Effect of phloretin on monosaccharide transport in erythrocyte ghosts

Beneš, I; Kolínská, J

doi:10.1007/bf01868107

Cited by 39 publications

(12 citation statements)

References 15 publications

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“…Phloretin is known to react rapidly, and it has been shown elsewhere (LeFevre & Marshall, 1959;Bene~, Kofinsk~i & Kotyk, 1972) that phloretin binds to the red cells immediately after it is mixed with a red cell suspension. This rapid binding has been shown to be exclusively on the membrane surface, and phloretin molecules do not penetrate the red cell after long incubation times (Bene~ et al, 1972). Our current experiments show that at "zero" incubation time, which denotes having the phloretin and nonelectrolyte together in one stopped-flow syringe and the red ceil suspension in another syringe, 0.25 m~ phloretin had essentially no effect on urea permeability; i.e., (Sphlore,in/S)~ 1.0.…”

Section: Resultsmentioning

confidence: 98%

The effect of phloretin on red cell nonelectrolyte permeability

1974

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Summary. The effect of phloretin on permeability of small nonelectrolytes into human red cells was shown to be bimodal for hydrophilic molecules and nonbimodal for lipophilic molecules. At low phloretin concentrations (<0.1 raM) hydrophilic as well as lipophilic nonelectrolyte permeation was increased. At high phloretin concentrations (> 0. lmM) the permeability of hydrophilic molecules was decreased, whereas lipophilic molecule permeability continued to be increased. These results suggest that the mechanism for phloretin acting on red cell nonelectrolyte pathways is different for hydrophilic molecules, as compared to lipophilic permeant molecules. The pH of the red cell buffer suspension also influenced the effect of phloretin on nonelectrolyte permeability and the keto or un-ionized form of phloretin (present at low pH), which is known to have a gTeater affinity for the membrane, had a larger effect on hydrophilic nonelectrolyte permeability than the ionized form of phloretin. Macey and Farmer (1970) reported that 0.5 mM phloretin inhibits urea, methylurea and glycerol permeability into human red blood cells. More recently, Owen and Solomon (1972) showed that 0.25 mM phloretin also inhibits urea permeability as well as other hydrophilic molecules such as formamide and acetamide, and that 0.25 mM phloretin enhances the permeability of lipophilic molecules such as propionamide and 2,3-butanediol. Macey and Farmer (1970) suggested the possibility that phloretin was inhibiting a specific facilitated transport system for urea. On the other hand, Owen and Solomon (1972) proposed that the effect of phloretin was a general one, influencing the red cell permeability to most nonelectrolytes and showed that 0.25 mM phloretin inhibits permeation of hydrophilic molecules indirectly proportionately to the ether partition coefficient, ko,~er, of the permeant molecule, whereas phloretin enhances permeation of lipophilic molecules in direct proportion to k,th~r. Similar observations were

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Section: Resultsmentioning

confidence: 98%

The effect of phloretin on red cell nonelectrolyte permeability

1974

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“…From experiments on ghosts, Ben~s et al [4] suggested that there are phloretin binding sites on both sides, but the observations are puzzling and the argument appears to be faulty: exit of xylose was inhibited by phloretin added to the external medium but not by phloretin trapped inside the ghosts, while entry was inhibited in the opposite way, i.e., by internal but not by external inhibitor. Believing, mistakenly as we now know [9,14], that phloretin cannot pass through the cell membrane, the authors concluded that transport is blocked only when the inhibitor is bound trans with respect to the substrate; but since phloretin would have had time to equilibrate [9], it should have been available on both sides, and therefore transport should have been inhibited in all cases.…”

Section: The Asymmetry Of the Glucose Carrier And Phloretin Bindingmentioning

confidence: 92%

“…According to an early report, phloretin does not penetrate the cell membrane [4] and therefore, when added to the suspending medium, could only add to the outer carrier form, regardless of whether the inner form possesses a phloretin binding site. Later studies demonstrated that it actually enters red cells rapidly and is strongly adsorbed both to the cell membrane and to hemoglobin [9,14].…”

Section: The Asymmetry Of the Glucose Carrier And Phloretin Bindingmentioning

confidence: 99%

Asymmetrical binding of phloretin to the glucose transport system of human erythrocytes

Krupka

1985

J. Membrain Biol.

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The sidedness of phloretin binding to the glucose carrier has been determined by comparing the type of inhibition produced in zero trans entry and zero trans exit experiments. Initial rates of zero trans entry were measured by the method of R.D. Taverna and R.G. Langdon (Biochim. Biophys. Acta 298:412-421, 1973), which involves pink ghosts loaded with glucose oxidase; this obviates the problem of rapid substrate accumulation inside the cells. With phloretin equilibrated across the membrane, the inhibition of entry was competitive, and the inhibition of exit noncompetitive. The experimental procedures were validated by showing that the inhibition by cytochalasin B, known to bind inside but not outside, was noncompetitive in entry and competitive in exit, as predicted. It was also demonstrated that even after pre-incubation of the cells with a relatively high concentration of phloretin, the phloretin adsorbed in the membrane did not significantly alter the rate of carrier reorientation. The results show that the outward-facing form of the glucose carrier, but not the inward-facing form, bears a phloretin binding site; thus phloretin, as well as cytochalasin B, is bound asymmetrically, phloretin outside and cytochalasin B inside.

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“…It has been shown that in contrast to sugar permeability, which could be inhibited from either surface (for the sidedness of the action of the aglycone, see Bene~, Kolinskfi and Kotyk, 1972), the equilibrium exchange for halide anions (Schnell, Gerhardt, Lepke & Passow, 1973) and sulfate could only be inhibited if the agent was present on the outer surface of the membrane. In view of the demonstrated asymmetry of the effect of phlorizin on anion exchange we included into the present work an investigation of the sidedness of the effect of this modifier on net salt movements.…”

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confidence: 95%

Effects of phlorizin on net chloride movements across the valinomycin-treated erythrocyte membrane

Kaplan

Passow

1974

J. Membrain Biol.

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Summary. The rate of valinomycin-induced KC1 efflux from human red cells and ghosts was measured in the presence and absence of phlorizin. Extracellular phlorizin accelerated the KC1 efflux. The effect depended on the phlorizin concentration and showed half saturation at about 0.4 mM phlorizin. Hunter's procedure was used to calculate C1 permeabilities (Pcl) by means of the Goldman equation from rate constants of K + loss in valinomycin-treated ghosts. For saturating phlorizin concentrations a 20-fold increase, approximately, of Pcl was calculated. The observed increase in Pcl is in contrast to the almost total inhibition of CI-equilibrium exchange. Similar to the effects on anion exchange permeability the effect on Pci is only observed when phlorizin is present at the outer surface of the erythrocyte membrane while internal phlorizin is without effect. A similar asymmetry was observed in the stimulation of valinomycin-induced K + exchange at identical K + concentrations on both sides Of the membrane. The effects of phlorizin were only observed if net KC1 flow was out of the cells but not if it was in the opposite direction. The effect of phlorizin on net KC1 movements and sugar transfer were unaltered when the phlorizin was subjected to several consecutive purifications. This indicates that the observed effects are due to the glycoside and not to contaminations with its aglycone.The penetration of anions across the erythrocyte membrane can be followed under two different experimental conditions (Harris & Pressman, 1967;Scarpa, Cecchetto & Azzone, 1970;Hunter, 1971): (1) in the absence of cation movements where one anion species from the red cell interior exchanges against another in the medium, and (2) by rendering the membrane permeable for cations and then following the net salt movements which include the movements of the accompanying anion. The results of the two types of measurements showed that exchange permeability for halides is many orders of magnitude higher than the net permeability. This was confirmed by comparisons of the permeabilities as measured by means of radioisotopes at Dorman equilibrium with electrical conductance measurements (Hoffman & Lassen, 1971 ;Lassen, 1972

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Effect of phloretin on monosaccharide transport in erythrocyte ghosts

Cited by 39 publications

References 15 publications

The effect of phloretin on red cell nonelectrolyte permeability

The effect of phloretin on red cell nonelectrolyte permeability

Asymmetrical binding of phloretin to the glucose transport system of human erythrocytes

Effects of phlorizin on net chloride movements across the valinomycin-treated erythrocyte membrane

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