bien à produire des fruits plus petits que lorsque l'oeuf est placé sur le pétale. Enfin, j'ai montré que la femelle du papillon pond plus souvent sur le pétale lorsque elle visite des fleurs dotées d'un long tube de corolle, et que les larves issues de ces oeufs ont moins de chances de réussir à pénétrer dans le fruit que les larves issues des oeufs placés à l'intérieur de la fleur. Aussi, la variation observée du site de ponte pourrait être causé par la morphologie de la fleur qui contraint le papillon à pondre sur le pétale. Vu dans leur ensemble, les résultats obtenus pendant ce doctorat suggèrent que la participation des mâles à la pollination, l'absence de développement des fruits et la profondeur du tube de corolle pourraient réduire les coûts que S. latifolia subit dans son interaction avec H. bicruris. Par ailleurs, je n'ai pas détecté de mécanismes qui permettraient au papillon de réduire les coûts que la plante pourrait lui imposer. La prochaine étape serait de déterminer l'effet des traits identifiés dans ce doctorat sur la fitness globale de la plante et du papillon pour estimer pleinement leur efficacité à réduire les coûts et à favoriser une issue mutualiste. De même, il faudrait évaluer l'effet de ces traits en populations naturelles pour identifier le rôle des facteurs environnementaux sur leur efficacité.AbstractAlthough mutualisms can be regarded as harmonious relationships between the interacting partners, they are best conceptualized as reciprocal exploitations in which each partner attempts to increase its own benefits and decrease its costs. To date, identifying the factors which promote or discourage mutualistic outcomes remains a major goal to reconstruct the ecological conditions leading to mutualisms. The aim of this PhD thesis was to identify phenotypic traits that may increase the net benefits of each partner in the interaction between the plant Silene latifolia (Caryophyllaceae) and its pollinator / seed predator, the moth Hadena bicruris (Noctuidae). This study system is particularly well suited because the fitness of both interacting species can be assessed. The female moth lays its egg in the flower it pollinated, and its offspring grows in the fruit, feeding on the seeds of the plant, which allows for the follow-up of both larva and fruit fates. Furthermore, the inherent conflict of interest over the seeds as plant progeny vs. larval resource may stimulate the evolution of traits that reduce overexploitation in both the moth and plant. In a first study, I show that male moths are efficient pollinators, hence increasing seed production without increasing oviposition. The contribution of male moths to pollination might thus improve the net benefits of the interaction for the host plant. Females of the H. bicruris moth lay a single egg per flower, and place it either inside the corolla tube or on the petal. My second study shows that plants are more likely to abort the infested flower or to produce a smaller fruit when the egg was experimentally placed inside the flower compared to plants that received an egg on the petal. Finally, female moths were found to lay their eggs more frequently on the petal when visiting a flower with a deep corolla tube, and larvae hatching from these eggs less likely to successfully attack the fruit. Variation in egg position on the flower may thus be the result of a constraint imposed by floral morphology. Overall, this PhD work suggests that the pollination by male moths, flower abortion, and deep corolla tube may efficiently reduce the costs experienced by S. latifolia in its interaction with H. bicruris. Interestingly, no apparent mechanism of costs reduction was detected for the moth. Further studies should focus on the effects of these traits (i) in the long term fitness of both the plant and the insect and (ii) their interactions with environmental factors (biotic and abiotic) that may affect their efficiency in natural populations.

G-protein coupled receptor-evoked glutamate exocytosis from astrocytes: role of prostaglandins.

Astrocytes are highly secretory cells, participating in rapid brain communication by releasing glutamate. Recent evidences have suggested that this process is largely mediated by Ca(2+)-dependent regulated exocytosis of VGLUT-positive vesicles. Here by taking advantage of VGLUT1-pHluorin and TIRF illumination, we characterized mechanisms of glutamate exocytosis evoked by endogenous transmitters (glutamate and ATP), which are known to stimulate Ca(2+) elevations in astrocytes. At first we characterized the VGLUT1-pHluorin expressing vesicles and found that VGLUT1-positive vesicles were a specific population of small synaptic-like microvesicles containing glutamate but which do not express VGLUT2. Endogenous mediators evoked a burst of exocytosis through activation of G-protein coupled receptors. Subsequent glutamate exocytosis was reduced by about 80% upon pharmacological blockade of the prostaglandin-forming enzyme, cyclooxygenase. On the other hand, receptor stimulation was accompanied by extracellular release of prostaglandin E2 (PGE2). Interestingly, administration of exogenous PGE2 produced per se rapid, store-dependent burst exocytosis of glutamatergic vesicles in astrocytes. Finally, when PGE2-neutralizing antibody was added to cell medium, transmitter-evoked exocytosis was again significantly reduced (by about 50%). Overall these data indicate that cyclooxygenase products are responsible for a major component of glutamate exocytosis in astrocytes and that large part of such component is sustained by autocrine/paracrine action of PGE2.

Enhanced diastolic reflections on arterial pressure pulse during exercise recovery

RESUMEDurant la phase de récupération d'un exercice de course à pied d'intensité maximale ou submaximale, une augmentation de la pression artérielle systolique centrale (aortique) résultant de la réflexion des ondes de pouls sur l'arbre vasculaire est constatée chez l'individu en bonne santé. En diastole cependant, l'impact de la réflexion de ces ondes de pouls sur la pression centrale demeure inconnu durant la récupération d'un exercice.Nous avons évalué les ondes de pouls centrales systolique et diastolique chez onze athlètes d'endurance durant la phase de récupération d'un exercice de course à pied dans des conditions d'effort maximal (sur tapis de course) et lors d'un effort submaximal lors d'une course à pied de 4000 mètres en plein air sur terrain mixte.Pour chaque sujet et lors des deux exercices, l'onde de pouls a été mesurée au niveau radial par tonométrie d'aplanation durant une phase de repos précédant l'exercice, puis à 5, 15, 25, 35 et 45 minutes après la fin de l'exercice. En utilisant une fonction mathématique de transfert, l'onde de pouls centrale a été extrapolée à partir de l'onde de pouls radiale. En compilant la forme de l'onde de pouls centrale avec une mesure simultanée de la pression artérielle brachiale, un index d'augmentation de l'onde de pouls en systole (Alx) et en diastole (Als) peut être calculé, reflétant l'augmentation des pressions résultant de la réflexion des ondes sur l'arbre vasculaire périphérique.A 5 minutes de la fin de l'exercice, les deux index ont été mesurés moindres que ceux mesurés lors de la phase précédant celui-ci. Lors des mesures suivantes, Alx est resté bas, alors que Aid a progressivement augmenté pour finalement dépasser la valeur de repos après 45 minutes de récupération. Le même phénomène a été constaté pour les deux modalités d'exercice (maximal ou submaximal). Ainsi, au-delà de quelques minutes de récupération après un exercice de course d'intensité maximale ou submaximale, nous avons montré par ces investigations que les ondes de pouls réfléchies en périphérie augmentent de façon sélective la pression centrale en diastole chez l'athlète d'endurance.ABSTRACTDuring recovery from a maximal or submaximal aerobic exercise, augmentation of central (aortic) systolic pressure by reflected pressure waves is blunted in healthy humans. However, the extent to which reflected pressure waves modify the central pulse in diastole in these conditions remains unknown. We evaluated systolic and diastolic central reflected waves in 11 endurance-trained athletes on recovery from a maximal running test on a treadmill (treadmill-max) and a 4000m run in field conditions. On both occasions in each subject, the radial pulse was recorded with applanation tonometry in the resting preexercise state and then 5, 15, 25, 35, and 45 minutes after exercise termination. From the central waveform, as reconstructed by application of a generalized transfer function, we computed a systolic (Alx) and a diastolic index (Aid) of pressure augmentation by reflections. At 5 minutes, both indices were below preexercise. At further time-points, Alx remained low, while Aid progressively increased, to overshoot above preexercise at 45 minutes. The same behavior was observed with both exercise types. Beyond the first few minutes of recovery following either maximal or submaximal aerobic exercise, reflected waves selectively augment the central pressure pulse in diastole, at least in endurance- trained athletes.

A combined ear sensor for pulse oximetry and carbon dioxide tension monitoring: accuracy in critically ill children.

IMPLICATIONS: A new combined ear sensor was tested for accuracy in 20 critically ill children. It provides noninvasive and continuous monitoring of arterial oxygen saturation, arterial carbon dioxide tension, and pulse rate. The sensor proved to be clinically accurate in the tested range.

Avoiding transcription factor competition at promoter level increases the chances of obtaining oscillations

Background: The ultimate goal of synthetic biology is the conception and construction of genetic circuits that are reliable with respect to their designed function (e.g. oscillators, switches). This task remains still to be attained due to the inherent synergy of the biological building blocks and to an insufficient feedback between experiments and mathematical models. Nevertheless, the progress in these directions has been substantial. Results: It has been emphasized in the literature that the architecture of a genetic oscillator must include positive (activating) and negative (inhibiting) genetic interactions in order to yield robust oscillations. Our results point out that the oscillatory capacity is not only affected by the interaction polarity but by how it is implemented at promoter level. For a chosen oscillator architecture, we show by means of numerical simulations that the existence or lack of competition between activator and inhibitor at promoter level affects the probability of producing oscillations and also leaves characteristic fingerprints on the associated period/amplitude features. Conclusions: In comparison with non-competitive binding at promoters, competition drastically reduces the region of the parameters space characterized by oscillatory solutions. Moreover, while competition leads to pulse-like oscillations with long-tail distribution in period and amplitude for various parameters or noisy conditions, the non-competitive scenario shows a characteristic frequency and confined amplitude values. Our study also situates the competition mechanism in the context of existing genetic oscillators, with emphasis on the Atkinson oscillator.

Evaluation of bedbath in critically ill patients: impact of water temperature on the pulse oximetry variation

This is a participant study, quasi-experimental, of a before and after type. A quantitative approach of biophysiological measures was used, represented by the saturation of oxygen measured by pulse oximeter (SpO2), and recorded on three occasions: before, during and after the bedbath in critically ill patients hospitalized at the ICU of a University Hospital in Brazil. Objective: to compare the SpO2 in various stages of the bath, with and without control of water temperature. Data collection was performed between December 2007 and April 2008 on a convenience sample consisting of 30 patients aged over 18 who had classification in TISS-28 from level II. Results show that water temperature control means a lower variation of SpO2 (p<0.05). No marked differences in variation of saturation between men and women or between age groups were established. In conclusion, heated and constant water temperature during the bedbath is able to minimize the fall of SpO2 that occurs while handling patients during procedures.

Wipe sampling procedure coupled to LC-MS/MS analysis for the simultaneous determination of 10 cytotoxic drugs on different surfaces.

A simple wipe sampling procedure was developed for the surface contamination determination of ten cytotoxic drugs: cytarabine, gemcitabine, methotrexate, etoposide phosphate, cyclophosphamide, ifosfamide, irinotecan, doxorubicin, epirubicin and vincristine. Wiping was performed using Whatman filter paper on different surfaces such as stainless steel, polypropylene, polystyrol, glass, latex gloves, computer mouse and coated paperboard. Wiping and desorption procedures were investigated: The same solution containing 20% acetonitrile and 0.1% formic acid in water gave the best results. After ultrasonic desorption and then centrifugation, samples were analysed by a validated liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in selected reaction monitoring mode. The whole analytical strategy from wipe sampling to LC-MS/MS analysis was evaluated to determine quantitative performance. The lowest limit of quantification of 10 ng per wiping sample (i.e. 0.1 ng cm(-2)) was determined for the ten investigated cytotoxic drugs. Relative standard deviation for intermediate precision was always inferior to 20%. As recovery was dependent on the tested surface for each drug, a correction factor was determined and applied for real samples. The method was then successfully applied at the cytotoxic production unit of the Geneva University Hospitals pharmacy.

Determination of Vasopressin and Desmopressin in urine by means of liquid chromatography coupled to quadrupole time-of-flight mass spectrometry for doping control purposes.

The anti-diuretic neurohypophysial hormone Vasopressin (Vp) and its synthetic analogue Desmopressin (Dp, 1-desamino-vasopressin) have received considerable attention from doping control authorities due to their impact on physiological blood parameters. Accordingly, the illicit use of Desmopressin in elite sport is sanctioned by the World Anti-Doping Agency (WADA) and the drug is classified as masking agent. Vp and Dp are small (8-9 amino acids) peptides administered orally as well as intranasally. Within the present study a method to determine Dp and Vp in urinary doping control samples by means of liquid chromatography coupled to quadrupole high resolution time-of-flight mass spectrometry was developed. After addition of Lys-Vasopressin as internal standard and efficient sample clean up with a mixed mode solid phase extraction (weak cation exchange), the samples were directly injected into the LC-MS system. The method was validated considering the parameters specificity, linearity, recovery (80-100%), accuracy, robustness, limit of detection/quantification (20/50 pg mL(-1)), precision (inter/intra-day<10%), ion suppression and stability. The analysis of administration study urine samples collected after a single intranasal or oral application of Dp yielded in detection windows for the unchanged target analyte for up to 20 h at concentrations between 50 and 600 pg mL(-1). Endogenous Vp was detected in concentrations of approximately 20-200 pg mL(-1) in spontaneous urine samples obtained from healthy volunteers. The general requirements of the developed method provide the characteristics for an easy transfer to other anti-doping laboratories and support closing another potential gap for cheating athletes.

Wide-pulse-high-frequency neuromuscular stimulation of triceps surae induces greater muscle fatigue compared with conventional stimulation.

We compared the extent and origin of muscle fatigue induced by short-pulse-low-frequency [conventional (CONV)] and wide-pulse-high-frequency (WPHF) neuromuscular electrical stimulation. We expected CONV contractions to mainly originate from depolarization of axonal terminal branches (spatially determined muscle fiber recruitment) and WPHF contractions to be partly produced via a central pathway (motor unit recruitment according to size principle). Greater neuromuscular fatigue was, therefore, expected following CONV compared with WPHF. Fourteen healthy subjects underwent 20 WPHF (1 ms-100 Hz) and CONV (50 μs-25 Hz) evoked isometric triceps surae contractions (work/rest periods 20:40 s) at an initial target of 10% of maximal voluntary contraction (MVC) force. Force-time integral of the 20 evoked contractions (FTI) was used as main index of muscle fatigue; MVC force loss was also quantified. Central and peripheral fatigue were assessed by voluntary activation level and paired stimulation amplitudes, respectively. FTI in WPHF was significantly lower than in CONV (21,717 ± 11,541 vs. 37,958 ± 9,898 N·s P<0,001). The reductions in MVC force (WPHF: -7.0 ± 2.7%; CONV: -6.2 ± 2.5%; P < 0.01) and paired stimulation amplitude (WPHF: -8.0 ± 4.0%; CONV: -7.4 ± 6.1%; P < 0.001) were similar between conditions, whereas no change was observed for voluntary activation level (P > 0.05). Overall, our results showed a different motor unit recruitment pattern between the two neuromuscular electrical stimulation modalities with a lower FTI indicating greater muscle fatigue for WPHF, possibly limiting the presumed benefits for rehabilitation programs.

Pulse wave analysis of aortic pressure: diastole should also be considered.

BACKGROUND: : The systolic augmentation index (sAix), calculated from the central aortic pulse wave (reconstructed from the noninvasive recording of the radial pulse with applanation tonometry), is widely used as a simple index of central arterial stiffness, but has the disadvantage of also being influenced by the timing of the reflected with respect to the forward pressure wave, as shown by its inverse dependence on heart rate (HR). During diastole, the central aortic pulse also contains reflected waves, but their relationship to arterial stiffness and HR has not been studied. METHODS: : In 48 men and 45 women, all healthy, with ages ranging from 19 to 70 years, we measured pulse wave velocity (PWV, patients supine), a standard evaluator of arterial stiffness, and carried out radial applanation tonometry (patients sitting and supine). The impact of reflected waves on the diastolic part of the aortic pressure waveform was quantified in the form of a diastolic augmentation index (dAix). RESULTS: : Across ages, sexes, and body position, there was an inverse relationship between the sAix and the dAix. When PWV and HR were added as covariates to a prediction model including age, sex and body position as main factors, the sAix was directly related to PWV (P < 0.0001) and inversely to HR (P < 0.0001). With the same analysis, the dAix was inversely related to PWV (P < 0.0001) and independent of HR (P = 0.52). CONCLUSION: : The dAix has the same degree of linkage to arterial stiffness as the more conventional sAix, while being immune to the confounding effect of HR. The quantification of diastolic aortic pressure augmentation by reflected waves could be a useful adjunct to pulse wave analysis.

Profound impact of uncomplicated pregnancy on diastolic, but not systolic pulse contour of aortic pressure

RESUME: L'objectif de cette étude était de déterminer l'impact de la grossesse non compliquée sur l'onde de pouls de la pression aortique centrale. Méthode 66 femmes au total avec une grossesse simple ont été réparties en trois groupes selon le stade de leur gestation: premier trimestre (T1, n=22), deuxième trimestre (T2, n=20) et troisième trimestre (T3, n=24). Le groupe contrôle (C, n=21) était constitué de femmes non enceintes, en bonne santé habituelle, prenant une contraception oestroprogestative. La tonométrie d'aplanation a été utilisée pour l'acquisition des ondes de pouls centrale un appareil disponible dans le commerce (SphygmoCor) permet l'enregistrement de l'onde de pouls périphérique avec un tonomètre d'aplanation de l'artère radiale au niveau du poignet, puis effectue sa transformation en sa forme centrale, grâce à une analyse de Fourrier et une fonction de transfert. L'influence des ondes réfléchies sur l'onde de pouls a été déterminée non seulement pendant la systole (augmentation systolique), comme on procède habituellement dans l'analyse de l'onde de pouls, mais aussi pendant la diastole (augmentation diastolique). Résultats Au cours de la grossesse, les pressions centrales systolique et diastolique sont restées inchangées et comparables aux valeurs mesurées chez les femmes qui ne sont pas enceintes. Dans le groupe contrôle, l'augmentation systolique s'élevait à 8.1±7.5% de la pression de pouls ; il n'y avait pas de différence statistiquement significative avec les valeurs obtenues chez les femmes enceintes, et ce, à n'importe quel stade de la grossesse (T1 : 4.6±11.4%, T2: 5.0±9.3%, T3 : 4.7±8.1%). Par contre, l'amplitude de l'augmentation diastolique diminuait avec la progression de la grossesse (C 6.5±2.4%, T1 : 5.2±3.1%, T2 : 3.8±2.6%; P=0.002 versus C; T3 : 2.3±2.0%; P<0.0001 versus C et P=0.004 versus T 1). Conclusion La grossesse ne modifie pas la forme de l'onde de pouls systolique centrale, ce qui implique de la part du système cardiovasculaire une adaptation fine à la demande croissante de flux sanguin, et ce, à tous les stades de la grossesse. Par contre, l'amplitude de l'onde de réflexion atteignant l'aorte pendant la diastole diminue progressivement au cours de la grossesse. Perspectives De récentes études montrent qu'une valeur anormalement haute de l'augmentation systolique de la pression centrale, comme on peut la déterminer avec la tonométrie d'aplanation, pourrait être un indice de trouble hypertensif de la grossesse débutant. Cette technique simple pourrait être d'autant plus facile à mettre en oeuvre si les valeurs normales pour l'augmentation systolique étaient indépendantes du stade de la grossesse, comme le suggèrent nos résultats, du moins pour les mesures prises en position assise.

An anti-CD19 antibody coupled to a tetanus toxin peptide induces efficient Fas ligand (FasL)-mediated cytotoxicity of a transformed human B cell line by specific CD4+ T cells.

Treatment of B cell lymphoma patients with MoAbs specific for the common B cell marker (CD20) has shown a good overall response rate, but the number of complete remissions is still very low. The use of MoAbs coupled to radioisotopes can improve the results, but induces undesirable myelodepression. As an alternative, we proposed to combine the specificity of MoAbs with the immunogenicity of T cell epitopes. We have previously shown that an anti-Ig lambda MoAb coupled to an MHC class II-restricted universal T cell epitope peptide P2 derived from tetanus toxin induces efficient lysis of a human B cell lymphoma by a specific CD4+ T cell line. Here we demonstrate that the antigen presentation properties of the MoAb peptide conjugate are maintained using a MoAb directed against a common B cell marker, CD19, which is known to be co-internalized with the B cell immunoglobulin receptor. In addition, we provide evidence that B cell lysis is mediated by the Fas apoptosis pathway, since Fas (CD95), but not tumour necrosis factor receptor (TNFr) or TNF-related receptors, is expressed by the target B cells, and FasL, but not perforin, is expressed by the effector T cells. These results show that B cell lymphomas can be 'foreignized' by MoAb-peptide P2 conjugates directed against the common B cell marker CD19 and eliminated by peptide P2-specific CD4+ T cells, via the ubiquitous Fas receptor. This approach, which bridges the specificity of passive antibody therapy with an active T cell immune response, may be complementary to and more efficient than the present therapy results with unconjugated chimeric anti-CD20 MoAbs.

In vivo targeting of an anti-tumor antibody coupled to antigenic MHC class I complexes induces specific growth inhibition and regression of established syngeneic tumor grafts.

The concept of antibody-mediated targeting of antigenic MHC/peptide complexes on tumor cells in order to sensitize them to T-lymphocyte cytotoxicity represents an attractive new immunotherapy strategy. In vitro experiments have shown that an antibody chemically conjugated or fused to monomeric MHC/peptide can be oligomerized on the surface of tumor cells, rendering them susceptible to efficient lysis by MHC-peptide restricted specific T-cell clones. However, this strategy has not yet been tested entirely in vivo in immunocompetent animals. To this aim, we took advantage of OT-1 mice which have a transgenic T-cell receptor specific for the ovalbumin (ova) immunodominant peptide (257-264) expressed in the context of the MHC class I H-2K(b). We prepared and characterized conjugates between the Fab' fragment from a high-affinity monoclonal antibody to carcinoembryonic antigen (CEA) and the H-2K(b) /ova peptide complex. First, we showed in OT-1 mice that the grafting and growth of a syngeneic colon carcinoma line transfected with CEA could be specifically inhibited by systemic injections of the conjugate. Next, using CEA transgenic C57BL/6 mice adoptively transferred with OT-1 spleen cells and immunized with ovalbumin, we demonstrated that systemic injections of the anti-CEA-H-2K(b) /ova conjugate could induce specific growth inhibition and regression of well-established, palpable subcutaneous grafts from the syngeneic CEA-transfected colon carcinoma line. These results, obtained in a well-characterized syngeneic carcinoma model, demonstrate that the antibody-MHC/peptide strategy can function in vivo. Further preclinical experimental studies, using an anti-viral T-cell response, will be performed before this new form of immunotherapy can be considered for clinical use.

The effect of vaccines based on ovalbumin coupled to gas-filled microbubbles for reducing infection by ovalbumin-expressing Listeria monocytogenes.

Gas-filled microbubbles (MB) are a very promising alternative to the currently evaluated lipid- or polymer-based particulate Ag delivery systems. We recently demonstrated the ability of MB to deliver associated Ag to DC, to activate them and thereby induce both humoral and cellular immune responses. We now extended the characterization of MB as antigen-delivery system by appraising the efficiency of MB-associated ovalbumin (OVA-MB) at protecting mice against pathogen infection. Ultrasound-mediated imaging demonstrated that the administration of OVA via MB generates a depot at the injection site that lasts for several hours. We found that OVA-MB injected subcutaneously is far more effective at inducing specific Ab and T cell immunity than immunization with free OVA. Moreover, a covalent link between MB and OVA causes a stronger bias towards a Th1-type of immune response than adsorption of the Ag or its covalent link to liposomes of the same lipid composition. Finally, vaccination of mice with OVA-MB partially protects against a systemic infection with OVA-expressing Listeria monocytogenes. The vaccine induces specific effector CD8 T cell responses capable of decreasing more than 100 fold the bacterial load. MB thus represent a potent Ag delivery system for vaccination against intracellular infectious agents.