Evolutionary dynamics of interlinked public goods traits: an experimental study of siderophore production in Pseudomonas aeruginosa.

Public goods cooperation is common in microbes, and there is much interest in understanding how such traits evolve. Research in recent years has identified several important factors that shape the evolutionary dynamics of such systems, yet few studies have investigated scenarios involving interactions between multiple public goods. Here, we offer general predictions about the evolutionary trajectories of two public goods traits having positive, negative or neutral regulatory influence on one another's expression, and we report on a test of some of our predictions in the context of Pseudomonas aeruginosa's production of two interlinked iron-scavenging siderophores. First, we confirmed that both pyoverdine and pyochelin siderophores do operate as public goods under appropriate environmental conditions. We then tracked their production in lines experimentally evolved under different iron-limitation regimes known to favour different siderophore expression profiles. Under strong iron limitation, where pyoverdine represses pyochelin, we saw a decline in pyoverdine and a concomitant increase in pyochelin - consistent with expansion of pyoverdine-defective cheats derepressed for pyochelin. Under moderate iron limitation, pyochelin declined - again consistent with an expected cheat invasion scenario - but there was no concomitant shift in pyoverdine because cross-suppression between the traits is unidirectional only. Alternating exposure to strong and moderate iron limitation caused qualitatively similar though lesser shifts compared to the constant-environment regimes. Our results confirm that the regulatory interconnections between public goods traits can significantly modulate the course of evolution, yet also suggest how we can start to predict the impacts such complexities will have on phenotypic divergence and community stability.

Levels and determinants of inflammatory biomarkers in a Swiss population-based sample (CoLaus Study).

OBJECTIVE: to assess the levels and determinants of interleukin (IL)-1ß, IL-6, tumour necrosis factor (TNF)-a and C-reactive protein (CRP) in a healthy Caucasian population.METHODS: population sample of 2884 men and 3201 women aged 35 to 75. IL-1ß, IL-6 and TNF-a were assessed by a multiplexed particle-based flow cytometric assay and CRP by an immunometric assay.RESULTS: Spearman rank correlations between duplicate cytokine measurements (N?=?80) ranged between 0.89 and 0.96; intra-class correlation coefficients ranged between 0.94 and 0.97, indicating good reproducibility. Among the 6085 participants, 2289 (37.6%), 451 (7.4%) and 43 (0.7%) had IL-1ß, IL-6 and TNF-a levels below detection limits, respectively. Median (interquartile range) for participants with detectable values were 1.17 (0.48-3.90) pg/ml for IL-1ß; 1.47 (0.71-3.53) pg/ml for IL-6; 2.89 (1.82-4.53) pg/ml for TNF-a and 1.3 (0.6-2.7) ng/ml for CRP. On multivariate analysis, greater age was the only factor inversely associated with IL-1ß levels. Male sex, increased BMI and smoking were associated with greater IL-6 levels, while no relationship was found for age and leisure-time PA. Male sex, greater age, increased BMI and current smoking were associated with greater TNF-a levels, while no relationship was found with leisure-time PA. CRP levels were positively related to age, BMI and smoking, and inversely to male sex and physical activity.CONCLUSION: Population-based levels of several cytokines were established. Increased age and BMI, and to a lesser degree sex and smoking, significantly and differentially impact cytokine levels, while leisure-time physical activity has little effect.

CELLULAR AND MOLECULAR STUDY OF SUBCORTICAL BAND HETEROTOPIA IN THE MOUSE MUTANT HECO

The HeCo mouse model is characterized by a subcortical heterotopia formed by misplaced neurons normally migrating into the superficial cortical layers. The mutant mouse has a tendency to epileptic seizures. In my thesis project we discovered the mutated Eml1 gene, a member of the echinoderm microtubule-associated protein (EMAP) family, in HeCo as well as in a family of three children showing complex malformation of cortical development. This discovery formed an important step in exploring the pathogenic mechanisms underlying the HeCo phenotype. In vitro results showed that during cell division the EML1 protein is associated with the midbody and a mutated version of Eml1 highlighted an important role of the protein in the astral MT array during cell cycle. In vivo, we found that already at an early age of cortical development (E13), ectopic progenitors such as RGs (PAX6) and IPCs (TBR2) accumulate in the IZ along the entire neocortex. We demonstrated that in the VZ of the HeCo mouse, spindle orientation and cell cycle exit are perturbed. In later stages (E17), RG fibers are strongly disorganized with deep layer (TBR1) and upper layer (CUX1) neurons trapped within an ectopic mass. At P3, columns of upper layer neurons were present between the heterotopia and the developing cortex; these columns were also present at P7 but at lesser extent. Time lapse video recording (E15.5) revealed that the parameters characterizing the migration of individual neurons are not disturbed in HeCo; however, this analysis showed that the density of migrating neuron was smaller in HeCo. In conclusion, truncated EML1 is likely to play a prominent role during cell cycle but also acts on the cytoskeletal architecture altering the shape of RG fibers thus influencing the pattern of neuronal migration. The signal transduction between external cues and intracellular effector pathways through MTs may be secondary but sustains the heterotopia development and further studies are needed to clarify the impact of EML1 in progenitors versus post-mitotic cells.

Photoreceptor types and distributions in the retinae of insectivores.

The retinae of insectivores have been rarely studied, and their photoreceptor arrangements and expression patterns of visual pigments are largely unknown. We have determined the presence and distribution of cones in three species of shrews (common shrew Sorex araneus, greater white-toothed shrew Crocidura russula, dark forest shrew Crocidura poensis; Soricidae) and in the lesser hedgehog tenrec Echinops telfairi (Tenrecidae). Special cone types were identified and quantified in flattened whole retinae by antisera/antibodies recognizing the middle-to-long-wavelength-sensitive (M/L-)cone opsin and the short-wavelength-sensitive (S-)cone opsin, respectively. A combination of immunocytochemistry with conventional histology was used to assess rod densities and cone/rod ratios. In all four species the rods dominate at densities of about 230,000-260,000/mm2. M/L- and S-cones are present, comprising between 2% of the photoreceptors in the nocturnal Echinops telfairi and 13% in Sorex araneus that has equal diurnal and nocturnal activity phases. This suggests dichromatic color vision like in many other mammals. A striking feature in all four species are dramatically higher S-cone proportions in ventral than in dorsal retina (0.5% vs. 2.5-12% in Sorex, 5-15% vs. 30-45% in Crocidura poensis, 3-12% vs. 20-50% in Crocidura russula, 10-30% vs. 40-70% in Echinops). The functional and comparative aspects of these structural findings are discussed.

Cellular immune responses against mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) infection is known to have two main outcomes: latent infection (LTBI) where the pathogen is in a dormant form or active tuberculosis disease (TB), which is, most of the time, highly transmissible. Over one-third of the world's population asymptomatically harbours a latent form of Mtb with a 10% risk of disease reactivation. Efficient vaccine strategies remain unknown and the existing BCG vaccine is believed to protect against only some forms of TB (extra-pulmonary TB in children). Moreover, timely identification of TB remains complex with the actual diagnosis based on clinical observations associated to low efficient tests. Furthermore, current therapies are expensive, heavy and long for patients, and present lesser and lesser efficiency against new drug-resistant strains of Mtb. It is thus important to develop our knowledge on host -Mtb relationship to propose new vaccines, diagnosis tools and medications for the future. This thesis aims at improving our understanding of human immunology in the field of TB. All along this work, the same algorithm has been used and points towards the discovery of new correlates of protection through the comparison of T-cell immune responses in patients with LTBI or TB. We performed a comprehensive analysis of T-cell immune responses to Mtb using polychromatic flow cytometiy to study the functional profile of Μ/ό-specific CD4 Τ cells. We observed a polyfunctional profile in LTBI where CD4 Τ cells mainly co-produced IFN-γ, TNF-α and IL-2. In contrast, in TB, Mtó-specific CD4 Τ cells were mostly single TNF-a positive. Thus, analysis of the cytokine profiles was a strong immunological measure discriminating TB and LTBI. We next analyzed Thl7 cells. Mtò-specific Thl7 cells lacked immediate {i.e. ex vivo) IL-17A effector function in both LTBI and TB individuals. Moreover, they were also absent in bronchoalveolar lavages (BALs). Interestingly, we noticed that Mtb- specific Thl7 cells from LTBI but not from TB subjects acquired the ability to produce IL- 17A following Mtb-specific T-cell expansion. We finally performed a comprehensive characterization of Mfè-specific CD8 Τ cells that were detected in most (60%) TB patients and few (15%) LTBI subjects. We observed differences in the phenotype, the cytotoxicity and the proliferative capacities but not in the cytokine profile of Mtò-specific CD8 Τ cells between LTBI and TB. We concluded that the activity of Mtb infection (i.e. latent versus active) and the clinical presentation were associated to distinct profiles of Mtó-specific CD8 T-cell responses. To conclude, a multiparametric analysis including both CD4 and CD8 T-cell responses to Mtb lead to the development of a significantly improved diagnostic test discriminating between LTBI and TB. All together, these results provide new insights into the interaction between Mtb and the host immune response and expand upon our prior knowledge of tuberculosis. - L'infection par Mycobacterium tuberculosis peut résulter en une infection tuberculeuse latente et asymptomatique ou encore en une forme active et la plupart du temps contagieuse, la tuberculose. Un tiers de la population mondiale serait infectée de manière chronique avec 10 % de risques de développer la maladie durant la vie. Il n'existe actuellement aucun vaccin efficace, le BCG ne conférant qu'une protection partielle contre certaines formes extrapulmonaires de la maladie chez l'enfant. D'autre part, il n'existe pas de méthode diagnostique fiable et rapide, celle-ci se basant dans un premier temps sur l'analyse de la situation clinique des patients. Enfin, les thérapies actuelles sont couteuses et contraignantes pour les patients et tendent à ne plus être efficaces contre les souches émergentes de mycobactérie multi-résistantes. Aussi, il est important de bien comprendre la relation hôte-pathogène de manière à pouvoir proposer de nouveaux outils vaccinaux, diagnostiques et thérapeutiques. Ce manuscrit s'inscrit dans cette direction et vise à améliorer nos connaissances de la réponse immunitaire humaine dans le cadre de la tuberculose. Nous avons suivi un algorithme similaire tout au long des études proposées en comparant les réponses immunes des patients latents à celles des patients actifs, et ce, dans le but de mettre en évidence de potentiels corrélats de protection. Nous avons réalisé par cytométrie en flux une analyse du profil fonctionnel des cellules lymphocytaires CD4 dans la réponse au pathogène. Dans le cas de la tuberculose active, les cellules CD4 sécrètent majoritairement du TNF-α quand, au contraire, elles sécrètent à la fois du TNF-α, de l'IFN-γ et de l'IL-2 (poly-fonctionnalité) dans l'infection latente. Cette observation nous a permis de proposer un nouveau test diagnostique de la maladie active. Nous avons aussi étudié les cellules CD4 Thl7, impliquées dans la réponse immunitaire cellulaire contre les pathogènes extracellulaires et les champignons. Nous avons souligné une variation dans la production d'IL-17 entre infection latente et tuberculose active qui pourrait être impliquée dans la protection de l'individu contre le pathogène. D'autre part, ce manuscrit propose une caractérisation des cellules Τ CD8 dites cytotoxiques dans la tuberculose. Des divergences dans la fréquence des réponses observées, le phénotype mais aussi les capacités prolifératives et cytotoxiques ont pu être mises en évidence entre latence et tuberculose active. Ces observations soulignent le rôle important de ce groupe cellulaire dans l'évolution de la maladie et permettent de proposer une amélioration de l'outil diagnostic précédemment proposé et se basant à la fois sur le profil fonctionnel des cellules Τ CD4 ainsi que sur la présence potentielle d'une réponse CD8 spécifique au pathogène. Ces diverses études réalisées sur les cellules Τ humaines répondant spécifiquement à Mtb nous permettent de faire un pas supplémentaire dans la compréhension de notre réponse immunitaire face à ce pathogène particulièrement dangereux qui continue à l'heure actuelle à tuer chaque année des millions de personnes. - La tuberculose (TB) résulte d'une infection bactérienne par Mycobacterium tuberculosis (Mtb) et existe sous deux formes majeures: une forme latente, lorsque la bactérie est en phase de dormance ainsi qu'une forme active durant laquelle la bactérie se divise activement, entraînant les symptômes de la maladie. La personne infectée devient alors contagieuse dans la plupart des cas. Aujourd'hui des études épidémiologiques assument que plus d'un tiers de la population mondiale serait infectée par la forme latente de la bactérie et que 10% des cas réactiveront donnant lieu à diverses présentations de la maladie. Il n'existe actuellement aucun vaccin réellement efficace chez l'adulte. D'autre part, les traitements antibiotiques utilisés sont très lourds pour les patients et les cliniciens doivent faire face à l'émergence de nouvelles souches bactériennes multi-résistantes non affectées par les thérapies existantes. Les autorités sanitaires sont, d'autre part, confrontées à l'absence d'un outil diagnostique rapide, fiable et efficace. En effet, la méthode de référence reste la culture microbiologique du pathogène qui prend généralement plusieurs semaines, pendant lesquelles le patient pourra contaminer d'autres personnes. En résumé, la lutte contre la tuberculose doit passer par l'élaboration d'un vaccin efficace, de nouvelles thérapies, mais aussi par la mise en place de nouveaux tests diagnostics plus rapides afin d'éviter la dissémination de la maladie. Aussi, la relation hôte-bactérie qui n'est actuellement que peu comprise doit être investiguée. Ce travail de thèse a pour but d'étudier la réponse immunitaire chez l'homme infecté par Mtb et vise plus particulièrement l'étude d'une population clé de cellules immunitaires: les lymphocytes T. L'étude des cellules Τ CD4 nous a permis dans un premier temps de proposer un nouveau test diagnostic de la maladie active. Nous avons aussi analysé plus en détail une population spécifique des cellules Τ CD4 (les cellules Thl7), nous permettant d'associer leur fonction avec un possible état physiologique de protection contre le pathogène. En second lieu nous avons réalisé une caractérisation des cellules Τ CD8, à la fois chez les personnes avec des infections latentes et chez les personnes malades. Nous avons mis en évidence des différences fonctionnelles chez les deux groupes de patients, nous permettant ainsi une meilleure compréhension de l'immunité contre Mtb. Enfin, nous avons combiné les différents profils immunologiques obtenus pour développer un test diagnostic plus performant et sensible que celui proposé antérieurement. Ces diverses études réalisées sur les cellules Τ humaines nous permettent de faire un pas supplémentaire dans la compréhension de la réponse immunitaire face à ce pathogène particulièrement dangereux qui continue à tuer chaque année des millions de personnes.

Structure of the ovaries of the Nimba otter shrew, Micropotamogale lamottei, and the Madagascar hedgehog tenrec, Echinops telfairi.

The otter shrews are members of the subfamily Potamogalinae within the family Tenrecidae. No description of the ovaries of any member of this subfamily has been published previously. The lesser hedgehog tenrec, Echinops telfairi, is a member of the subfamily Tenrecinae of the same family and, although its ovaries have not been described, other members of this subfamily have been shown to have ovaries with non-antral follicles. Examination of these two species illustrated that non-antral follicles were characteristic of the ovaries of both species, as was clefting and lobulation of the ovaries. Juvenile otter shrews range from those with only small follicles in the cortex to those with 300- to 400-microm follicles similar to those seen in non-pregnant and pregnant adults. As in other species, most of the growth of the oocyte occurred when follicles had one to two layers of granulosa cells. When larger follicles became atretic in the Nimba otter shrew, hypertrophy of the theca interna produced nodules of glandular interstitial tissue. In the tenrec, the hypertrophying theca interna cells in most large follicles appeared to undergo degeneration. Both species had some follicular fluid in the intercellular spaces between the more peripheral granulosa cells. It is suggested that this fluid could aid in separation of the cumulus from the remaining granulosa at ovulation. The protruding follicles in lobules and absence of a tunica albuginea might also facilitate ovulation of non-antral follicles. Ovaries with a thin-absent tunica albuginea and follicles with small-absent antra are widespread within both the Eulipotyphla and in the Afrosoricida, suggesting that such features may represent a primitive condition in ovarian development. Lobulated and deeply crypted ovaries are found in both groups but are not as common in the Eulipotyphla making inclusion of this feature as primitive more speculative.

Methadone enantiomer plasma levels, CYP2B6, CYP2C19, and CYP2C9 genotypes, and response to treatment.

BACKGROUND AND OBJECTIVE: Recent in vitro studies have suggested an important role of cytochrome P450 (CYP) 2B6 and CYP2C19 in methadone metabolism. We aimed to determine the influence of CYP2B6, CYP2C9, and CYP2C19 genetic polymorphism on methadone pharmacokinetics and on the response to treatment. METHODS: We included 209 patients in methadone maintenance treatment on the basis of their response to treatment and their daily methadone dose. Patients were genotyped for CYP2B6, CYP2C9, and CYP2C19. Steady-state trough and peak (R)-, (S)-, and (R,S)-plasma levels and peak-to-trough plasma level ratios were measured. RESULTS: CYP2B6 genotype influences (S)-methadone and, to a lesser extent, (R)-methadone plasma levels, with the median trough (S)-methadone plasma levels being 105, 122, and 209 ng . kg/mL . mg for the noncarriers of allele *6, heterozygous carriers, and homozygous carriers (*6/*6), respectively (P = .0004). CYP2C9 and CYP2C19 genotypes do not influence methadone plasma levels. Lower peak and trough plasma levels of methadone and higher peak-to-trough ratios were measured in patients considered as nonresponders [median (R,S)-methadone trough plasma levels of 183 and 249 ng . kg/mL . mg (P = .0004) and median peak-to-trough ratios of 1.82 and 1.58 for high-dose nonresponders and high-dose responders, respectively (P = .0003)]. CONCLUSION: Although CYP2B6 influences (S)-methadone plasma levels, given that only (R)-methadone contributes to the opioid effect of this drug, a major influence of CYP2B6 genotype on response to treatment is unlikely and has not been shown in this study. Lower plasma levels of methadone in nonresponders, suggesting a higher clearance, and higher peak-to-trough ratios, suggesting a shorter elimination half-life, are in agreement with the usual clinical measures taken for such patients, which are to increase methadone dosages and to split the daily dose into several intakes.

Osteoid osteoma-like lesions: MDCT features and treatment by radiofrequency ablation : 46

Purpose: To assess the MDCT features of bone lesions that mimic osteoid osteoma (OO-like lesions) and evaluate their treatment by radiofrequency (RF) ablation. Methods and materials: All percutaneous RF ablations performed between May 2002 and June 2009 for a presumed (clinical and MDCT features) diagnosis of OO were retrospectively reviewed. Per-procedural biopsies were always performed and histopathological diagnoses were noted. The following MDCT features of all bone lesions were assessed by two musculoskeletal radiologists in consensus: skeletal distribution and location within the bone, size, central calcification, surrounding osteosclerosis and periosteal reaction. Clinical success was also evaluated. Results: Eighty patients (54 males, 26 females, mean age 24.1 years, range 5-48) underwent RF ablation. The histopathological diagnoses were: 54 non-contributory biopsies, 16 OO, 10 OO-like lesions (5 chronic osteomyelitis, 3 chondroblastoma, 1 eosinophilic granuloma, 1 fibrous dysplasia). The OO-like lesions were significantly greater in size (p = 0.001) and exhibited trends toward medullary location within the bone, moderate surrounding osteosclerosis and less periosteal reaction, compared to OO. Primary clinical success for OO-like lesions was 100% at 1 month, 85.7% at 6 and 12 months, and 66.7% at 24 months. Secondary success was 100%. Conclusion: Greater size, medullary location within the bone, lesser surrounding osteosclerosis and periosteal reaction on MDCT may help differentiate OO-like lesions from OO. OO-like lesions are safely and successfully treated by RF ablation.

An essential role for transmembrane TNF in the resolution of the inflammatory lesion induced by Leishmania major infection.

TNF is an essential player in infections with Leishmania major, contributing to the control of the inflammatory lesion and, to a lesser degree, to parasite killing. However, the relative contribution of the soluble and transmembrane forms of TNF in these processes is unknown. To investigate the role of transmembrane TNF (mTNF) in the control of L. major infections, mTNF-knock-in (mTNF(Delta/Delta)) mice, which express functional mTNF but do not release soluble TNF, were infected with L. major, and the development of the inflammatory lesion and the immune response was compared to that occurring in L. major-infected TNF(-/-) and wild-type mice. mTNF(Delta/Delta) mice controlled the infection and resolved their inflammatory lesion as well as wild-type mice, a process associated with the early clearance of neutrophils at the site of parasite infection. In contrast, L. major-infected TNF(-/-) mice developed non-healing lesions, characterized by an elevated presence of neutrophils at the site of infection and partial control of parasite number within the lesions. Altogether, the results presented here demonstrate that mTNF, in absence of soluble TNF, is sufficient to control infection due to L. major, enabling the regulation of inflammation, and the optimal killing of Leishmania parasites at the site of infection.

Effects of 2 different prior endurance exercises on whole-body fat oxidation kinetics: light vs. heavy exercise.

This study aimed to compare the effects of 2 different prior endurance exercises on subsequent whole-body fat oxidation kinetics. Fifteen men performed 2 identical submaximal incremental tests (Incr2) on a cycle ergometer after (i) a ∼40-min submaximal incremental test (Incr1) followed by a 90-min continuous exercise performed at 50% of maximal aerobic power-output and a 1-h rest period (Heavy); and (ii) Incr1 followed by a 2.5-h rest period (Light). Fat oxidation was measured using indirect calorimetry and plotted as a function of exercise intensity during Incr1 and Incr2. A sinusoidal equation, including 3 independent variables (dilatation, symmetry and translation), was used to characterize the fat oxidation kinetics and to determine the intensity (Fat(max)) that elicited the maximal fat oxidation (MFO) during Incr. After the Heavy and Light trials, Fat(max), MFO, and fat oxidation rates were significantly greater during Incr2 than Incr1 (p < 0.001). However, Δ (i.e., Incr2-Incr1) Fat(max), MFO, and fat oxidation rates were greater in the Heavy compared with the Light trial (p < 0.05). The fat oxidation kinetics during Incr2(Heavy) showed a greater dilatation and rightward asymmetry than Incr1(Heavy), whereas only a greater dilatation was observed in Incr2(Light) (p < 0.05). This study showed that although to a lesser extent in the Light trial, both prior exercise sessions led to an increase in Fat(max), MFO, and absolute fat oxidation rates during Incr2, inducing significant changes in the shape of the fat oxidation kinetics.

Hypoxia: unique myocardial morphology?

OBJECTIVE: The objective of this study was to investigate the effects of chronic and intermittent hypoxia on myocardial morphology. METHODS: Rats randomly divided into 3 groups (n = 14 per group) were exposed to room air (Fio(2) = 0.21), chronic hypoxia (Fio(2) = 0.10), and intermittent hypoxia (chronic hypoxia with 1 hour per day of room air) for 2 weeks. Weight, blood gas analysis, hematocrit, hemoglobin, red cells, and right and left ventricular pressures were measured. Hearts excised for morphologic examination were randomly divided into 2 groups (9 per group for gross morphologic measurements and 5 per group for histologic and morphometric analysis). The weight ratio of right to left ventricles plus interventricular septum, myocyte diameter, cross-sectional area, and free wall thickness in right and left ventricles were measured. RESULTS: Despite the same polycythemia, the right ventricle pressure (P <.05) and ratio of right to left ventricle pressures (P <.02) were higher after chronic hypoxia than intermittent hypoxia. The ratio of heart weight to total body weight and the ratio of right to left ventricles plus interventricular septum was higher (P <.01) in chronic and intermittent hypoxia than in normoxia. Myocyte diameter was not different between the right and left ventricles in normoxia, whereas right ventricle myocytes were larger than left ventricle myocytes in chronic hypoxia (P <.05) and intermittent hypoxia (P <.0005). There was marked dilatation of right ventricle size (P <.001) and marked reduction of left ventricle (P <.001) size in chronic and intermittent hypoxia compared with normoxia. The total ventricular area (right ventricle plus left ventricle area) remained the same in all groups. The wall thickness ratio in chronic hypoxia and intermittent hypoxia was increased (P <.001) compared with normoxia in the right ventricle but not in the left ventricle. CONCLUSIONS: Intermittent reoxygenation episodes do not induce a lesser ventricular hypertrophic response than observed with chronic hypoxia. The functional myocardial preconditioning consequence of intermittent reoxygenation is not supported by structural differences evident with the available techniques.

Chirurgie du pied et de la cheville rhumatoïdes [Surgery of the rheumatoid foot and ankle]

Surgery of the rheumatoid foot and ankle Initial presentation of rheumatoid arthritis (RA) will be foot and ankle symptoms in 20% of cases. Eventually, 50-90% of patients will develop foot and ankle alterations. Typical deformities of the forefoot include hallux valgus and dorso-lateral dislocation of the lesser metatarso-phalangeal joints. RA involvement of the hindfoot frequently leads to plano-valgus deformity. Approximately 20% of surgery performed for RA is done at the foot and ankle. Early surgery is efficient against pain and deformity. Patient's satisfaction rate after surgery is high despite a slight higher complication rate compared to conventional foot and ankle surgery.

Statistical significance of quantitative PCR.

BACKGROUND: PCR has the potential to detect and precisely quantify specific DNA sequences, but it is not yet often used as a fully quantitative method. A number of data collection and processing strategies have been described for the implementation of quantitative PCR. However, they can be experimentally cumbersome, their relative performances have not been evaluated systematically, and they often remain poorly validated statistically and/or experimentally. In this study, we evaluated the performance of known methods, and compared them with newly developed data processing strategies in terms of resolution, precision and robustness. RESULTS: Our results indicate that simple methods that do not rely on the estimation of the efficiency of the PCR amplification may provide reproducible and sensitive data, but that they do not quantify DNA with precision. Other evaluated methods based on sigmoidal or exponential curve fitting were generally of both poor resolution and precision. A statistical analysis of the parameters that influence efficiency indicated that it depends mostly on the selected amplicon and to a lesser extent on the particular biological sample analyzed. Thus, we devised various strategies based on individual or averaged efficiency values, which were used to assess the regulated expression of several genes in response to a growth factor. CONCLUSION: Overall, qPCR data analysis methods differ significantly in their performance, and this analysis identifies methods that provide DNA quantification estimates of high precision, robustness and reliability. These methods allow reliable estimations of relative expression ratio of two-fold or higher, and our analysis provides an estimation of the number of biological samples that have to be analyzed to achieve a given precision.

Minerais, métaux, isotopes : recherches archéométriques sur les mines de plomb et d'argent en Valais, Suisse

RESUME Les nombreuses mines de plomb et d'argent du Valais témoignent d'une activité minière importante par le passé, sans toutefois dévoiler ni l'importance des minéralisations, ni l'ancienneté de l'exploitation. La présente recherche a pour but de comprendre pourquoi les grandes mines sont concentrées dans une région, et de déterminer la chronologie de leur exploitation. L'originalité de ce travail réside dans son interdisciplinarité, plus précisément dans l'application des méthodes minéralogiques pour résoudre une problématique historique. Afin d'évaluer les ressources minières en plomb et en argent du Valais, 57 mines et indices ont été repérés et échantillonnés. Les signatures isotopiques du Pb (74 analyses) et les compositions chimiques élémentaires (45 analyses) ont été déterminées. Les plus grandes exploitations se situent dans la nappe de Siviez-Mischabel, au Sud d'une ligne Vallée du Rhône / Val de Bagnes ainsi que dans le Lötschental. Elles sont liées, d'après leur signature isotopique de plomb, à des minéralisations d'âge calédonien (408 à 387 Ma) ou tardi-hercynien (333 à 286 Ma). À ces périodes, l'ancien continent est très lourd et subit une subsidence thermique. Des premières fractures d'extrême importance se forment. Comme il s'agit d'accidents tectoniques majeurs, des gisements de grande extension peuvent se former dans ce contexte. D'autres minéralisations se situent dans les domaines helvétiques (Massif des Aiguilles Rouges, Massif du Mont Blanc et couverture sédimentaire), couvrant une région au Nord de la Vallée du Rhône et du Val d'Entremont. D'âge post-hercynien à tardi-alpin (notons qu'il n'y a pas de minéralisations d'âge tertiaire), elles sont pour la plupart liées à des intrusions granitiques, sources de plomb juvénile. Les mines situées dans ces unités tectoniques sont nettement moins étendues que celles de la nappe de Siviez-Mischabel, ce qui permet de penser que les minéralisations correspondantes le sont également. Les périodes d'exploitation des mines peuvent être déterminées par quatre approches différentes l'archéologie minière, la lecture des textes historiques, l'étude des déchets métallurgiques et la comparaison de la signature isotopique du plomb, que l'on mesure dans un objet archéologique bien daté (monnaie, bijoux etc.), avec celles des minerais. Cette dernière méthode a été appliquée et développée dans le cadre de la présente recherche. Pour ce faire, 221 échantillons d'objet en plomb ou en argent datés entre l'Âge du Fer et le Moyen Age ont été analysés par la méthode des isotopes de plomb et comparés à environ 1800 signatures isotopiques de minerais des gisements les plus importants en Suisse et en Europe. Avant l'époque romaine et jusqu'au 1 er siècle de cette époque, le plomb provient principalement des mines de la péninsule ibérique alors en pleine activité. Un apport des mines d'Europe tempérée, notamment des Vosges, reste à confirmer. A partir du 1" siècle de notre ère, le plomb a principalement été importé en Suisse occidentale de grands centres de productions situées en Allemagne du Nord (région d'Eifel). Les mines de plomb valaisannes, notamment celles de Siviez, débutent leur exploitation en même temps, principalement pour couvrir les besoins locaux, mais également pour l'exportation jusque dans l'arc lémanique et, dans une moindre importance, au-delà. À partir du 4ème siècle, le besoin en plomb a été couvert par un apport des mines locales et par la refonte d'objets anciens. Ce changement d'approvisionnement est probablement lié aux tensions créées par les invasions germaniques durant la seconde moitié du 3' siècle ; le marché suisse n'est dès lors plus approvisionné par le nord, c'est-à-dire par la vallée du Rhin. Quant à l'argent, l'exploitation de ce métal est attestée à partir de la fin du La Tène, peu après l'apparition de ce métal dans la région valaisanne. L'échantillonnage ne couvrant pas l'époque romaine, rien n'est connu pour cette période. A partir du 5" siècle, une exploitation d'argent est de nouveau attestée. Cependant, l'exploitation d'argent des mines locales ne gagne en importance qu'à partir du Moyen Âge avec les frappes monétaires, notamment les frappes carolingiennes et épiscopales valaisannes. Les sources d'argent sont différentes selon leur utilisation : à part quelques exceptions notamment vers la fin du La Tène et au tardo-antique, les bijoux et objets de cultes ont été souvent créés à partir d'argent refondu, contrairement aux monnaies pour lesquelles l'argent provient des mines locales. On note un approvisionnement différent de ce métal pour les objets, notamment les monnaies, selon leur lieu de fabrication : on peut clairement distinguer les objets valaisans de ceux du Plateau Suisse. SUMMARY The many lead and silver mines of the Valais testify of an important mining activity in the past, without however revealing neither the importance of the mineralizations, nor the era of the exploitation. The purpose of this research is to understand why the large mines are concentrated in one region, and to determine the history of their exploitation. The uniqueness of this work lies in its interdisciplinarity, more precisely in the application of mineralogical methods to solve historical problems. In order to evaluate the lead and silver mining resources of the Valais region, 57 mines and ore deposits were located and sampled. The isotope signatures of Pb (74 analyses) and the compositions of the chemical elements (45 analyses) were determined. The largest activities are in the Siviez-Mischabel area, located in the South of the boundary formed by the Rhone, Bagnes and Lotschental valleys. According to their lead isotope signatures, they are linked to mineralizations of the Caledonian (408 to 387 my) or tardi-Hercynian (333 to 286 my) orogenies. In those times, the old continent was very heavy and underwent a thermal subsidence. First fractures of great significance were formed. Through these major tectonic events, large extended ore deposits can be formed. Other mineralizations are found in the helvetic regions situated north of the Rhone and the Entremont valley (the Aiguilles Rouges basement, Mount Blanc basement and the covering sediment). Because they are from post-hercynien to tardi-alpine age (there are no mineralizations of tertiary age), they are mainly linked to granite intrusions, the sources of juvenile lead. The mines found in these tectonic units are significantly less extensive than those of the Siviez-Mischabel area, leading to the assumption that the respective mineralizations extend accordingly. The history of exploitation of the mines can be determined by four different sources: mining archaeology, historical texts, metallurgical waste, and the comparison of the isotope signature of the lead from accurately dated archaeological objects (currency, jewels etc), with those of the ores. This last approach was applied and developed within the framework of this research. The lead isotope signatures of 221 lead or silver objects from the Iron Age to the Middle Age were compared with approximately 1800 samples of ore of the most important ore deposits in Switzerland and Europe. Before the Roman time up to the 1st century, lead comes mainly from the mines of the Iberian Peninsula then in full activity. A contribution of the mines of Central Europe, in particular of the Vosges, remains to be confirmed. From the 1st century on, lead was mainly imported into Western Switzerland from Northern Germany (Eiffel region). The lead mines in the Valais region, in particular those of Siviez, begin their exploitation at the same time, mainly to meet the local needs, but also for export to the lemanic basin and of lesser importance, beyond. As from the 4th century, the need of lead was met by the production from local mines and the recycling of old objects. This change of supply is probably related to the tensions created by the Germanic invasions during second half of the 3rd century; as a consequence, the Swiss market is not supplied any more by the north, i.e. the Rhine valley. Silver production is confirmed starting from the end of La Tene, shortly after the appearance of this metal in the Valais region. Since no objects of Roman origin were analyzed, nothing is known for this period. From the 5th century on, silver production is again confirmed. However, significant silver production from local mines starts only in the Middle Age with the coinage, in particular Carolingian and Episcopal minting from the Valais region. The sources of silver differ according to their use: besides some exceptions in particular towards the end of La Tene and the tardi-Roman, the jewels and objects of worships were often created from recycled silver, contrary to the coins the silver for which comes from the local mines. A different source of silver is observed according to the location of coin manufacture: Objects originating from the Valais region are clearly distinguished from those from the Plateau Suisse. ZUSAMMENFASSUNG Die grosse Zahl von Blei- und Silberminen im Wallis ist Zeugnis einer bedeutenden Bergbautätigkeit, es fehlen aber Hinweise über ihren Umfang und den Zeitraum ihrer Ausbeutung. Die vorliegende Arbeit sucht zu ergründen, warum grosse Minen sich in einer eng begrenzten Region häufen und in welchem Zeitraum sie genutzt wurden. Die Besonderheit der Studie liegt in ihrer Interdisziplinarität, genauer in der Anwendung von mineralogischen Methoden zur Beantwortung historischer Fragestellungen. Zur Beurteilung der Lagerstätten wurden von 57 Minen und Aufschlüssen Proben entnommen oder Nachweise erbracht und mittels 74 Isotopen-Analysen von Blei und 45 chemischen Gesamtanalysen ausgewertet. Die wichtigsten Vorkommen liegen in der Siviez- Mischabel- Decke südlich der Linie Rhonetal- Val de Bagnes, sowie im Lötschental. Die Bleiisotopen- Alter weisen ihre Entstehung der kaledonischen (408 - 387 Mio. J.) oder der spät- herzynischen (333 - 286 Mio. J.) Gebirgsbildungsphase zu. In dieser Periode ist die kompakte Landmasse sehr schwer und erfairt eine thermische Absenkung. Es bilden sich tektonische Brüche von kontinentaler Ausdehnung. Die grossen tektonischen Bewegungen ermöglichen die Bildung von ausgedehnten Lagerstätten. Andere Vorkommen finden sich im Bereich der Helvetischen Alpen (Aiguilles Rouges Massiv, Mont-Blanc-Massiv und Sediment-Decken) im Gebiet nördlich des Rhonetales bis zum Val d'Entremont. Altersmässig sind sie der nach-hercynischen bis zur spät-alpidischen Orogenese zuzuweisen (auffällig ist das Fehlen von Vorkommen im Tertiär) und haben sich meist in der Folge von Granit- Intrusion, dem Ursprung von primärem Blei ausgebildet. Die Bergwerke in diesem Bereich sind deutlich weniger ausgedehnt als jene in der Siviez-Mischabel-Decke und entsprechen wahrscheinlich dem geringen Umfang der zugehörigen Vorkommen. Die Nutzungsperioden der Minen können mit vier verschiedenen Methoden bestimmt werden: Minenarchäologie, Historische Quellen, Auswertung von metallischen Abfällen (Schlacken) und Vergleich der Bleiisotopen-Zusammensetzung von Erzen mit jener von zeitlich gut datierbaren archäologischen Gegenständen (Münzen, Schmuckstücke). Die letztere Methode wurde im Rahmen der vorliegenden Forschungsarbeit entwickelt und angewendet. Zu diesem Zweck wurden an 221 Proben von Blei- oder Silberobjekten, die in die Periode zwischen Eisenzeit und Mittelalter eingestuft werden können, Bleiisotopen- Analysen durchgeführt und mit ca. 1800 Proben aus den wichtigsten Lagerstätten der Schweiz und Europas verglichen. Vor der Römerzeit und bis ins 1. Jahrh. stammt das Blei vornehmlich aus den in jener Zeit in voller Ausbeutung begriffenen Minen der Iberischen Halbinsel. Der Beitrag von Mitteleuropa, besonders der Vogesen, muss noch bestätigt werden. Ab dem 1. Jahrh. nach Chr. wurde die Westschweiz hauptschlich mit Blei aus den grossen Produktionszentren Norddeutschlands, vorwiegend der Eifel, versorgt. In dieser Periode setzt die Ausbeutung der Bleiminen des Wallis, besonders von Siviez, ein. Sie dienen der Deckung des örtlichen Bedarfs aber auch der Ausfuhr in das Gebiet des Genfersees und in einem bescheidenen Rahmen sogar darüber hinaus. Ab dem 4. Jahrhundert wurden vermehrt alte Objekte eingeschmolzen. Dieser Wechsel der Versorgungsquellen war vermutlich eine Folge der Wölkerwanderung in der zweiten Hälfte des 3. Jahrhunderts. Ab diesem Zeitpunkt war Helvetien der Zugang zu den Versorgungsquellen des Nordens, besonders des Rheinlandes, verwehrt. Der Abbau von Silber ist ab dem Ende des La Tène nachgewiesen, nur wenig nach dem Auftreten dieses Metalls im Wallis. Über die Römerzeit können wegen dem Fehlen entsprechender Proben keine Aussagen gemacht werden. Eine erneute Abbauperiode ist ab dem 5. Jahrhundert nachgewiesen. Die Produktion der örtlichen Minen erreicht aber erst im Mittelalter eine gewisse Bedeutung mit der Prägung von Mnzen durch die Karolinger und die Walliser Bischöfe. Die Herkunft des Silbers ist abhängig von dessen Verwendung. Mit wenigen Ausnahmen in der Zeit des La Tène und der späteren Römerzeit wurde für Kunst- und Kult- Gegenstände rezykliertes Silber verwendet, für Münzprägungen neues Silber aus den örtlichen Minen. Von Einfluss auf die Herkunft war auch der Produktionsstandort: Die Objekte aus dem Wallis unterscheiden sich deutlich von jenen des Mittellandes.

Cell-permeable peptides induce dose- and length-dependent cytotoxic effects.

We have explored the threshold of tolerance of three unrelated cell types to treatments with potential cytoprotective peptides bound to Tat(48-57) and Antp(43-58) cell-permeable peptide carriers. Both Tat(48-57) and Antp(43-58) are well known for their good efficacy at crossing membranes of different cell types, their overall low toxicity, and their absence of leakage once internalised. Here, we show that concentrations of up to 100 microM of Tat(48-57) were essentially harmless in all cells tested, whereas Antp(43-58) was significantly more toxic. Moreover, all peptides bound to Tat(48-57) and Antp(43-58) triggered significant and length-dependent cytotoxicity when used at concentrations above 10 microM in all but one cell types (208F rat fibroblasts), irrespective of the sequence of the cargo. Absence of cytotoxicity in 208F fibroblasts correlated with poor intracellular peptide uptake, as monitored by confocal laser scanning fluorescence microscopy. Our data further suggest that the onset of cytotoxicity correlates with the activation of two intracellular stress signalling pathways, namely those involving JNK, and to a lesser extent p38 mitogen-activated protein kinases. These responses are of particular concern for cells that are especially sensitive to the activation of stress kinases. Collectively, these results indicate that in order to avoid unwanted and unspecific cytotoxicity, effector molecules bound to Tat(48-57) should be designed with the shortest possible sequence and the highest possible affinity for their binding partners or targets, so that concentrations below 10 microM can be successfully applied to cells without harm. Considering that cytotoxicity associated to Tat(48-57)- and Antp(43-58) bound peptide conjugates was not restricted to a particular type of cells, our data provide a general framework for the design of cell-penetrating peptides that may apply to broader uses of intracellular peptide and drug delivery.