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Introduction: Pulmonary arterial hypertension (PAH) is a devastating lung disease with no curative treatments available. Cell-based therapies are a promising regenerative approach, including the direct application of stem cell derived extracellular vesicles (EVs). Mesenchymal stromal cells (MSCs) have shown promise as a treatment for many cardiovascular diseases; their therapeutic effects are predominately mediated by paracrine mechanisms, in particular the release of EVs. Yet, systemically administered EVs are rapidly cleared to the liver and spleen, with minimal lung accumulation. Hypothesis: We hypothesized that encapsulation of EVs within nanoporous microgels would provide a novel therapeutic product to improve EV lung delivery and retention and therefore enhance their therapeutic benefits. Methods & Results: MSC-derived EVs were purified using tangential flow filtration resulting in a median size of 108±53nm by nanoparticle tracking analysis, and positive for CD63 by Western blot. EV-loaded microgels were prepared using a microfluidic device producing agarose-gelatin (1%-1%) microgels of 47.2±0.4um diameter. In vitro uptake assays were performed with DiR or pkh26 labelled EVs and quantified by microscopy. Encapsulated EVs demonstrated a delayed-release profile in vitro with reduced HUVEC uptake over 24h (9.7±1.1% encap. vs. 46±5.3% free). In vivo biodistribution studies were performed in the monocrotaline model of PAH using central intravenous delivery. EV-loaded microgels were efficiently retained within the lungs after 24h (81±12% encap vs. 7.5±0.4% free), while free EVs accumulated in the liver (74±2.3% free vs 10±2.5% encap). Furthermore, lung digestion and flow cytometry analysis revealed that encapsulated EVs were readily taken up by CD45+ immune cells and CD31+ endothelial cells to a greater extent than free EVs, indicating enhanced interaction with the local lung microenvironment. Conclusion: Microencapsulation of EVs results in significantly enhanced local retention within the lung, leading to increased local cellular uptake compared to systemically administered EVs.

Sensing performance of solid-state nanopores is limited by the fast kinetics of small molecular targets. To address this challenge, we translate the presence of a small target to a large conformational change of a long polymer. In this work, we explore the performance of solid-state nanopores for sensing the conformational states of molecular nanoswitches assembled using the principles of DNA origami. These programmable single-molecule switches show great potential in molecular diagnostics and long-term information storage. We investigate the translocation properties of linear and looped nanoswitch topologies using nanopores fabricated in thin membranes, ultimately comparing the performance of our nanopore platform for detecting the presence of a DNA analogue to a sequence found in a Zika virus biomarker gene with that of conventional gel electrophoresis. We found that our system provides a high-throughput method for quantifying several target concentrations within an order of magnitude by sensing only several hundred molecules using electronics of moderate bandwidth that are conventionally used in nanopore sensing systems.

Endothelial progenitor cells and human mesenchymal stem cells (hMSCs) have shown great regenerative potential to repair damaged tissue; however, their injection in vivo results in low retention and poor cell survival. Early clinical research has focussed on cell encapsulation to improve viability and integration of delivered cells. However, this strategy has been limited by the inability to reproduce large volumes of standardized microcapsules and the lack of information on cell-specific egress and timed release from hydrogel microcapsules. Here, we address both of these limitations. First, we use a droplet microfluidic platform to generate monodisperse agarose microcapsules, and second we encapsulate and characterize egress of therapeutically relevant cells (human umbilical vein endothelial cells, endothelial progenitor cells, and hMSCs). With increased temporal resolution, we demonstrate distinct differences in egress between cell types. Importantly, therapeutic cells (hMSCs) egress quickly, in

Assays targeting biomarkers for the early diagnosis of disease demand a sensing platform with a high degree of specificity and sensitivity. In this work, we developed and characterized a solid-state nanopore-based sensing assay for the detection of short nucleic acid targets with readily customizable nanostructured DNA probe sets. We explored the electrical signatures of three DNA nanostructures to determine their performance as probe sets in a digital counting scheme to quantify the concentration of targets. With these probes, we demonstrate the specific, simultaneous detection of two different DNA targets in a 2-plex assay, and separately that of microRNA-155, a biomarker linked to various human cancers. In addition to specific target detection, our scheme demonstrated the ability to quantify at least six different microRNA concentrations. These results highlight the potential for solid-state nanopores as single-molecule counters for future digital diagnostic technologies.

Integrating nanopore sensors within microfluidic architectures is key to providing advanced sample processing capabilities upstream of the biosensor. When confined in a microchannel, the nanopore capture and translocation characteristics are altered when subjected to cross-flow, affecting sensor performance. Here, we study the capture rate and translocation of 1-5 kbp double-stranded DNA molecules through solid-state nanopores in the presence of tangential fluid flow over the nanopore aperture. Experiments reveal a trend of increased capture rate with cross-flow, reaching a 5-fold enhancement (dependent on DNA length) at moderate flow rates, before decreasing at higher flow rates. By modeling DNA dynamics in microchannels under the combined effect of laminar flow, Brownian motion and electrophoretic drift, it is shown that the observed trend is the result of two competing mechanisms: enhanced DNA transport by convection and reduction in the nanopore's capture volume with increased flow velocity. Moreover, it is shown that the viscous drag force exerted by flow on a translocating DNA can be exploited to tune the kinetics of DNA translocation.

An imaging-integrated microfluidic cell volume sensor was used to evaluate the volumetric growth rate of single cells from a Saccharomyces cerevisiae population exhibiting two phenotypic expression states of the PDR5 gene. This gene grants multidrug resistance by transcribing a membrane transporter capable of pumping out cytotoxic compounds from the cell. Utilizing fluorescent markers, single cells were isolated and trapped, then their growth rates were measured in two on-chip environments: rich media and media dosed with the antibiotic cycloheximide. Approximating growth rates to first-order, we assessed the fitness of individual cells and found that those with low PDR5 expression had higher fitness in rich media whereas cells with high PDR5 expression had higher fitness in the presence of the drug. Moreover, the drug dramatically reduced the fitness of cells with low PDR5 expression but had comparatively minimal impact on the fitness of cells with high PDR5 expression. Our experiments show the utility of this imaging-integrated microfluidic cell volume sensor for high-resolution, single-cell analysis, as well as its potential application for studies that characterize and compare the fitness and morphology of individual cells from heterogeneous populations under different growth conditions.

The identification of molecular tags along nucleic acid sequences has many potential applications in bionanotechnology, disease biomarker detection, and DNA sequencing. An attractive approach to this end is the use of solid-state nanopores, which can electrically detect molecular substructure and can be integrated into portable lab-on-a-chip sensors. We present here a DNA origami-based approach of molecular assembly in which solid-state nanopores are capable of differentiating 165 bp scaffolds containing zero, one, and two dsDNA protrusions. This highly scalable technique requires minimal sample preparation and is customizable for a wide range of targets and applications. As a proof-of-concept, an aptamer-based DNA displacement reaction is performed in which a dsDNA protrusion is formed along a 255 bp scaffold in the presence of ATP. While ATP is too small to be directly sensed using conventional nanopore methods, our approach allows us to detect ATP by identifying molecular substructure along the DNA scaffold.

International audience; Epidemiological data clearly show that high blood pressure is associated with cardiovascular mortality and progression of chronic kidney disease. Although several randomized controlled trials performed over the last decades have demonstrated a cardiovascular benefit of reducing blood pressure in hypertensive patients, solid evidence for general and specific blood pressure targets, especially in the population with chronic kidney disease, is lacking. In this context, recently published trials have provided novel insights in the management of hypertension. Here, we review and discuss the existing evidence and the current guidelines on blood pressure targets in both the general and the population with chronic kidney disease.

Fibrosis is a leading cause of death in developed countries that is characterized by a progressive deterioration of tissue mechanical behavior. Developing methods to study tissue mechanics and myofibroblast activation may lead to new targets for therapeutic treatments that are urgently needed. Microtissue arrays are a promising approach to conduct relatively high throughput research into fibrosis as they recapitulate key biomechanical aspects of the disease through a relevant 3D extracellular environment. In early work, our group developed a device called the MVAS-force to stretch microtissues while enabling simultaneous assessment of their dynamic mechanical behavior. Here we investigated TGF-β1 induced fibroblast to myofibroblast differentiation in microtissue cultures using our MVAS-force device through assessing α-SMA expression, contractility and stiffness. By doing so, we linked cell-level phenotypic changes to functional changes that characterize the clinical manifestation of fibrotic disease. As expected, TGF-β1 treatment promoted a myofibroblastic phenotype and microtissues became stiffer and possessed increased contractility. Furthermore, these changes were partially reversible upon TGF-β1 withdrawal. In contrast, however, long-term cyclic stretching maintained myofibroblast activation. Furthermore stretching had no effect compared static cultures when TGF-β1 receptors were inhibited and stretching promoted myofibroblast differentiation when given latent TGF-β1. Together these results suggest that external mechanical stretch may activate latent TGF-β1 and might be a powerful stimulus for continued myofibroblast activation to progress fibrosis. Further exploration of this pathway with our approach may yield new insights into myofibroblast activation and more effective therapeutic treatments for fibrosis.Insight boxUsing a novel high-throughput approach, we quantified the effects of dynamic mechanical stretching on the phenotype and function of cells in 3D microtissue cultures during myofibroblast activation with TGF-β1 treatment and subsequent withdrawal. Our findings show that mechanical stretch may activate endogenously produced latent TGF-β1 to maintain the presence and activity of myofibroblasts after tissue injury. Importantly, through this feed forward mechanism, mechanical stretch might be a powerful stimulus that directs tissues away from recovery and towards the development of fibrosis.

Characterizing the time-dependent mechanical properties of cells is not only necessary to determine how they deform, but also to fully understand how external forces trigger biochemical-signaling cascades to govern their behavior. Presently mechanical properties are largely assessed by applying local shear or compressive forces on single cells in isolation grown on non-physiological 2D surfaces. In comparison, we developed the microfabricated vacuum actuated stretcher to measure tensile loading of 3D multicellular ‘microtissue’ cultures. With this approach, we assessed here the time-dependent stress relaxation and recovery responses of microtissues, and quantified the spatial remodeling that follows step length changes. Unlike previous results, stress relaxation and recovery in microtissues measured over a range of step amplitudes and pharmacological treatments followed a stretched exponential behavior describing a broad distribution of inter-related timescales. Furthermore, despite a performed compendium of experiments, all responses led to a single linear relationship between the residual elasticity and degree of stress relaxation, suggesting that these mechanical properties are coupled through interactions between structural elements and the association of cells with their matrix. Lastly, although stress relaxation could be quantitatively and spatially linked to recovery, they differed greatly in their dynamics; while stress recovery behaved as a linear process, relaxation time constants changed with an inverse power law with step size. This assessment of microtissues offers insights into how the collective behavior of cells in a 3D collagen matrix generate the dynamic mechanical properties of tissues, which is necessary to understanding how cells deform and sense mechanical forces in the body.

When mechanically stretched, cells cultured on 2D substrates share a universal softening and fluidization response that arises from poorly understood remodeling of well-conserved cytoskeletal elements. It is known, however, that the structure and distribution of these cytoskeletal elements are profoundly influenced by the dimensionality of a cell’s environment (ie. on a 2D surface vs. within a 3D matrix). Therefore, in this study we aimed to determine whether cells cultured in a 3D extracellular matrix also follow the same softening response and to link this mechanical change to direct evidence of cytoskeletal remodeling. To achieve this, we developed a new high-throughput approach to measure the dynamic mechanical properties of cells and allow for sub-cellular imaging of physiologically relevant 3D microtissue cultures. We found that fibroblast, smooth muscle and skeletal muscle microtissues strain softened but did not fluidize, and upon loading cessation, they fully regained their initial mechanical properties. These responses required a filamentous actin cytoskeleton, and were mirrored by changes to actin remodeling rates, and direct visual evidence of actin depolymerization during stretching and repolymerization after stretch cessation. On the other hand, the response could not be attributed to either remodeling of microtubules or myosin motor activity. Our new approach for assessing cell mechanics has linked behaviors seen in 2D cultures to a soft 3D extracellular matrix, and connected visual remodeling of the cytoskeleton to changes in mechanical properties at the tissue-level. Significance Statement With every breath and movement, cells in our body are subjected to mechanical forces. These forces are key regulators of normal development and function, as well as disease progression. To understand how cells “feel” mechanical cues in their microenvironment, we have previously relied on two-dimensional experimental approaches and often assessed single cells in isolation. Here, we present a novel lab-on-chip device, which enables simultaneous mechanical stimulation and sub-cellular imaging of three-dimensional multi-cellular microtissues. In this article with this device, we quantitatively linked force-induced mechanical changes in microtissues to specific molecular remodelling pathways in the cytoskeleton. The approaches and insights presented in this study will deepen our understanding of the mechanobiological pathways governing tissue development and function in health and disease.

Previous work has shown that cocooning heart explant-derived cells (EDCs) within protective nanoporous gel capsules before intra-myocardial injection increases the retention of transplanted cells and the paracrine production of nanoparticles to improve post infarct cardiac function. In this study, we investigated the influence of cocoon size and intracapsular cell number on cell-treatment outcomes using a newly developed microfluidic-based (MF) cellular cocooning platform. Methods/Results: Traditional vortex-based encapsulation (Vx) inherently provides cocoons of varying diameters (30-100 μm; 68±5 μm). By altering the flow pressure ratios and the nozzle diameters within the MF chip, we encapsulated human EDCs within small (51±1 μm, MF50) and large (90±1 μm, MF90) diameter nanoporous gel cocoons for comparison with standard Vx-defined capsules (71±1 μm, MF70). MF cocooning mirrored the expected Poisson distribution with smaller cocoons having a greater proportion of single cells while larger diameter cocoons contained greater proportions of multicellular aggregates. Immunodeficient mice underwent left coronary artery ligation 1 week before randomization to echocardiographic guided intra-myocardial injection of EDCs (suspended or variable diameter cocoons) or vehicle. Increasing cocoon diameter stimulated progressive salutary effects on post-infarct function (ejection fraction), scar burden and newly generated peri-infarct blood vessels (isolectin B4+) and cardiomyocytes (BrdU+/TNT+) 4 weeks after treatment. Bioluminescent imaging of luciferase tagged cells revealed increasing cocoon diameter reduced the rate of cell clearance from injured tissues. Disrupting cell-cell contact within the capsules (using a custom antibody cocktail to block E/P-selectin and N-cadherin) reduced the amount and profile of pro-healing cytokines + nanoparticles delivered to injured myocardium. Conclusions: Increasing cocoon diameter and cell occupancy within protective nanoporous gel cocoons boosts paracrine-mediated repair of damaged myocardium by slowing clearance of cells from injured tissues and the number of cytokines + nanoparticles secreted by micro-encapsulated cells.

While encapsulation of cells within protective nanoporous gel cocoons increases cell retention and pro-survival integrin signaling, the influence of cocoon size and intra-capsular cell-cell interactions on therapeutic repair are unknown. Here, we employ a microfluidic platform to dissect the impact of cocoon size and intracapsular cell number on the regenerative potential of transplanted heart explant-derived cells. Deterministic increases in cocoon size boosted the proportion of multicellular aggregates within cocoons, reduced vascular clearance of transplanted cells and enhanced stimulation of endogenous repair. The latter being attributable to cell-cell stimulation of cytokine and extracellular vesicle production while also broadening of the miRNA cargo within extracellular vesicles. Thus, by tuning cocoon size and cell occupancy, the paracrine signature and retention of transplanted cells can be enhanced to promote paracrine stimulation of endogenous tissue repair.

International audience; Light chain proximal tubulopathy (LCPT) is a rare kidney disease associated with plasma cell dyscrasias, characterized by light chain deposits in the proximal tubular cells, with or without crystal formation. We describe an exceptional case of LCPT without crystal formation in a kidney allograft, in a patient who underwent two renal transplants for a light chain deposition disease (LCDD) complicating smoldering myeloma. This is the first description of this association in successive kidney allografts. We concisely describe pathology of LCDD and LCPT and discuss potential pathophysiological mechanisms relating these two conditions.

International audience; Immunosuppression in kidney transplant recipients with decreased graft function and severe histological vascular changes can be particularly challenging. Belatacept could be a valuable option, as a rescue therapy in this context. We report a retrospective case control study comparing a CNI to belatacept switch in 17 patients with vascular damage and low eGFR to a control group of 18 matched patients with CNI continuation. Belatacept switch was performed on average 51.5 months after kidney transplantation (6.2-198 months). There was no difference between the two groups regarding eGFR at inclusion, and 3 months before inclusion. In the "CNI to belatacept switch group," mean eGFR increased significantly from 23.5 ± 6.7 mL/min/1.73m2 on day 0, to 30.4 ± 9.1 mL/min/1.73 m2 on month 6 (p < 0.001) compared to the control group, in which no improvement was observed. These results were still significant on month 12. Two patients experienced biopsy-proven acute rejection. One was effectively treated without belatacept discontinuation. Two patients needed belatacept discontinuation for infection. In conclusion, the remplacement of CNI with belatacept in patients with decreased allograft function and vascular lesions is associated with an improvement in eGFR.

The strain-induced reorientation response of cyclically stretched cells has been well characterized in uniform strain fields. In the present study, we comprehensively analyse the behaviour of human fibroblasts subjected to a highly non-uniform strain field within a polymethylsiloxane microdevice. Our results indicate that the strain gradient amplitude and direction regulate cell reorientation through a coordinated gradient avoidance response. We provide critical evidence that strain gradient is a key physical cue that can guide cell organization. Specifically, our work suggests that cells are able to pinpoint the location under the cell of multiple physical cues and integrate this information (strain and strain gradient amplitudes and directions), resulting in a coordinated response. To gain insight into the underlying mechanosensing processes, we studied focal adhesion reorganization and the effect of modulating myosin-II contractility. The extracted focal adhesion orientation distributions are similar to those obtained for the cell bodies, and their density is increased by the presence of stretching forces. Moreover, it was found that the myosin-II activity promoter calyculin-A has little effect on the cellular response, while the inhibitor blebbistatin suppresses cell and focal adhesion alignment and reduces focal adhesion density. These results confirm that similar internal structures involved in sensing and responding to strain direction and amplitude are also key players in strain gradient mechanosensing and avoidance.

We combine a high-resolution resistive pulse sensor with hydrodynamic flow focusing.We quantify sorting purity, throughput and resolution with polystyrene microspheres.We size/sort a population of Saccharomyces cerevisiae (yeast) cells.We integrate PDMS valves, both for sensitivity enhancement and cell collection. We demonstrate a microfluidic device that integrates high-sensitivity volume sensing with active pressure-driven flow sorting. Label-free size-based sorting of microparticles and cells is achieved using hydrodynamic flow focusing combined with a resistive pulse sensor with tunable sensitivity that utilizes the Coulter principle. Measurements are performed in real time, and pressure-driven flows are automatically adjusted in feedback to direct microparticles into one of multiple outlet sorting channels at 97.3% efficiency and with a resolution of ~3µm3. In a subsequent test, cells were sorted by volume from a single culture of Saccharomyces cerevisiae (yeast) at 100% efficiency. Next, we demonstrate the device's ability to size a population of cells at high-throughput and identify features of interest prior to sorting. Subpopulations of arrested G1 and M-phase cells were successfully resolved from a single yeast culture. This integrated on-chip sizing and sorting method is ideal for sorting small numbers of particles/cells at very high resolution.

The prevalence of hospital acquired infections and antibiotic resistant pathogens necessitates the development of bacteria sensing systems that do not require sample amplification via conventional cell culturing, which can be prohibitively time-consuming. To meet this need, we designed an optofluidic Raman detection platform which utilized a microfluidic driven hollow-core photonic crystal fiber, which in combination with silver nanoparticles, provides a large enhancement to the Raman signal. By confining both light and cells within this fiber, spectral events generated by the flowing cells facilitates a novel method of cell counting to simultaneously quantify and qualify infections. Counting is performed automatically by a genetically optimized support vector machine learning algorithm that was previously developed by our group. The microfluidic system can be regenerated multiple times, and allows for online detection of planktonic bacteria to levels as low as 4 CFU/mL in 15 min. This compares favourably to other methods currently under development such as qPCR and biosensing techniques. Furthermore, Raman spectral differences between bacteria allow for inherent multiplexed detection in serum, by adding another layer to the learning algorithm. Further development of this device has promising potential as a rapid point-of-care system for infection management in the clinic.

Cellular organization, migration and proliferation in three-dimensions play a critical role in numerous physiological and pathological processes. Nano- and micro-fabrication approaches have demonstrated that nano- and micro-scale topographies of the cellular microenvironment directly impact organization, migration and proliferation. In this study, we investigated these dynamics of two cell types (NIH3T3 fibroblast and MDCK epithelial cells) in response to microscale grooves whose dimensions exceed typical cell sizes. Our results demonstrate that fibroblasts display a clear preference for proliferating along groove ridges whereas epithelial cells preferentially proliferate in the grooves. Importantly, these cell-type dependent behaviours were also maintained when in co-culture. We show that it is possible to spatially separate a mixed suspension of two cell types by allowing them to migrate and proliferate on a substrate with engineered microtopographies. This ability may have important implications for investigating the mechanisms that facilitate cellular topographic sensing. Moreover, our results may provide insights towards the controlled development of complex three-dimensional multi-cellular constructs.

A multi-layered polydimethylsiloxane microfluidic device with an integrated suspended membrane has been fabricated that allows dynamic and multi-axial mechanical deformation and simultaneous live-cell microscopy imaging. The transparent membrane’s strain field can be controlled independently along two orthogonal directions. Human foreskin fibroblasts were immobilized on the membrane’s surface and stretched along two orthogonal directions sequentially while performing live-cell imaging. Cyclic deformation of the cells induced a reversible reorientation perpendicular to the direction of the applied strain. Cells remained viable in the microdevice for several days. As opposed to existing microfluidic or macroscale stretching devices, this device can impose changing, anisotropic and time-varying strain fields in order to more closely mimic the complexities of strains occurring in vivo. Electronic supplementary material The online version of this article (doi:10.1007/s10529-013-1381-5) contains supplementary material, which is available to authorized users.

We present gravitational field-flow fractionation and hydrodynamic chromatography of colloids eluting through 18 μm microchannels. Using video microscopy and mesoscopic simulations, we investigate the average retention ratio of colloids with both a large specific weight and neutral buoyancy. We consider the entire range of colloid sizes, including particles that barely fit in the microchannel and nanoscopic particles. Ideal theory predicts four operational modes, from hydrodynamic chromatography to Faxén-mode field-flow fractionation. We experimentally demonstrate, for the first time, the existence of the Faxén-mode field-flow fractionation and the transition from hydrodynamic chromatography to normal-mode field-flow fractionation. Furthermore, video microscopy and simulations show that the retention ratios are largely reduced above the steric-inversion point, causing the variation of the retention ratio in the steric- and Faxén-mode regimes to be suppressed due to increased drag. We demonstrate that theory can accurately predict retention ratios if hydrodynamic interactions with the microchannel walls (wall drag) are added to the ideal theory. Rather than limiting the applicability, these effects allow the microfluidic channel size to be tuned to ensure high selectivity. Our findings indicate that particle velocimetry methods must account for the wall-induced lag when determining flow rates in highly confining systems.

Background Kidney graft survival in simultaneous pancreas–kidney (SPK) recipients is known to decrease after pancreas graft failure. Methods Sixty-three consecutive SPK recipients were retrospectively reviewed. Kidney graft function and proteinuria were evaluated at three months after the transplantation and at last follow-up. Histopathologic findings of protocol biopsies performed three months and one yr after transplantation were analyzed. Results Twelve patients lost the pancreas graft. Donors' characteristics were similar in patients with or without pancreas failure. After a median follow-up of 36 months, mean eGFR with a functional pancreas was 69.5 mL/min/1.73 m² vs. 56.3 mL/min/1.73 m² (p = 0.01) after pancreas loss. Patients who lost pancreas had a median proteinuria of 0.28 g vs. 0.13 g per 24 h (p = 0.02). Analysis of three-month protocol biopsies revealed more frequent isolated glomerulitis after pancreas failure (p = 0.0001), without peritubular capillaritis or C4d deposition. No donor-specific anti-HLA antibodies were detectable in these patients. Chronic tubulointerstitial changes were more frequent in patients with pancreas loss. There was no evidence of diabetic nephropathy recurrence. Conclusion SPK recipients develop an early kidney graft dysfunction after pancreas failure. Histopathologic findings revealed frequent glomerulitis without antibody-mediated rejection and early chronic changes.

International audience; Evidence before this study We searched PubMed on June 30, 2016, for English-language cohort studies published since Jan 1, 1980, of patients with invasive listeriosis worldwide with the keywords " listeria " , " listeriosis " , " maternal " , and " neurolisteriosis ". Studies had to include epidemiological or clinical data on listeriosis. All clinical forms of infection were included (bacteraemia, neurolisteriosis, and maternal–neonatal infection). Host risk factors for listeriosis have been well identified, but the clinical features and prognostic factors of the disease are based on retrospective studies compiling heterogeneous data or random collected cases. Furthermore, no clinical trial has ever been done and medical management is not evidence based. Added value of the study Our study is the first prospective clinical study focusing on all forms of invasive listeriosis. The study is based on a national mandatory system that allowed the nearly complete capture of microbiologically proven cases. The study shows a higher burden of listeriosis than reported before: more than 80% of infected mothers experienced major fetal or neonatal complications (fetal loss, very high prematurity, early or late onset disease); only 39% of patients with neurolisteriosis survived and fully recovered. The study provides important new data to improve management and predict outcome in listeriosis, such as determination of the time window for fetal losses (

On-chip microvalves regulate electrical and fluidic access to an array of nanopores integrated within microfluidic networks. This configuration allows for on-chip sequestration of biomolecular samples in various flow channels and analysis by independent nanopores.

Naftidrofuryl (CAS 31329-57-4) is used, mainly in elderly patients, in the treatment of various vascular disorders. The aim of this study was to evaluate and compare the pharmacokinetics of naftidrofuryl after single oral administration of a 200 mg naftidrofuryl tablet (Praxilene) in caucasian male and female subjects with renal impairment versus healthy volunteers. This prospective and open study was conducted in three parallel groups: Group A = healthy subjects with a Cl(CR) > 80 ml/min, Group B = uraemic patients with a 20 < or = Cl(CR) < 40 ml/min, Group C = uraemic patients with a Cl(CR) < 20 ml/min. Blood samples were taken over a period of 32 h after dosing. The mean values (+/-SD) of the pharmacokinetic parameters of naftidrofuryl for group A were as follows: tmax: 1.3 h (median), Cmax: 174 +/- 46 ng/ml, t(1/2 beta): 4.4 +/- 1.1 h, AUC(0-infinity): 1541 +/- 384 ng x h/ml; for group B: tmax: 2.5 h (median), Cmax: 239 +/- 94 ng/ml, t(1/2 beta): 5.0 +/- 1.2 h, AUC(0-infinity): 2361 +/- 751 ng x h/ml; for group C: tmax: 3.0 h (median), Cmax: 236 +/- 104 ng/ml, t(1/2 beta): 5.0 +/- 2.1 h, AUC(0-infinity): 2488 +/- 2003 ng x h/ml. The statistical analysis was performed on the pharmacokinetic parameters with one-way ANOVA in order to compare each group. No significant difference between each group was observed. In conclusion, renal insufficiency did not appear to influence the pharmacokinetic profile of oral naftidrofuryl.

Background Whether mycophenolate mofetil (MMF) can prevent the vascular endothelial dysfunction related to the administration of calcineurin inhibitor after organ transplantation remains unknown. Methods Four groups of Lewis rats, grafted with Brown Norway donor aortic abdominal allograft, received since the transplantation cyclosporine A (CsA, 5 mg/kg/day), MMF (40 mg/kg/day), CsA+MMF, or vehicle (control) for 2 weeks. Results Fifteen days after transplantation, all immunosuppressive regimens were equally effective in preventing graft rejection. When compared with control rats, the endothelium-dependent relaxation to acetylcholine was reduced, and the vasoconstrictor effect of phenylephrine was enhanced in thoracic aorta of CsA-treated rats but not in rats treated with MMF alone or combined with CsA without difference for the endothelium-independent relaxation to sodium nitroprusside. The relaxation to acetylcholine was abolished by the nitric oxide (NO)-synthase inhibitor N-nitro-l-arginine in all groups. Moreover, the endothelial NO-synthase protein dimer:monomer ratio in the thoracic aorta and the plasma nitrites concentrations, an indicator of NO availability, were decreased in CsA-treated rats but not in rats treated with MMF alone or combined with CsA. Conclusions This study demonstrates that MMF prevents systemic endothelial dysfunction and the enhanced sensitivity to vasoconstrictors related to CsA administration in a rat allograft aortic model through an increase in NO availability related to the improvement of endothelial NO-synthase functionality.

Mutations in the PKD1 (polycystin 1) and PKD2 (polycystin 2) genes cause autosomal dominant polycystic kidney disease (ADPKD). Most Pkd2-null mouse embryos present with left-right laterality defects. For the first time, we report the association of ADPKD resulting from a mutation in PKD2 and left-right asymmetry defects. PKD1 and PKD2 were screened for mutations or large genomic rearrangements in 3 unrelated patients with ADPKD presenting with laterality defects: dextrocardia in one and situs inversus totalis in 2 others. A large gene deletion, a single-exon duplication, and an in-frame duplication respectively, were found in the 3 patients. These polymorphisms were found in all tested relatives with ADPKD, but were absent in unaffected related individuals. No left-right anomalies were found in other members of the 3 families. A possible association between heterotaxia and a PKD2 mutation in our 3 patients is suggested by: (1) the existence of laterality defects in Pkd2-null mouse and zebrafish models and (2) detection of a pathogenic PKD2 mutation in the 3 probands, although PKD2 mutations account for only 15% of ADPKD families. The presence of left-right laterality defects should be systematically screened in larger cohorts of patients with ADPKD harboring PKD2 mutations.

Summary This study attempted to establish whether a calcineurin inhibitor (CNI)-free immunosuppressant regimen based on sirolimus (SRL) is associated with a preservation of conduit arteries endothelial function in kidney recipients or not. Twenty-nine kidney recipients were randomized to receive since transplantation SRL (n = 15) or cyclosporin A (CsA, n = 14) associated with mycophenolate mofetil (MMF) and steroids (6 months) in a parallel prospective study. Systolic, diastolic blood pressures, glomerular filtration rate (GFR) and radial artery flow-mediated dilatation (FMD) induced by postischaemic hyperaemia were assessed in a blind manner at one (M1) and 7 months (M7) after transplantation. Endothelium-independent dilatation was assessed by glyceryl trinitrate spray. There was no difference between the groups for all vascular parameters at M1. At M7, systolic blood pressure was lower (SRL: 119 ± 3 vs. CsA: 138 ± 4 mmHg, P

We used a suspended microchannel resonator (SMR) combined with picoliter-scale microfluidic control to measure buoyant mass and determine the 'instantaneous' growth rates of individual cells. The SMR measures mass with femtogram precision, allowing rapid determination of the growth rate in a fraction of a complete cell cycle. We found that for individual cells of Bacillus subtilis, Escherichia coli, Saccharomyces cerevisiae and mouse lymphoblasts, heavier cells grew faster than lighter cells.

Background. Although cyclosporine maintenance therapy reduces the risk of acute rejection and increases short-term graft survival in renal transplant recipients, its associated nephrotoxicity increases the risk of chronic graft dysfunction. The dose that allows an optimal risk-to-benefit ratio has not been established. Methods. This multicentre study enrolled stable renal allograft recipients receiving cyclosporine and mycophenolate mofetil without corticosteroids in their second year post-transplant. Patients were randomized to a cyclosporine dose targeted to a standard area under the concentration–time curve (AUC)0–12 h (usual exposure, n =1 04) or 50% of the study standard AUC0–12 h (low exposure, n = 108) using a three-point pharmacokinetic sampling. The primary endpoint was the percentage of patients with treatment failure at 24 months (graft loss/acute rejection/ nephrotoxicity/>15% serum creatinine level increase). Results. Treatment failure was reported in 37 out of 101 (37%) patients in the usual-exposure and 19 out of 106 (18%) patients in the low-exposure groups (P = 0.003). Mean estimated glomerular filtration rate decreased from baseline to 2 years with usual exposure and increased with low exposure (P < 0.001). Mean systolic and diastolic blood pressures were lower with low exposure (P = 0.03 and P = 0.008, respectively). Conclusion. In renal transplant recipients receiving maintenance therapy without corticosteroids, a minimization strategy using three-point pharmacokinetic sampling to reduce and maintain cyclosporine exposure to 50% of the usual levels is safe and reduces the risk of graft dysfunction.

Until now there are few data in the literature describing psychiatric comorbidity in patients waiting for renal transplantation. We have conducted a cross sectional study estimating the prevalence of anxiety and depressive disorders in three groups of renal transplant patients, before transplantation, six months and one year after. The MINI was used to estimate the prevalence of anxiety and depressive disorders. Anxiety and depressive symptoms were assessed using the HAD. Patients' quality of life was also assessed using the SF-36. This study did not find any major impact of renal transplantation on the prevalence of structured psychiatric disorders. Indeed, the prevalence of depressive and anxiety disorders did not differ significantly between the three groups. The mean scores of anxiety did not differ significantly between the three groups in contrast to the mean scores of depression, which differed significantly between the group "before transplantation" and the group "one year after transplantation". We did not find any significant difference concerning the scores of patient's quality of life between the three groups, except for the item "health perceived by the patients themselves". Health perceived by the patients was greater in the group "after transplantation". The quality of life of dialysed or transplant patients was strongly correlated with anxiety and depressive symptoms scores, emphasizing the major interest of a multidisciplinary approach for these patients.

Long-term survival of patients with chronic lymphocytic leukemia (CLL) is over 10 years, and such patients are thus potential kidney recipients in the case of superimposed end-stage renal disease. However, the renal and patient outcome in this condition is unknown. We report the charts of four patients with CLL who were engrafted in France with a deceased-donor kidney and underwent routine triple immunosuppressive therapy. The results show that these patients developed severe infectious episodes (fatal in one case) and tumoral complications including rapid progression of CLL in two cases. Moreover, the graft may be infiltrated and damaged by monoclonal B cells: one patient lost his graft 14 months after transplantation. Various therapeutic options (modifications of the immunosuppressive regimen, anti-CD20 antibodies, irradiation of the graft) showed little (if any) efficacy. Therefore, we believe that CLL is a too hazardous condition to envisage solid organ transplantation with a routine immunosuppressive regimen, and we propose a more appropriate approach.

BACKGROUND Fucoidan, a new low molecular weight sulfated polysaccharide (LMWF), has previously been shown to mobilize bone marrow-derived progenitors cells via stimulation of stromal derived factor (SDF)-1 release. Mobilized progenitor cells have been suggested to repair intimal lesions after immune-mediated endothelial injury and thus prevent intimal proliferation. The aim of this study was to evaluate the effect of LMWF treatment in a rat aortic allograft model of transplant arteriosclerosis (TA). METHODS Aortic grafts were performed in Brown Norway (BN, donor) and Lewis (Lew, recipient) rats. The recipient rats were treated with LMWF (5 mg/kg/day) and sacrificed at 30 days. To determine the role of SDF-1 in mediating the effects of LMWF, a specific inhibitor of the SDF-1 receptor CXCR4, AMD 3100 (20 microg/kg/day), was used. The grafted segments were evaluated by morphometric (histochemical) analyses. RESULTS Untreated aortic allografts exhibited severe intimal proliferation, indicative of TA. In contrast, LMWF treatment significantly prevented allograft intimal proliferation as compared with controls (5.7+/-3 vs. 66.2+/-6 microm, P

A working model has been developed which can be used to significantly increase the accuracy of cantilever deflection measurements using optical beam techniques (used in cantilever-based sensors and atomic force microscopes), while simultaneously simplifying their use. By using elementary geometric optics and standard vector analysis it is possible, without any fitted or adjustable parameters, to completely and accurately describe the relationship between the cantilever deflection and the signal measured by a position sensitive photo-detector. By arranging the geometry of the cantilever/optical beam, it is possible to tailor the detection system to make it more sensitive at different stages of the cantilever deflection or to simply linearize the relationship between the cantilever deflection and the measured detector signal. Supporting material and software has been made available for download at http://www.physics.mun.ca/beaulieu_lab/papers/cantilever_analysis.htm so that the reader may take full advantage of the model presented herein with minimal effort.

We report a case of a hemodialysed patient who developed a cutaneous Mycobacterium chelonae infection. This infection was only localised on the left upper limb, downstream from the arteriovenous fistula. M. chelonae is an unusual human pathogen, which is present in soil, dust, and stagnant water. Various factors, especially immunosuppression can favour this sort of infection in humans. Because of the ubiquity of the mycobacterium, the source of the inoculation sometimes remains unknown. However, a great number of cases are related to nosocomial infections. The preferential localizations are cutaneous, ocular, and pulmonary. Some cases of cutaneous infections due to M. chelonae, or caused by other atypical mycobacterium, are described in renal transplantation, peritoneal dialysis and hemodialysis. In the case we describe here, the source of contamination was not identified. The outcome was favourable with clarithromycin. This treatment is still continued because of a reappearance of the lesions when treatment was discontinued.

We present a robust and simple method for direct, label-free PCR product quantification using an integrated microelectronic sensor. The field-effect sensor can sequentially detect the intrinsic charge of multiple unprocessed PCR products and does not require sample processing or additional reagents in the PCR mixture. The sensor measures nucleic acid concentration in the PCR relevant range and specifically detects the PCR products over reagents such as Taq polymerase and nucleotide monomers. The sensor can monitor the product concentration at various stages of PCR and can generate a readout that resembles that of a real-time fluorescent measurement using an intercalating dye but without its potential inhibition artifacts. The device is mass-produced using standard semiconductor processes, can be reused for months, and integrates all sensing components directly on-chip. As such, our approach establishes a foundation for the direct integration of PCR-based in vitro biotechnologies with microelectronics.

We report on a differential microcantilever-based system capable of measuring surface stress changes which occur during electrochemical reactions. Surface stress changes induced by sub-monolayer ionic adsorption on metal surfaces and/or by electromechanical transformations in thin films can be measured. Our system is composed of two microcantilever sensors. The first active microcantilever serves as the working electrode (in a conventional three-probe electrochemical cell configuration) and as the mechanical transducer (bending of the microcantilever), yielding simultaneous, real-time, in situ measurements of the current and interfacial stress changes. The second reference microcantilever serves as a reference sensor to detect any unwanted cantilever deflection resulting from temperature variations, mechanical vibrations and/or uncontrolled chemical reactions. A technique for isolating the electrical contact made to the microcantilever from the electrolyte solution is presented. A method for creating a working electrode with a reproducible area of 1.0 mm 2 is also described. This micromechanical cantilever sensor has a deflection sensitivity of 0.2 nm, which translates to a surface stress sensitivity of 1 × 10 −4 N/m with a dynamic range of 5× 10 5 . The differential mode of this system has been characterized by measuring the potential-induced surface stress, at the Au(1 1 1) solid–liquid (HClO 4 electrolyte) interface, during anion (ClO 4 − ) adsorption on gold, while varying the solution temperature. Furthermore, we have measured the surface stress induced during ion doping/dedoping of dodecyl benzenesulfonate-doped polypyrrole (PPy(DBS)) films in an aqueous solution