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Y. M. Khalifa, R. Thakar, L. Peng, N. Acharya, T. Desai; Microtopography Enhances Keratocyte Phenotype for Therapeutic Applications. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4534.
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To examine a set of three-dimensional topographical constructs with features on the microscale to determine the optimal topographical feature and configuration that evokes organized collagen and extracellular matrix synthesis.
1. Precisely defined poly-(dimethylsiloxane) (PDMS) arrays of micropegs and microchannels on the order of ≤ 30 microns were fabricated. 2. Scaffold topography and morphology were characterized by light microscopy and scanning electron microscopy (SEM). 3. Bovine keratocyte morphology and proliferation was characterized with F-actin immunostaining and EdU (5-ethynyl-2’-deoxyuridine) cell cycle assay. 4. Bovine keratocyte phenotype was characterized via immunostaining for the extracellular matrix (ECM) proteins collagens I, V, VI, laminin, elastin, fibronectin, and proteoglycans.
Circularity and Orientation: Microchannel topography results in a circularity that is more linear, and keratocytes orient parallel to the microchannels. Wider channels yield a higher keratocyte circularity and a more disorganized array of keratocytes. Micropegs result in a more stretched morphology of the keratocyte but little control on the orientation of the cell. Proliferation: Microtopographies, especially more narrow microchannels, cause proliferation to decrease. ECM production: The production of ECM proteins varies in quantity and type based on microtopographical feature and dimension.
To aid in the development of a more optically clear and mechanically strong tissue engineered corneal stroma, we have developed microfabrication techniques to control the structural niches of cells. Our recent findings demonstrate that shallow, microtextured substrata in the form of microchannels and micropegs with dimensions ≤ 25 microns resulted in control over cell circularity, orientation, proliferation, and ECM production. This data validates the notion that a microtopography-based system can be used to successfully direct keratocyte orientation, proliferation, and ECM production.
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