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Principal Investigator: DAVID W SCHMIDTKE
Organization: UNIVERSITY OF TEXAS DALLAS
Fiscal Year: 2024
Award: $40,290
Funding agency: National Eye Institute
Abstract
Corneal scarring is a leading cause of blindness worldwide. Through their interactions with the extracellular
matrix (ECM), stromal keratocytes play a central role in fundamental biological processes such as developmental
morphogenesis and wound healing. Recent studies suggest that feedback from the ECM can have a significant
impact stromal cell behavior. Specifically, ECM stiffness, topography and protein composition have all been
shown to modulate keratocyte differentiation, contractility and patterning. However, there are still significant gaps
in our knowledge of how multiple stimuli are integrated to produce specific cell phenotypes. The parent R01
project focuses on the development and testing of novel 2D and 3D experimental platforms which allow us to
systematically incorporate combinations of both soluble and non-soluble cues. In particular, in Aim 1 we are
determining how substrate elasticity modulates the keratocyte response to soluble biochemical cues; in Aim 2
we are determining how substrate topography and composition modulates the keratocyte response to
biochemical and biomechanical cues; and in Aim 3 we are determining how substrate dimensionality (2-D versus
3-D) modulates the keratocyte response to biochemical, biomechanical and topographic cues.
In this proposal we propose the acquisition of a Zeiss microscope stage CO2 incubator system that will be
installed on a motorized Zeiss Axio Observer Z1 inverted microscope. This upgrade will allow us to perform live-
cell analysis, time lapse imaging, and provide us with multi-modal information. Based on experimental data
collected over the first 4 years of the project, our needs for live-cell and time lapse imaging have expanded
dramatically. This instrumentation will allow us quantitatively (i) determine and measure dynamic cell morphology
and transient cell activation information with improved temporal resolution; (ii) characterize the impact of collagen
fibril alignment and topography on keratocyte migration in response to novel wound injury models we have
developed; (iii) allow us to identify the relative roles of proliferation and migration in response to secreted and/or
immobilized growth factors and extracellular matrix proteins; and (iv) allow us to determine the effects of 3D
confinement on keratocyte migration. The acquisition of this live-cell imaging system is expected to provide
unprecedented insights into the role of key biophysical signaling pathways on the differentiation of keratocytes
(e.g. quiescent, migratory, regenerative and repair phenotypes). Knowledge obtained using these novel
constructs should also aid in the development of targeted anti-fibrosis therapies, as well as guide tissue
engineering approaches to developing stromal tissue replacements.
Terms: <3-D><3-Dimensional><3D><Biochemical><Biological Function><Biological Process><Biomechanics><Biophysics><Blindness><CO2><Carbon Dioxide><Carbonic Anhydride><Cell Body><Cell-Extracellular Matrix><Cells><Cellular Morphology><Collagen Fibril><Cornea><Cues><Data><Development><Dimensions><ECM><Elasticity><Extracellular Matrix><Extracellular Matrix Proteins><Feedback><Fibrosis><Funding><Growth Agents><Growth Factor><Growth Substances><Image><Immobilization><Incubators><Knowledge><Measures><Microscope><Modeling><Morphogenesis><Motor><Parents><Pattern><Phenotype><Play><Proliferating><Proteins><Proteins Growth Factors><Regulation><Role><Signal Pathway><Stimulus><Stromal Cells><System><Testing><Time><Tissue Engineering><Wound Repair><bioengineered tissue><biomechanical><biophysical foundation><biophysical principles><biophysical sciences><cell behavior><cell morphology><cellular behavior><cicatrix corneal><corneal><corneal fibrosis><corneal scar><developmental><engineered tissue><imaging><imaging system><improved><insight><instrumentation><live cell image><live cell imaging><live cellular image><live cellular imaging><migration><morphogenetic process><multi-modality><multimodality><novel><orthopedic freezing><parent><regenerative><repair><repaired><replacement tissue><response><social role><temporal measurement><temporal resolution><three dimensional><time measurement><vision loss><visual loss><wound healing><wound injury><wound recovery><wound resolution>