Atrial assist device, a new alternative to lifelong anticoagulation?

OBJECTIVE: Atrial fibrillation is a very common heart arrhythmia, associated with a five-fold increase in the risk of embolic strokes. Treatment strategies encompass palliative drugs or surgical procedures all of which can restore sinus rhythm. Unfortunately, atria often fail to recover their mechanical function and patients therefore require lifelong anticoagulation therapy. A motorless volume displacing device (Atripump) based on artificial muscle technology, positioned on the external surface of atrium could avoid the need of oral anticoagulation and its haemorrhagic complications. An animal study was conducted in order to assess the haemodynamic effects that such a pump could provide. METHODS: Atripump is a dome-shape siliconecoated nitinol actuator sewn on the external surface of the atrium. It is driven by a pacemaker-like control unit. Five non-anticoagulated sheep were selected for this experiment. The right atrium was surgically exposed, the device sutured and connected. Haemodynamic parameters and intracardiac ultrasound (ICUS) data were recorded in each animal and under three conditions; baseline; atrial fibrillation (AF); atripump assisted AF (aaAF). RESULTS: In two animals, after 20 min of AF, small thrombi appeared in the right atrial appendix and were washed out once the pump was turned on. Assistance also enhanced atrial ejection fraction. 31% baseline; 5% during AF; 20% under aaAF. Right atrial systolic surfaces (cm2) were; 5.2 +/- 0.3 baseline; 6.2 +/- 0.1 AF; 5.4 +/- 0.3 aaAF. CONCLUSION: This compact and reliable pump seems to restore the atrial "kick" and prevents embolic events. It could avoid long-term anticoagulation therapy and open new hopes in the care of end-stage heart failure.

Tumor necrosis factor-alpha and alphaB-crystallin up-regulation during antibody-mediated demyelination in vitro: a putative protective mechanism in oligodendrocytes.

By using an in vitro model of antibody-mediated demyelination, we investigated the relationship between tumor necrosis factor-alpha (TNF-alpha) and heat shock protein (HSP) induction with respect to oligodendrocyte survival. Differentiated aggregate cultures of rat telencephalon were subjected to demyelination by exposure to antibodies against myelin oligodendrocyte glycoprotein (MOG) and complement. Cultures were analyzed 48 hr after exposure. Myelin basic protein (MBP) expression was greatly decreased, but no evidence was found for either necrosis or apoptosis. TNF-alpha was significantly up-regulated. It was localized predominantly in neurons and to a lesser extent in astrocytes and oligodendrocytes, and it was not detectable in microglial cells. Among the different HSPs examined, HSP32 and alphaB-crystallin were up-regulated; they may confer protection from oxidative stress and from apoptotic death, respectively. These results suggest that TNF-alpha, often regarded as a promoter of oligodendroglial death, could alternatively mediate a protective pathway through alphaB-crystallin up-regulation.

A modular magnetic anastomotic device for minimally invasive digestive anastomosis: proof of concept and preliminary data in the pig model.

BACKGROUND: The aim of our study was to assess the feasibility of minimally invasive digestive anastomosis using a modular flexible magnetic anastomotic device made up of a set of two flexible chains of magnetic elements. The assembly possesses a non-deployed linear configuration which allows it to be introduced through a dedicated small-sized applicator into the bowel where it takes the deployed form. A centering suture allows the mating between the two parts to be controlled in order to include the viscerotomy between the two magnetic rings and the connected viscera. METHODS AND PROCEDURES: Eight pigs were involved in a 2-week survival experimental study. In five colorectal anastomoses, the proximal device was inserted by a percutaneous endoscopic technique, and the colon was divided below the magnet. The distal magnet was delivered transanally to connect with the proximal magnet. In three jejunojejunostomies, the first magnetic chain was injected in its linear configuration through a small enterotomy. Once delivered, the device self-assembled into a ring shape. A second magnet was injected more distally through the same port. The centering sutures were tied together extracorporeally and, using a knot pusher, magnets were connected. Ex vivo strain testing to determine the compression force delivered by the magnetic device, burst pressure of the anastomosis, and histology were performed. RESULTS: Mean operative time including endoscopy was 69.2 ± 21.9 min, and average time to full patency was 5 days for colorectal anastomosis. Operative times for jejunojejunostomies were 125, 80, and 35 min, respectively. The postoperative period was uneventful. Burst pressure of all anastomoses was ≥ 110 mmHg. Mean strain force to detach the devices was 6.1 ± 0.98 and 12.88 ± 1.34 N in colorectal and jejunojejunal connections, respectively. Pathology showed a mild-to-moderate inflammation score. CONCLUSIONS: The modular magnetic system showed enormous potential to create minimally invasive digestive anastomoses, and may represent an alternative to stapled anastomoses, being easy to deliver, effective, and low cost.

α-Ketoglutarate regulates acid-base balance through an intrarenal paracrine mechanism.

Paracrine communication between different parts of the renal tubule is increasingly recognized as an important determinant of renal function. Previous studies have shown that changes in dietary acid-base load can reverse the direction of apical α-ketoglutarate (αKG) transport in the proximal tubule and Henle's loop from reabsorption (acid load) to secretion (base load). Here we show that the resulting changes in the luminal concentrations of αKG are sensed by the αKG receptor OXGR1 expressed in the type B and non-A-non-B intercalated cells of the connecting tubule (CNT) and the cortical collecting duct (CCD). The addition of 1 mM αKG to the tubular lumen strongly stimulated Cl--dependent HCO3- secretion and electroneutral transepithelial NaCl reabsorption in microperfused CCDs of wild-type mice but not Oxgr1-/- mice. Analysis of alkali-loaded mice revealed a significantly reduced ability of Oxgr1-/- mice to maintain acid-base balance. Collectively, these results demonstrate that OXGR1 is involved in the adaptive regulation of HCO3- secretion and NaCl reabsorption in the CNT/CCD under acid-base stress and establish αKG as a paracrine mediator involved in the functional coordination of the proximal and the distal parts of the renal tubule.

Mechanism, regulation, and ecological role of bacterial cyanide biosynthesis.

A few bacterial species are known to produce and excrete hydrogen cyanide (HCN), a potent inhibitor of cytochrome c oxidase and several other metalloenzymes. In the producer strains, HCN does not appear to have a role in primary metabolism and is generally considered a secondary metabolite. HCN synthase of proteobacteria (especially fluorescent pseudomonads) is a membrane-bound flavoenzyme that oxidizes glycine, producing HCN and CO2. The hcnABC structural genes of Pseudomonas fluorescens and P. aeruginosa have sequence similarities with genes encoding various amino acid dehydrogenases/oxidases, in particular with nopaline oxidase of Agrobacterium tumefaciens. Induction of the hcn genes of P. fluorescens by oxygen limitation requires the FNR-like transcriptional regulator ANR, an ANR recognition sequence in the -40 region of the hcn promoter, and nonlimiting amounts of iron. In addition, expression of the hcn genes depends on a regulatory cascade initiated by the GacS/GacA (global control) two-component system. This regulation, which is typical of secondary metabolism, manifests itself during the transition from exponential to stationary growth phase. Cyanide produced by P. fluorescens strain CHA0 has an ecological role in that this metabolite accounts for part of the biocontrol capacity of strain CHA0, which suppresses fungal diseases on plant roots. Cyanide can also be a ligand of hydrogenases in some anaerobic bacteria that have not been described as cyanogenic. However, in this case, as well as in other situations, the physiological function of cyanide is unknown.

Control of thrombin signaling through PI3K is a mechanism underlying plasticity between hair follicle dermal sheath and papilla cells.

In hair follicles, dermal papilla (DP) and dermal sheath (DS) cells exhibit striking levels of plasticity, as each can regenerate both cell types. Here, we show that thrombin induces a phosphoinositide 3-kinase (PI3K)-Akt pathway-dependent acquisition of DS-like properties by DP cells in vitro, involving increased proliferation rate, acquisition of ;myofibroblastic' contractile properties and a decreased capacity to sustain growth and survival of keratinocytes. The thrombin inhibitor protease nexin 1 [PN-1, also known as SERPINE2) regulates all those effects in vitro. Accordingly, the PI3K-Akt pathway is constitutively activated and expression of myofibroblastic marker smooth-muscle actin is enhanced in vivo in hair follicle dermal cells from PN-1(-/-) mice. Furthermore, physiological PN-1 disappearance and upregulation of the thrombin receptor PAR-1 (also known as F2R) during follicular regression in wild-type mice also correlate with such changes in DP cell characteristics. Our results indicate that control of thrombin signaling interferes with hair follicle dermal cells plasticity to regulate their function.

Validity of an arterial pressure waveform analysis device: does the puncture site play a role in the agreement with intermittent pulmonary artery catheter thermodilution measurements?

OBJECTIVE: The measurement of cardiac output is a key element in the assessment of cardiac function. Recently, a pulse contour analysis-based device without need for calibration became available (FloTrac/Vigileo, Edwards Lifescience, Irvine, CA). This study was conducted to determine if there is an impact of the arterial catheter site and to investigate the accuracy of this system when compared with the pulmonary artery catheter using the bolus thermodilution technique (PAC). DESIGN: Prospective study. SETTING: The operating room of 1 university hospital. PARTICIPANTS: Twenty patients undergoing cardiac surgery. INTERVENTIONS: CO was determined in parallel by the use of the Flotrac/Vigileo systems in the radial and femoral position (CO_rad and CO_fem) and by PAC as the reference method. Data triplets were recorded at defined time points. The primary endpoint was the comparison of CO_rad and CO_fem, and the secondary endpoint was the comparison with the PAC. MEASUREMENTS AND MAIN RESULTS: Seventy-eight simultaneous data recordings were obtained. The Bland-Altman analysis for CO_fem and CO_rad showed a bias of 0.46 L/min, precision was 0.85 L/min, and the percentage error was 34%. The Bland-Altman analysis for CO_rad and PAC showed a bias of -0.35 L/min, the precision was 1.88 L/min, and the percentage error was 76%. The Bland-Altman analysis for CO_fem and PAC showed a bias of 0.11 L/min, the precision was 1.8 L/min, and the percentage error was 69%. CONCLUSION: The FloTrac/Vigileo system was shown to not produce exactly the same CO data when used in radial and femoral arteries, even though the percentage error was close to the clinically acceptable range. Thus, the impact of the introduction site of the arterial catheter is not negligible. The agreement with thermodilution was low.

The use of the vac in complex wounds - does it work and how?

Background: Complex wounds pose a major challenge in reconstructive and trauma surgery. Several approaches to increase the healing process have been proposed in the last decades. In this study we study the mechanism of action of the Vacuum Assisted Closure device in diabetic wounds. Methods: Full-thickness wounds were excised in diabetic mice and treated with the VAC device or its isolated components: an occlusive dressing (OD) alone, subathmospheric pressure at 125 mm Hg (Suction), and a polyurethane foam without (Foam) and with (Foamc) downward compression of approximately 125 mm Hg. The last goups were treated with either the complete VAC device (VAC) or with a silicne interface that alows fluid removel (Mepithel-VAC). The effects of the treatment modes on the wound surface were quantified by a two-dimensional immunohistochemical staging system based on vasculature, as defined by blood vessel density (CD31) and cell proliferation (defined by ki67 positivity), 7 days post wounding. Finite element modelling was used to predict wound surface deformation under dressing modes and cross sections of in situ fixed tissues were used to measure actual microstrain. Results: The foam-wound interface of the Vacuum Assisted Closure device causes significant wound stains (60%) causing a deformation of the single cell level leading to a profound upregulation of cell proliferation (4-fold) and angiogenisis (2.2-fold) compared to OD treated wounds. Polyurethane foam exposure itself causes a frather unspecific angiogenic response (Foamc, 2 - fold, Foam, 2.2 - fold) without changes of the cell proliferation rate of the wound bed. Suction alone without a specific interface does not have an effect on meassured parameters, showing similar results to untreated wounds. A perforated silicone interface caused a significant lower microdeforamtion of the wound bed correlating to changes of the wound tissues. Conclusion: The Vacuum Assisted Closure device induce significanttissue growth in diabetic wounds. The wound foam interface under suction causes profound macrodeformation that stimulates tissue growth by angiogenesis and cell proliferation. It needs to be taken in consideration that in the clinical setting different wound types may profit from different elements of this suction device.

"In vitro" study of the molecular mechanism of the "E.Coli" Hsp 70/Hsp40/NEF chaperone system and its interactions with ?-sinuclein

SUMMARY Under stressful conditions, mutant or post-translationally modified proteins may spontaneously misfold and form toxie species, which may further assemble into a continuum of increasingly large and insoluble toxic oligomers that may further condense into less toxic, compact amyloids in the cell Intracellular accumulation of aggregated proteins is a common denominator of several neurodegenerative diseases. To cope with the cytotoxicity induced by abnormal, aggregated proteins, cells have evolved various defence mechanisms among which, the molecular chaperones Hsp70. Hsp70 (DnaK in E. coii) is an ATPase chaperone involved in many physiological processes in the cell, such as assisting de novo protein folding, dissociating native protein oligomers and serving as pulling motors in the import of polypeptides into organelles. In addition, Hsp70 chaperones can actively solubilize and reactivate stable protein aggregates, such as heat- or mutation-induced aggregates. Hsp70 requires the cooperation of two other co-chaperones: Hsp40 and NEF (Nucleotide exchange factor) to fulfil its unfolding activity. In the first experimental section of this thesis (Chapter II), we studied by biochemical analysis the in vitro interaction between recombinant human aggregated α-synuclein (a-Syn oligomers) mimicking toxic a-Syn oligomers species in PD brains, with a model Hsp70/Hsp40 chaperone system (the E. coii DnaK/DnaJ/GrpE). We found that chaperone-mediated unfolding of two denatured model enzymes were strongly affected by α-Syn oligomers but, remarkably, not by monomers. This in vitro observed dysfunction of the Hsp70 chaperone system resulted from the sequestration of the Hsp40 proteins by the oligomeric α-synuclein species. In the second experimental part (Chapter III), we performed in vitro biochemical analysis of the co-chaperone function of three E. coii Hsp40s proteins (DnaJ, CbpA and DjlA) in the ATP-fuelled DnaK-mediated refolding of a model DnaK chaperone substrate into its native state. Hsp40s activities were compared using dose-response approaches in two types of in vitro assays: refolding of heat-denatured G6PDH and DnaK-mediated ATPase activity. We also observed that the disaggregation efficiency of Hsp70 does not directly correlate with Hsp40 binding affinity. Besides, we found that these E. coii Hsp40s confer substrate specificity to DnaK, CbpA being more effective in the DnaK-mediated disaggregation of large G6PDH aggregates than DnaJ under certain conditions. Sensibilisées par différents stress ou mutations, certaines protéines fonctionnelles de la cellule peuvent spontanément se convertir en formes inactives, mal pliées, enrichies en feuillets bêta, et exposant des surfaces hydrophobes favorisant l'agrégation. Cherchant à se stabiliser, les surfaces hydrophobes peuvent s'associer aux régions hydrophobes d'autres protéines mal pliées, formant des agrégats protéiques stables: les amyloïdes. Le dépôt intracellulaire de protéines agrégées est un dénominateur commun à de nombreuses maladies neurodégénératives. Afin de contrer la cytotoxicité induite par les protéines agrégées, les cellules ont développé plusieurs mécanismes de défense, parmi lesquels, les chaperonnes moléculaires Hsp70. Hsp70 nécessite la collaboration de deux autres co-chaperonnes : Hsp40 et NEF pour accomplir son activité de désagrégation. Hsp70 (DnaK, chez E. coli) est impliquée par ailleurs dans d'autres fonctions physiologiques telles que l'assistanat de protéines néosynthétisées à la sortie du ribosome, ou le transport transmembranaire de polypeptides. Par ailleurs, les chaperonnes Hsp70 peuvent également solubiliser et réactiver des protéines agrégées à la suite d'un stress ou d'une mutation. Dans la première partie expérimentale de cette thèse (Chapter II), nous avons étudié in vitro l'interaction entre les oligomères d'a-synucleine, responsables entre autres, de la maladie de Parkinson, et le système chaperon Hsp70/Hsp40 (système Escherichia coli DnaK/DnaJ/GrpE). Nous avons démontré que contrairement aux monomères, les oligomères d'a-synucleine inhibaient le système chaperon lors du repliement de protéines agrégées. Cette dysfonction du système chaperon résulte de la séquestration des chaperonnes Hsp40 par les oligomères d'a-synucleine. La deuxième partie expérimentale (Chapitre III) est consacrée à une étude in vitro de la fonction co-chaperonne de trois Hsp40 d'is. coli (DnaJ, CbpA, et DjlA) lors de la désagrégation par DnaK d'une protéine pré-agrégée. Leurs activités ont été comparées par le biais d'une approche dose-réponse au niveau de deux analyses enzymatiques: le repliement de la protéine agrégée et l'activité ATPase de DnaK. Par ailleurs, nous avons mis en évidence que l'efficacité de désagrégation d'Hsp70 et l'affinité des chaperonnes Hsp40 vis-à-vis de leur substrat n'étaient pas corrélées positivement. Nous avons également montré que ces trois chaperonnes Hsp40 étaient directement impliquées dans la spécificité des fonctions accomplies par les chaperonnes Hsp70. En effet, DnaK en présence de CbpA assure la désagrégation de large agrégats protéiques avec une efficacité nettement plus accrue qu'en présence de DnaJ.

AMP-activated protein kinase mediates glucocorticoid-induced metabolic changes: a novel mechanism in Cushing's syndrome.

Chronic exposure to glucocorticoid hormones, resulting from either drug treatment or Cushing's syndrome, results in insulin resistance, central obesity, and symptoms similar to the metabolic syndrome. We hypothesized that the major metabolic effects of corticosteroids are mediated by changes in the key metabolic enzyme adenosine monophosphate-activated protein kinase (AMPK) activity. Activation of AMPK is known to stimulate appetite in the hypothalamus and stimulate catabolic processes in the periphery. We assessed AMPK activity and the expression of several metabolic enzymes in the hypothalamus, liver, adipose tissue, and heart of a rat glucocorticoid-excess model as well as in in vitro studies using primary human adipose and primary rat hypothalamic cell cultures, and a human hepatoma cell line treated with dexamethasone and metformin. Glucocorticoid treatment inhibited AMPK activity in rat adipose tissue and heart, while stimulating it in the liver and hypothalamus. Similar data were observed in vitro in the primary adipose and hypothalamic cells and in the liver cell line. Metformin, a known AMPK regulator, prevented the corticosteroid-induced effects on AMPK in human adipocytes and rat hypothalamic neurons. Our data suggest that glucocorticoid-induced changes in AMPK constitute a novel mechanism that could explain the increase in appetite, the deposition of lipids in visceral adipose and hepatic tissue, as well as the cardiac changes that are all characteristic of glucocorticoid excess. Our data suggest that metformin treatment could be effective in preventing the metabolic complications of chronic glucocorticoid excess.

EZ-IO(®) intraosseous device implementation in a pre-hospital emergency service: A prospective study and review of the literature.

INTRODUCTION: Intraosseous access is increasingly recognised as an effective alternative vascular access to peripheral venous access. We aimed to prospectively study the patients receiving prehospital intraosseous access with the EZ-IO(®), and to compare our results with those of the available literature. METHODS: Every patient who required an intraosseous access with the EZ-IO from January 1st, 2009 to December 31st, 2011 was included. The main data collected were: age, sex, indication for intraosseous access, localisation of insertion, success rate, drugs and fluids administered, and complications. All published studies concerning the EZ-IO device were systematically searched and reviewed for comparison. RESULTS: Fifty-eight patients representing 60 EZ-IO procedures were included. Mean age was 47 years (range 0.5-91), and the success rate was 90%. The main indications were cardiorespiratory arrest (74%), major trauma (12%), and shock (5%). The anterior tibia was the main route. The main drugs administered were adrenaline (epinephrine), atropine and amiodarone. No complications were reported. We identified 30 heterogeneous studies representing 1603 EZ-IO insertions. The patients' characteristics and success rate were similar to our study. Complications were reported in 13 cases (1.3%). CONCLUSION: The EZ-IO provides an effective way to achieve vascular access in the pre-hospital setting. Our results were similar to the cumulative results of all studies involving the use of the EZ-IO, and that can be used for comparison for further studies.

Role of the GNOM gene in Arabidopsis apical-basal patterning--From mutant phenotype to cellular mechanism of protein action.

How the apical-basal axis of polarity is established in embryogenesis is still a mystery in plant development. This axis appeared specifically compromised by mutations in the Arabidopsis GNOM gene. Surprisingly, GNOM encodes an ARF guanine-nucleotide exchange factor (ARF-GEF) that regulates the formation of vesicles in membrane trafficking. In-depth functional analysis of GNOM and its closest relative, GNOM-LIKE 1 (GNL1), has provided a mechanistic explanation for the development-specific role of a seemingly mundane trafficking regulator. The current model proposes that GNOM is specifically involved in the endosomal recycling of the auxin-efflux carrier PIN1 to the basal plasma membrane in provascular cells, which in turn is required for the accumulation of the plant hormone auxin at the future root pole through polar auxin transport. Thus, the analysis of GNOM highlights the importance of cell-biological processes for a mechanistic understanding of development.

Bacterial-induced protection against allergy through a novel multi-component immunoregulatory mechanism

Airborne microbial products have been reported to promote immune responses that suppress asthma, yet how these beneficial effects take place remains controversial and poorly understood. We have found that pulmonary exposure with the bacterium Escherichia coli leads to a suppression of allergic airway inflammation, characterized by reduced airway-hyperresponsiveness, eosinophilia and cytokine production by T cells in the lung. This immune modulation was neither mediated by the induction of a Th1 response nor regulatory T cells; was dependent on TLR-4 but did not involve TLR-desensitization. Dendritic cell migration to the draining lymph nodes and subsequent activation of T cells was unaffected by prior exposure to E.coli indicating that the immunomodulation was limited to the lung environment. In non-treated control mice ovalbumin was primarily presented by airway CD11b+ CD11c+ DCs expressing high levels of MHC class II molecules whilst the DCs in E.coli-treated mice displayed a less activated phenotype and had impaired antigen presentation capacity. Consequently, in situ Th2 cytokine production by ovalbuminspecific effector T cells recruited to the airways was significantly reduced. The suppression of airways hyper responsiveness was mediated through the recruitment of IL-17-producing gd-T cells; however, the suppression of dendritic cells and T cells was mediated through a distinct mechanism that could not be overcome by the local administration of activated dendritic cells, or by the in vivo administration of TNF-alpha. Taken together, these data reveal a novel multi-component immunoregulatory pathway that acts to protect the airways from allergic inflammation.