Development and applications of a DNA labeling method with magnetic nanoparticles to study the role of horizontal gene transfer events between bacteria in soil pollutant bioremediation processes.

Horizontal gene transfers are critical mechanisms of bacterial evolution and adaptation that are involved to a significant level in the degradation of toxic molecules such as xenobiotic pesticides. However, understanding how these mechanisms are regulated in situ and how they could be used by man to increase the degradation potential of soil microbes is compromised by conceptual and technical limitations. This includes the physical and chemical complexity and heterogeneity in such environments leading to an extreme bacterial taxonomical diversity and a strong redundancy of genes and functions. In addition, more than 99 % of soil bacteria fail to develop colonies in vitro, and even new DNA-based investigation methods (metagenomics) are not specific and sensitive enough to consider lysis recalcitrant bacteria and those belonging to the rare biosphere. The objective of the ANR funded project "Emergent" was to develop a new culture independent approach to monitor gene transfer among soil bacteria by labeling plasmid DNA with magnetic nanoparticles in order to specifically capture and isolate recombinant cells using magnetic microfluidic devices. We showed the feasibility of the approach by using electrotransformation to transform a suspension of Escherichia coli cells with biotin-functionalized plasmid DNA molecules linked to streptavidin-coated superparamagnetic nanoparticles. Our results have demonstrated that magnetically labeled cells could be specifically retained on micromagnets integrated in a microfluidic channel and that an efficient selective separation can be achieved with the microfluidic device. Altogether, the project offers a promising alternative to traditional culture-based approaches for deciphering the extent of horizontal gene transfer events mediated by electro or natural genetic transformation mechanisms in complex environments such as soil.

nDEP-driven cell patterning and bottom-up construction of cell aggregates using a new bioelectronic chip.

Creating cell aggregates of controlled size and shape and patterning cells on substrates using a bottom-up approach constitutes important challenges for tissue-engineering applications and studies of cell-cell interactions. In this paper, we report nDEP (negative dielectrophoresis) driven assembly of cells as compact aggregates or onto defined areas using a new bioelectronic chip. This chip is composed of a quadripolar electrode array obtained using coplanar electrodes partially covered with a thin, micropatterned PDMS membrane. This thin PDMS layer was coated with poly-L-lysine and played the role of adhesive substrate for cell patterning. For the formation of detachable cell aggregates, the PDMS was not pretreated and cells were simply immobilized into assemblies maintained by cell-cell adhesion after the electric field removal. Cell viability after exposition to DEP buffer was also assessed, as well as cell spreading activity following DEP-driven assembly.

[Management of the cardiovascular complications of treatment in thoracic oncology]. / Prise en charge des complications cardiovasculaires des traitements en oncologie thoracique.

The management of patients suffering from bronchial and lung tumors depends on conventional chemotherapy and/or targeted molecular therapies. The prescription of these chemotherapies may be accompanied by cardiovascular complications, principally congestive heart failure, arterial hypertension and arterial or venous thrombo-embolism, the frequency of which varies with the molecule administered. The management of these complications is currently poorly standardized and should take account of the patient's oncological prognosis.

Assessment of 0.5 T static field exposure effect on yeast and HEK cells using electrorotation.

This study aims to examine the influence of a 0.5 T static magnetic field (SMF) on yeast and human embryonic kidney (HEK) 293 cell using electrorotation (ROT). Following 48 h exposition to the SMF, no difference was noted between ROT spectra of unexposed and exposed yeast cells, which extend previous reports on the absence of SMF effects on yeast. We also compared the ROT spectrum and the extracted electrical characteristics of HEK cells exposed during 72 h to a 0.5 T uniform static magnetic field with those of unexposed cells. SMF potential effects on HEK proliferation kinetics and cell morphology were also assessed by using the trypan blue exclusion method and scanning electron microscopy, respectively. At last, no significant differences were observed between control and exposed HEK cells concerning electrical properties, growth, and morphology.

Microfluidic immunomagnetic cell separation using integrated permanent micromagnets.

In this paper, we demonstrate the possibility to trap and sort labeled cells under flow conditions using a microfluidic device with an integrated flat micro-patterned hard magnetic film. The proposed technique is illustrated using a cell suspension containing a mixture of Jurkat cells and HEK (Human Embryonic Kidney) 293 cells. Prior to sorting experiments, the Jurkat cells were specifically labeled with immunomagnetic nanoparticles, while the HEK 293 cells were unlabeled. Droplet-based experiments demonstrated that the Jurkat cells were attracted to regions of maximum stray field flux density while the HEK 293 cells settled in random positions. When the mixture was passed through a polydimethylsiloxane (PDMS) microfluidic channel containing integrated micromagnets, the labeled Jurkat cells were selectively trapped under fluid flow, while the HEK cells were eluted towards the device outlet. Increasing the flow rate produced a second eluate much enriched in Jurkat cells, as revealed by flow cytometry. The separation efficiency of this biocompatible, compact micro-fluidic separation chamber was compared with that obtained using two commercial magnetic cell separation kits.

Monitoring the endocytosis of magnetic nanoparticles by cells using permanent micro-flux sources.

Trapping of cells is essential to perform basic handling operations in cell-based microsystems, such as media exchange, concentration, cell isolation and cell sorting. Cell trapping by magnetophoresis typically requires cell labeling with magnetic nanoparticles. Here we report on endocytotic uptake of 100 nm magnetic nanoparticles by Human Embryonic Kidney 293 cells. The attraction of labeled cells by micro-magnet arrays characterised by very high magnetic field gradients (≤106 T/m) was studied as a function of labeling conditions (nanoparticle concentration in the extracellular medium, incubation time). The threshold incubation conditions for effective magnetophoretic trapping were established. This simple technique may be exploited to minimise the quantity of magnetic nanoparticles needed for efficient cell trapping, thus reducing stress or nanoparticle-mediated toxicity. Nanoparticle internalization into cells was confirmed using both confocal and Transmission Electron Microscopy (TEM).

A new microfluidic device for electric lysis and separation of cells.

This paper demonstrates the potential use of a new microfluidic device embedding thick electrodes for cell lysis and cell separation applications. The system consists of a microfluidic channel featuring conductive walls made of a polydimethylsiloxane (PDMS) matrix mixed with carbon nanoparticles. Cell lysis was performed electrically by applying square pulses across the channel width, which was monitored by fluorimetry. Lysed and unlysed cells showed different dielectrophoretic behavior under appropriate experimental conditions, which suggests that the developed device is suitable to perform both cell lysis and subsequent sorting of viable and dead cells.

Tunable and label-free bacteria alignment using standing surface acoustic waves.

This paper describes a new technique for focusing bacteria in a microfluidic channel and subsequently controlling their trajectory. Bacteria alignment is obtained using standing surface acoustic waves (SSAW) generated by two interdigitated transducer electrodes (IDTs) patterned on a piezoelectric wafer. The bacteria are focused in the standing wave pressure nodes, separated by half a wavelength, the electrode geometry and applied voltage frequency being chosen accordingly. Interestingly, the position of a pressure node can be modulated by introducing a phase shift between the electrical signals applied to both IDTs. The bacteria, trapped in this node, follow it and can therefore be deflected. This technique works with label-free bacteria in their culture medium and induces low power consumption, which is very interesting for portable devices.

[Focus on managing AF and preventing stroke]. / Mise au point sur la prise en charge de la FA et prévention des AVC.

Guidelines published from the European Society of Cardiology/American Heart Association, and from the American College of Chest Physicians, clarified the strategy of antithrombotic treatment in AF, which is based on the presence of risk factors for thromboembolism. The European guidelines have been updated in 2010 and differ from the ACCP guidelines. They integrated the CHA(2)DS(2)Vasc score and modified accordingly the previous recommendations based on the CHADS(2)score, which allows physicians to classify patients as at low, moderate or high risk, according to their individual risk characteristics. Recently published registries confirm under-prescription of VKA treatment in the 'real world', even in patients at high thromboembolic risk, and over-prescription for at least one-third of patients at low risk. Therefore reducing the risk of thromboembolism should be the physicians'primary aim, particularly with the advent of alternative treatments and the development of new antithrombotic drugs such as oral thrombin and factor Xa inhibitors, which are currently being evaluated in clinical trials.

Contactless diamagnetic trapping of living cells onto a micromagnet array.

This paper focuses on the application of magnetophoresis to a new cell patterning method. The principle was demonstrated by using a CoPt micromagnet array, producing regularly spaced magnetic traps where cells were confined without any contact under the effect of negative magnetophoresis. To obtain this effect, yeast cells (Saccharomyces cerevisiae), which are diamagnetic, were placed in an aqueous solution enriched in paramagnetic ions. Unlabeled (non-magnetic) cell manipulation by magnetophoresis requires the production of high magnetic field gradients, ensuring significant forces. Therefore, micromagnets are particularly interesting for our application, since the field gradient increases as magnet dimensions are reduced.

[Postpartum cardiomyopathy revealed by acute lower limb ischemia]. / Cardiomyopathie du péripartum révélée par l'ischémie aiguë d'un membre inférieur.

Peripartum cardiomyopathy is an uncommon disease defined as a dilated cardiomyopathy during puerperium, with left ventricular dysfunction (ejection fraction < 45%) without any other etiology. The etiology of this disease remains uncertain and it can be revealed in a variety of ways. Thrombo-embolic complications may be, although infrequently, the initial manifestation of peripartum cardiomyopathy, which is usually an intracardiac thrombosis. Lower extremity embolism is uncommon. The case reported is about a 39-year-old woman, multiparous, who presented, 40 days after delivery, a global heart failure with atrial fibrillation, revealed by left lower extremity thromboembolism. After echocardiographic and etiologic examinations, the diagnosis was established as peripartum cardiomyopathy. It evolved favourably after 2 months of medical treatment: the symptoms and cardiomegaly decreased, left ventricular systolic function was improved.

[Tuberous sclerosis and cardiac tumor (case report)]. / Sclérose tubéreuse de Bourneville et tumeur cardiaque (à propos d'un cas).

We report the case of a cardiac tumor, which is found in a systematic cardiac examination of an 11 year old girl who had a tuberous sclerosis. The interest of this observation is to show the importance of a general examination, especially cardiac, within this pathology. Tuberous sclerosis is an hereditary disease, associated with rhabdomyomas in 50% of cases or more. Cardiac rhabdomyomas are frequently multiple and detected in utero in some cases. Conversely, 60% of these tumors are seen in a context of tuberous sclerosis. Rhabdomyomas are the most common cardiac tumors of infants and children, the large majority occurring in patients younger than one year and are diagnosed more and more in foetal echocardiography.