The cornea serves as a barrier for the eye and its transparency is required for its function to refract and transmit light to the retina. The normal cornea is avascular. Examples of conditions associated with corneal angiogenesis include chemical burns, chronic contact lens hypoxia, Stevens-Johnson syndrome, pterygia, graft rejection and infectious keratitis (2-4). The normal cornea synthesizes a number of anti-angiogenic molecules, one of which is maspin, a non-inhibitory Serpin that regulates adhesion, migration and invasion of corneal fibroblasts as well as other cell types, including carcinoma cells. Maspin also inhibits in-growth of vessels in the corneal micropocket model and inhibits tube formation, proliferation and migration of microvascular endothelial cells. The overall goal of this project is to understand the mechanism of maspin inhibition of corneal angiogenesis and to develop forms of maspin for the prevention and treatment of angiogenesis. The Specific Objectives are: 1) To test the hypothesis that maspin alters heme and lymph-angiogenesis by multiple mechanisms involving both direct effects on vascular and lymph endothelial cells and indirect effects on corneal cells. The ability of maspin to alter lymph-angiogenesis will be determined and compared with its activity against heme-angiogenesis. The effect of maspin on the levels of pro-and anti-angiogenic proteins synthesized by corneal epithelial and stromal fibroblast cells and heme- and lymph-endothelial cells will be determined. 2) To test the hypothesis that the phosphorylated form of the N-terminal and/or C-terminal region of maspin inhibits angiogenesis, with peptides of these regions more potent than the intact molecule, and to develop maspin peptides that have potential as anti-angiogenic drugs. Identification of the region(s) and post-translational modifications of maspin required for its anti-angiogenic properties will utilize chimeric proteins constructed from maspin and ovalbumin, phosphorylation mimics, phosphorylation null forms and maspin peptides. An organ culture model of angiogenesis will be developed to test the maspin forms. 3) To test the hypothesis that maspin plays a major in vivo regulatory role of heme- and lymph- angiogenesis in the cornea. The effect of loss of one maspin allele on heme- and lymph-angiogenesis will be tested using maspin mice. Maspin reconstitution and depletion experiments with antibodies will be used. The proposed studies focus on understanding heme- and lymph-angiogenesis, a major public health issue that threatens vision, and aims to generate new treatment methods. The studies will also extend our understanding of factors that control corneal angiogenesis through identification of the mechanisms by which the anti-angiogenic molecule maspin alters angiogenesis and will characterize the properties of maspin required for this activity. The results will guide the generation and initial testing of maspin mutants and peptides as potential modalities for the treatment of neovascular diseases.

Public Health Relevance

A critical feature of the cornea is its transparency, which is required for the passage and focusing of light onto the retina for normal vision. Injury and disease can cause the inappropriate formation of blood and lymph vessels in the cornea that can block light transmission. Maspin is a protein that inhibits vessel growth in the cornea. Studies will be conducted towards developing this protein as a therapeutic for the prevention of corneal vascularization.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY021152-03
Application #
8500301
Study Section
Anterior Eye Disease Study Section (AED)
Program Officer
Mckie, George Ann
Project Start
2011-08-01
Project End
2015-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
3
Fiscal Year
2013
Total Cost
$363,375
Indirect Cost
$125,875
Name
Medical College of Wisconsin
Department
Biochemistry
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
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Barnes, Jarrod; Warejcka, Debra; Simpliciano, Jennifer et al. (2012) Latency-associated peptide of transforming growth factor-?1 is not subject to physiological mannose phosphorylation. J Biol Chem 287:7526-34
Warejcka, Debra J; Narayan, Malathi; Twining, Sally S (2011) Maspin increases extracellular plasminogen activator activity associated with corneal fibroblasts and myofibroblasts. Exp Eye Res 93:618-27