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Sally A. Meiners, Ph.D. Office: RWJMS/Piscataway R527 I |
| Research Description:
The FnA-D Region of Tenascin-C and Neuronal Growth A major focus of my laboratory has been to understand how axonal growth and regeneration are modulated by interactions with extracellular matrix molecules, in particular, tenascin-C. We have found that the alternately spliced region of tenascin-C, called fnA-D, when used by itself, increases neurite outgrowth in culture. FnA-D also provides directional cues to growing neurites, a function defined as neurite guidance. Neurites demonstrate a strong preference for growth surfaces coated with fnA-D when given a choice at an interface. FnA-D even influences extension onto surfaces coated with normally repulsive chondroitin sulfate proteoglycans, a major type of inhibitory molecule found in the glial scar. This suggests that fnA-D or motifs within fnA-D could be incorporated into a therapeutic cocktail to facilitate axonal regeneration in the injured CNS. We have localized the outgrowth promoting activity to the amino acid sequence VFDNFVLK and the guidance activity to the amino acid sequence DINPYGFTVSWMASE, both found within the fnA-D region of tenascin-C. We identified the a7ß1 integrin as the receptor for the outgrowth promoting activity and are continuing to work to identify the receptor that mediates the guidance activity. Our observations have lead us to hypothesize that neurite outgrowth and guidance mediated by fnA-D are distinct processes, each of which can be manipulated independently to encourage directed neuronal regrowth. Engineering Nanofibrillar Surfaces for Spinal Cord Repair The goal of this work is to test the effects of VFDNFVLK and DINPYGFTVSWMASE, in conjunction with a scaffold made of electospun nanofibers, in an animal model of neuronal regeneration. The nanofibers are composed of a slowly biodegradable form of polyamide and mimic the porosity and three dimensional geometry of the extracellular matrix on which neurons grow in vivo. We have developed techniques to covalently modify the nanofibers with VFDNFVLK and DINPYGFTVSWMASE. The peptide-modified nanofibers will be surgically implanted at the site of a spinal cord injury in a rat to provide a biomimetic surface, structurally reminiscent of the extracellular matrix, for neuronal attachment and growth, as well as a bridge for axonal elongation across the glial scar. In early experiments, the unmodified nanofibers permitted some axonal regrowth, even in the absence of peptides, and reduced glial scarring. They also promoted activation of Rac, a signaling molecule involved in axon extension. Our hypothesis is that the peptide/nanofiber combination will be an even more favorable substrate than nanofibers alone for axonal regrowth in vivo, resulting in longer or more regenerating axons, earlier regeneration, or improved functional recovery. Engineering Nanofibrillar Surfaces for Cell Culture Research focused on deciphering the biochemical mechanisms that regulate
cell function has largely depended on the use of tissue culture methods
in which cells are grown on two dimensional plastic or glass surfaces.
However, the flat surface of the tissue culture plate is a poor topological
approximation of the more complex three dimensional architecture of
the extracellular matrix and the basement membrane, a structurally compact
form of the extracellular matrix. The three dimensional nanostructures
formed by the nanofibrils of the extracellular matrix/basement membrane
are essential for the reproduction of physiological patterns of cell
adherence, cytoskeletal organization, migration, signal transduction,
morphogenesis, proliferation, and differentiation in cell culture. The
goal of this work is to develop and test a series of completely synthetic
three dimensional nanofibrillar growth surfaces that structurally resemble
the extracellular matrix/basement membrane. Such chemical and physical
parameters as matrix compliancy and surface chemistry (covalent attachment
of specific peptide recognition motifs derived from extracellular matrix
proteins) will be modified to evaluate their role in the promoting specific
neuronal, astrocytic, fibroblastic, and stem cell functions. We will
also evaluate the causal relationship between changes in cellular function
and morphology with the activation of the Rho family of small GTPases,
Rho, Rac, and Cdc42, the mediators and propagators of ECM-cytoskeleton
signaling. Our overall aim will be to develop a number of structurally
defined, completely synthetic three dimensional nanofibrillar surfaces
that can be utilized for more physiologically relevant studies of cell/tissue
function in culture and also to provide matrices that are optimized
for specific applications in regenerative medicine, such as wound healing.
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