We have described a protein from human placenta which supports growth of keratinocytes in culture (O'Keefe and Payne, 1985). This material has now been partially purified and partially characterized. It is precipitated by ammonium sulfate, destroyed by heat, acid, or trypsin, does not compete for binding with EGF, and migrates on gel filtration with a Mr of 41,000. The partially purified material also stimulates thymocyte growth but is not IL- 1, and it is uncertain whether the same factor stimulates both keratinocytes and thymocytes. In order to continue to purify and characterize this factor, we will test new purification methods and try to obtain enough purified material to produce antibody or to obtain a partial amino acid sequence which can be used to synthesize a polypeptide which can be used for immunization. The antibody will be used to further purify the material by western blotting. We will determine whether the more purified material continues to stimulate thymocytes or is separable from the IL-l-like activity. We will continue to validate a rapid 3H-thymidine assay that has facilitated our efforts at purification for use in assessing promotion of proliferation growth in cultured keratinocytes. We will characterize the biologic activities of the purified material by testing its ability to phosphorylate target cell membranes, to stimulate protein kinase C, to stimulate polyphosphoinositide metabolism, to alter intracellular calcium ion concentrations, and to interact with matrix components in enhancing keratinocyte growth. We will study interaction of the radioactively labelled factor with target cell including keratinocytes and other cell types and begin to study the receptor for the factor. Finally, we will use a radioimmunoassay for the factor to measure its concentration in plasma and tissues and to determine the cell of origin of the factor. These studies will elucidate the structure and function of this unique growth factor for keratinocytes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
2R01AR025871-09
Application #
3155356
Study Section
General Medicine A Subcommittee 2 (GMA)
Project Start
1979-07-01
Project End
1990-06-30
Budget Start
1987-07-01
Budget End
1988-06-30
Support Year
9
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Type
Schools of Medicine
DUNS #
078861598
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Hu, P; O'Keefe, E J; Rubenstein, D S (2001) Tyrosine phosphorylation of human keratinocyte beta-catenin and plakoglobin reversibly regulates their binding to E-cadherin and alpha-catenin. J Invest Dermatol 117:1059-67
O'Keefe, E J; Hamilton, E H; Lee, S C et al. (1993) Trichohyalin: a structural protein of hair, tongue, nail, and epidermis. J Invest Dermatol 101:65S-71S
Chiu, M L; Jones, J C; O'Keefe, E J (1992) Restricted tissue distribution of a 37-kD possible adherens junction protein. J Cell Biol 119:1689-700
Petersen, M J; Woodley, D T; Stricklin, G P et al. (1992) Synthesis and regulation of keratinocyte collagenase. Matrix Suppl 1:192-7
Hamilton, E H; Sealock, R; Wallace, N R et al. (1992) Trichohyalin: purification from porcine tongue epithelium and characterization of the native protein. J Invest Dermatol 98:881-9
Feldman, S R; O'Keefe, E J (1991) Isolation of a partial clone of desmoplakin-1 by antibody screening of a lambda gt11 library. Am J Med Sci 301:151-6
Hamilton, E H; Payne Jr, R E; O'Keefe, E J (1991) Trichohyalin: presence in the granular layer and stratum corneum of normal human epidermis. J Invest Dermatol 96:666-72
O'Keefe, E; Payne Jr, R E (1991) Minoxidil: inhibition of proliferation of keratinocytes in vitro. J Invest Dermatol 97:534-6
Woodley, D T; Briggaman, R A; Herzog, S R et al. (1990) Characterization of ""neo-dermis"" formation beneath cultured human epidermal autografts transplanted on muscle fascia. J Invest Dermatol 95:20-6
Petersen, M J; Woodley, D T; Stricklin, G P et al. (1990) Enhanced synthesis of collagenase by human keratinocytes cultured on type I or type IV collagen. J Invest Dermatol 94:341-6

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