The primary objective of this proposal is to gain the necessary training to develop into an independent investigator, integrating my background in the vision sciences with the fundamentals of molecular biology. This objective will be met by a combination of didactic and practical laboratory training under the guidance of mentors Dr. James V. Jester, Dr. H.Dwight Cavanagh, and Dr. Jerry W. Shay. The second objective of this proposal focuses on acquiring the knowledge and molecular techniques required to answer the hypothesis that: nuclear localization of the pro-survival oncogene, BCL-2, is responsible for regulating the final stages of corneal epithelial differentiation and apoptosis. Surface cell apoptosis is a critical component to the maintenance of corneal epithelial homeostasis and is thus a prerequisite for clear vision. In the normal human, the anti-apoptotic protein, BCL-2, localizes to the nucleus and disappears prior to surface cell apoptosis, suggesting a role for BCL-2 in mediating surface cell shedding. Telomerized human corneal epithelial (hTCEpi) cells have also been shown in culture to exhibit disappearance of nuclear BCL-2 prior to apoptosis, thus providing a novel model for studying the mechanism(s) responsible for regulating epithelial desquamation in vitro. In this proposal, this model will be used to examine four specific aims: 1) That an hTCEpi construct recapitulates normal human corneal epithelial cell differentiation and shedding regarding expression, localization, and disappearance of BCL-2 prior to TUNEL labeling. 2) That enhanced expression of BCL-2 results in increased cell survival, persistence of nuclear BCL-2 localization, and decreased surface cell shedding, whereas reduced expression of BCL-2 results in decreased cell survival, loss of nuclear BCL-2 localization, and increased surface cell shedding. 3) That the regulatory loop of BCL-2 and the serine 70 phosphorylation site are critical to regulating surface cell shedding. 4) That events leading to increased apoptosis such as UV light signal surface cell apoptosis through the phosphorylation of BCL-2.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08EY015713-03
Application #
7118968
Study Section
Special Emphasis Panel (ZEY1-VSN (09))
Program Officer
Shen, Grace L
Project Start
2004-09-01
Project End
2009-08-31
Budget Start
2006-09-01
Budget End
2007-08-31
Support Year
3
Fiscal Year
2006
Total Cost
$93,658
Indirect Cost
Name
University of Texas Sw Medical Center Dallas
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
800771545
City
Dallas
State
TX
Country
United States
Zip Code
75390
Robertson, Danielle M; Petroll, W Matthew; Cavanagh, H Dwight (2008) The effect of nonpreserved care solutions on 12 months of daily and extended silicone hydrogel contact lens wear. Invest Ophthalmol Vis Sci 49:7-15
Robertson, Danielle M; Cavanagh, H Dwight (2008) The Clinical and Cellular Basis of Contact Lens-related Corneal Infections: A Review. Clin Ophthalmol 2:907-917
Robertson, Danielle M; Ho, Su-Inn; Cavanagh, H Dwight (2008) Characterization of DeltaNp63 isoforms in normal cornea and telomerase-immortalized human corneal epithelial cells. Exp Eye Res 86:576-85
Robertson, Danielle M; Ho, Su-Inn; Hansen, Baranda S et al. (2007) Insulin-like Growth Factor Binding Protein-3 expression in the human corneal epithelium. Exp Eye Res 85:492-501
Robertson, Danielle M; McCulley, James P; Cavanagh, H Dwight (2007) Severe acanthamoeba keratitis after overnight orthokeratology. Eye Contact Lens 33:121-3
Robertson, Danielle M; Petroll, W Matthew; Jester, James V et al. (2007) The role of contact lens type, oxygen transmission, and care-related solutions in mediating epithelial homeostasis and pseudomonas binding to corneal cells: an overview. Eye Contact Lens 33:394-8;discussion 399-400
Robertson, Danielle M; Petroll, W Matthew; Jester, James V et al. (2007) Current concepts: contact lens related Pseudomonas keratitis. Cont Lens Anterior Eye 30:94-107