Cette étude a été financée en partie par la subvention la NASA Nebraska espace et la National Science Foundation sous octroi n ° 0926880.

1. Préparation du bateau Latex <ol><li> Fixer les deux extrémités de l&#39;enceinte de latex à des connexions flexibles cannelés, qui sont fixés sur une table de travail solide. </li><li> Mesurer la zone d&#39;intérêt sur le navire en latex afin de déterminer le champ de vision. La zone d&#39;intérêt pour un test de stent doit être centré entre les raccords cannelés et comprennent environ un pouce de chaque côté du stent afin d&#39;observer les souches en dehors de la zone stent. </li><li> Enregistrer la distance entre le bord extérieur d&#39;un raccord de tuyau cannelé à l&#39;emplacement central entre les connecteurs, qui est également le centre approximatif de la cuve de latex. Traduire la distance sur le cathéter en mesurant à partir du centre du stent jusqu&#39;à l&#39;cathéter. Ensuite, marquez le cathéter avec un marqueur. </li><li> Retirez le récipient du latex provenant des raccords de tuyaux de fer barbelé. </li><li> Préparer le navire en latex par pulvérisation de la zone d&#39;intérêt avec un modèle stochastique de la peinture en aérosol blanc et le noir ou le marquagela zone d&#39;intérêt avec des points aléatoires en utilisant un marqueur permanent. Pour les plus petits échantillons et modèle stochastique fine est nécessaire. </li></ol> 2. Système de test in vitro et d&#39;étalonnage du système ARAMIS <ol><li> Sélectionner le panneau d&#39;étalonnage qui est légèrement plus grande que la zone d&#39;intérêt mesuré à l&#39;étape 1. </li><li> Placez le panneau d&#39;étalonnage entre les raccords cannelés à la zone d&#39;intérêt et de veiller à ce que la zone d&#39;intérêt est bien éclairée. </li><li> Régler la distance entre deux caméras, la distance de l&#39;échantillon, et la hauteur de la caméra sur la base du panneau d&#39;étalonnage sélectionné. Chaque panneau d&#39;étalonnage est différent, donc le mode d&#39;emploi ARAMIS devra être consulté afin de déterminer ces distances. </li><li> Ouvrez un nouveau projet dans ARAMIS en sélectionnant &quot;Fichier&quot;, puis &quot;Nouveau Projet&quot;. Cliquez ensuite sur le &quot;capteur&quot; et sélectionnez &quot;Calibration&quot;, puis &quot;d&#39;étalonnage complet&quot;. </li><li> Le logiciel ARAMIS va maintenant guider l&#39;utilisateur à travers les étapes pour calibrer t<html> il caméras. </li><li> Avec l&#39;ouverture de l&#39;objectif totalement ouverte, le focus de la caméra sur le panneau d&#39;étalonnage en desserrant la vis de réglage sur l&#39;appareil photo et la rotation de la lentille. Une fois la mise, resserrer la vis de réglage et fermer l&#39;ouverture. </li><li> Prenez la première image du processus d&#39;étalonnage. Déplacer ou faire pivoter le panneau d&#39;étalonnage selon la démonstration sur l&#39;ordinateur, jusqu&#39;à ce que l&#39;image est centrée sur l&#39;écran de l&#39;ordinateur. Prenez la deuxième image. Répétez cette procédure pour le reste des images de calibration. </li><li> Une fois que toutes les images d&#39;étalonnage sont prises, le logiciel ARAMIS analyse d&#39;imagerie va calculer les paramètres d&#39;étalonnage. Le processus d&#39;étalonnage doit être répétée si l&#39;écart d&#39;étalonnage est supérieure à 0,04. Les ajustements apportés à la mise au point de la caméra ou la distance entre les caméras rendra le vide processus d&#39;étalonnage. </li><li> Retirez le panneau d&#39;étalonnage et de placer le navire en latex peint en arrière sur les raccords de tuyaux de fer barbelé. </li></ol>titre &quot;&gt; 3. Prétest pour éviter le bruit de fond excessif <ol><li> Déterminer le nombre d&#39;images par seconde que l&#39;on souhaite pour le test. Augmentation des cadres par seconde sera de produire des résultats plus uniformes de contrainte. </li><li> Réglez la vitesse d&#39;obturation inférieure à 1 image par seconde et que personne ne le rouge est affiché sur l&#39;image. </li><li> Prenez 5 images. </li><li> Ajouter les points de départ sur la série d&#39;images et de calculer le test. </li><li> Tout en maintenant la touche &quot;ctrl&quot;, cliquez sur le centre de l&#39;échantillon à observer le bruit de fond. Si le pré-test de bruit est supérieur à 75 microdéformation le processus de calibrage doit être refait. </li></ol> 4. Le déploiement du stent <ol><li> Sélectionnez le nombre d&#39;images souhaitées à prendre lors de l&#39;essai. 200 images suffiront pour l&#39;expansion du stent. </li><li> Progressivement insérer le cathéter dans le vaisseau de latex, et en utilisant l&#39;indicateur de marqueur sur le cathéter pour guider l&#39;insertion de l&#39;extenseur jusqu&#39;à ce qu&#39;il atteigne l&#39;emplacement central. </li><li> Commencez à prendre des images avec ARAMIS. </li><li> Pour le stent expansible par ballonnet, augmenter progressivement la pression de ballon pour étendre stent jusqu&#39;à ce que le ballonnet est entièrement étendu, puis diminue progressivement la pression de ballonnet à zéro et le ballonnet est dégonflé et retirée en même temps que le cathéter. </li><li> Pour le stent auto-expansible, progressivement enlever la gaine jusqu&#39;à ce que le stent est complètement déployée, puis progressivement rétracter le cathéter. </li></ol> 5. Analyse des Images <ol><li> Historique de la souche d&#39;un point spécifique sur le navire <ol><li> Créer un point d&#39;étape en maintenant la touche &quot;ctrl&quot; et en cliquant sur la zone d&#39;intérêt. </li><li> Sélectionner le type de souche qui est désiré, la souche à-dire dans les directions X, Y, XY, souche majeure, la souche mineure, ou souche Mises. </li><li> L&#39;intrigue dans le coin en bas à droite affiche la souche au point sélectionné sur la durée du test. </li></ol></li><li> Souche spatiale long d&#39;un trajet spécifique du récipient <ol><li> Crmangera une ligne point de plusieurs étapes en cliquant sur le &quot;sections&quot;, puis &quot;créer une section&quot;. Sélectionner une ligne sur la parallèle d&#39;image à l&#39;axe X à Y est égal à zéro. Cela va créer un certain nombre de points d&#39;étape en ligne. </li><li> Après la ligne à plusieurs étages est créé la parcelle dans le coin inférieur affichera une série de lignes sur une parcelle. Chaque ligne représente la souche moins une instance dans le temps le long de la longueur de la section. </li></ol></li><li> La création d&#39;un cylindre meilleur ajustement pour analyser le taux d&#39;expansion et le rayon de la cuve <ol><li> Dans la barre d&#39;outils supérieure sélectionnez &quot;primitives&quot;, puis &quot;cylindre meilleur ajustement&quot;. </li><li> Sélectionnez une petite section de l&#39;image en utilisant le &quot;sélectionner à travers la surface&quot; outil sur la barre d&#39;outils droite. </li><li> Le logiciel ARAMIS va générer un trois cylindres dimensions meilleur ajustement. </li><li> Les images peuvent ensuite être recyclés dans d&#39;observer la façon dont le diamètre du vaisseau latex est variable. </li></ol></li><li> Évaluation de la distance entredeux points <ol><li> Dans le cadre du clic &quot;analyse&quot; sur l&#39;onglet &quot;point à point à distance&quot;. </li><li> Sélectionner une longueur de l&#39;image qui est désiré pour l&#39;analyse en sélectionnant deux points. </li><li> Les images peuvent être recyclés dans d&#39;observer le changement de distance entre les deux points dans le temps. </li></ol></li></ol> 6. Les résultats représentatifs Les entretoises d&#39;endoprothèse élargir vers l&#39;extérieur de la paroi vasculaire, les souches seront généralement supérieures à environ Lieu de stent. Figure 1 est un exemple de la cartographie souche au cours du processus de recul de ballon-stent expansible, ainsi que l&#39;histoire contrainte majeure à un moment spécifique. Les points noirs de la figure 1 sont des points de référence, qui ont été utilisés par les caméras à haute vitesse pour saisir et de suivre les déplacements de ces points de référence sur le conduit. Basé sur le mouvement enregistré des points de référence, le logiciel sera alors utilisée pour calculer les souches de la conduite ou unny autre objet ciblé. Souche Major, appelée aussi la souche maximum en principal, est calculé comme suit: <img alt="L&#39;équation 1" src="/files/ftp_upload/3945/3945eq1.jpg" /> Il est clair que le stent implanté conduit à la distribution des déformations non-uniforme sur la surface du vaisseau. Cela pourrait s&#39;expliquer par le recul de chargement à partir des extrémités-contrainte conduit latex et la structure en treillis de l&#39;endoprothèse. Ce champ de déformation correspond à la phase initiale de recul stent, comme identifié par le marqueur de la croix rouge dans l&#39;image en bas de la figure 1. La majeure souche histoire courbe d&#39;un point spécifique de 10 étapes distinctes démontré de l&#39;implantation du stent. L&#39;expansion se produit ballon d&#39;environ 10 à 12 secondes et le recul de stent de la déflation du ballon se produit entre 12 et 14 secondes. <img alt="Figure 1" src="/files/ftp_upload/3945/3945fig1.jpg" /> Figure 1. </strong&gt; Dispositif expérimental (en haut); la distribution des déformations non-uniforme sur la surface conduit stent (au milieu); l&#39;historique de la souche majeure au point 10 (en bas).

<ol>
	<li>Fischman, D. L., Leon, M. B., Baim, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+randomized+comparison+of+coronary-stent+placement+and+balloon+angioplasty+in+the+treatment+of+coronary+artery+disease.+Stent+Restenosis+Study+Investigators.">A randomized comparison of coronary-stent placement and balloon angioplasty in the treatment of coronary artery disease. Stent Restenosis Study Investigators.</a> N. Engl. J. Med. 331, 496-501 (1994).</li><li>Abul Hasan Muhammad Bashar, T. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+Properties+of+Various+Z-Stent+Designs:+An+Endovascular+Stent-Grafting+Perspective.">Mechanical Properties of Various Z-Stent Designs: An Endovascular Stent-Grafting Perspective.</a> Artificial Organs. 27, 714-721 (2003).</li><li>Nuutinen, J. u. h. a. -. P. e. k. k. a. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+properties+and+in+vitro+degradation+of+bioabsorbable+self-expanding+braided+stents.">Mechanical properties and in vitro degradation of bioabsorbable self-expanding braided stents.</a> Journal of Biomaterials Science -- Polymer Edition. , 255-266 (2003).</li><li>C. Schulz, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coronary+stent+symmetry+and+vascular+injury+determine+experimental+restenosis.">Coronary stent symmetry and vascular injury determine experimental restenosis.</a> Heart. 83, 462-467 (2000).</li><li>Jiménez, J. M., Davies, P. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hemodynamically+Driven+Stent+Strut+Design.">Hemodynamically Driven Stent Strut Design.</a> Annals of Biomedical Engineering. 1483, (2009).</li><li>Johnston, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Mechanical+Properties+of+Endovascular+Stents:+An+In+Vitro+Assessment.">The Mechanical Properties of Endovascular Stents: An In Vitro Assessment.</a> Cardiovascular Engineering: An International Journal. 10, 128-135 (2010).</li><li>Mejia, J. u. a. n. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+the+effect+of+stent+strut+profile+on+shear+stress+distribution+using+statistical+moments.">Evaluation of the effect of stent strut profile on shear stress distribution using statistical moments.</a> Biomedical Engineering Online. , 1-10 (2009).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> ARAMIS User Manual. , (2009).</li><li>Chesler, N. C., Thompson-Figueroa, J., Millburne, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurements+of+Mouse+Pulmonary+Biomechanics.">Measurements of Mouse Pulmonary Biomechanics.</a> Journal of Biomechanical Engineering. 126, 309-314 (2004).</li><li>de Korte, C. L., Sierevogel, M. J., Mastik, F., Strijder, C., Schaar, J. A., Velema, E., Pasterkamp, G., Serruys, P. W., van der Steen, A. F. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+Atherosclerotic+Plaque+Components+With+Intravascular+Ultrasound+Elastography+In+Vivo+A+Yucatan+Pig+Study.">Identification of Atherosclerotic Plaque Components With Intravascular Ultrasound Elastography In Vivo A Yucatan Pig Study.</a> Circulation. 105, 1627-1630 (2002).</li></ol>

Pas de conflits d&#39;intérêt déclarés.

Le système stéréo de mesure optique de surface souche est utilisée pour mesurer les déformations locales sur la surface de déformation pour les deux en-et out-of-avion mouvements sans contact avec l&#39;échantillon. Ce système utilise deux caméras haute vitesse optiques pour prendre des photos d&#39;un motif aléatoire contrastant mettre sur la surface à construire des mesures précises des mouvements de chaque point, avec une grande précision de résoudre les souches de surface. ...

<table border="1"><tbody><tr><td> Matériels d&#39;occasion </td><td> Entreprise </td><td> Numéro de catalogue </td><td> Commentaires </td></tr><tr><td> ARAMIS système de caméra </td><td> GOM: techniques de mesure optiques </td><td></td><td></td></tr><tr><td> STENT PALMAZ Genesis transhépatique BILIAIRE </td><td> Cordis Corporation </td><td> PG5910B </td><td> Ballon-stent expansible </td></tr><tr><td> Z-MED cathéter de dilatation à ballonnet </td><td> B. Braun Medical Inc </td><td> PDZ336 </td><td> Cathéter de dilatation à ballonnet </td></tr></tbody></table>

monitoring the wall mechanics during stent deployment in a vessel

Les essais cliniques ont signalé des taux de resténose différents pour différents modèles de stent 1. Il est spéculé que les concentrations de contraintes induites par stent sur ​​la paroi artérielle conduire à des lésions tissulaires, qui initie la resténose 2-7. Cette hypothèse nécessite des investigations supplémentaires, y compris de meilleures quantifications de la distribution des déformations non-uniforme sur l&#39;artère la suite de l&#39;implantation du stent. Une méthode non-contact de surface souche de mesure pour l&#39;artère stent est présenté dans ce travail. Stéréo ARAMIS surface optique de mesure de déformation système utilise deux caméras optiques à haute vitesse pour capturer le mouvement de chaque point de référence, et de résoudre les trois souches dimensions sur la surface déformer 8,9. En tant que maillage de stent est déployé dans un vaisseau latex avec un motif aléatoire de contraste pulvérisé ou tiré sur sa surface extérieure, la souche de surface est enregistré à chaque instant de la déformation. Les distributions de contrainte calculées peuvent ensuite être utilisées pour comprendre la loréponse lésion cal, de valider les modèles de calcul, et de formuler des hypothèses pour des études in vivo.

Watch this Scientific Journal Video about Monitoring the Wall Mechanics During Stent Deployment in a Vessel at JoVE.com

Monitoring the Wall Mechanics During Stent Deployment in a Vessel

Stent-induites des distributions de contrainte artérielles sont caractérisés à l&#39;aide d&#39;un système de surface optique souche de mesure. Cette technique de visualisation est utilisé pour obtenir un aperçu de l&#39;impact de l&#39;implantation du stent sur le navire hôte.

Suivi des Mécanique mur pendant le déploiement du stent dans un vaisseau

Clinical trials have reported different restenosis rates for various stent designs1. It is speculated that stent-induced strain concentrations on the ...

monitoring-the-wall-mechanics-during-stent-deployment-in-a-vessel

Research

JoVE Journal

Bioengineering

9.2K vues.  University of Nebraska-Lincoln. Stent-induites des distributions de contrainte artérielles sont caractérisés à l'aide d'un système de surface optique souche de mesure. Cette technique de visualisation est utilisé pour obtenir un aperçu de l'impact de l'implantation du stent sur le navire hôte.

Vidéo: Monitoring the Wall Mechanics During Stent Deployment in a Vessel

Magnetic Resonance Elastography Methodology for the Evaluation of Tissue Engineered Construct Growth

Traditional mechanical testing often results in the destruction of the sample, and in the case of long term tissue engineered construct studies, the use of destructive assessment is not acceptable. A proposed alternative is the use of an imaging process called magnetic resonance elastography. Elastography is a nondestructive method for determining the engineered outcome by measuring local mechanical property values (i.e., complex shear modulus), which are essential markers for identifying the structure and functionality of a tissue. As a noninvasive means for evaluation, the monitoring of engineered constructs with imaging modalities such as magnetic resonance imaging (MRI) has seen increasing interest in the past decade1. For example, the magnetic resonance (MR) techniques of diffusion and relaxometry have been able to characterize the changes in chemical and physical properties during engineered tissue development2. The method proposed in the following protocol uses microscopic magnetic resonance elastography (&mu;MRE) as a noninvasive MR based technique for measuring the mechanical properties of small soft tissues3. MRE is achieved by coupling a sonic mechanical actuator with the tissue of interest and recording the shear wave propagation with an MR scanner4. Recently, &mu;MRE has been applied in tissue engineering to acquire essential growth information that is traditionally measured using destructive mechanical macroscopic techniques5. In the following procedure, elastography is achieved through the imaging of engineered constructs with a modified Hahn spin-echo sequence coupled with a mechanical actuator. As shown in Figure 1, the modified sequence synchronizes image acquisition with the transmission of external shear waves; subsequently, the motion is sensitized through the use of oscillating bipolar pairs. Following collection of images with positive and negative motion sensitization, complex division of the data produce a shear wave image. Then, the image is assessed using an inversion algorithm to generate a shear stiffness map6. The resulting measurements at each voxel have been shown to strongly correlate (R2&gt;0.9914) with data collected using dynamic mechanical analysis7. In this study, elastography is integrated into the tissue development process for monitoring human mesenchymal stem cell (hMSC) differentiation into adipogenic and osteogenic constructs as shown in Figure 2.

The procedure demonstrates the methodology of magnetic resonance elastography for monitoring the engineered outcome of adipose and osteogenic tissue engineered constructs through noninvasive local assessment of the mechanical properties using microscopic magnetic resonance elastography (&mu;MRE).

Méthodologie élastographie par résonance magnétique pour l&#39;évaluation des tissus Conçu croissance Construct

Traditional mechanical testing often results in the destruction of the sample, and in the case of long term tissue engineered construct studies, the use ...

Recherche

Bio-ingénierie

Mass Cytometry: Protocol for Daily Tuning and Running Cell Samples on a CyTOF Mass Cytometer

In recent years, the rapid analysis of single cells has commonly been performed using flow cytometry and fluorescently-labeled antibodies. However, the issue of spectral overlap of fluorophore emissions has limited the number of simultaneous probes. In contrast, the new CyTOF mass cytometer by DVS Sciences couples a liquid single-cell introduction system to an ICP-MS.1 Rather than fluorophores, chelating polymers containing highly-enriched metal isotopes are coupled to antibodies or other specific probes.2-5 Because of the metal purity and mass resolution of the mass cytometer, there is no "spectral overlap" from neighboring isotopes, and therefore no need for compensation matrices. Additionally, due to the use of lanthanide metals, there is no biological background and therefore no equivalent of autofluorescence. With a mass window spanning atomic mass 103-203, theoretically up to 100 labels could be distinguished simultaneously. Currently, more than 35 channels are available using the chelating reagents available from DVS Sciences, allowing unprecedented dissection of the immunological profile of samples.6-7
Disadvantages to mass cytometry include the strict requirement for a separate metal isotope per probe (no equivalent of forward or side scatter), and the fact that it is a destructive technique (no possibility of sorting recovery). The current configuration of the mass cytometer also has a cell transmission rate of only ~25%, thus requiring a higher input number of cells.
Optimal daily performance of the mass cytometer requires several steps. The basic goal of the optimization is to maximize the measured signal intensity of the desired metal isotopes (M) while minimizing the formation of oxides (M+16) that will decrease the M signal intensity and interfere with any desired signal at M+16. The first step is to warm up the machine so a hot, stable ICP plasma has been established. Second, the settings for current and make-up gas flow rate must be optimized on a daily basis. During sample collection, the maximum cell event rate is limited by detector efficiency and processing speed to 1000 cells/sec. However, depending on the sample quality, a slower cell event rate (300-500 cells/sec) is usually desirable to allow better resolution between cells events and thus maximize intact singlets over doublets and debris. Finally, adequate cleaning of the machine at the end of the day helps minimize background signal due to free metal.

The steps necessary for daily tuning and optimization of the performance of a CyTOF mass cytometer are described. Comments on optimal sample preparation and flow rate are discussed

Cytométrie de masse: Protocole pour Tuning quotidien et échantillons cellulaires course sur un cytomètre de masse CyTOF

In recent years, the rapid analysis of single cells has commonly been performed using flow cytometry and fluorescently-labeled antibodies. However, the ...

A Versatile Automated Platform for Micro-scale Cell Stimulation Experiments

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in vitro analysis are well suited to study of large numbers of cells (&ge;105) in milliliter-scale volumes (&ge;0.1 ml). However, there are many instances in which it is necessary or desirable to scale down culture size to reduce consumption of the cells of interest and/or reagents required for their culture, stimulation, or processing. Unfortunately, conventional approaches do not support precise and reproducible manipulation of micro-scale cultures, and the microfluidics-based automated systems currently available are too complex and specialized for routine use by most laboratories. To address this problem, we have developed a simple and versatile technology platform for automated culture, stimulation, and recovery of small populations of cells (100 - 2,000 cells) in micro-scale volumes (1 - 20 &mu;l). The platform consists of a set of fibronectin-coated microcapillaries ("cell perfusion chambers"), within which micro-scale cultures are established, maintained, and stimulated; a digital microfluidics (DMF) device outfitted with "transfer" microcapillaries ("central hub"), which routes cells and reagents to and from the perfusion chambers; a high-precision syringe pump, which powers transport of materials between the perfusion chambers and the central hub; and an electronic interface that provides control over transport of materials, which is coordinated and automated via pre-determined scripts. As an example, we used the platform to facilitate study of transcriptional responses elicited in immune cells upon challenge with bacteria. Use of the platform enabled us to reduce consumption of cells and reagents, minimize experiment-to-experiment variability, and re-direct hands-on labor. Given the advantages that it confers, as well as its accessibility and versatility, our platform should find use in a wide variety of laboratories and applications, and prove especially useful in facilitating analysis of cells and stimuli that are available in only limited quantities.

We have developed an automated cell culture and interrogation platform for micro-scale cell stimulation experiments. The platform offers simple, versatile, and precise control in cultivating and stimulating small populations of cells, and recovering lysates for molecular analyses. The platform is well suited to studies that use precious cells and/or reagents.

Une plate-forme automatisée polyvalent pour micro-échelle expériences de stimulation cellulaire

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in ...

Experimental Investigation of Secondary Flow Structures Downstream of a Model Type IV Stent Failure in a 180&#176; Curved Artery Test Section

The arterial network in the human vasculature comprises of ubiquitously present blood vessels with complex geometries (branches, curvatures and tortuosity). Secondary flow structures are vortical flow patterns that occur in curved arteries due to the combined action of centrifugal forces, adverse pressure gradients and inflow characteristics. Such flow morphologies are greatly affected by pulsatility and multiple harmonics of physiological inflow conditions and vary greatly in size-strength-shape characteristics compared to non-physiological (steady and oscillatory) flows 1 - 7.
Secondary flow structures may ultimately influence the wall shear stress and exposure time of blood-borne particles toward progression of atherosclerosis, restenosis, sensitization of platelets and thrombosis 4 - 6, 8 - 13. Therefore, the ability to detect and characterize these structures under laboratory-controlled conditions is precursor to further clinical investigations.
A common surgical treatment to atherosclerosis is stent implantation, to open up stenosed arteries for unobstructed blood flow. But the concomitant flow perturbations due to stent installations result in multi-scale secondary flow morphologies 4 - 6. Progressively higher order complexities such as asymmetry and loss in coherence can be induced by ensuing stent failures vis-&#224;-vis those under unperturbed flows 5. These stent failures have been classified as &#34;Types I-to-IV&#34; based on failure considerations and clinical severity 14.
This study presents a protocol for the experimental investigation of the complex secondary flow structures due to complete transverse stent fracture and linear displacement of fractured parts (&#34;Type IV&#34;) in a curved artery model. The experimental method involves the implementation of particle image velocimetry (2C-2D PIV) techniques with an archetypal carotid artery inflow waveform, a refractive index matched blood-analog working fluid for phase-averaged measurements 15 - 18. Quantitative identification of secondary flow structures was achieved using concepts of flow physics, critical point theory and a novel wavelet transform algorithm applied to experimental PIV data 5, 6, 19 - 26.

Experimental Investigation of Secondary Flow Structures Downstream of a Model Type IV Stent Failure in a 180&amp;#176; Curved Artery Test Section

Stent implants in stenosed arterial curvatures are prone to "Type IV" failures involving the complete transverse fracture of stents and linear displacement of the fractured parts. We present a protocol for detection of secondary flow (vortical) structures in a curved artery model, downstream of clinically relevant "Type IV" stent failures.

Etude expérimentale des structures de flux secondaire en aval d&#39;un échec Modèle Type IV Stent dans une section d&#39;essai de 180 ° Artère Curved

The arterial network in the human vasculature comprises of ubiquitously present blood vessels with complex geometries (branches, curvatures and ...

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to intrinsic parameters within the process allow a high degree of control over scaffold characteristics including fiber diameter, alignment and porosity. By developing scaffolds with similar dimensions and topographies to organ- or tissue-specific extracellular matrices (ECM), micro-environments representative to those that cells are exposed to in situ can be created. 
The airway bronchiole wall, comprised of three main micro-environments, was selected as a model tissue. Using decellularized airway ECM as a guide, we electrospun the non-degradable polymer, polyethylene terephthalate (PET), by three different protocols to produce three individual electrospun scaffolds optimized for epithelial, fibroblast or smooth muscle cell-culture. Using a commercially available bioreactor system, we stably co-cultured the three cell-types to provide an in vitro model of the airway wall over an extended time period.
This model highlights the potential for such methods being employed in in vitro diagnostic studies investigating important inter-cellular cross-talk mechanisms or assessing novel pharmaceutical targets, by providing a relevant platform to allow the culture of fully differentiated adult cells within 3D, tissue-specific environments.

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Advancements in biomaterial technologies enable the development of three-dimensional multi-cell-type constructs. We have developed electrospinning protocols to produce three individual scaffolds to culture the main structural cells of the airway to provide a 3D in vitro model of the airway bronchiole wall.

Adapter le processus Électrofilage de fournir trois environnements uniques pour un Tri-couches<em&gt; In Vitro</em&gt; Modèle du mur Airway

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to ...

Ferromagnetic Bare Metal Stent for Endothelial Cell Capture and Retention

Rapid endothelialization of cardiovascular stents is needed to reduce stent thrombosis and to avoid anti-platelet therapy which can reduce bleeding risk. The feasibility of using magnetic forces to capture and retain endothelial outgrowth cells (EOC) labeled with super paramagnetic iron oxide nanoparticles (SPION) has been shown previously. But this technique requires the development of a mechanically functional stent from a magnetic and biocompatible material followed by in-vitro and in-vivo testing to prove rapid endothelialization. We developed a weakly ferromagnetic stent from 2205 duplex stainless steel using computer aided design (CAD) and its design was further refined using finite element analysis (FEA). The final design of the stent exhibited a principal strain below the fracture limit of the material during mechanical crimping and expansion. One hundred stents were manufactured and a subset of them was used for mechanical testing, retained magnetic field measurements, in-vitro cell capture studies, and in-vivo implantation studies. Ten stents were tested for deployment to verify if they sustained crimping and expansion cycle without failure. Another 10 stents were magnetized using a strong neodymium magnet and their retained magnetic field was measured. The stents showed that the retained magnetism was sufficient to capture SPION-labeled EOC in our in-vitro studies. SPION-labeled EOC capture and retention was verified in large animal models by implanting 1 magnetized stent and 1 non-magnetized control stent in each of 4 pigs. The stented arteries were explanted after 7 days and analyzed histologically. The weakly magnetic stents developed in this study were capable of attracting and retaining SPION-labeled endothelial cells which can promote rapid healing.

Our goals were to design, manufacture and test ferromagnetic stents for endothelial cell capture. Ten stents were tested for fracture and 10 more stents were tested for retained magnetism. Finally, 10 stents were tested in-vitro and 8 more stents were implanted in 4 pigs to show cell capture and retention.

Ferromagnétique Bare Metal Stent pour endothéliale capture de cellules et de la conservation

Rapid endothelialization of cardiovascular stents is needed to reduce stent thrombosis and to avoid anti-platelet therapy which can reduce bleeding risk. ...

Mammalian Cell Encapsulation in Alginate Beads Using a Simple Stirred Vessel

Cell encapsulation in alginate beads has been used for immobilized cell culture in vitro as well as for immunoisolation in vivo. Pancreatic islet encapsulation has been studied extensively as a means to increase islet survival in allogeneic or xenogeneic transplants. Alginate encapsulation is commonly achieved by nozzle extrusion and external gelation. Using this method, cell-containing alginate droplets formed at the tip of nozzles fall into a solution containing divalent cations that cause ionotropic alginate gelation as they diffuse into the droplets. The requirement for droplet formation at the nozzle tip limits the volumetric throughput and alginate concentration that can be achieved. This video describes a scalable emulsification method to encapsulate mammalian cells in 0.5% to 10% alginate with 70% to 90% cell survival. By this alternative method, alginate droplets containing cells and calcium carbonate are emulsified in mineral oil, followed by a decrease in pH leading to internal calcium release and ionotropic alginate gelation. The current method allows the production of alginate beads within 20 min of emulsification. The equipment required for the encapsulation step consists in simple stirred vessels available to most laboratories.

This video and manuscript describe an emulsion-based method to encapsulate mammalian cells in 0.5% to 10% alginate beads which can be produced in large batches using a simple stirred vessel. The encapsulated cells can be cultured in vitro or transplanted for cellular therapy applications.

Encapsulation de cellules de mammifères dans des perles d&#39;alginate à l&#39;aide d&#39;un récipient agité simple

Cell encapsulation in alginate beads has been used for immobilized cell culture in vitro as well as for immunoisolation in vivo. Pancreatic islet ...

Simulating the Mechanics of Lens Accommodation via a Manual Lens Stretcher

The goal of this protocol is to mimic the biomechanics of physiological accommodation in a cost-efficient, practical manner. Accommodation is achieved through the contraction of the ciliary body and relaxation of zonule fibers, which results in the thickening of the lens necessary for near vision. Here, we present a novel, simple method in which accommodation is replicated by tensing the zonules connected to the lens capsule via a manual lens stretcher (MLS). This method monitors the radial stretching achieved by a lens when subjected to a consistent force and allows for a comparison of accommodating lenses, which can be stretched, to non-accommodating lenses, which cannot be stretched. Importantly, the stretcher couples to the zonules directly, and not to the sclera of the eye, thus only requiring the lens, zonules, and ciliary body rather than the entire globe sample. This difference can significantly decrease the cost of acquiring donor cadaver lenses by about 62% compared to acquiring an entire globe.

We present an efficient method of studying lens accommodation by using a manual lens stretcher. The protocol mimics physiological accommodation by pulling the zonules connected around the lens capsule, thereby, stretching the lens.

Simulation de la mécanique de la lentille hébergement via une civière objectif manuel

The goal of this protocol is to mimic the biomechanics of physiological accommodation in a cost-efficient, practical manner. Accommodation is achieved ...

Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial bypass. Characterization of the growth dynamics and remodeling process of degradable polymer scaffold-based tissue-engineered blood vessels (TEBVs) with pulsatile stimulation is crucial for vascular tissue engineering. Optical imaging techniques stand out as powerful tools for monitoring vascularization of engineered tissue enabling high-resolution imaging in real-time culture. This paper demonstrates a nondestructive and fast real-time imaging strategy to monitor the growth and remodeling of TEBVs in long-term culture by using optical coherence tomography (OCT). Geometric morphology is evaluated, including vascular remodeling process, wall thickness, and comparison of TEBV thickness in different culture time points and presence of pulsatile stimulation. Finally, OCT provides practical possibilities for real-time observation of the degradation of polymer in the reconstructing tissues under pulsatile stimulation or not and in each vessel segment, by compared with the assessment of polymer degradation using scanning electron microscopic(SEM) and polarized microscope.

A step by step protocol for nondestructive and long-period monitoring the process of vascular remodeling and scaffold degradation in real-time culture of biodegradable polymeric scaffold-based tissue-engineered blood vessels with pulsatile stimulation using optical coherence tomography is described here.

Contrôles non destructifs suivi du développement de dégradable axée sur l’échafaud tissulaire des vaisseaux sanguins à l’aide de la tomographie à cohérence optique

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial ...

Patterning the Geometry of Human Embryonic Stem Cell Colonies on Compliant Substrates to Control Tissue-Level Mechanics

Human embryonic stem cells demonstrate a unique ability to respond to morphogens in vitro by self-organizing patterns of cell fate specification that correspond to primary germ layer formation during embryogenesis. Thus, these cells represent a powerful tool with which to examine the mechanisms that drive early human development. We have developed a method to culture human embryonic stem cells in confined colonies on compliant substrates that provides control over both the geometry of the colonies and their mechanical environment in order to recapitulate the physical parameters that underlie embryogenesis. The key feature of this method is the ability to generate polyacrylamide hydrogels with defined patterns of extracellular matrix ligand at the surface to promote cell attachment. This is achieved by fabricating stencils with the desired geometric patterns, using these stencils to create patterns of extracellular matrix ligand on glass coverslips, and transferring these patterns to polyacrylamide hydrogels during polymerization. This method is also compatible with traction force microscopy, allowing the user to measure and map the distribution of cell-generated forces within the confined colonies. In combination with standard biochemical assays, these measurements can be used to examine the role mechanical cues play in fate specification and morphogenesis during early human development.

Extracellular matrix ligands can be patterned onto polyacrylamide hydrogels to enable the culture of human embryonic stem cells in confined colonies on compliant substrates. This method can be combined with traction force microscopy and biochemical assays to examine the interplay between tissue geometry, cell-generated forces, and fate specification.

Modelage de la géométrie des colonies de cellules souches embryonnaires humaines sur des substrats conformes pour contrôler la mécanique de niveau tissulaire

Human embryonic stem cells demonstrate a unique ability to respond to morphogens in vitro by self-organizing patterns of cell fate specification that ...