The specific aims of this proposal are to identify pigment epithelial (PE) cell surface receptors which participate in the recognition and attachment of rod outer segments (ROS) and receptors which are responsible for initiating and controlling the ingestion phase of phagocytosis. These goals are directed towards understanding the biochemical and molecular events which participate in the phagocytosis of rod outer segments by the pigment epithelium, and towards elucidating the molecular cause of the failure of this process in the RCS rate model of retinal degeneration. Because of the reduced capability of the RCS rat PE to phagocytize ROS, this animal provides a convenient model for probing a fundamental cellular interaction, the maintenance of which is vital to the persistence of vision. By unravelling the cause of a retinal degeneration in the rat, we may better understand one of the possible causes of retinal degeneration in man. These specific goals are possible because of the discovery that the mutation in the RCS rat PE affects the ingestion phase of phagocytosis. Additionally, I have developed an antiserum to rat PE cells which inhibits the phagocytosis of ROS by these cells. Thus it is now possible to design experiments to identify the cell surface receptors which participate in the several separate steps of phagocytosis, and to identify those receptors which are absent or defective in the RCS rat PE. Monoclonal antibodies (MCA) will be produced against PE cell plasma membranes, and the specific antibodies which inhibit the phagocytosis of ROS will be selected. PE cell surface antigens will be separated by 2D-SDS PAGE and transblotting. After reaction with antiserum or MCA and 125I-protein A, specific antigens will be identified by autoradiography. Cell surface receptors for ROS binding will be radiolabeled and cross-linked to ROS surface proteins. The receptors will then be identified as outlined above. The role of actin in ROS disk shedding will be explored using actin antibodies immunoferritin labeling, and electron microscopy.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY000046-18
Application #
3255093
Study Section
(VID)
Project Start
1978-03-01
Project End
1987-11-30
Budget Start
1986-12-01
Budget End
1987-11-30
Support Year
18
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of California Los Angeles
Department
Type
Schools of Medicine
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Gregory, C Y; Abrams, T A; Hall, M O (1994) Stimulation of A2 adenosine receptors inhibits the ingestion of photoreceptor outer segments by retinal pigment epithelium. Invest Ophthalmol Vis Sci 35:819-25
Hall, M O; Abrams, T A; Mittag, T W (1993) The phagocytosis of rod outer segments is inhibited by drugs linked to cyclic adenosine monophosphate production. Invest Ophthalmol Vis Sci 34:2392-401
Gregory, C Y; Hall, M O (1992) The phagocytosis of ROS by RPE cells is inhibited by an antiserum to rat RPE cell plasma membranes. Exp Eye Res 54:843-51
Gregory, C Y; Abrams, T A; Hall, M O (1992) cAMP production via the adenylyl cyclase pathway is reduced in RCS rat RPE. Invest Ophthalmol Vis Sci 33:3121-4
Hall, M O; Abrams, T A; Mittag, T W (1991) ROS ingestion by RPE cells is turned off by increased protein kinase C activity and by increased calcium. Exp Eye Res 52:591-8
Hall, M O; Abrams, T A (1991) The phagocytosis of ROS by RPE cells is not inhibited by mannose-containing ligands. Exp Eye Res 53:167-70
Hall, M O; Abrams, T A (1991) RPE cells from normal rats do not secrete a factor which enhances the phagocytosis of ROS by dystrophic rat RPE cells. Exp Eye Res 52:461-4
Hall, M O; Burgess, B L; Arakawa, H et al. (1990) The effect of inhibitors of glycoprotein synthesis and processing on the phagocytosis of rod outer segments by cultured retinal pigment epithelial cells. Glycobiology 1:51-61
Hall, M O; Abrams, T (1987) Kinetic studies of rod outer segment binding and ingestion by cultured rat RPE cells. Exp Eye Res 45:907-22
Colley, N J; Clark, V M; Hall, M O (1987) Surface modification of retinal pigment epithelial cells: effects on phagocytosis and glycoprotein composition. Exp Eye Res 44:377-92

Showing the most recent 10 out of 14 publications