摘要:

A number of investigators have studied the effect of metabolic inhibitors on the facility of aqueous outflow from the enucleated eye perfused eye at“room temperature”. Some inhibitors such as iodoacetate and iodoacetamide increased the facility, others such asp-chloromercuri-benzoate decreased it. Most however, including sodium fluoride, were reported to have had no effect.We have found that 10 mM sodium fluoride had a very small effect on facility of aqueous outflow at room temperature (22°C), but reduced the facility as much as 50% (average 30%) at 35°C. The magnitude of the sodium fluoride effect did not appear to be dependent on calcium levels in the medium, but the presence of Ca2+significantly prolonged its duration.Iodoacetamide and iodoacetate both increased facility at 35°C in a manner similar to previous findings at room temperature except that at 8–10 mM, iodoacetate caused a 55% decrease; under these conditions, there was considerable release of tissue debris into the anterior chamber which probably blocked the TM. A number of other agents, such as 2,4-dinitrophenol, KCN, and 2-deoxyglucose were studied, but none produced any marked effect on facility.These results emphasize that temperature may profoundly modify the effect of metabolically active agents. The mechanism of the fluoride effect has not been determined but it is consistent with the action of F on the G protein associated with phospholipase C.

ISSN:0271-3683    

DOI:10.3109/02713689308999489

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

A low molecular size peroxidase with a high affinity for phospholipid hydroperoxide was purified from bovine retina by sequential extraction with low and high ionic strength buffer, followed by ammonium sulfate fractionation, chromatography on an ultraspherogel column and Protein PAK-SP column. The purified enzyme has a low Km, (0.011 mmol/L) for phospholipid hydroperoxide, and a high Km(1.37 mmol/L) for glutathione. Glutathione oxidation was competitively inhibited by vitamin E, Ki0.019 mmol/L. The retinal PHGPX is different from the PHGPX purified by others from heart and liver in molecular size. The molecular size estimated by gel filtration chromatography is below 6 kDa.

ISSN:0271-3683    

DOI:10.3109/02713689308999490

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

The present study provides the first evidence forδ13-reduction of 15-ketoprostaglandins (15-keto-PGs) by bovine ocular tissues. The 9,000xg supernatants of cornea, iris, ciliary body, retina, and RPE-choroid except lens exhibitedδ13-reductase activity toward 15-keto-PG E2and F2αin the presence of NADPH or NADH as an electron donor. Among the tissues tested, the highest activity was observed in ciliary body and iris, followed by RPE-choroid, retina, and cornea. The NADPH- and NADH-linked double bond reductase activities were inhibited by dicumarol, quercitrin, indomethacin and disulfiram, but not by potassium cyanide.NADPH-linked 15-keto-PG F2αδ13-reductase was purified from bovine iris-ciliary body cytosol by fractionation with ammonium sulfate and high performance liquid chromatography (HPLC) with TSK gel DEAE-5PW, and TSK gel Blue-5PW. The purified enzyme was homogenous by the criterion of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Its molecular weight was estimated to be about 57,000 by electrophoresis, and about 55,000 by gel filtration HPLC with Superose 12.

ISSN:0271-3683    

DOI:10.3109/02713689308999491

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

The fractal dimension of the retinal vasculature and isolated venous and arterial trees down to a caliber of 40μm was estimated in 23 routine fluorescein angiograms of normal retinas. Fractal dimension was determined with a method based on the box counting theorem. This method is less susceptible to the radial architecture of the retinal vascular tree than those previously reported (mass-radius relation and density-density correlation function). Two scale ranges with different fractal dimension were consistently present. The estimated fractal dimensions showed no significant difference between isolated arterial and venous trees which is not supported by previous reports. This method was designed for simple application in a clinical setting.

ISSN:0271-3683    

DOI:10.3109/02713689308999492

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

The characteristics of taurine transport were studied in a human retinal pigment epithelial cell line (HRPE). Uptake of taurine into monolayer cultures of the HRPE cells was markedly stimulated by the presence of NaCl in the uptake medium whereas the uptake was negligible in its absence. This NaCl-dependent uptake was an active process as the cells were able to accumulate taurine against a concentration gradient. The uptake rate of taurine was found to be many-fold greater than that ofγ-aminobutyric acid (GABA). Unlabeled taurine and GABA competed with radiolabeled taurine for the uptake process, the former being more effective than the latter. However, uptake of radiolabeled GABA was not affected by unlabeled taurine and GABA. Substrate specificity studies revealed strong interaction ofβ-amino acids with the transport system responsible for taurine uptake.α-Amino acids failed to inhibit taurine uptake. A specific anion requirement was observed for optimal activity of the taurine transport system and CI-was the most supportive among several anions tested. Kinetic analyses showed that multiple Na+and one CI-were involved in transfer of one taurine molecule. The transport process consisted of a single saturable system with a Michaelis-Menten constant of 2.0±0.1μM. These results show that the HRPE cell line expresses a high-affinity taurine transport system. This is the first demonstration of the presence of the taurine transporter in the human retinal pigment epithelium and the HRPE cell line may provide a useful model system for future studies involving taurine transport in the retinal pigment epithelium.

ISSN:0271-3683    

DOI:10.3109/02713689308999493

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

Myeloid bodies (MBs) are distinct lamellar regions of the normally branched tubular smooth endoplasmic reticulum of retinal pigment epithelial (RPE) cells. These organelles are present in the retinas of many lower vertebrates. Previous investigations have reported a relationship between outer segment disk shedding and phago-cytosis by the RPE, and the formation of MBs. The current morphometric study was undertaken to establish if a temporal relationship existed between MB occurrence and the phagocytosis of shied photoreceptor outer segment tips in the chick retina. We report on the occurrence of phagosomes, MBs, and for the first time, MB precursors (templates) in the RPE over a 24-hr diurnal cycle.Phagosome numbers were observed to be highest within 2 hr following lights on, and again following lights off, while MB precursors were most prevalent at two time points, immediately prior to the times for rod and cone outer segment shedding at lights on and lights off respectively. Traditional MBs of the small (0.06 to 0.239μm2) and medium (0.24 to 0.99μm2) variety increased both in size and number from 01:00 hr of a 24 hr time period, with smaller and fewer MBs being present late in the dark part of the diurnal cycle, and larger and more numerous MBs in the later part of the light portion and into the early part of the dark portion of the cycle.This study has demonstrated a very different diurnal pattern of occurrence in both the size (organelle area as a percentage of total cell area) and number of MB precursors as compared to either small or medium-sized MBs, suggesting that MB precursors may represent a functionally, as well as morphologically distinct population of MBs.

ISSN:0271-3683    

DOI:10.3109/02713689308999494

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

The effect of monensin on the cytotoxic effect of antitransferrin receptor immunotoxin (IT) was determined on cultured, human retinal pigment epithelial (hRPE) cells. Human RPE cells were treated with 0.1 - 10,000 ng/ml IT with and without 0.01 - 0.1μM monensin, a lysosomotropic reagent that can influence IT activity. Monensin (0.01μM) shortened the onset of cell kill with IT (10,000 ng/ml) from 48 to 24 hours (p = 0.0016). Although 0.01μM monensin alone was not cytotoxic to hRPE cells, a single 7 - day treatment with monensin caused up to a 4.1 - fold increase in antiproliferative potency of IT on proliferating hRPE cells (p≤0.0001). Enhancement was obtained with only a 1 - hour exposure to 0.1μM monensin (p = 0.0001). In contrast, IT (0.1 - 10,000 ng/ml) combined with monensin (0.01μM) had minimal effect on density - arrested cells. IT with or without monensin did not inhibit proliferation of Rhesus monkey RPE cells. Our results indicate that monensin enhances the selective cytotoxic effect of IT on proliferating hRPE cells in culture.

ISSN:0271-3683    

DOI:10.3109/02713689308999495

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

The effect of the inhibitors of aerobic metabolism on the transport of rubidium and ATP content in rat lens has been studied under organ culture system. The inhibitors used were malonate, monofluoroacetate, cyanide, antimycin A, 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO), 2,4-dinitrophenol (DNP) and dicumarol. All these compounds inhibited the transport of rubidium, reflecting a decrease in the ability of the lens to transport potassium. The levels of ATP also decreased. The results were similar to those obtained under anaerobic conditions where the lenses were incubated in a nitrogen atmosphere without the inhibitors. The degree of inhibition observed by the metabolic inhibitors as well as by anaerobiosis indicate that the rat lens depends substantially on aerobic oxidation for its energy needs, unlike bovine and rabbit lenses studied before.

ISSN:0271-3683    

DOI:10.3109/02713689308999496

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

To test the hypothesis that myeloid body (MB) formation results from the shedding of retinal photoreceptor outer segments and the consequent degradation of lipids derived from outer segment disk membranes, the effect of massive outer segment shedding and the disruption of such outer segment shedding on MB formation were examined in the leopard frog. Light entrained frogs were first placed in constant light (700 lux) for 48 hours to inhibit shedding, followed by a 1.5 hours dark priming eitherin vivoorin vitro, and then returned to light for an additional 4 hours which results in massive outer segment shedding. To serve as a control, the effect of shedding disruption on MB formation was assessedin vivousing light manipulation to inhibit shedding, or mechanical removal of the neurosensory retinain vitro.The results indicate that although the phagosome numbers were clearly elevated in the samples taken from eitherin vivoorin vitroeye-cup preparations where outer segment shedding had been stimulated, there was no significant concomitant increase in Mbs number over controls kept under constant light for 48 hr or constant light 48 hr plus 1.5 hr in dark, where MBs represent approximately 5% of the total RPE cell area. In contrast, when shedding was interrupted either by removal of the neural retina immediately afterin vitroeye-cups were returned to light or by maintaining frogs in dark without light stimulation, the RPE cells contained very few phagosome, yet in both conditions RPE cells showed a two-fold increase in MB area over the shedding-stimulated controls (p=0.0001). In conclusion, MB formation appears not to be simply a direct result of photoreceptor outer segment disk shedding and membrane degradation since the interruption of photoreceptor outer segment shedding results in an increase in MB occurrence in the RPE. Based on this new evidence we now speculate that MBs in the frog RPE may be actively involved in lipid recycling related to photoreceptor outer segment membrane turnover.

ISSN:0271-3683    

DOI:10.3109/02713689308999497

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

Amiloride analogues with N5alkyl substitutions are specific high-affinity ligands for the Na+-H+exchanger in various tissues. As a means to characterize the Na+-H+exchanger in the bovine corneal epithelium, we determined the binding properties of [3H] methylisobutylamiloride (MIA) to a fraction enriched in plasma membrane from this tissue. [H]MIA bound to these membranes in a time,-a temperature-, and -a pH-dependent manner. The binding was optimal at 4°C and at pH 8.5 and it reached equilibrium at 60 min. Under these conditions, specific binding, which was inhibitable by excess unlabeled MIA, was about 85%. Scatchard analysis of this specific binding revealed a single saturable binding component with a Kdof 61 nM and a Bmaxof 271 pmoles/mg protein. Inhibition of [3H]MIA specific binding by amiloride analogues showed the following order of potency: MIA>dimethylamiloride (DMA)>benzamil>amiloride. Na+did not compete with MIA for binding. The effectiveness of clonidine, an alpha2agonist, and cimetidine, an H2receptor antagonist, as inhibitors of Na+-H+exchange activity was also determined because these compounds are used to distinguish between the exchanger subtypes. At concentrations higher than those needed for receptor interaction, clonidine was more effective than cimetidine in decreasing MIA binding.The activity of Na+-H+exchanger, which was measured as the uptake of Na+in the presence of an outwardly directly H+gradient, was also inhibited by DMA, benzamil and amiloride with the same order of potency as obtained in the binding studies. As seen with the MIA binding studies, clonidine was more potent than cimetidine in inhibiting 22+Nauptake. We conclude that the specific binding of MIA observed in these membrane preparations represents the binding of the ligand to the Na+-H+exchanger and that the MIA binding site on the exchanger is distinct from the Na+binding site (substrate site). The relative potencies of clonidine and cimetidine to inhibit MIA binding and22Na+uptake indicate that the Na+-H+exchanger present in the corneal epithelial plasma membrane belongs to the amiloride-insensitive type that has been described in the brush-border membrane of the kidney and intestine.

ISSN:0271-3683    

DOI:10.3109/02713689308999498

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor