Increased vulnerability of neurones and glial cells to low concentrations of methylmercury in a prooxidant situation.

Using reaggregating rat brain cell cultures at two different stages of differentiation, we examined the biochemical effects of a 10-day treatment with nanomolar concentrations of methylmercuric chloride (monomethylmercury), in the presence or absence of promoters of hydroxyl radical formation (10 microM copper sulphate plus 100 microM ascorbate). A decrease in total protein content accounted for the general cytotoxicity of these compounds, whereas selective effects were assessed by determining the activities of cell type-specific enzymes. Methylmercury, up to 100 nM, as well as the copper ascorbate mixture, when applied separately, induced no general cytotoxicity, and only slight effects on neuronal parameters. However, when applying 100 nM methylmercury and the copper-ascorbate mixture together, a drastic decrease in neuronal and glial parameters was found. Under these conditions, the content of reactive oxygen species, assessed by 2',7'-dichlorofluorescin oxidation, increased greatly, while the activities of antioxidant enzymes decreased. In the presence of copper and ascorbate, differentiated cultures appeared more resistant than immature ones to low methylmercury concentrations (1-10 mM), but did undergo similar changes in both cell type-specific and antioxidant enzyme activities at 100 nM methylmercury. These results suggest that in prooxidant conditions low doses of mercury can become much more deleterious for the central nervous system.

Aufftrittsangst : kardiorespiratorische Aktivität bei ängstlichen und nichtängstlichen Musikstudenten in einer Auftrittssituation = Performance anxiety : cardiorespiratory activity in high- and low-anxious professional music students in a performance situation

Fragebogenstudien haben gezeigt, dass ängstliche Musiker vor und/oder während eines Auftritts möglicherweise unter Hyperventilationssymptomen leiden. Berichtete Symptome beinhalten Kurzatmigkeit, schnelles oder tiefes Einatmen, Schwindelgefühl und Herzklopfen. Bisher hat jedoch noch keine Studie getestet, ob diese selbstberichteten Symptome tatsächlich kardiorespiratorische Veränderungen widerspiegeln. Das Hauptziel dieser Studie ist es, zu bestimmen, ob sich Auftrittsangst bei Musikern physiologisch über kardiorespiratorische Muster äußert. Wir haben insgesamt 74 Musikstudenten von Schweizer Musikhochschulen getestet. Diese Studenten wurden aufgrund ihrer selbstberichteten Auftrittsangst (STAI-S) in zwei Gruppen unterteilt: ängstliche Musiker und nichtängstliche Musiker. Die Studenten wurden in drei unterschiedlichen Situationen getestet: Ausgangszustand, Auftritt ohne Publikum, Auftritt mit Publikum. Wir haben folgende Parameter gemessen: a) kardiorespiratorische Muster und endexpiratorisches CO2, welches eine gute nichtinvasive Schätzung des Hyperventilationsgrades liefert und b) subjektiv wahrgenommene Emotionen und subjektiv wahrgenommene physiologische Aktivität. Das Poster zeigt die ersten Resultate der 15 ängstlichsten und der 15 am wenigsten ängstlichen Musiker. Das Hauptinteresse gilt den folgenden Punkten: Herz- und Atemfrequenz, subjektiv wahrgenommenes Herzklopfen, subjektiv wahrgenommene Kurzatmigkeit und subjektiv wahrgenommenes Angstgefühl. Die Resultate dieser Studie zeigen erstens, dass ängstliche und nichtängstliche Musikstudenten zu den verschiedenen Messzeitpunkten eine vergleichbare physiologische Aktivität aufweisen und zweitens, dass ängstliche Musikstudenten ein signifikant höheres Angstgefühl haben und signifikant mehr Herzklopfen und Kurzatmigkeit wahrnehmen vor und/oder während eines Auftritts mit Publikum. Dies deutet darauf hin, dass sich ängstliche und nichtängstliche Musikstudenten a) bezüglich der subjektiv wahrgenommenen physiologischen Symptome und des selbst berichteten Angstgefühls vor und/oder während eines öffentlichen Auftritts unterscheiden und sich b) bezüglich der untersuchten physiologischen Reaktionen nicht unterscheiden.

Nighttime blood pressure: a target for therapy?

Ambulatory blood pressure (BP) monitoring is increasingly used in the evaluation of hypertensive patients. The ability to monitor BP throughout the day and night allows the detection of abnormal nocturnal BP patterns, the most common being a "nondipping" pattern, which is associated with increased cardiovascular risk; its correction appears to have a positive impact on cardiovascular outcome. Antihypertensive treatment should be individually adjusted to control BP during both daytime and nighttime. However, drug-induced lowering of nocturnal BP, if excessive, could amplify the morning BP surge in patients with daytime BP elevation, increasing the risk of developing a cardiovascular event. Ambulatory BP monitoring therefore represents a unique tool to establish the most appropriate antihypertensive drug regimen for the individual patient.

The G0/G1 switch gene 2 is a novel PPAR target gene.

PPARs (peroxisome-proliferator-activated receptors) alpha, beta/delta and gamma are a group of transcription factors that are involved in numerous processes, including lipid metabolism and adipogenesis. By comparing liver mRNAs of wild-type and PPARalpha-null mice using microarrays, a novel putative target gene of PPARalpha, G0S2 (G0/G1 switch gene 2), was identified. Hepatic expression of G0S2 was up-regulated by fasting and by the PPARalpha agonist Wy14643 in a PPARalpha-dependent manner. Surprisingly, the G0S2 mRNA level was highest in brown and white adipose tissue and was greatly up-regulated during mouse 3T3-L1 and human SGBS (Simpson-Golabi-Behmel syndrome) adipogenesis. Transactivation, gel shift and chromatin immunoprecipitation assays indicated that G0S2 is a direct PPARgamma and probable PPARalpha target gene with a functional PPRE (PPAR-responsive element) in its promoter. Up-regulation of G0S2 mRNA seemed to be specific for adipogenesis, and was not observed during osteogenesis or myogenesis. In 3T3-L1 fibroblasts, expression of G0S2 was associated with growth arrest, which is required for 3T3-L1 adipogenesis. Together, these data indicate that G0S2 is a novel target gene of PPARs that may be involved in adipocyte differentiation.

Insulin and IGF-1 enhance the expression of the neuronal monocarboxylate transporter MCT2 by translational activation via stimulation of the phosphoinositide 3-kinase-Akt-mammalian target of rapamycin pathway.

MCT2 is the main neuronal monocarboxylate transporter essential for facilitating lactate and ketone body utilization as energy substrates. Our study reveals that treatment of cultured cortical neurons with insulin and IGF-1 led to a striking enhancement of MCT2 immunoreactivity in a time- and concentration-dependent manner. Surprisingly, neither insulin nor IGF-1 affected MCT2 mRNA expression, suggesting that regulation of MCT2 protein expression occurs at the translational rather than the transcriptional level. Investigation of the putative signalling pathways leading to translation activation revealed that insulin and IGF-1 induced p44- and p42 MAPK, Akt and mTOR phosphorylation. S6 ribosomal protein, a component of the translational machinery, was also strongly activated by insulin and IGF-1. Phosphorylation of p44- and p42 MAPK was blocked by the MEK inhibitor PD98058, while Akt phosphorylation was abolished by the PI3K inhibitor LY294002. Phosphorylation of mTOR and S6 was blocked by the mTOR inhibitor rapamycin. In parallel, it was observed that LY294002 and rapamycin almost completely blocked the effects of insulin and IGF-1 on MCT2 protein expression, whereas PD98059 and SB202190 (a p38K inhibitor) had no effect on insulin-induced MCT2 expression and only a slight effect on IGF-1-induced MCT2 expression. At the subcellular level, a significant increase in MCT2 protein expression within an intracellular pool was observed while no change at the cell surface was apparent. As insulin and IGF-1 are involved in synaptic plasticity, their effect on MCT2 protein expression via an activation of the PI3K-Akt-mTOR-S6K pathway might contribute to the preparation of neurons for enhanced use of nonglucose energy substrates following altered synaptic efficacy.

Pregnancy-associated changes in host permissiveness to metastasis

THESIS SUMMARY : Metastasis is a multistep process involving tumour cell-autonomous features, the host tissue stroma of the primary tumour, the blood or lymphatic system as well as a receptive target organ. Most studies on factors influencing metastasis have concentrated on the characteristics of the disseminating tumour cell and on early steps of metastasis including invasion and angiogenesis. Although these steps are necessary for tumour cells to disseminate, it is the challenges encountered in the later steps of metastasis -survival while in the circulation and engraftment and outgrowth in the target organ -that account for the inefficiency of circulating tumour cells in establishing secondary lesions. Full understanding of the metastatic process therefore requires elucidation of the mechanisms that regulate these late steps, and in particular that determine what makes any given tissue permissive for metastatic tumour growth. To address this issue, we assessed the mechanisms whereby a physiological situation -pregnancy -can alter host permissiveness toward metastasis. We show that pregnant NOD/SCID mice -injected intravenously with tumour cells -develop more metastases than their non-pregnant counterparts irrespective of the tumour cell type. There was no direct effect of pregnancy-related circulating factors on tumour cell proliferation, and subcutaneous tumour growth does not vary between pregnant and nonpregnant animals. However, decreased elimination of tumour cells from the lung microvasculature was observed in pregnant mice, prompting us to assess whether pregnancy-related adaptations in innate immunity could account for this differential clearing. We found that natural killer (NK) cell fractions are decreased in blood and spleen of pregnant mice and that NK cell cytotoxicity is impaired, as reported previously. The use of NK-deficient mice or tumour cell lines resistant to NK killing abrogates the difference in metastasis load between pregnant and virgin mice. CD11 b+ Gr-1+ myeloid-derived suppressor cells (MDSC) have previously been shown to accumulate in tumour-bearing mice and to down-modulate NK activity. Accordingly, we show an increase in MDSC in pregnant mouse blood, spleen, lungs and liver. Depletion of MDSC prior to tumour cell injection decreased metastasis load in pregnant NOD/SCID mice but had no effect on virgin mice. Similarly, adoptive transfer of MDSC extracted from pregnant mice into virgin mice lead to increased metastasis take. In parallel, we investigated whether the lung and liver microenvironments are modified during pregnancy thereby providing a more "permissive soil" for the establishment of metastases. A comparative analysis of microarray data of pregnant mouse lungs and liver with "premetastatic niche" gene expression profiles of these organs shows that similar mechanisms could mediate an increase in lung and liver metastasis in pregnant mice and in mice harbouring an aggressive primary tumour. Several commonly up-regulated genes point towards the recruitment of myeloid cells, consistent with the accumulation of MDSC observed in pregnant mice. MDSC have never been evoked in the context of pregnancy before. Although the role of MDSC in pregnancy requires further investigation we suggest that MDSC accumulation constitutes an important and hitherto unrecognised common denominator of maternal immune tolerance and cancer immune escape. RESUME DE THESE : La métastatisation est un processus en plusieurs étapes qui implique des compétences particulières chez les cellules tumorales, le stroma de la tumeur primaire, les vaisseaux sanguins ou lymphatiques ainsi qu'un organe cible' réceptif. Jusqu'alors, la recherche s'est principalement intéressée aux facteurs qui influencent les étapes précoces de la métastatisation donc aux caractéristiques de la cellule métastatique, et aux processus tels que l'invasion et l'angiogenèse, tandis que peu d'études traitent des étapes tardives tel que la survie dans la circulation sanguine et l'établissement d'une lésion dans l'organe cible. En particulier, l'élucidation des facteurs qui déterminent la permissivité d'un tissu à la greffe de cellules disséminantes est indispensable à la compréhension de ce processus complexe qu'est la métastatisation. Nous proposons ici un modèle de souris récapitulant les étapes tardives de la métastatisation dans un contexte d'une permissivité accrue aux métastases chez la souris gravide, et nous évaluons les mécanismes impliqués. Les souris gestantes développent plus de métastases après l'injection intraveineuse de cellules tumorales, indépendamment du type de tumeur d'origine. Les taux élevés d'hormones et de facteurs de croissance chez la souris gravide n'inflúencent pas la prolifération des cellules tumorales et fa croissance de tumeurs sous-cutanées n'est pas non plus accélérée par la gestation. En revanche, une fois injectées, les cellules tumorales sont éliminées ` moins rapidement des vaisseaux pulmonaires chez la souris gravide que chez les contrôles. Cette observation est compatible avec un effet de la gestation sur l'immunité innée et nous avons mis en évidence une diminution des proportions de cellules NK (natural killer) dans le sang et la rate en particulier, ainsi qu'une cytotoxicité moindre envers des cellules tumorales. En utilisant des souris déficientes en cellules NK ou en injectant des cellules résistantes à l'attaqué par des cellules NK, la différence entre souris gestantes et non-gestantes disparaît. Il a été démontré chez des souris porteuses de tumeurs, que l'accumulation de cellules immunosuppressives de la lignée myélo-monocytaire (ou MDSC pour myeloid-derived suppressor tells) pouvait être responsable d'une inhibition de l'activité de cellules NK. Des nombres augmentés de ces cellules, caractérisées par les marqueurs de surface CD11b et Gr-1, ont été trouvés dans le sang, la rate, les poumons et le foie de souris gravides. Leur rôle dans la métastatisation est démontré par le fait que leur dépletion diminue le nombre de lésions secondaires chez la souris gestante, tandis que leur transfert dans des souris non-gestantes augmente le taux de métastases. L'utilisation de puces à ADN sur les foies et poumons de souris gravides a permis de mettre en évidence des différences d'expression génique proches de celles observées dans l'établissement de niches pré-métastatiques. Ceci suggère que des mécanismes similaires pourraient être responsables d'une permissivité accrue aux métastases chez la souris gravide et chez la souris porteuse d'une tumeur primaire agressive, telle que, en particulier, l'accumulation de cellules immunosuppressives dans les organes cibles. C'est la première fois que l'accumulation de MDSC est évoquée chez la souris gravide et nous proposons ici que celles-ci jouent un rôle dans la tolérance immunitaire envers le foetus et sont responsables de l'échappement de cellules tumorales injectées à la surveillance immunitaire par des cellules NK.

Improving cardiovascular and renal outcomes in gout: what should we target?

Epidemiological and experimental studies have shown that hyperuricaemia and gout are intricately linked with hypertension, metabolic syndrome, chronic kidney disease and cardiovascular disease. A number of studies suggest that hyperuricaemia and gout are independent risk factors for the development of these conditions and that these conditions account, in part, for the increased mortality rate of patients with gout. In this Review, we first discuss the links between hyperuricaemia, gout and these comorbidities, and present the mechanisms by which uric acid production and gout might favour the development of cardiovascular and renal diseases. We then emphasize the potential benefit of urate-lowering therapies on cardiovascular and renal outcomes in patients with hyperuricaemia. The mechanisms that link elevated serum uric acid levels and gout with these comorbidities seem to be multifactorial, implicating low-grade systemic inflammation and xanthine oxidase (XO) activity, as well as the deleterious effects of hyperuricaemia itself. Patients with asymptomatic hyperuricaemia should be treated by nonpharmacological means to lower their SUA levels. In patients with gout, long-term pharmacological inhibition of XO is a treatment strategy that might also reduce cardiovascular and renal comorbidities, because of its dual effect of lowering SUA levels as well as reducing free-radical production during uric acid formation.

SEPT4, a p53 target gene product, implicates p 53 in the regulation of exocytosis

Résumé : Le cancer, qui est responsable d'un quart des décès en Suisse, exhibe un état cellulaire désordonné, qui lui-même, est la conséquence d'un dérèglement des gènes. Le gène le plus fréquemment altéré, dans les cas de cancers humains, est p53. Ce gène encode un facteur de transcription, impliqué dans la régulation de nombreux gènes impliqués dans le cycle cellulaire, l'apoptose ou la différenciation. Notre laboratoire a récemment identifié seize nouveaux gènes, dont l'expression est régulée par p53, parmi lesquels sept4, su jet de cette thèse. La protéine 5EPT4 appartient à la famille des septines, qui est impliquée dans la cytokinèse. Dans ce travail, nous avons confirmé la régulation de l'expression de sept4 par p53 dans des tissus de souris, et étonnamment, seul un des deux promoteurs du gène sept4 est contrôlé par p53. En outre, l'approche immunohistologique nous a permis de supposer une implication de la protéine SEPT4 dans le mécanisme de l'exocytose. Cette hypothèse a été confirmée par l'interaction de SEPT4 avec la protéine syntaxine 1A, et par son activité inhibitrice sur la sécrétion stimulée. En élargissant l'étude de la protéine SEPT4, nous avons découvert que celle-ci avait comme partenaire fonctionnel, la protéine Pinl, une enzyme qui catalyse l'isomérisation cis-trans du lien peptidique précédant une proline. bans ce contexte, nous avons démontré que l'interaction entre ces deux protéines reposait sur le domaine WW de Pinl, un type de domaine reconnaissant les motifs phosphoséryl-prolyl et phosphothréonyl-prolyl. Ce dernier résultat nous a conduit à examiner la phosphorylation de 5EPT4. Nous avons démontré que la partie N-terminale de SEPT4 était phosphorylée par la kinase Cdk5. La co¬expression de Cdk5 et de SEPT4 stimule la dégradation de SEPT4, indépendamment de la voie du protéasome. Ainsi, l'ensemble de nos observations fournissent l'évidence de l'engagement de la protéine SEPT4 dans la régulation de l'exocytose, et soutiennent le rôle de p53 dans le contrôle de l'exocytose, via SEPT4, ce qui constituerait un nouveau rôle fonctionnel pour ce gardien du génome. Summary: Cancer, which is responsible for a quarter of the deaths in Switzerland, exhibits a disordered cellular state, which itself, is the consequence of an altered state of genes. The most frequently altered gene in human cancer is p53. This gene encodes a transcription factor, implicated in the regulation of numerous genes involved in cell cycle, apoptosis or differentiation. Our laboratory has recently identified sixteen new genes whose expression is regulated by p53, amongst them septin 4, which is the subject of this thesis. The SEPT4 protein belongs to the septin family which is implicated in cytokinesis. In the present work, we have confirmed the regulation of sept4 expression by p53 in mouse tissues, and surprisingly, only one of the two sept4 promoters is regulated by p53. In addition, the immunohistologic approach enabled us to suppose a role of SEPT4 in exocytosis. This assumption was confirmed by the interaction of SEPT4 with syntaxin 1A, and by its inhibiting activity on stimulated secretion. By widening the analysis of SEPT4, we identified Pin1 as an interacting protein. Pin1 is an enzyme which catalyzes the cis-trans isomerization of the peptide bond preceding a proline residue. In this context, we showed that the interaction between these two proteins depends on the WW domain of Pin 1. This domain has been shown to function as a phosphoserine- or phosphothreonine¬binding module. This last result prompted us to examine phosphorylation of SEPT4. We demonstrated that the N-terminal portion of SEPT4 was phosphorylated by the Cdk5 kinase. The co-expression of Cdk5 with 5EPT4 stimulates SEPT4 degradation, independently of the proteasome pathway. Thus, these observations provide evidence for the engagement of SEPT4 in the regulation of exocytosis, and supports the role of p53 in the control of exocytosis, via SEPT4, which constitutes a new functional role for this guardian of the genome.

LIGHT/HVEM/LTβR interaction as a target for the modulation of the allogeneic immune response in transplantation.

The exchange of information during interactions of T cells with dendritic cells, B cells or other T cells regulates the course of T, B and DC-cell activation and their differentiation into effector cells. The tumor necrosis factor superfamily member LIGHT (homologous to lymphotoxin, exhibits inducible expression and competes with HSV glycoprotein D for binding to herpesvirus entry mediator, a receptor expressed on T lymphocytes) is transiently expressed upon T cell activation and modulates CD8 T cell-mediated alloreactive responses upon herpes virus entry mediator (HVEM) and lymphotoxin β receptor (LTβR) engagement. LIGHT-deficient mice, or WT mice treated with LIGHT-targeting decoy receptors HVEM-Ig, LTβR-Ig or sDcR3-Ig, exhibit prolonged graft survival compared to untreated controls, suggesting that LIGHT modulates the course and severity of graft rejection. Therefore, targeting the interaction of LIGHT with HVEM and/or LTβR using recombinant soluble decoy receptors or monoclonal antibodies represent an innovative therapeutic strategy for the prevention and treatment of allograft rejection and for the promotion of donor-specific tolerance.

Identification of ectodysplasin target genes reveals the involvement of chemokines in hair development.

Ectodysplasin (Eda), a member of the tumor necrosis factor (Tnf) family, regulates skin appendage morphogenesis via its receptor Edar and transcription factor NF-κB. In humans, inactivating mutations in the Eda pathway components lead to hypohidrotic ectodermal dysplasia (HED), a syndrome characterized by sparse hair, tooth abnormalities, and defects in several cutaneous glands. A corresponding phenotype is observed in Eda-null mice, where failure in the initiation of the first wave of hair follicle development is a hallmark of HED pathogenesis. In an attempt to discover immediate target genes of the Eda/NF-κB pathway, we performed microarray profiling of genes differentially expressed in embryonic skin explants after a short exposure to recombinant Fc-Eda protein. Upregulated genes included components of the Wnt, fibroblast growth factor, transforming growth factor-β, Tnf, and epidermal growth factor families, indicating that Eda modulates multiple signaling pathways implicated in skin appendage development. Surprisingly, we identified two ligands of the chemokine receptor cxcR3, cxcl10 and cxcl11, as new hair-specific transcriptional targets of Eda. Deficiency in cxcR3 resulted in decreased primary hair follicle density but otherwise normal hair development, indicating that chemokine signaling influences the patterning of primary hair placodes only.

Interleukin-1 as a therapeutic target in gout.

PURPOSE OF REVIEW: To give an overview of current evidence for interleukin (IL)-1 blockade in the management of gout. RECENT FINDINGS: Three IL-1 blockers are currently available for clinical use: anakinra, rilonacept and canakinumab. Recent studies have focused on drugs with a long half-life: rilonacept and canakinumab. For treatment of acute gouty arthritis, three randomized controlled trials (RCTs) showed efficacy of canakinumab with some safety concerns and one RCT failed to show efficacy of rilonacept. For prevention of gout flare when starting uric acid lowering therapy (ULT), four RCTs showed efficacy of rilonacept and one RCT showed efficacy of canakinumab. SUMMARY: There is sufficient evidence supporting the use of IL-1 blockers for treatment of acute gouty arthritis or for prevention of gout flares when starting ULT in selected patients, with contraindications or intolerance to conventional therapy. More data are needed to assess safety and to specify their use in routine practice.

Music Performance Anxiety (MPA) : cardiorespiratory activity in high- and low-anxious professional music students before a performance situation

Descriptors: music performance anxiety, respiration, hyperventilation Surveys indicate that high-anxious musicians may suffer from hyperventilation (HV) before or during performance. Reported symptoms include shortness of breath, fast/ deep breathing and thumping heart. However, no study has yet tested if these selfreported symptoms reflect actual cardiorespiratory activity. Themain goal of this study was to determine if MPA is manifested physiologically in specific correlates of cardiorespiratory activity associated with HV.We studied 74 professional music students from Swiss Music Academies. In this study, we compared the most anxious students (highanxious; n 5 20) with the least anxious students (low-anxious; n 5 23) based on their self-reported performance anxiety. We measured cardiorespiratory patterns with the Lifeshirt system, end-tidal CO2 with a capnograph (EtCO2, a good non-invasive estimator of HV), self-perceived physiological activation and affective experience in three situations on different days: baseline, performance without audience, and performance with audience. Comparing measures for the private vs. the public concert, high- compared to low-anxious students showed a significant drop in EtCO2 before the public concert and reported larger increases in anxiety, tension, palpitations and breathing difficulties. In contrast, heart rate, respiratory rate and volume did not differ significantly between groups. The results of this study support the hypothesis thatMPA may be associated with a tendency to hyperventilate and, thus, point to a potential hyperventilation problem in high-anxious music students.

Nanomedicines for active targeting: physico-chemical characterization of paclitaxel-loaded anti-HER2 immunonanoparticles and in vitro functional studies on target cells.

Paclitaxel (Tx)-loaded anti-HER2 immunonanoparticles (NPs-Tx-HER) were prepared by the covalent coupling of humanized monoclonal anti-HER2 antibodies (trastuzumab, Herceptin) to Tx-loaded poly (dl-lactic acid) nanoparticles (NPs-Tx) for the active targeting of tumor cells that overexpress HER2 receptors. The physico-chemical properties of NPs-Tx-HER were compared to unloaded immunonanoparticles (NPs-HER) to assess the influence of the drug on anti-HER2 coupling to the NP surface. The immunoreactivity of sulfo-MBS activated anti-HER2 mAbs and the in vitro efficacy of NPs-Tx-HER were tested on SKOV-3 ovarian cancer cells that overexpress HER2 antigens. Tx-loaded nanoparticles (NPs-Tx) obtained by a salting-out method had a size of 171+/-22 nm (P.I.=0.1) and an encapsulation efficiency of about of 78+/-10%, which corresponded to a drug loading of 7.8+/-0.8% (w/w). NPs-Tx were then thiolated and conjugated to activated anti-HER2 mAbs to obtain immunonanoparticles of 237+/-43 nm (P.I.=0.2). The influence of the activation step on the immunoreactivity of the mAbs was tested on SKOV-3 cells using 125I-radiolabeled mAbs, and the activity of the anti-HER2 mAbs was minimally affected after sulfo-MBS functionalization. Approximately 270 molecules of anti-HER2 mAbs were bound per nanoparticle. NPs-Tx-HER exhibited a zeta potential of 0.2+/-0.1 mV. The physico-chemical properties of the Tx-loaded immunonanoparticles were very similar to unloaded immunonanoparticles, suggesting that the encapsulation of the drug did not influence the coupling of the mAbs to the NPs. No drug loss was observed during the preparation process. DSC analysis showed that encapsulated Tx is in an amorphous or disordered-crystalline phase. These results suggest that Tx is entrapped in the polymeric matrix and not adsorbed to the surface of the NPs. In vitro studies on SKOV-3 ovarian cancer cells demonstrated the greater cytotoxic effect of NPs-Tx-HER compared to other Tx formulations. The results showed that at 1 ng Tx/ml, the viability of cells incubated with drug encapsulated in NP-Tx-HER was lower (77.32+/-5.48%) than the viability of cells incubated in NPs-Tx (97.4+/-12%), immunonanoparticles coated with Mabthera, as irrelevant mAb (NPs-Tx-RIT) (93.8+/-12%) or free drug (92.3+/-9.3%).

Configurations de proches et adaptation dans le parcours de vie d'individus en situation de vulnérabilité

L'importance des proches et le rôle primordial des relations familiales pour l'adaptation et l'ajustement psychologique des individus sont largement reconnus. Cependant, peu d'études ont considéré que, dans la modernité avancée, les réseaux de proches ou familiaux sont diversifiés, caractérisés par des interdépendances complexes. La littérature sur les relations interpersonnelles et l'adaptation a négligé cette diversité et omis que les interdépendances structurales, de soutien mais aussi de conflit, pouvaient s'avérer importantes à considérer en regard de l'adaptation et de l'ajustement psychologique des individus. La présente thèse apporte des éléments de compréhension sur le lien entre les configurations de proches et l'adaptation individuelle dans le parcours de vie, en se centrant sur des individus en situation de vulnérabilité, que ce soit parce qu'ils font l'expérience d'une transition importante de leur parcours, ou en raison d'un manque de ressources psychologiques, cognitives et/ou sociales. Des résultats empiriques éclairent quatre grands axes de questionnement portant sur les ressources et le stress présents dans les interdépendances au sein des configurations familiales et de proches, sur leur association avec l'ajustement psychologique des individus, sur leur évolution au cours du temps, ainsi que sur la façon dont ces interdépendances sont façonnées par les trajectoires de vie. Les résultats mettent en évidence une diversité de configurations familiales ou de proches, qui donne lieu à des degrés inégaux de ressources, significativement associées à l'ajustement psychologique. Cependant, les interdépendances au sein des configurations ne constituent que rarement des ressources ou un facteur protecteur pour les individus en situation de vulnérabilité mais, au contraire, elles génèrent du stress supplémentaire. L'exploration longitudinale met en évidence une importante instabilité dans les configurations familiales et de proches des individus en situation de vulnérabilité qui compromet leur adaptation et leur ajustement psychologique.