The effect of rimantadine on the structure of model and biological membranes

The action of the antiviral drug rimantadine on the structure of bilayer lipid membranes (BLM) and RBC membranes was investigated. Structural changes in BLM were recorded by ionophore conductivity changes and by changes in the third harmonic of capacity current signal due to lateral compression of BLM in an electric field. It was shown that the adsorption of rimantadine on BLM results in an increase in ionophore mobility in bilayer membranes of dioleolyllecithin (DOL) and common lipids of bovine brain (CL) and in a decrease in those of azolectin (A). Relative changes in the third harmonic signal also depend on the membrane composition and have different signs. The results may be explained by the rimantadine action on the lipid bilayer structure: "rigidification" of A-membranes and "fluidization" of BLM from DOL and CL. Structural reorganization of RBC membranes as investigated by the ability of the cells to enter a micropipette (inner diameter greater than or equal to 3 microns) thereby undergoing deformation. It was shown that rimantadine influences RBC deformability due to drug induced inhomogenous mechanical membrane properties. Also, rimantadine accelerated the process of artificially induced aggregation of erythrocytes. The relation of the effects on artificial and biological membranes, and the structural changes in the lipid phase of membrane are discussed.     

Separation of red blood cells at high volume concentration under low centrifugal accelerations

The separation process of blood and RBC suspensions in a hematocrit range between 0.3-0.7 was investigated with a centrifuge allowed to run at low accelerations (100 xg-1000 xg). The position of the interface between the supernatant of plasma and the RBC column was continuously recorded by a new optoelectronic measuring system. The separation process could be mathematically described by an exponential decrease of the cell column approaching a final packing. At a given centrifugal acceleration the time constant is influenced by hematocrit, aggregation, deformation and plasma viscosity. The final packing depends linearly on the starting hematocrit (0.3-0.7) and can be used as a measure of deformability.     

Electrostatic fixed charge distribution in the RBC-glycocalyx and their influence upon the total free interaction energy

On the basis of a recently developed biophysical model of cell-cell interaction, including electrostatic, electrodynamic, steric and bonding/bridging interaction energies the influence of different fixed charge (dissociated groups of the glycocalyx) density distributions in red blood cell (RBC) glycocalyces on the total free interaction energy was investigated. An analytical equation of electrostatic free energy on the basis of the linear Poisson-Boltzmann approach taking into account arbitrary distributions of fixed glycocalyx charges was obtained and corresponding free electrostatic energies of three example distributions were calculated. The electrodynamic, steric and bonding/bridging energies were computed as usual. It was shown that the free energy as a function of interaction distances strongly depends on the charge distribution and, correspondingly, the "weight" of this energy term in the total free interaction energy balance equation. Generally, it can be stated that as more charges are assumed to be fixed in the outer layer of RBC glycocalyx as more important becomes the electrostatic energy in contrast to the remaining three terms.     

In vitro crosslinking of DNA by 8-methoxypsoralen visualized by electron microscopy

By electron microscopic visualisation of totally denatured DNA, we have detected photochemically induced 8-methoxypsoralen crosslinks in vitro after irradiation at 360 nm. The amount of crosslinks was expressed as the percentage of DNA length which was kept in double-stranded appearance by closely situated crosslinks. This percentage correlated well with irradiation time, irradiation intensity, and the concentration of 8-methoxypsoralen. These parameters have also been correlated with the mean size and the size distribution of non-crosslinked regions of DNA, so called bubbles. For a comparison with another psoralen type, we have carried out a similar set of experiments using 4,5,8-trimethylpsoralen.     

Identification of specific pore residues mediating KCNQ1 inactivation. A novel mechanism for long QT syndrome

KCNQ1 inactivation bears electrophysiological characteristics different from classical N- and C-type inactivation in Shaker-like potassium channels. However, the molecular site of KCNQ1 inactivation has not yet been determined. KCNQ2 channels do not exert a fast inactivation in contrast to KCNQ1 channels. By expressing functional chimeras between KCNQ1 and KCNQ2 in Xenopus oocytes, we mapped the region of this inactivation to transmembrane domain S5 and the pore loop H5 and finally narrowed down the site to positions Gly(272) and Val(307) in KCNQ1. Exchanging these two amino acids individually with the analogous KCNQ2 residue abolished inactivation. Furthermore, a KCNQ1-like inactivation was introduced into KCNQ2 by mutagenesis in the corresponding region, confirming its relevance for the inactivation process. As KCNQ1 inactivation involves the regions S5 and H5, it exhibits a geography distinct from N- or C-type inactivation. Native cardiac I(Ks) channels comprising KCNQ1 and accessory MinK subunits do not inactivate because of the functional interaction of KCNQ1 with MinK. Mutations in KCNQ1 can lead to long QT1 syndrome, an inherited form of arrhythmia. The long QT1 mutant KCNQ1(L273F) displays a pronounced KCNQ1 inactivation. Here we show that when expressing mutant I(Ks) channels formed from KCNQ1(L273F) and MinK, MinK association no longer eliminates KCNQ1 inactivation. This results in smaller repolarizing currents in the heart and therefore represents a novel mechanism leading to long QT syndrome.     

Effect of weaning time and associated management practices on postweaning chronic diarrhea in captive rhesus monkeys (Macaca mulatta)

OBJECTIVE: Our purpose was to assess the extent to which early weaning and other weaning-management factors affect development of postweaning chronic diarrhea in captive rhesus monkeys at the California Regional Primate Research Center between 1992 and 1995. METHODS: Data for weaning, management, and onset of diarrhea were obtained from daily records. The Cox proportional hazard model was used to assess whether the risk of chronic diarrhea was related to early weaning. RESULTS: Monkeys that were lighter at weaning had a threefold increase in risk of postweaning chronic diarrhea (P = 0.07), compared with that in heavier monkeys. An episode of preweaning diarrhea increased the risk of postweaning chronic diarrhea twofold (P = 0.08). Relocation of monkeys to outdoor facilities in the fall was associated with a fivefold decrease in risk (P < 0.001), compared with that of other seasons, and weaning in 1993 was associated with a twofold decrease in risk, compared with that of other years (P = 0.04). CONCLUSIONS: Multiple factors need to be considered for prevention of postweaning chronic diarrhea, including weaning weight, preweaning diarrhea, season weaned, and weaning conditions that change from year to year.