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M O Hall

Publications and source records attributed to M O Hall.

13 recordsLinked to original sources
PubMed1992

The phagocytosis of ROS by RPE cells is inhibited by an antiserum to rat RPE cell plasma membranes.

A polyclonal antiserum to a rat retinal pigment epithelium (RPE) plasma membrane-enriched fraction has been utilized to identify candidate receptor proteins which may be involved in the phagocytosis of rod outer segments (ROS) by the RPE. Immunoblots of RPE cell extracts show that the the antiserum recognizes a number of glycoproteins, including two with M(r)s of 174 and 75 kDa. The antiserum also recognizes their non-glycosylated counterparts, with M(r)s of 169 and 65 kDa, respectively, which are synthesized after treatment of the cells with tunicamycin B2. Immuno-precipitation of [35S]-methionine-labeled RPE cell extracts also demonstrates the presence of antibodies to these same glycoproteins as well as to other proteins. The antiserum inhibits the binding of ROS to the RPE, which subsequently results in a decrease in the ingestion of ROS. ROS phagocytosis by the RPE is inhibited by 97% in the presence of a 1:10 dilution of the IgG fraction of the antiserum. Phagocytosis recovers to normal levels after 4-6 hr of chase in the absence of antibodies. After sequential adsorption of the IgG fraction to monolayers of fixed RPE cells, which removes RPE surface-specific IgGs, the extent of inhibition of ROS phagocytosis produced by the IgG fraction is reduced. Using immunoblotting we have identified a number of surface-specific immunoreactive bands which are adsorbed out of the antiserum, including the 174 and 75 kDa bands. These data give further support to the hypothesis that ROS phagocytosis is a receptor-mediated process, which occurs via specific cell surface glycoprotein receptors.

Animals↗
PubMed1992

cAMP production via the adenylyl cyclase pathway is reduced in RCS rat RPE.

cAMP production was investigated in retinal pigment epithelium (RPE) cells isolated from normal rats and from rats with an inherited retinal dystrophy (Rdy/p+). In normal RPE cells, 5'-[N-Ethylcarboxamido]-adenosine (A2 receptors) produced a fivefold increase in the level of cyclic adenosine monophosphate (cAMP) over basal levels. However, only a onefold increase in cAMP was observed in dystrophic cells. cAMP production by prostaglandins E1 and E2 (prostaglandin receptors) in dystrophic RPE cells was only 29-38% of the level observed in normal cells. Direct stimulation of adenylyl cyclase by 10 mumol/l forskolin increased cAMP levels in normal RPE cells by 90 fold over basal, but only by sixfold in the dystrophic cells. These data suggest there may be a defect in the adenylyl cyclase signaling pathway in dystrophic RPE cells.

Adenosine↗
PubMed1991

RPE cells from normal rats do not secrete a factor which enhances the phagocytosis of ROS by dystrophic rat RPE cells.

Retinal pigment epithelial (RPE) cells from normal and dystrophic rats were grown separately and in mixed culture for 7 days, without a change of growth medium. Isolated rod outer segments (ROS) were suspended in the conditioned medium from these cells, and were fed to the mixed or pure RPE cell cultures. No increase or decrease in the phagocytosis of ROS by dystrophic or normal RPE cells, respectively, was observed. These results suggest that normal RPE cells do not secrete a diffusible factor(s) which enhances the phagocytosis of ROS by dystrophic RPE cells.

Animals↗
PubMed1991

ROS ingestion by RPE cells is turned off by increased protein kinase C activity and by increased calcium.

The activation of protein kinase C (PKC) by phorbol myristate acetate (PMA) rapidly inhibits the phagocytosis of rod outer segments (ROS) by cultured rat retinal pigment epithelial (RPE) cells. PMA, at a concentration between 3.3 and 10 nM, blocks ROS ingestion by 50%, but does not inhibit the binding of ROS. The Ca2+ ionophore, A23187, also inhibits ROS phagocytosis, with an IC50 of about 0.5-1.0 microM and interferes with the ability of RPE cells to bind ROS. The effects of both of these drugs are reversible after drug washout. When PMA and A23187 are applied to cells consecutively, the effects are additive. These results suggest either that PMA and A23187, act upon the same proteins in the pathway which controls ROS ingestion, or that A23187 affects phagocytosis at the ROS binding level, while PKC affects steps further along the ingestion path. The effect of this process is to shut down the ingestion of ROS, as is seen during the prolonged feeding of ROS to RPE cells in culture.

Animals↗
PubMed1991

The phagocytosis of ROS by RPE cells is not inhibited by mannose-containing ligands.

We have examined the ability of mannose and the mannose-rich ligands, mannan and mannosylated BSA, to inhibit the phagocytosis of rod outer segments (ROS) by cultured rat retinal pigment epithelial (RPE) cells. Mannose, at concentrations up to 0.25 M, had no effect on either the binding or the ingestion of ROS. At concentrations above 0.25 M, the cells were rounded and showed detachment from the substrate, and phagocytosis was markedly inhibited. Neither mannan (2 mg ml-1), nor mannosylated BSA(0.8 mg ml-1), affected the phagocytosis of ROS. These results suggest that the phagocytosis of ROS is probably not mediated by a mannose receptor on the surface of the RPE cells.

Animals↗
PubMed1990

The effect of inhibitors of glycoprotein synthesis and processing on the phagocytosis of rod outer segments by cultured retinal pigment epithelial cells.

Retinal pigment epithelial cells selectively phagocytize rod outer segments by a process that may be mediated by specific cell surface receptors. Since many receptors are glycoproteins, we have studied the effect of tunicamycin, an inhibitor of N-linked oligosaccharide synthesis, and of castanospermine and swainsonine, which are inhibitors of oligosaccharide processing, on the ability of cultured retinal pigment epithelial cells to phagocytize rod outer segment. Tunicamycin inhibits the glycosylation of newly synthesized glycoproteins by 85-90%; concomitantly, the phagocytosis of rod outer segments is inhibited by 70-80%. The effect of tunicamycin is to initially reduce rod outer segments binding, and therefore the subsequent ingestion of rod outer segments. SDS-PAGE analysis and autoradiography of [35S]methionine labelled extracts of tunicamycin-treated cells, demonstrates the disappearance of a number of glycoprotein bands, and the appearance of a number of protein bands of lower Mr. Kinetic analysis of the disappearance and reappearance of specific glycoproteins suggests that the lower Mr bands are the non-glycosylated forms of the higher Mr bands. By contrast, castanospermine and swainsonine have no effect on the ability of retinal pigment epithelial cells to phagocytize rod outer segments, or on the SDS-PAGE pattern of treated cells, although they were shown to inhibit oligosaccharide processing as expected. These results support the hypothesis that rod outer segment phagocytosis by retinal pigment epithelial cells is mediated by specific glycoprotein receptors. N-Glycosylation of these receptors is required for their function, or for their insertion into the plasma membrane, whereas processing of the N-linked oligosaccharide chains of these receptors is not crucial for rod outer segment phagocytosis by retinal pigment epithelial cells.

Animals↗
PubMed1987

Kinetic studies of rod outer segment binding and ingestion by cultured rat RPE cells.

Retinal pigment epithelial (RPE) cells selectively phagocytize rod outer segments (ROS) by a process which may be mediated by specific cell surface receptors. We have studied the kinetics of this process using rat RPE cells grown in tissue culture. By cooling RPE cells to 17 degrees C, the binding and ingestion phases of phagocytosis can be separated. Maximum ROS binding with minimum ingestion occurs at 17 degrees C; above 17 degrees C the rate of ingestion increases markedly. Thus it is possible to measure the kinetics of ROS binding to RPE cells at 17 degrees C and of ROS ingestion at 37 degrees C. At 17 degrees C, ROS binding is saturable, both with respect to time and to ROS concentration. ROS ingestion saturates after 4 hr of incubation at 37 degrees C, after which the cells are refractory to further ROS ingestion for 1-2 hr. During this recovery period, rapid digestion of the internalized ROS takes place. Cycloheximide, when present at a concentration (2 x 10(-5) M) which inhibits protein synthesis by 92%, has no effect on ROS phagocytosis or on the recovery of ROS ingestion at 37 degrees C. This suggests that if receptors mediate the ingestion of ROS by RPE cells, they are not degraded after the ROS are internalized. Dystrophic rat (RCS-p+) RPE cells exhibit normal binding, but very limited ingestion of ROS at 37 degrees C. The rate and amount of ROS binding to these cells at 37 degrees C is comparable with that occurring to normal cells at 17 degrees C. These observations support the hypothesis that there are a limited number of receptors which are specific for ROS binding on the surface of normal and dystrophic rat RPE cells.

Animals↗
PubMed1979

Donor eye program.

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Academic Medical Centers↗
PubMed1978

Phagocytosis of light- and dark-adapted rod outer segments by cultured pigment epithelium.

Pigment epithelial cells in culture retain their ability to phagocytize rod outer segments. These cells phagocytize rod outer segments isolated from light-adapted rats, or from dark-adapted rats killed after the time at which the lights would normally be turned on. However, they phagocytize for fewer rod outer segments prepared in the dark from the retinas of rats killed before the onset of the normal light cycle. Phagocytosis of dark rod outer segments is variable, but that of light outer segments is reproducible. It is postulated that the effect of light is to synchronize the chemical events that occur at the surface of the rods to prepare them for phagocytosis. These processes also occur in the dark, but more slowly and irregularly than in the light.

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