Temperature dependence of the exchange of monovalent anions in human red blood cells.

The temperature dependence of anion exchange across the red cell membrane was studied between 5 degrees C and 55 degrees C by measuring the rate of shrinkage of cells when transferred from a medium of pH 7.6 to one of pH 9.3 (as measured at 22 degrees C). The rates of shrinkage varied with the anion studied, the order being F-> Cl-> Br-> I-> SCN- but were faster in the presence of trace amounts of carbon dioxide than in its absence. NO3- was as fast as Cl- when carbon dioxide was present but comparable with I- when it was removed. Arrhenius plots of the rates were linear for all anions over the temperature range studied and gave the following apparent activation energies in kJ mol-1; F-, 67.7; NO3-, 68.4; Cl-, 70.2; Br-, 79.6; SCN-, 87.4 and I-, 95.1 in the presence of carbon dioxide. Inhibition of carbonic anhydrase with 5 microns ethoxzolamide and the removal of the carbon dioxide by degassing raised the activation energies to; F-, 124.8; NO3-, 129.0; Cl-, 141.5: Br-, 159.4; SCN-, 150.0 and I-, 185.6 kJ. mol-1. With the exception of F-, the apparent activation energies of the anions were linearly related to their thermochemical (dehydrated) radii in both cases. The relationship between the ionic radii and the energy of transfer suggests that anion exchange involves transfer through a hydrophobic pathway and that additional energy is required to overcome restrictions experienced in passing through the pathway. It is proposed that this, rather than a conformational change in the protein determines the activation energy of the process.

The effect of ATP-depletion on the inhibition of glucose exits from human red cells.

The effect of ATP-depletion or its consequence, by metabolic inhibition, on the inhibition of glucose transport by various inhibitors was studied in human red cells. In cells depleted of ATP, glucose exit times were longer than in normal cells and the times increased with the duration of depletion. The Km for external glucose was higher in ATP-depleted cells than in normal undepleted cells (3.0 mM c.f. 2.5 mM at 30 degrees C). In contrast, the apparent Ki for cytochalasin B decreased from 0.85 microM in the normal cells to 0.5 microM after ATP-depletion. Half-maximal rates of glucose exit in the absence, and in the presence of 2 microM cytochalasin B were found at ATP concentrations of 0.43 and 0.68 microM, respectively. Although glucose exits from ATP-depleted cells exposed to the irreversible inhibitor of glucose transport, 1-fluoro-2,4-dinitrobenzene (FDNB) were slower than in normal cells, the relative degrees of inhibition were not significantly different. However, normal and ATP-depleted cells responded differently to treatment with 1,2-cyclohexanedione, a modifier of arginine residues which inhibits glucose exit. While normal cells were markedly inhibited, depleted cells were much less affected and the inhibitory effect of cytochalasin B seen in normal cells was reduced. These findings demonstrate that the glucose transport system of human red cells is affected by intracellular ATP and that ATP alters the affinity of the transporter for certain inhibitors. The implications of these findings are discussed.

Evidence of multiple operational affinities for D-glucose inside the human erythrocyte membrane.

1. The Michaelis-Menten parameters of labelled D-glucose exit from human erythrocytes at 2 degrees C into external solution containing 50 mM D-galactose were obtained. The Km is 3.4 +/0 0.4 mM, V 17.3 +/- 1.4 MMOL . 1(-1) cell water . min-1 for this infinite-trans exit procedure. 2. The kinetic parameters of equilibrium exchange of D-glucose at 2 degrees C are Km = 25 +/- 3.4 mM, V 30 +/- 4.1 mmol . 1(-1) cell water . min-1. 3. The Km for net exit of D-glucose into solutions containing zero sugar is 15.8 +/- 1.7 mM, V 9.3 +/- 3.3 mmol . 1(-1) cell water . min-1. 4. This experimental evidence corroborates the previous finding of Hankin, B.L., Lieb, W.R. and Stein, W.D. [(1972) Biochim. Biophys. Acta 255, 126--132] that there are sites with both high and low operational affinities for D-glucose at the inner surface of the human erythrocyte membrane. This result is inconsistent with current asymmetric carrier models of sugar transport.

Pinworm granuloma of the liver.

A granuloma containing Enterobius vermicularis was found in the liver of a 62-year old woman from Arkansas who had a 4-year history of recurrent surgery for adenocarcinoma of the colon. Hematogenous spread and direct penetration of the liver preceded by either invasion of the peritoneum through unhealthy or traumatized intestinal tissue, or from migration up the genital tract are the infective mechanisms considered. This type of parasitic liver lesion is the third reported case, the first in North America, and the first in a female.

Clonal chromosome aberrations in a case of nodular fasciitis.

We report a case of nodular fasciitis with clonal chromosome aberrations including a reciprocal t(3;15)(q21;q22) and interstitial deletion (13)(q14q21).

Melanoma today does not require radical surgery.

From this study we conclude that melanoma has a more favorable outcome today than 20 years ago, 2 cm margins around the tumor achieve acceptable survival and local recurrence rates, primary closure is preferred when possible, elective regional node dissection is of questionable value, and further trials are warranted to determine optimal therapy for intermediate thickness melanoma.

Chronic fixed drug eruption caused by acetaminophen.

A case of chronic fixed drug eruption resembling parapsoriasis en plaques is reported, which presented with persistent, stable lesions that were present for seven months before the diagnosis was established. The patient's skin cleared totally with avoidance of acetaminophen and flared prominently with re-exposure. Distribution of the recurrence was different: some previous sites had apparently become refractory and remained clear, some involvement had recurred in the same site, and new areas of involvement had appeared, causing the eruption to "wander," as is often seen in acute fixed drug eruption due to acetaminophen.

Mechanism of action of iontophoresis in the treatment of palmar hyperhidrosis.

Sweat glands from the palm of a patient with hyperhidrosis were examined by light and electron microscopy before and after treatment with tap water iontophoresis. No changes in structure were noted after treatment, disclaiming the currently accepted theory that mechanical ductal obstruction is the mechanism of action of this method. Furthermore, we believe the safety and effectiveness of this treatment are supported by our experience.

The polyguanidinium-ring-complex cation shield in the human red cell glucose transporter.

Widdas and Baker (1991) have described a possible structure and functioning of the human erythrocyte glucose transporter, based on its amino acid sequence. It was noted that eight positively charged side chains surrounded the proposed transfer cleft on the inside. These could form a ring shield which prevented the loss of K+ ions. Theoretical and model-making considerations point to this shield being made up of a polyguanidinium-ring-complex cation with eight (or ten) positive charges distributed over two identical rings of hydrogen bonds and normal linkages.

The polyguanidinium complex of the glucose transporter: a possible basis for bistable cationic gates and anionic channels in biology.

The detailed molecular structure and mechanism of action of the red cell glucose transporter would endow it with the ability to transfer the less hydrated potassium ion. This is prevented by a polyguanidinium-ring-complex cation which provides an effective electrical energy barrier. The ring complex has structural features which could form a bistable state of ring charges; in one form the electrical repulsions would be fully effective (gate 'shut'), whereas in the other form they would be ineffective (gate 'open'). It is also suggested that the polyguanidinium linkages could form a linear complex which could be the basis for anionic channels in biology. The theoretical basis for both these hypotheses is described.

The pH volume changes of human red cells in vitro due to the exchange of chloride and hydroxyl anions.

In glucose exit experiments measured photoelectrically, the excursions on the chart recorder were found to be larger for exits in media of alkaline pH. This was shown to be due to the addition of a pH volume effect to that of the osmotic shrinkage resulting from the glucose efflux. The pH-dependent volume change also occurred in glucose-free cells and was a linear function of the pH of the medium between pH 6.8 and 9.0. The effect is consistent with the loss (or gain) of chloride in exchange for hydroxyl anions on the band 3 anion transporter and with the buffering of the hydroxyl anions by haemoglobin. The implications for the working of the anion exchanger and for respiratory physiology are discussed.

Parameters for 3-O-methyl glucose transport in human erythrocytes and fit of asymmetric carrier kinetics.

1. Equilibrium exchanges in the range of 2-40 mM-3-O-methyl glucose at 16 degrees C suggested that the half-saturation concentration for exchange was 22 mM and that the maximum velocity (Vmax) was ca. 149 mmol l-1 min-1. 2. Initial rates of exchange influx from 1, 2, 4 and 8 mM into 76 mM solution gave a half-saturation value of 3.6 mM and a Vmax of 122 mmol-1 min-1. 3. The non-transportable inhibitor 4,6-O-ethylidene-alpha-D-glucopyranose (ethylidene glucose) acting on the outside of the cells inhibited 3-O-methyl glucose exchanges at 16 degrees C with an inhibition constant (KI) of ca. 11 mM. 4. Sen-Widdas exit experiments gave the half-saturation for 3-O-methyl glucose at 16 degrees C as only ca. 2 mM and the KI for ethylidene glucose as ca. 4 mM. 5. Efflux inhibitions by ethylidene glucose are satisfactorily predicted by the asymmetric carrier kinetics of Regen & Tarpley (1974) when using the parameters derived from the exchange experiments but not with parameters from Sen-Widdas exits. 6. Uphill transfer by counterflow experiments and Sen-Widdas exits cannot be fitted by the Regen and Tarpley kinetics (using the same parameters) unless the kinetics are modified to provide for an extra exchange element which replaces some of the net exit component in the equations. 7. At present the modification to the kinetics is only possible in computer simulations and data handling, but with it the fit to experimental results is good. The nature of the modification is described and in the light of it a revised interpretation of the significance of the Km derived from Sen-Widdas exits is discussed.