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Results: 1 to 10 of 11

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Substrate stiffness heterogeneities disrupt endothelial barrier integrity in a micropillar model of heterogeneous vascular stiffening.
VanderBurgh JA, Hotchkiss H, Potharazu A, Taufalele PV, Reinhart-King CA
(2018) Integr Biol (Camb) 10: 734-746
MeSH Terms: Adherens Junctions, Animals, Aorta, Atherosclerosis, Cattle, Cell Adhesion, Cell Communication, Cell Movement, Dimethylpolysiloxanes, Endothelial Cells, Endothelium, Vascular, Focal Adhesions, Human Umbilical Vein Endothelial Cells, Humans, Leukocytes, Materials Testing, Neutrophils, Phenotype, Tunica Intima, Vascular Stiffness, Vinculin
Show Abstract · Added April 10, 2019
Intimal stiffening has been linked with increased vascular permeability and leukocyte transmigration, hallmarks of atherosclerosis. However, recent evidence indicates age-related intimal stiffening is not uniform but rather characterized by increased point-to-point heterogeneity in subendothelial matrix stiffness, the impact of which is much less understood. To investigate the impact of spatially heterogeneous matrix rigidity on endothelial monolayer integrity, we develop a micropillar model to introduce closely-spaced, step-changes in substrate rigidity and compare endothelial monolayer phenotype to rigidity-matched, uniformly stiff and compliant substrates. We found equivalent disruption of adherens junctions within monolayers on step-rigidity and uniformly stiff substrates relative to uniformly compliant substrates. Similarly, monolayers cultured on step-rigidity substrates exhibited equivalent percentages of leukocyte transmigration to monolayers on rigidity-matched, uniformly stiff substrates. Adherens junction tension and focal adhesion density, but not size, increased within monolayers on step-rigidity and uniformly stiff substrates compared to more compliant substrates suggesting that elevated tension is disrupting adherens junction integrity. Leukocyte transmigration frequency and time, focal adhesion size, and focal adhesion density did not differ between stiff and compliant sub-regions of step-rigidity substrates. Overall, our results suggest that endothelial monolayers exposed to mechanically heterogeneous substrates adopt the phenotype associated with the stiffer matrix, indicating that spatial heterogeneities in intimal stiffness observed with age could disrupt endothelial barrier integrity and contribute to atherogenesis.
0 Communities
1 Members
0 Resources
21 MeSH Terms
Standard Reticle Slide To Objectively Evaluate Spatial Resolution and Instrument Performance in Imaging Mass Spectrometry.
Zubair F, Prentice BM, Norris JL, Laibinis PE, Caprioli RM
(2016) Anal Chem 88: 7302-11
MeSH Terms: Dimethylpolysiloxanes, Epoxy Compounds, Gentian Violet, Ions, Microscopy, Atomic Force, Polymers, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Show Abstract · Added August 17, 2016
Spatial resolution is a key parameter in imaging mass spectrometry (IMS). Aside from being a primary determinant in overall image quality, spatial resolution has important consequences on the acquisition time of the IMS experiment and the resulting file size. Hardware and software modifications during instrumentation development can dramatically affect the spatial resolution achievable using a given imaging mass spectrometer. As such, an accurate and objective method to determine the working spatial resolution is needed to guide instrument development and ensure quality IMS results. We have used lithographic and self-assembly techniques to fabricate a pattern of crystal violet as a standard reticle slide for assessing spatial resolution in matrix-assisted laser desorption/ionization (MALDI) IMS experiments. The reticle is used to evaluate spatial resolution under user-defined instrumental conditions. Edgespread analysis measures the beam diameter for a Gaussian profile and line scans measure an "effective" spatial resolution that is a convolution of beam optics and sampling frequency. The patterned crystal violet reticle was also used to diagnose issues with IMS instrumentation such as intermittent losses of pixel data.
1 Communities
3 Members
0 Resources
7 MeSH Terms
A microfluidic platform for chemical stimulation and real time analysis of catecholamine secretion from neuroendocrine cells.
Ges IA, Brindley RL, Currie KP, Baudenbacher FJ
(2013) Lab Chip 13: 4663-73
MeSH Terms: Animals, Calcium, Carbachol, Catecholamines, Cattle, Cells, Cultured, Chromaffin Cells, Chromatography, High Pressure Liquid, Dimethylpolysiloxanes, Electrochemical Techniques, Electrodes, Kinetics, Microfluidic Analytical Techniques, Pituitary Adenylate Cyclase-Activating Polypeptide, Platinum, Potassium Chloride, Silicon, Stimulation, Chemical
Show Abstract · Added November 19, 2013
Release of neurotransmitters and hormones by calcium-regulated exocytosis is a fundamental cellular process that is disrupted in a variety of psychiatric, neurological, and endocrine disorders. As such, there is significant interest in targeting neurosecretion for drug and therapeutic development, efforts that will be aided by novel analytical tools and devices that provide mechanistic insight coupled with increased experimental throughput. Here, we report a simple, inexpensive, reusable, microfluidic device designed to analyze catecholamine secretion from small populations of adrenal chromaffin cells in real time, an important neuroendocrine component of the sympathetic nervous system and versatile neurosecretory model. The device is fabricated by replica molding of polydimethylsiloxane (PDMS) using patterned photoresist on silicon wafer as the master. Microfluidic inlet channels lead to an array of U-shaped "cell traps", each capable of immobilizing single or small groups of chromaffin cells. The bottom of the device is a glass slide with patterned thin film platinum electrodes used for electrochemical detection of catecholamines in real time. We demonstrate reliable loading of the device with small populations of chromaffin cells, and perfusion/repetitive stimulation with physiologically relevant secretagogues (carbachol, PACAP, KCl) using the microfluidic network. Evoked catecholamine secretion was reproducible over multiple rounds of stimulation, and graded as expected to different concentrations of secretagogue or removal of extracellular calcium. Overall, we show this microfluidic device can be used to implement complex stimulation paradigms and analyze the amount and kinetics of catecholamine secretion from small populations of neuroendocrine cells in real time.
0 Communities
1 Members
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18 MeSH Terms
Microfabricated scaffold-guided endothelial morphogenesis in three-dimensional culture.
Liu Y, Markov DA, Wikswo JP, McCawley LJ
(2011) Biomed Microdevices 13: 837-46
MeSH Terms: Basement Membrane, Bioreactors, Cell Culture Techniques, Collagen, Dimethylpolysiloxanes, Endothelium, Vascular, Fibroblasts, Humans, Microvessels, Morphogenesis, Reproducibility of Results, Surface Properties, Tissue Scaffolds
Show Abstract · Added March 27, 2014
Morphogenesis is a fundamental process by which new blood vessels are formed during angiogenesis. The ability to control angiogenesis would lead to improvements in tissue engineering constructions; indeed, the study of angiogenesis has numerous clinical applications, for example, in the investigation of metastatic cancer, peripheral and coronary vascular disease, and wound healing. Conventional in vitro organotypic cell culture approaches to these studies are limited primarily by their reliance on microvascular vessel formation through a random process of morphogenesis that lacks the spatial reproducibility and orientation needed for high-throughput drug testing. We have developed a bioreactor system for scaffold-guided tubulogenesis coupled with 3-D organotypic culture to spatially control vessel formation and its orientation. To create microchannels to guide microvessel formation, we fabricated rigid scaffolds using photolithography and light curing epoxy, and soft scaffolds formed by a polydimethylsiloxane (PDMS) stamp directly into collagen. Scaffolds seeded with dermal microvascular endothelial cells were placed between gelled layers of collagen containing dermal fibroblasts within a Transwell filter system and cultured for up to 2 weeks to allow for vessel maturation. Morphological analysis of thin tissue sections following standard histology and immunohistochemical detection of endothelial cells, fibroblasts, and basement membrane confirmed vessel formation along the microchannel walls with either scaffold. This system may also provide a means to explore revascularization within decellularized extracellular matrices, the culture of microvessel networks with controlled geometries, and possibly the spatial guidance of angiogenesis for interfacing with an external microfluidic supply network. As a new tool for guided angiogenesis, our approach introduces new possibilities for identification of anti-angiogenic therapeutics.
1 Communities
2 Members
0 Resources
13 MeSH Terms
A versatile valve-enabled microfluidic cell co-culture platform and demonstration of its applications to neurobiology and cancer biology.
Gao Y, Majumdar D, Jovanovic B, Shaifer C, Lin PC, Zijlstra A, Webb DJ, Li D
(2011) Biomed Microdevices 13: 539-48
MeSH Terms: Animals, Cell Communication, Cell Line, Tumor, Cell Movement, Coculture Techniques, Dimethylpolysiloxanes, Endothelial Cells, Humans, Hydrodynamics, Mice, Microfluidic Analytical Techniques, Neurons, Pressure, Sepharose
Show Abstract · Added June 24, 2013
A versatile microfluidic platform allowing co-culture of multiple cell populations in close proximity with separate control of their microenvironments would be extremely valuable for many biological applications. Here, we report a simple and compact microfluidic platform that has these desirable features and allows for real-time, live-cell imaging of cell-cell interactions. Using a pneumatically/hydraulically controlled poly(dimethylsiloxane) (PDMS) valve barrier, distinct cell types can be cultured in side-by-side microfluidic chambers with their optimum culture media and treated separately without affecting the other cell population. The platform is capable of both two-dimensional and three-dimensional cell co-culture and through variations of the valve barrier design, the platform allows for cell-cell interactions through either direct cell contact or soluble factors alone. The platform has been used to perform dynamic imaging of synapse formation in hippocampal neurons by separate transfection of two groups of neurons with fluorescent pre- and post-synaptic protein markers. In addition, cross-migration of 4T1 tumor cells and endothelial cells has been studied under normoxic and hypoxic conditions, which revealed different migration patterns, suggesting the importance of the microenvironments in cell-cell interactions and biological activities.
0 Communities
2 Members
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14 MeSH Terms
A metering rotary nanopump for microfluidic systems.
Darby SG, Moore MR, Friedlander TA, Schaffer DK, Reiserer RS, Wikswo JP, Seale KT
(2010) Lab Chip 10: 3218-26
MeSH Terms: Automation, Biocompatible Materials, Dimethylpolysiloxanes, Equipment Design, Membranes, Artificial, Microfluidic Analytical Techniques, Nanotechnology, Perfusion, Pressure, Silicon
Show Abstract · Added April 23, 2013
We describe the design, fabrication, and testing of a microfabricated metering rotary nanopump for the purpose of driving fluid flow in microfluidic devices. The miniature peristaltic pump is composed of a set of microfluidic channels wrapped in a helix around a central camshaft in which a non-cylindrical cam rotates. The cam compresses the helical channels to induce peristaltic flow as it is rotated. The polydimethylsiloxane (PDMS) nanopump design is able to produce intermittent delivery or removal of several nanolitres of fluid per revolution as well as consistent continuous flow rates ranging from as low as 15 nL min(-1) to above 1.0 µL min(-1). At back pressures encountered in typical microfluidic devices, the pump acts as a high impedance flow source. The durability, biocompatibility, ease of integration with soft-lithographic fabrication, the use of a simple rotary motor instead of multiple synchronized pneumatic or mechanical actuators, and the absence of power consumption or fluidic conductance in the resting state all contribute to a compact pump with a low cost of fabrication and versatile implementation. This suggests that the pump design may be useful for a wide variety of biological experiments and point of care devices.
1 Communities
3 Members
0 Resources
10 MeSH Terms
Window on a microworld: simple microfluidic systems for studying microbial transport in porous media.
Markov DA, Samson PC, Schaffer DK, Dhummakupt A, Wikswo JP, Shor LM
(2010) J Vis Exp :
MeSH Terms: Dimethylpolysiloxanes, Ecosystem, Environmental Monitoring, Green Fluorescent Proteins, Microbiological Techniques, Microfluidic Analytical Techniques, Vibrio
Show Abstract · Added March 7, 2014
Microbial growth and transport in porous media have important implications for the quality of groundwater and surface water, the recycling of nutrients in the environment, as well as directly for the transmission of pathogens to drinking water supplies. Natural porous media is composed of an intricate physical topology, varied surface chemistries, dynamic gradients of nutrients and electron acceptors, and a patchy distribution of microbes. These features vary substantially over a length scale of microns, making the results of macro-scale investigations of microbial transport difficult to interpret, and the validation of mechanistic models challenging. Here we demonstrate how simple microfluidic devices can be used to visualize microbial interactions with micro-structured habitats, to identify key processes influencing the observed phenomena, and to systematically validate predictive models. Simple, easy-to-use flow cells were constructed out of the transparent, biocompatible and oxygen-permeable material poly(dimethyl siloxane). Standard methods of photolithography were used to make micro-structured masters, and replica molding was used to cast micro-structured flow cells from the masters. The physical design of the flow cell chamber is adaptable to the experimental requirements: microchannels can vary from simple linear connections to complex topologies with feature sizes as small as 2 microm. Our modular EcoChip flow cell array features dozens of identical chambers and flow control by a gravity-driven flow module. We demonstrate that through use of EcoChip devices, physical structures and pressure heads can be held constant or varied systematically while the influence of surface chemistry, fluid properties, or the characteristics of the microbial population is investigated. Through transport experiments using a non-pathogenic, green fluorescent protein-expressing Vibrio bacterial strain, we illustrate the importance of habitat structure, flow conditions, and inoculums size on fundamental transport phenomena, and with real-time particle-scale observations, demonstrate that microfluidics offer a compelling view of a hidden world.
1 Communities
2 Members
0 Resources
7 MeSH Terms
Rapid and inexpensive fabrication of polymeric microfluidic devices via toner transfer masking.
Easley CJ, Benninger RK, Shaver JH, Steven Head W, Piston DW
(2009) Lab Chip 9: 1119-27
MeSH Terms: Animals, Cell Culture Techniques, Cells, Cultured, Cost-Benefit Analysis, Dimethylpolysiloxanes, Equipment Design, Islets of Langerhans, Mice, Microfluidic Analytical Techniques, Printing, Time Factors
Show Abstract · Added December 6, 2012
An alternative fabrication method is presented for production of masters for single- or multi-layer polymeric microfluidic devices in a standard laboratory environment, precluding the need for a cleanroom. This toner transfer masking (TTM) method utilizes an office laser printer to generate a toner pattern which is thermally transferred to a metal master to serve as a mask for etching. With master fabrication times as little as one hour (depending on channel depth) using commercially-available equipment and supplies, this approach should make microfluidic technology more widely accessible to the non-expert-even the non-scientist. The cost of fabrication consumables was estimated to be < $1 per master, over an order of magnitude decrease in consumable costs compared to standard photolithography. In addition, the use of chemical etching allows accurate control over the height of raised features (i.e., channel depths), allowing the flexibility to fabricate multiple depths on a single master with little added time. Resultant devices are shown capable of pneumatic valving, three-dimensional channel formation (using layer-connecting vias), droplet fluidics, and cell imaging and staining. The multiple-depth capabilities of the method are proven useful for cellular analysis by fabrication of handheld, disposable devices used for trapping and imaging of live murine pancreatic islets. The precise fluidic control provided by the microfluidic platform allows subsequent fixing and staining of these cells without significant movement, thus spatial correlation of imaging and staining is attainable-even with rare alpha cells that constitute only approximately 10% of the islet cells.
0 Communities
1 Members
0 Resources
11 MeSH Terms
Simultaneous orientation and cellular force measurements in adult cardiac myocytes using three-dimensional polymeric microstructures.
Zhao Y, Lim CC, Sawyer DB, Liao R, Zhang X
(2007) Cell Motil Cytoskeleton 64: 718-25
MeSH Terms: Animals, Cell Culture Techniques, Cells, Cultured, Dimethylpolysiloxanes, Male, Muscle Contraction, Myocytes, Cardiac, Nylons, Rats, Rats, Wistar
Show Abstract · Added May 28, 2014
A number of techniques have been developed to monitor contractile function in isolated cardiac myocytes. While invaluable observations have been gained from these methodologies in understanding the contractile processes of the heart, they are invariably limited by their in vitro conditions. The present challenge is to develop innovative assays to mimic the in vivo milieu so as to allow a more physiological assessment of cardiac myocyte contractile forces. Here we demonstrate the use of a silicone elastomer, poly(dimethylsiloxane) (PDMS), to simultaneously orient adult cardiac myocytes in primary culture and measure the cellular forces in a three-dimensional substrate. The realignment of adult cardiac myocytes in long-term culture (7 days) was achieved due to directional reassembly of the myofibrils along the parallel polymeric sidewalls. The cellular mechanical forces were recorded in situ by observing the deformation of the micropillars embedded in the substrate. By coupling the cellular mechanical force measurements with on-chip cell orientation, this novel assay is expected to provide a means of a more physiological assessment of single cardiac myocyte contractile function and may facilitate the future development of in vitro assembled functional cardiac tissue.
(c) 2007 Wiley-Liss, Inc.
0 Communities
1 Members
0 Resources
10 MeSH Terms
Seeding of corneal wounds by epithelial cell transfer from micropatterned PDMS contact lenses.
Pino CJ, Haselton FR, Chang MS
(2005) Cell Transplant 14: 565-71
MeSH Terms: Cell Movement, Cells, Cultured, Contact Lenses, Cornea, Corneal Injuries, Dimethylpolysiloxanes, Epithelium, Corneal, Humans, Male, Middle Aged, Silicones, Time Factors, Wound Healing
Show Abstract · Added May 27, 2014
Persistent corneal wounds result from numerous eye disorders, and to date, available treatments often fail to accelerate reepithelialization, the key initial step in wound healing. To speed reepithelialization, we explored a cell-transfer transplant method utilizing polydimethylsiloxane (PDMS) contact lenses to deliver epithelial cells derived from limbal explants directly within a corneal wound. Human primary epithelial cells and an immortalized corneal epithelial cell line (HCE-SV40) grew well on PDMS contact lenses and their morphology and growth rates where similar to cells grown on tissue culture polystyrene. To initially study cell transfer from PDMS, HCE-SV40 cells were seeded onto PDMS with or without micropatterned posts. After a day in culture, HCE-SV40 cells attached to the unpatterned PDMS uniformly, whereas on micropatterned PDMS they appeared to attach primarily between posts. The cell-covered PDMS contacts were then placed cell-side down onto tissue culture plastic and, after 1, 2, or 3 days, the PDMS contact was removed and the transferred cells were trypsinized and counted. Micropatterned PDMS contact lenses with 100-microm-diameter posts and a post height of 40 microm transferred three times as many cells as unpatterned PDMS. Cell transfer to a wounded cornea was tested in a pig cornea organ culture model de-epithelialized by alkali treatment. Post micropatterned PDMS contact lenses were seeded with labeled HCE-SV40 cells at a density 50,000 cells/cm2 and applied to the wounded pig corneas. After 24, 48, or 96 h of application, PDMS contact lenses were removed, corneas fixed with formaldehyde, and sectioned. After 48 h, epithelial cells transferred from post micropatterned contact lenses to provide 35% epithelial coverage of denuded pig corneas; after 96 h coverage was 65%. We conclude that cell transfer from epithelial-coated PDMS contact lenses micropatterned with posts provides a promising approach to reepithelialize corneal surfaces.
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13 MeSH Terms