Bu çalışma Nebraska NASA Uzay Grant ve hibe No 0926880 altındaki Ulusal Bilim Vakfı tarafından kısmen desteklenmiştir.

1. Lateks Gemi hazırlanması <ol><li> Sağlam bir tezgah üzerine sabitlenmiş olan dikenli hortum bağlantıları için lateks geminin iki ucunda Fix. </li><li> Görüş alanını belirlemek için lateks gemide ilgi alanı ölçün. Stent test için ilgi alanı dikenli hortum konnektörleri arasında ortalanmış ve stentli alanı dışında suşları gözlemlemek için stent her tarafında yaklaşık bir inç içermelidir. </li><li> Bir dikenli hortum bağlantısı dış kenarından da lateks geminin yaklaşık merkezi konektörler arasındaki merkezi konumu olan mesafeyi kaydedin. Kateter kadar stentin merkezine ölçerek kateter üzerine mesafe çevirir. Sonra bir işaretleyici ile kateter işaretleyin. </li><li> Dikenli hortum konektörlerinden lateks gemi çıkarın. </li><li> Beyaz ve siyah sprey boya bir stokastik deseni ile ilgi alanı püskürtme veya işaretleyerek lateks geminin hazırlayınkalıcı bir kalem kullanarak rastgele noktalar ile ilgi alanı. Numuneler için daha küçük ve daha ince stokastik kalıp gerekir. </li></ol> Aramis Sistem 2. In vitro Test Sistemi ve Kalibrasyon <ol><li> 1. adımda ölçülen ilgi alanından biraz daha büyük olan kalibrasyon paneli seçin. </li><li> Ilgi alanında dikenli hortum konnektörleri arasındaki kalibrasyon paneli yerleştirin ve ilgi alanı iyi aydınlatılmış olduğundan emin olun. </li><li> Iki kamera, örnek olan uzaklığı ve seçilen kalibrasyon paneli dayalı kamera yükseklik arasındaki mesafeyi ayarlayın. Her kalibrasyon paneli farklıdır, bu nedenle Aramis kullanım kılavuzu bu mesafeleri belirlemek için başvurulan gerekecektir. </li><li> &quot;New Project&quot;, sonra &quot;Dosya&quot; seçerek Aramis yeni bir proje açın. Sonraki &quot;Sensör&quot; sekmesine tıklayın ve sonra, &quot;Tam Kalibrasyon&quot; &quot;Kalibrasyon&quot; seçeneğini seçin. </li><li> Aramis yazılım şimdi t kalibre adımlarında kullanıcıya yürüyeceksin<html> diye kameralar. </li><li> Tamamen açık diyafram ile kamera ayar vidası gevşeterek ve objektif çevirerek kalibrasyon panelindeki kamera odaklanmak. Bir kez odaklı, set vida sıkın ve diyafram kapatın. </li><li> Kalibrasyon sürecinin ilk görüntü atın. Yerinden veya görüntü bilgisayar ekranında odaklanmıştır kadar, bilgisayar üzerinde gösteri göre kalibrasyon paneli döndürün. İkinci görüntü çekmek. Kalibrasyon görüntüleri geri kalanı için bu işlemi tekrarlayın. </li><li> Bir kez kalibrasyon tüm görüntüleri alınır, Aramis görüntü analiz yazılımı, kalibrasyon ayarlarını hesaplayacaktır. Kalibrasyon sapma 0.04 büyükse kalibrasyon işlemi tekrarlanmalıdır. Kamera veya kameralar arasındaki mesafenin odak yapılan tüm ayarlamalar kalibrasyon işlemini geçersiz kılacaktır. </li><li> Kalibrasyon paneli çıkarın ve dikenli hortum konnektörleri üzerine geri boyalı lateks gemi yerleştirin. </li></ol>Aşırı Ortam Gürültüsü kaçının başlığı &quot;&gt; 3. Öntest <ol><li> Test için istenen saniyedeki kare sayısını belirleyin. Saniyede Artan kare daha düzgün gerginlik sonuçlar üretecektir. </li><li> Hiç kırmızı görüntü üzerinde görüntülenecek şekilde az 1 saniye başına kare ve deklanşör hızını ayarlayın. </li><li> 5 fotoğraf çekebilirsiniz. </li><li> Resim serisi noktaları başlatmak ve test hesaplamak ekleyin. </li><li> &quot;Ctrl&quot; tutarken, arka plan gürültü gözlemlemek için örnek merkezinde tıklayın. Öntest gürültü Microstrain 75 üstünde ise kalibrasyon işlemini yeniden yapılması gerekir. </li></ol> 4. Stent Dağıtım <ol><li> Test sırasında çekmek için istenilen görüntülerin miktarını seçin. 200 fotoğraf stent genişlemesi için yeterli olacaktır. </li><li> Yavaş yavaş lateks damar içine kateter sokun ve merkezi bir konuma ulaşana kadar stent yerleştirme kılavuzunu kateter işareti göstergesi kullanarak. </li><li> Aramis görüntüleri alarak başlayın. </li><li> Balon ile genişleyen stent için, yavaş yavaş balon tam genişletilmiş kadar stent genişletmek için balon basıncı artırmak; sonra yavaş yavaş sıfıra balon basıncı azaltmak ve balon söndürülmeli ve kateter ile birlikte geri çekilir. </li><li> Stent tam genişletilmiş kadar kendine genişleyen stent için, yavaş yavaş kılıf kaldırmak; sonra yavaş yavaş kateter geri çekin. </li></ol> 5.. Görüntü Analizi <ol><li> Gemide belli bir noktaya Gerilme geçmişi <ol><li> Ilgi alanı &quot;ctrl&quot; tuşuna basılı tutarak ve tıklayarak bir aşama noktası oluşturun. </li><li> Istenilen gerginlik türünü seçin, X, Y, XY, büyük gerilim, küçük gerilim, ya da Mises gerilme yani süzün. </li><li> Sağ alt köşesinde arsa test süresi boyunca seçilen noktada zorlanma gösterecektir. </li></ol></li><li> Kabın belirli bir yol boyunca mekansal suşu <ol><li> Cr&quot;bölümler&quot; sekmesini tıklayarak çok aşamalı bir nokta çizgi eate, ardından &quot;bölümü oluşturun&quot;. Y, X ekseni için resmin paralel bir çizgi seçin sıfıra eşittir. Bu aşama, bir hat noktaları bir dizi yaratacaktır. </li><li> Kademeli hattı oluşturulduktan sonra alt köşesinde arsa bir arsa üzerinde çizgiler serisinin gösterecektir. Her bir hat bölümüne uzunluğu boyunca zamanlı olarak bir örneği olarak suşu temsil eder. </li></ol></li><li> Geminin genişleme oranını ve yarıçapı analiz etmek için en uygun silindir oluşturma <ol><li> Üst araç çubuğunda, sonra &quot;en uygun cylinder&quot; &quot;ilkel&quot; seçeneğini seçin. </li><li> Doğru araç çubuğunda &quot;yüzey aracılığıyla seçin&quot; aracını kullanarak resmin küçük bir bölümünü seçin. </li><li> Aramis yazılım üç boyutlu en uygun silindir oluşturur. </li><li> Görüntüler daha sonra lateks damarın çapı ne kadar değişen gözlemlemek için yoluyla Siklustaki edilebilir. </li></ol></li><li> Arasındaki mesafe değerlendirilmesiiki nokta <ol><li> &quot;Mesafe noktadan noktaya&quot; üzerinde &quot;analiz&quot; sekmesini tıklayın. </li><li> Iki nokta seçerek analizi için arzu edilen görüntü ile ilgili bir uzunluğa seçmek. </li><li> Görüntüler daha sonra zaman içinde iki nokta arasındaki mesafe değişimi gözlemlemek yoluyla dolaşılan edilebilir. </li></ol></li></ol> 6. Temsilcisi Sonuçlar Stent dikmeler damar duvarı dışa doğru genişletin suşları genellikle stent konum etrafında daha yüksek olacaktır. Şekil 1 belirli bir noktada balon genişleyen stent gibi büyük gerginlik tarihin geri tepme işlemi sırasında gerginlik haritalama bir örnektir. Şekil 1&#39;de görülen siyah noktalar boru üzerinde bu referans noktaları deplasmanları yakalamak ve izlemek için yüksek hızlı kameralar tarafından kullanılan referans noktaları vardır. Referans noktaları kaydedilmiş hareketine bağlı olarak, daha sonra yazılımı kanalın suşları veya bir hesaplamak için kullanılacaktırny diğer hedeflenen nesne. Ayrıca, maksimum asal suşu olarak anılacaktır büyük bir deformasyon, aşağıdaki gibi hesaplanır: <img alt="Denklem 1" src="/files/ftp_upload/3945/3945eq1.jpg" /> Bu implante stent damar yüzeyinde üniform olmayan gerilme dağılımının neden olduğu açıktır. Bu uçlar-kısıtlı lateks boru ve stent kafes yapısı geri tepme yükleme ile açıklanabilir. Bu alanda suşu olarak, Şekil 1 &#39;in altındaki görüntü kırmızı bir çapraz işaretleyici tarafından tanımlanan stent geri tepme, başlangıç ​​aşamasında karşılık gelir. Belirli bir noktaya 10 majör uzama-geçmişi eğrisi stent ayırt edilebilir aşamaları gösterdi. Balon genişletme balonun sönmesi 12 ve 14 saniye arasında gerçekleşir sonra yaklaşık 10 ila 12 saniye ve stent geri tepme meydana gelir. <img alt="Şekil 1" src="/files/ftp_upload/3945/3945fig1.jpg" /> Şekil 1.. </strong&gt; Deneysel kurulumu (üstte); stentli boru yüzeyi (orta) üzerinde üniform olmayan gerilme dağılımı; noktası 10 (altta) de büyük gerilim öyküsü.

<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>

Çıkar çatışması ilan etti.

Stereo optik yüzey gerilim ölçüm sistemi örnek temas etmeden de-ve out-of-düzlemsel hareketler hem de deforme yüzey üzerinde yerel suşların ölçmek için kullanılır. Bu sistem yüzey suşları çözme yeteneği yüksek, her noktasının hareketlerinin doğru ölçümler oluşturmak için yüzeye koyarak rasgele zıt desen fotoğrafını çekmek için iki adet yüksek hızlı optik kameralar kullanır. Bu, istenen zıt doğru ölçüm örüntü sağlamak için yeterli ölçüde y...

<table border="1"><tbody><tr><td> Kullanılan Cihaz </td><td> Şirket </td><td> Katalog numarası </td><td> Yorumlar </td></tr><tr><td> Aramis Kamera Sistemi </td><td> GOM: Optik Ölçüm Teknikleri </td><td></td><td></td></tr><tr><td> Palmaz Genesis transhepatik BİLİYER STENT </td><td> Cordis Şirketi </td><td> PG5910B </td><td> Balon genişleyen stent </td></tr><tr><td> Z-MED Balon Dilatasyon Kateter </td><td> B. Braun Medical Inc </td><td> PDZ336 </td><td> Balon dilatasyon kateteri </td></tr></tbody></table>

monitoring the wall mechanics during stent deployment in a vessel

Klinik çalışmalar, çeşitli stent tasarımlar 1 için farklı restenoz oranları bildirdin. Bu arter duvarına stent bağlı gerilme konsantrasyonu restenoz 2-7 başlatır doku yaralanması, yol speküle edilmektedir. Bu hipotez stent implantasyonu sonrasında arter üzerinde homojen olmayan deformasyon dağılımının daha iyi unsurlarını da dahil olmak üzere ileri araştırmalar gerekir. Stentli arter için temassız yüzey gerilim ölçüm yöntemi bu çalışmada sunulmuştur. Aramis stereo optik yüzey gerilim ölçüm sistemi, her bir referans noktası hareketini yakalamak ve deforme yüzeyi 8,9 üzerinden üç boyutlu suşları gidermek için iki optik yüksek hızlı kamera kullanır. Bir kafes stent kendi dış yüzeyi üzerine püskürtülür ve çekilmiş bir rastgele zıt desenli bir lateks damar içine dağıtılır gibi, yüzey gerilim deformasyon her an kaydedilir. Hesaplanan suşu dağılımlar sonra lo anlamak için kullanılabilirlercal lezyon yanıtı, hesaplama modelleri doğrulamak, ve in vivo çalışmada daha için hipotezler formüle.

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 bağlı arteriyel suş dağılımları optik yüzey gerilim ölçüm sistemi kullanılarak karakterize edilmiştir. Bu görselleştirme tekniği ana gemide stent etkisini içgörü kazanmak için kullanılır.

Bir Gemi Stent Dağıtım sırasında Duvar Mekaniği izlenmesi

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 Görüntüleme.  University of Nebraska-Lincoln. Stent bağlı arteriyel suş dağılımları optik yüzey gerilim ölçüm sistemi kullanılarak karakterize edilmiştir. Bu görselleştirme tekniği ana gemide stent etkisini içgörü kazanmak için kullanılır.

Video: 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).

Doku Değerlendirilmesinde Manyetik Rezonans Elastography Metodoloji Construct Büyüme Tasarlandı

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

Biyomühendislik

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

Kütle Sitometri: Bir CyTOF Kütle Sitometre Daily Tuning ve Koşu Hücre Örnekleri için protokol

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.

Mikro ölçekli Hücre Stimülasyon deneyler için Çok Yönlü Otomatik Platformu

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.

180 ° Kavisli Arter Testi Bölüm Model Tip IV Stent Başarısızlık Mansap İkincil Akış Yapıların Deneysel İncelenmesi

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.

Elektrospinning Süreci Uyarlama bir Tri-katmanlı için üç Benzersiz ortamlar sağlamak için<em&gt; In Vitro</em&gt; Havayolu Wall Modeli

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.

Endotel Hücre yakalama ve Saklanmasına İlişkin ferromanyetik Bare Metal Stent

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.

Alginat Boncuklarda Memeli Hücre Kapsüllenmesinin Basit Bir Karıştırılmış Geminin Kullanımı

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.

Manuel Lens sedye ile Lens konaklama mekaniği simüle

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.

Geri dönüşlü optik koherens tomografi kullanarak parçalanabilir iskele tabanlı doku mühendisliği kan damarı gelişimi izleme

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.

Doku Seviyesi Mekaniğinin Kontrol Altına Almak İçin Uyumlu Yüzeylerde İnsan Embriyonik Kök Hücre Kolonilerinin Geometrisinin Desenlemi

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

Methods Article

Bir Gemi Stent Dağıtım sırasında Duvar Mekaniği izlenmesi