Assessment of seven culture media for the growth and isolation of Capnocytophaga spp.

International audienc

Les plasmocytes et leur niche : étude de la génération de plasmocytes humains in vitro

Tableau d'honneur de la Faculté des études supérieures et postdoctorales, 2016-2017Chez l’humain, les lymphocytes B mémoires IgG+ et IgA+ sont des cellules clés de l’immunité humorale. Ces cellules mémoires sont maintenues à long-terme dans notre organisme. Elles représentent une défense rapide et efficace contre toutes les infections que nous avons déjà vaincues pendant notre vie. Ces cellules mémoires qui rencontrent à nouveau leur antigène se différencient rapidement en plasmocytes à courte vie, et permettent la sécrétion massive d’immunoglobuline (Ig). La contrepartie mémoire de ces cellules sont les plasmocytes à longue vie qui sont présents dans les niches de la moelle osseuse et y sécrètent en permanence des anticorps protecteurs qui circulent dans le sang. Ces cellules sécrétrices peuvent avoir une durée de vie allant de dizaines d’années à la vie entière de l’individu. Les patients qui reçoivent des traitements de chimiothérapie ou de radiothérapie sont privés de ces cellules mémoires détruites par ces traitements au même titre que les cellules cancéreuses. Ces patients deviennent vulnérables aux infections et leur survie dépend de la régénération rapide de leur système hématopoïétique. Notre équipe a déjà mis au point une méthode pour préparer de grandes quantités des cellules mémoires capables de sécréter des IgG et des IgA. Les présents travaux visent à générer des plasmocytes fonctionnels et capables de survivre à long terme in vitro. La stratégie expérimentale visait à établir des conditions permettant de se rapprocher de l’environnement de la moelle osseuse. Dans un premier temps, nous avons étudié les paramètres permettant la différenciation des lymphocytes B mémoires en plasmocytes. Étant donné l’importance du potentiel redox dans l’environnement de la moelle osseuse, nous avons d’abord tenté d’en contrôler l’impact avec un antioxydant, le N-acétyle cystéine (NAC). Nos résultats ont démontré que le NAC avait un effet significatif et diminuait la phosphorylation de la protéine STAT3 en raison d’une inhibition des kinases JAK2 et JAK3. Étonnamment, cet antioxydant retardait la différenciation de nos lymphocytes B qui étaient stimulés avec une forte interaction CD40-CD154. Par la suite, la comparaison des interactions CD40-CD154 et CD27-CD70 a permis de conclure qu’il était essentiel de réduire à son minimum l’interaction CD40-CD154 et qu’il fallait ajouter les cytokines IL-6 et IL-10. Les cellules CD31+CD38+CD138+ générées présentaient un phénotype similaire à celui des plasmocytes de la moelle osseuse. Malheureusement la fréquence de ces cellules était faible et leur viabilité insuffisante. Afin d’augmenter la survie de ces cellules le dernier volet de nos travaux visait à se rapprocher des niches de la moelle osseuse. Notre but a été atteint en ajoutant des cellules mésenchymateuses issues de la moelle osseuse en présence de 8% de dioxygène (O2). Les cellules CD31+CD38+CD138+ générées ont une excellente viabilité et représentent plus de 50% des cellules totales en culture. De plus, le modèle de culture est maintenant établi dans un milieu exempt de sérum et de protéines animales. Dans l’ensemble, nos résultats permettent de proposer la production ex vivo de plasmocytes autologues avec une perspective thérapeutique pour réduire les risques d’infections des patients devenues immunodéficients, suite à un traitement de radiothérapie ou de chimiothérapie.In Humans, IgG+- and IgA+ memory B lymphocytes are key cells for the maintenance of humoral immunity. Memory B lymphocytes are long-lived cells maintained as a memory repertoire throughout our lives. Memory B lymphocytes can establish an efficient and rapid defense against previously encountered infections. These cells rapidly differentiate into short-lived plasma cells secreting high levels of antibodies. Their counterpart, the long-lived plasma cells are the memory populations present in the bone marrow microenvironment. The long-lived plasma cells release protective antibodies into the peripheral blood, maintaining a permanent immune protection. Plasma cells can remain active for years or even for the entire life of an individual. Conversely, patients treated for cancer receive massive doses of chemotherapy or radiotherapy, which are detrimental for immune memory cells as well as cancerous cells. Those patients become highly vulnerable to infectious diseases and their survival depends on the regeneration of their hematopoietic system. Our research group has established an in vitro model enabling to prepare large quantities of IgG and IgA-secreting memory B lymphocytes. The present study aims to further investigate in vitro conditions to generate plasma cells with high survival capacity and able to secrete antibodies. Our experimental strategy intends to establish culture conditions similar to the bone marrow environment. The first step was to study parameters involved into differentiation of memory B lymphocytes into plasma cells. The importance of the redox balance in the bone marrow environment led us to measure the impact of Nacetyl cysteine (NAC), an antioxidant. Our results showed that NAC decreased STAT3 activation by inhibiting the phosphorylation of two kinases namely JAK2 and JAK3. Surprisingly, the addition of NAC had a negative effect on the differentiation of memory B lymphocytes in our culture conditions using a high level of CD40 stimulation. By comparing the levels of CD40-CD154 and CD27-CD70 interactions, we then confirmed that a low level of CD40-CD154 interaction was essential. We also established that addition of IL-6 and IL-10 was better to favor plasma cell generation. The cells obtained in this model were CD31+CD38+CD138+, showing a phenotype close to that of plasma cells found in the bone marrow. Unfortunately, those cells were produced in low frequency and were characterized by a low viability. To increase the survival of these in vitro generated plasma cells, we tried to generate culture conditions that resemble the bone marrow environment. We have achieved this by adding mesenchymal stem cells from bone marrow and maintained the cells at a low O2 level (8%). Cells CD31+CD38+CD138+ obtained at the end of the culture periods showed a high viability, and corresponded to more than 50% of total cultured cells. As expected, those cells were non-proliferating and able to secrete IgG. Furthermore, this in vitro culture model was established with a serum free media. In conclusion, our findings pave the way for the ex vivo production of autologous plasma cell for therapeutic purposes in order to reduce the risks of infection of immune-deficient patients

Un chapiteau du temple oraculaire d’Apollon à Didymes

En 2018, de fructueux échanges entre le département Jardins des résidences présidentielles et le département des Antiquités grecques, étrusques et romaines du musée du Louvre ont conduit à l’identification d’un chapiteau colossal du temple d’Apollon à Didymes : celui-ci avait été rapporté en France en 1873 par Olivier Rayet, à la suite de la grande fouille menée dans la région de Milet, en Asie Mineure, grâce au généreux mécénat d’Edmond et de Gustave de Rothschild. La mémoire de ce bloc s’ét..

INVOLVEMENT OF RESPIRATORY CHAIN IN BIOFILM FORMATION IN PORPHYROMONAS GINGIVALIS

Oral Communication presented at the ";Forum des Jeunes Chercheurs";, Brest (France) 2011

Bone Marrow Mesenchymal Stem Cells Enhance the Differentiation of Human Switched Memory B Lymphocytes into Plasma Cells in Serum-Free Medium

The differentiation of human B lymphocytes into plasma cells is one of the most stirring questions with regard to adaptive immunity. However, the terminal differentiation and survival of plasma cells are still topics with much to be discovered, especially when targeting switched memory B lymphocytes. Plasma cells can migrate to the bone marrow in response to a CXCL12 gradient and survive for several years while secreting antibodies. In this study, we aimed to get closer to niches favoring plasma cell survival. We tested low oxygen concentrations and coculture with mesenchymal stem cells (MSC) from human bone marrow. Besides, all cultures were performed using an animal protein-free medium. Overall, our model enables the generation of high proportions of CD38+CD138+CD31+ plasma cells (≥50%) when CD40-activated switched memory B lymphocytes were cultured in direct contact with mesenchymal stem cells. In these cultures, the secretion of CXCL12 and TGF-β, usually found in the bone marrow, was linked to the presence of MSC. The level of oxygen appeared less impactful than the contact with MSC. This study shows for the first time that expanded switched memory B lymphocytes can be differentiated into plasma cells using exclusively a serum-free medium

Main flexible pavement and mix design methods in Europe and challenges for the development of an european method

Pavement and mix design represent one of the key components within the life cycle of a road infrastructure, with links to political, economic, technical, societal and environmental issues. Recent researches related to the characteristics of materials and associated behavior models both for materials and pavement, made it appropriate to consider updating current pavement design methods, and especially in the USA this has already been in process while in Europe uses of the methods developed in the early 1970s. Thus, this paper firstly presents a brief historical overview of pavement design methods, highlighting early limitations of old empirical methods. Afterwards, French, UK and Shell methods currently in use in Europe will be presented, underlining their main components in terms of methodology, traffic, climatic conditions and subgrade. The asphalt mix design and modeling in Europe are presented with their inclusion in the pavement design methods. Finally, the main challenges for the development of a European pavement design method are presented as well as the recent research developments that can be used for that methodThe second author would like to express the support of Portuguese National Funding Agency for Science, Research and Technology (FCT) through scholarship SFRH/BSAB/114415/ 2016. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.info:eu-repo/semantics/publishedVersio

Human CD38 hi

B lymphocyte differentiation into long-lived plasma cells is the keystone event for the production of long-term protective antibodies. CD40-CD154 and CD27-CD70 interactions are involved in human B lymphocyte differentiation into CD38hiCD138+ cells in vivo as well as in vitro. In this study, we have compared these interactions in their capacity to drive switched-memory B lymphocytes differentiation into CD38hiCD138+ plasma cells. The targeted B lymphocytes were isolated from human peripheral blood, expanded for 19 days, and then submitted to CD70 or CD154 interactions for 14 days. The expanded B lymphocytes were constitutively expressing CD39, whereas CD31’s expression was noticed only following the in vitro differentiation step (day 5) and was exclusively present on the CD38hi cell population. Furthermore, the generated CD38hiCD138+ cells showed a higher proportion of CD31+ cells than the CD38hiCD138- cells. Besides, analyses done with human blood and bone marrow plasma cells showed that in vivo and de novo generated CD38hiCD138+ cells have a similar CD31 expression profile but are distinct according to their reduced CD39 expression level. Overall, we have evidences that in vitro generated plasma cells are heterogeneous and appear as CD39+ precursors to the ones present in bone marrow niches

Evaluación del comportamiento a macro-fisuración por fatiga de mezclas bituminosas modificadas con polvo de neumático

The use of crumb rubber modified bitumen (CRMB) in asphalt mixes is a road engineering technology that has become increasingly important in recent years. Given the many economic and environmental benefits of this type of binder, the goal is to give CRMB the same level of performance as conventional polymermodified bitumen. The appearance and propagation of cracks due to fatigue phenomena is one of the most common distresses affecting road pavements. Since crumb rubber enhances the mechanical properties of asphalt mixes, it can provide a viable solution for fatigue cracking. This paper presents the results of a comparative analysis of the fatigue-cracking behavior of asphalt mixtures manufactured with crumb rubber modified bitumen and polymer-modified bitumen.El empleo de betunes modificados con polvo de neumático usado en la fabricación de mezclas bituminosas es una de las técnicas que mayor auge está teniendo en los últimos años en la ingeniería de carreteras. Dadas sus grandes ventajas económicas y ambientales, este tipo de ligantes pretende conseguir prestaciones similares a la de los betunes modificados con polímeros utilizados habitualmente. La aparición de fisuras debido a fenómenos de fatiga es una de las patologías más comunes en firmes de carretera. Debido a las mejoras de las propiedades mecánicas del betún aportadas tras la incorporación de polvo de neumático, las mezclas fabricadas con estos ligantes se postulan como una posible solución a dicho problema. En este artículo se lleva a cabo un análisis comparativo del comportamiento a fisuración por fatiga realizado sobre mezclas bituminosas fabricadas con betún modificado con polvo de neumático y con polímeros.This research was carried out within the framework of the R + D + i project entitled Proyecto Integrado de Investigación, Desarrollo y Demostración de Tecnologías para la aplicación de neumáticos fuera de uso en firmes de carretera resistentes a la propagación de grietas (ref. IDI-20091076), funded by the Center for Industrial Technological Development (CDTI) of the Ministry of Science and Innovation in Spain

Prevalence of oropharyngeal beta-lactamase-producing Capnocytophaga spp. in pediatric oncology patients over a ten-year period

BACKGROUND: The aim of this study was to evaluate the prevalence of beta-lactamase-producing Capnocytophaga isolates in young children hospitalized in the Pediatric Oncology Department of Hôpital Sud (Rennes, France) over a ten-year period (1993–2002). METHODS: In neutropenic children, a periodic survey of the oral cavity allows a predictive evaluation of the risk of systemic infections by Capnocytophaga spp. In 449 children with cancer, 3,053 samples were collected by oral swabbing and plated on TBBP agar. The susceptibility of Capnocytophaga isolates to five beta-lactams was determined. RESULTS: A total of 440 strains of Capnocytophaga spp. were isolated, 309 (70%) of which were beta-lactamase producers. The beta-lactamase-producing strains were all resistant to cefazolin, 86% to amoxicillin, and 63% to ceftazidime. The proportion of strains resistant to third-generation cephalosporins remained high throughout the ten-year study, while susceptibility to imipenem and amoxicillin combined with clavulanic acid was always conserved. CONCLUSION: These results highlight the risk of antibiotic failure in Capnocytophaga infections and the importance of monitoring immunosuppressed patients and testing for antibiotic susceptibility and beta-lactamase production