Double-mode impedance analysis of epithelial cell monolayers cultured on shear wave resonators

The viscoelastic behavior of epithelial cells (MDCK-I and MDCK-II) grown on AT-cut quartz crystals with a fundamental resonance at 5 MHz was investigated by impedance spectroscopy. Using the electromechanical model recently derived by Martin et al. [(1991) Anal Chem 63: 2272 – 2281] for Newtonian liquids in contact with shear wave resonators we quantified the viscous damping arising from the adherent cells by fitting the impedance data with a modified Butterworth-Van Dyke circuit in the region of the resonance frequency. Impedance spectroscopy was additionally performed in the frequency range from 1 Hz to 1 MHz to scrutinize the passive electrical properties of the epithelial cell layers using an additional platinum electrode. These data allow one to document the cell layers' integrity as well as the electrode coverage. We were able to confirm that the presence of a cell-layer mainly increases damping of the shear wave and does not exhibit a pure mass-load behavior. These findings were supported by the discovery that the inductance L in the electromechanical model was less influenced by the cell-layer than the resistance R. The apparent cell-viscosities determined by our method are 0.097 poise for MDCK-I and 0.142 poise for MDCK-II cell-layers. These low apparent viscosities may be explained in terms of a considerable spacing between the cells immobilized via their focal contacts and the quartz surface. Received: 5 June 1996 / Accepted: 6 August 1996     

Ti plasmid-encoded octopine and nopaline catabolism in Agrobacterium: specificities of the LysR-type regulators OccR and NocR,and protein-induced DNA bending

The occ and noc regions in octopine and nopaline Ti plasmids, respectively, are responsible for the catabolism of octopine and nopaline in Agrobacterium. The functions are activated in the presence of the opines by OccR and NocR, two related regulatory proteins, and the promoters contain common sequence motifs. We have investigated heterologous interactions between the regulators and the promoters. Previous experiments using all possible heterologous combinations of opines, regulators, and promoters in vivo had demonstrated that only the combination of nopalme, NocR, and the occ promoter led to limited promoter activation. We now show that OccR and NocR bind to the heterologous promoters in vitro and in vivo. The weak or non-existent promoter activation actually observed could be explained by the assumption that OccR and NocR use different activation mechanisms; we investigated protein-induced DNA bending because of reports that the two regulators differ in this respect. Analysis with a bending vector showed that both OccR and NocR induced a DNA bend that is relaxed in the presence of the respective opine. The data suggest that subtle differences in regulator/promoter interactions are responsible for the inactivity of the heterologous combinations. Investigations with a chimeric NocR/OccR protein indicated that it induced a DNA bend in both promoters. No opine-induced relaxation was detectable with the hybrid, and the inducible promoter was not activated. These findings suggest that bend relaxation may be an integral part of promoter activation.     

Aerosol dispersion in human lung: comparison between numerical simulations and experiments for bolus tests

Darquenne, Chantal, Peter Brand, Joachim Heyder, and ManuelPaiva. Aerosol dispersion in human lung: comparison between numerical simulations and experiments for bolus tests.J. Appl. Physiol. 83(3): 966-974, 1997.Bolus inhalations of 0.87-µm-diameter particles wereadministered to 10 healthy subjects, and data were compared withnumerical simulations based on a one-dimensional model of aerosoltransport and deposition in the human lung (J. Appl.Physiol. 77: 2889-2898, 1994). Aerosol boluseswere inhaled at a constant flow rate into various volumetric lungdepths up to 1,500 ml. Parameters such as bolus half-width, mode shift, skewness, and deposition were used to characterize the bolus and todisplay convective mixing. The simulations described the experimental results reasonably well. The sensitivity of the simulations to different parameters was tested. Simulated half-width appeared to beinsensitive to altered values of the deposition term, whereas it wasgreatly affected by modified values of the apparent diffusion in thealveolar zone of the lung. Finally, further simulations were comparedin experiments with a fixed penetration volume and various flow rates.Comparison showed good agreement, which may be explained by the factthat half-width, mode shift, and skewness were little affected by theflow rate.

     

On the role of glucocorticoid receptors in brain plasticity

Summary 1. The mapping of glucocorticoid receptors (GR) in the rat central nervous system (CNS) has demonstrated their widespread presence in large numbers of nerve and glial cell populations also outside the classical stress regions.2. The present paper summarizes the evidence that glucocorticoids via GR in the CNS can act as lifelong organizing signals from development to aging. The following examples are given. (a) In the prepubertal and adult offspring, prenatal corticosterone treatment can produce long-lasting changes in striatal dopaminergic communication. (b) In adulthood, the evidence suggests complex regulation by adrenocortical hormones of neurotrophic factors and their receptors in the hippocampal formation. (c) In aging, the strongly GR-immunoreactive pyramidal cell layer of the CA1 hippocampal area appears to be preferentially vulnerable to neurotoxic actions of glucocorticoids, especially in some rat strains.3. Strong evidence suggests that each nerve cell in the CNS is supported by a trophic unit, consisting of other nerve cells and glial cells, blood vessels, and extracellular matrix molecules. Due to multiple actions on nerve and glial cell populations of the different trophic units, the glucocorticoids may exert either an overall trophic or a neurotoxic action. It seems likely that with increasing age, the endangering actions of glucocorticoids on nerve cells prevail over the neurotrophic ones, leading to reduced nerve cell survival in some trophic units.     

Polymer-bound alkyltriazenes for mild racemization-free esterification of amino acid and peptide derivatives.

A novel tool for polymer-assisted solution phase (PASP) esterification of amino acid and peptide derivatives has been developed. When treated with carboxylic acids, polymer-bound alkyltriazenes react with a loss of nitrogen and transfer of the alkyl moiety to the carboxylate anion to form the corresponding alkyl esters. There are no limitations with regard to either the protecting groups or the nature of the amino acid. Furthermore no racemization occurs at the chiral centers of the amino acids as demonstrated by chiral GC-MS analyses. Alkyltriazene-resins were also applied successfully to the esterification of peptide acids and other peptidic structures, such as tripalmitoyl-S-glyceryl-cysteine (Pam3Cys). The triazene-mediated esterification reaction is exceptionally mild, and there is no need for prior activation of the carboxy groups. This method is therefore particularly suitable for the alkylation of complex peptidomimetic structures prone to racemization and for acid-sensitive structures.     

The evolution of optimal emergence times: bet hedging and the quest for an ideal free temporal distribution of individuals

Proper timing of activities is one of the principal challenges faced by most organisms. Organisms need to account for various aspects in decision making like avoiding inordinate risks, synchronizing with resource availability, or finding mates. We provide analytical and simulation models to investigate the influence of life expectancy, resource competition and unpredictable environmental conditions (environmental uncertainty) on the evolutionarily stable distribution of emergence times in organisms depending on seasonally available resources. We focus on the partitioning of total phenotypic variance in emergence times into 1) genetic variance in mean emergence times between lineages and 2) environmental trait variance that determines the intra‐lineage variance in the timing of emergence. Both, life expectancy of organisms and intensity of competition severely influence the evolutionary response to environmental uncertainty. Our main findings can be summarized as follows: 1) in general diversifying bet hedging (environmental trait variance) is the adequate mechanism to reduce the risk arising from environmental uncertainty while conservative bet hedging, i.e. delaying emergence into ‘safe’ phases of the season is restricted to short lived organisms and to situations with vanishing competition. 2) Environmental trait variance increases with increasing environmental uncertainty whereas 3) significant genetic variance evolves only under severe resource competition; it is driven by selection for an ideal free distribution of emergence times. 4) The level of genetic variance evolving declines with increasing life expectancy of organisms. 5) With sufficiently short life expectancy evolutionary branching and coexistence of distinctly different emergence strategies occurs; the number of co‐occurring strategies is determined by the level of environmental uncertainty. Our model provides cues for understanding how different ecological factors contribute and interact to shape the evolution of emergence strategies.