In vivo Characterization of Deformation Fields in Hypertense Amphibian Hearts Using the Speckle Image Photogrammetry Technique

Hypertension, a major cardiovascular disease, is one of the most prevalent and death causing disease worldwide. In the US, over 70 million adults have high blood pressure, that is, 1 in 3. A prolonged state of hypertension cause damages to the brain cells, the kidneys and can lead to a stroke. This research studies the biomechanical response of amphibian hearts in vivo when in hypertensive. A normotensive state is used as the control. Hypertension is induced by injecting a saline fluid directly into the heart ventricle using a syringe. The response of the heart is characterized by monitoring surface deformation fields using the speckle image photogrammetry technique. This is a non-contact optical full field strain technique that uses the ARAMIS photogrammetry software to monitor surface deformations over the entire three dimensional heart over several heart beats. The ARAMIS software is connected to two high speed cameras capable of taking more than 500 frames per second at full field. This technique was successfully used before to characterize myocardial infarction on amphibian hearts. The internal ventricle pressure is measured using a catheter pressure transducer located in the heart. The pressure inside the heart is directly related to the stress on the heart walls which are made of myocardia. During hypertension, the ventricle fluid volume is increased leading to the heart muscles contracting more forcefully. Preliminary results verified this concept with an increase in the deformations of the ventricle from a normotensive state to a hypertensive state. The results obtained showed a displacement increase of 0.18 mm corresponding to 12%. Major strains increased by 6%. The study also investigated whether the ventricle contraction mechanism is a sphere chamber model or a peristaltic tube model. The collected data supports the sphere chamber model. However, more experimentation is required to make these conclusions concrete

Combinatorial nanodot stripe assay to study cell haptotaxis

Haptotaxis, a cellular migration response to surface-bound biochemical cues, is essential for life processes such as angiogenesis, tissue repair, and embryonic development. In vitro, haptotaxis signaling processes are typically studied using protein gradient patterns. However, while these gradients provide global cell distributions for haptotaxis results, they do not easily and clearly provide precise information about the choices that cells make locally on different protein surface densities that exist within the gradient. Herein, we introduce the nanodot stripe assay (NSA) in order to (i) easily examine cell migration choices to different protein surface densities and (ii) to investigate the effect of protein nanodot cluster sizes on cell migration behavior. Each NSA design consists of a pair of alternating nanodot arrays of discrete and non-continuous surface density of either 0, 1, 3, 10, 30, 44 or 100% coverage. The NSA configuration challenges cells with a binary choice of low versus high surface densities of all possible combinations at once. The cell-surface affinity of the reference surface (RS), the area between patterned cues was adjusted towards maximizing cell response to the nanopatterned guidance cue. The RS was backfilled with a mixture of polyethylene glycol (PEG) and poly-D-lysine (PDL) with a low and high cell-surface affinity, respectively, and three RS combinations of 100:0, 90:10 and 75:25 %PEG:%PDL were tested with the NSA. The 90:10 %PEG:%PDL RS resulted in optimal C2C12 myoblasts haptotaxis responses to patterned netrin-1 nanodots. To study the effect of nanodot size, netrin was patterned as 200 nm × 200 nm, 400 nm × 400 nm and 800 nm × 800 nm dots. The migration response was found to be indifferent to the nanodot size. The NSA revealed that the myoblasts preferentially migrated onto higher netrin-1 density stripes when challenged with nanodot stripes with a threefold and greater density difference. Finally, by comparing the cell migration on NSA and stepped gradients, a response consistent with directional persistence could be identified on the stepped gradients. The NSA provides a powerful haptotaxis platform to advance the understanding of contact-mediated migration and signaling of motile cells to surface-bound protein cues.L'haptotaxie, une réponse de migration cellulaire a la biochimie des surfaces, est essentielle pour des processus vitaux tels que l'angiogenèse, la réparation tissulaire et le développement embryonnaire. In vitro, les processus de signalisation de l'haptotaxie sont généralement étudiés en utilisant des motifs de gradients de protéines sur des surfaces. Cependant, alors que ces gradients fournissent des distributions de cellules pour des résultats d'haptotaxie, ils ne fournissent pas facilement et clairement des informations précises sur les choix que font les cellules sujettes à des différences locales de densités de protéines sur la surface. Ici, nous introduisons le test de la bande de nanopoint (BNP) afin de (i) examiner facilement les choix de migration cellulaire à différentes densités de surface protéiques, et (ii) étudier l'effet de la taille des nanopoint de protéines sur la migration cellulaire. Chaque BNP consiste en une paire de matrice de nanopoint discret et alternant de façon discontinue entre des densités de soit 0, 1, 3, 10, 30, 44 ou 100%. La configuration du BNP présente ainsi les cellules avec un choix binaire de densités de surface faibles ou élevées couvrant toutes les combinaisons possibles. Pour étudier l'effet de la dimension des nanopoint de protéines, des nanopoint de 200 nm × 200 nm, 400 nm × 400 nm et 800 nm × 800 nm ont été testés. De plus, pour révéler la migration cellulaire, l'affinité de la surface cellulaire a la SR a été ajustée pour maximiser la réponse cellulaire au guidage du motif de nanopoint. Le SR a été remblayé avec un mélange de polyéthylène glycol (PEG) et de poly-D-lysine (PDL) avec une affinité faible et élevée à la surface des cellules, respectivement. Trois combinaisons de SR ont été testées; 100:0, 90:10 et 75:25 %PEG:%PDL. Le PEG 90%:10% PDL SR a résulté à des réponses optimales de l'haptotaxie des myoblastes C2C12 spécifiques aux nanopoints de netrin-1, et ont été conservés avec toutes les tailles de nanopoints. Les myoblastes ont migré préférentiellement vers les bandes de haute densité de nétrine-1 lorsque présentés avec des bandes de nanopoint avec une différence de densité de triple et plus. Par rapport aux résultats du test par gradient, les choix des cellules présentées au bande du BNP étaient exempts d'effet persistant de la migration cellulaire. Le BNP fournit une plate-forme d'haptotaxie puissante pour améliorer la compréhension de la migration et de la communication cellulaire face à des motifs de proteines sur les surfaces

Non-equilibrium Inertial Separation Array for High-throughput, Large-volume Blood Fractionation

Microfluidic blood processing is used in a range of applications from cancer therapeutics to infectious disease diagnostics. As these applications are being translated to clinical use, processing larger volumes of blood in shorter timescales with high-reliability and robustness is becoming a pressing need. In this work, we report a scaled, label-free cell separation mechanism called non-equilibrium inertial separation array (NISA). The NISA mechanism consists of an array of islands that exert a passive inertial lift force on proximate cells, thus enabling gentler manipulation of the cells without the need of physical contact. As the cells follow their size-based, deterministic path to their equilibrium positions, a preset fraction of the flow is siphoned to separate the smaller cells from the main flow. The NISA device was used to fractionate 400 mL of whole blood in less than 3 hours, and produce an ultrapure buffy coat (96.6% white blood cell yield, 0.0059% red blood cell carryover) by processing whole blood at 3 mL/min, or ∼300 million cells/second. This device presents a feasible alternative for fractionating blood for transfusion, cellular therapy and blood-based diagnostics, and could significantly improve the sensitivity of rare cell isolation devices by increasing the processed whole blood volume

Continuous Flow Microfluidic Bioparticle Concentrator

Innovative microfluidic technology has enabled massively parallelized and extremely efficient biological and clinical assays. Many biological applications developed and executed with traditional bulk processing techniques have been translated and streamlined through microfluidic processing with the notable exception of sample volume reduction or centrifugation, one of the most widely utilized processes in the biological sciences. We utilize the high-speed phenomenon known as inertial focusing combined with hydraulic resistance controlled multiplexed micro-siphoning allowing for the continuous concentration of suspended cells into pre-determined volumes up to more than 400 times smaller than the input with a yield routinely above 95% at a throughput of 240 ml/hour. Highlighted applications are presented for how the technology can be successfully used for live animal imaging studies, in a system to increase the efficient use of small clinical samples, and finally, as a means of macro-to-micro interfacing allowing large samples to be directly coupled to a variety of powerful microfluidic technologies