Thesis (Ph. D. in Applied Biosciences)--Massachusetts Institute of Technology, Biological Engineering Division, 2004. / Includes bibliographical references (leaves 119-130). / Tissue inhibitors of metalloproteinases (TIMPs) regulate tumor growth, progression and angiogenesis in a variety of experimental cancer models and in human malignancies. However, numerous studies have revealed important differences between TIMP family members in their ability to inhibit angiogenic processes in vitro and angiogenesis in vivo despite their universal ability to inhibit matrix metalloproteinase (MMP) activity. To address these differences, structure-function studies were conducted to identify and characterize the anti-angiogenic domains of TIMP-2, the endogenous MMP inhibitor that uniquely inhibits capillary endothelial cell (EC) proliferation and angiogenesis in vivo. Only the carboxy-terminal domain of TIMP-2 (T2C) and not the MMP-inhibitory N-terminal domain (T2N), inhibited capillary EC proliferation. Although both T2N and T2C inhibited embryonic angiogenesis, only T2C potently inhibited mitogen-stimulated angiogenesis. These findings demonstrate that TIMP-2 possesses two distinct types of anti-angiogenic activities which can be uncoupled from each other. The anti-proliferative activity of T2C was further mapped to the 24-amino acid peptide, Loop 6, which proved to be a potent inhibitor of both embryonic and nitogen-stimulated angiogenesis in vivo. Initial studies into the mechanism(s) by which Loop 6 inhibits angiogenesis revealed that the anti-proliferative effects of Loop 6 are due, at least in part, to the inhibition of cell cycle progression and not to the induction of apoptosis. This inhibition was associated with increased levels of cell cycle inhibitor p27. Although Loop 6 did not compete with bFGF for binding to its receptor, / (cont.) five potential cell surface complexes were observed in crosslinking studies of capillary EC treated with ¹²⁵I-labeled T2C or Loop 6. Finally, given the high degree of homology between TIMP-2 and TIMP-4, we hypothesized that TIMP-4 might share anti-proliferative and MMP inhibition- independent anti-angiogenic activities with TIMP-2. Our results demonstrate that although TIMP-4 inhibits capillary EC migration, it does not inhibit capillary EC proliferation. Furthermore, TIMP-4 did not result in significant inhibition of embryonic angiogenesis in the CAM. These results suggest that TIMP-2 is unique among TIMP family members in its ability to inhibit angiogenesis via two distinct pathways. One of these activities, housed within Loop 6, results in the potent inhibition of angiogenesis in vivo. / by Cecilia A. Fernández. / Ph.D.in Applied Biosciences
| Identifer | oai:union.ndltd.org:MIT/oai:dspace.mit.edu:1721.1/31110 |
| Date | January 2004 |
| Creators | Fernández, Cecilia A., 1969- |
| Contributors | Robert S. Langer and Marsha A. Moses., Massachusetts Institute of Technology. Biological Engineering Division., Massachusetts Institute of Technology. Biological Engineering Division. |
| Publisher | Massachusetts Institute of Technology |
| Source Sets | M.I.T. Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 130 leaves, 8163231 bytes, 8180083 bytes, application/pdf, application/pdf, application/pdf |
| Rights | MIT theses are protected by copyright. They may be viewed, downloaded, or printed from this source but further reproduction or distribution in any format is prohibited without written permission., http://dspace.mit.edu/handle/1721.1/7582 |
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