Browsing by Subject "Schwann cell"
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Item Applications of micro-3D printing to microfluidic cell dosing(2014-08) Robinson, Michael Mayes; Shear, Jason B.Cellular growth, development, differentiation, and death are mediated to some degree by the interaction of soluble factors with plasma membrane receptors. Traditionally the cellular response to chemical cues has been studied by exposing entire culture dishes to a desired reagent. While the addition of soluble reagents homogenously to cell culture dishes provides a basis for understanding much of cell biology, greater spatial resolution of reagent delivery is necessary in order to elucidate mechanisms on the subcellular scale. This dissertation explores techniques that may improve the quality and precision of delivering soluble factors to cultured cells in order to better understand the complex processes of cell biology. These advancements were made possible by applying high intensity, focused laser light to soluble materials to achieve microscopic three-dimensional (µ-3D) printing. In combination with a previously developed microfluidic cell dosing platform, microstructures were designed and µ-3D printed to hydrodynamically focus reagent streams for cell dosing. Structures were also µ-3D printed within micrometers of living cells from a solution of gelatin and bovine serum albumin with minimal cytotoxicity. When µ-3D printed, these proteins displayed both temperature and pH-responsive properties. In order to allow for on-the-fly control of reagent stream size and temporal pulse width, microstructures were µ-3D printed from temperature-responsive N- isoproplyacrylamide. To further improve the temporal resolution of the system, a technique for cycling between reagents with millisecond exchange times using laminar flow microfluidics was developed. The utility of these techniques was demonstrated by staining rat Schwann cells and mouse neuroblastoma rat glioma hybrid cells (NG108-15) with focused streams of fluorescent dyes. These advancements may allow future experiments to determine the placement of soluble factors necessary for bacterial quorum sensing or stem cell differentiation.Item Controlling neural cell behavior with electric field stimulation across a conductive substrate(2012-12) Nguyen, Hieu Trung 1980-; Schmidt, Christine E.Electrical stimulation of tissues induces cell alignment, directed migration, extended processes, differentiation, and proliferation, but the mechanisms involved remain largely unknown. To reveal effects of electric fields (EF) through the media on cell behavior, voltage (7.45 – 22 V), current density (36 – 106 mA/cm2), duration (2 – 24 hrs), and alternating currents (AC, 2 – 1000 Hz) were varied independently when exposed to cell cultures. It was determined that current density and duration are the primary attribute Schwann cells respond to when an EF is applied through the media. This implies that the number of charges moving across the cell surface may play a key role in EF-induced changes in cell behavior. Identical conditions were used to stimulate cells grown on the surface of a conductive substrate to examine if a scaffold can provide structural and EF cues. The effects of an EF through the substrate were examined by placing a protein gel on the surface during stimulation and observing the morphology of subsequent cell cultures and the physical topology of the gel. EFs were shown to create Ca2+ redistribution across gels and subtle changes in collagen I fibril banding. Stimulated gels were able to induce perpendicular Schwann cell alignment on newly seeded cultures days after initial EF exposure, and the cell response decreased when seeded at longer times, indicating the effects of EF on the matrix environment has a relaxation time. These findings were then integrated into a biodegradable, electrically conductive polypyrrole-poly-ε-caprolactone polymer developed by collaborators. Dorsal root ganglia placed in matrix gels on top of conducting polymer exhibited significantly longer axons when stimulated with DC and AC signals. The overall results demonstrate that EFs have a significant effect on the extracellular environment. The broad implication of this data grants researchers with the ability to physically and metabolically control cell behavior with EFs, including improved wound healing or reduced cancer metastasis.