Effects of phorbol esters on endothelial cell microfilaments: Laser scanning confocal microscopy and quantitative morphometry of dose dependent changes

Phorbol esters are known to alter microfilaments but it is not clear if the changes correspond to modulation of the phosphoinositide turnover/protein kinase C system. The novel technique of laser scanning confocal epifluorescence was used to study fiber orientation in phorbol ester treated cells. We treated endothelial cells with control agents and agents known to stimulate protein kinase C: 4 alpha-phorbol, phorbol 12-myristate 13-acetate (PMA), phorbol dibutyrate (PDB), or lipopolysaccharide. After incubation with the test agents, the endothelial cell microfilaments were stained with rhodamine pholloidin and viewed by conventional epifluorescence and by laser scanning confocal epifluorescence microscopy. The images obtained by the confocal microscopy corresponded to a thin optical section through the cells, 300 nm or more in thickness. The microfilaments extended predominantly in the plane of focus. After exposure of the cells to phorbol esters, the stress fibers became more nearly parallel in arrangement or were shortened, but remained in the plane of focus. The modification of microfilaments in response to phorbol esters was quantitated by a single blind analysis. In order to compare the morphological changes with a biochemical action of the phorbol esters, we measured phosphoinositide turnover. The dose-dependence of morphological changes was compared and contrasted to the dose-dependent effect of phorbol esters on bradykinin-stimulated phosphoinositide turnover. PMA had about the same EC50 (1-5 nM) for both biochemical and morphological processes. PDB was less potent in inducing the disruption of microfilament structure than in inhibiting phosphoinositide turnover. Lipopolysaccharide was ineffective in inducing a morphological change under these conditions. A simple activation of protein kinase C is insufficient to explain the dose-dependent effects of phorbol esters. Thus a morphometric analysis can help distinguish the potency of cytoskeleton modulators.     

Linear and non-linear regression analysis of genotype X environment interactions in pearl millet

Summary Regression analysis was computed on the grain yield of 15 single cross F₁ hybrids of pearl millet (Pennisetum typhoides (Burm.) S. & H.) evaluated in 20 environments at 19 sites in India to assess the nature of genotype X environment interactions. Linear, quadratic, cubic, twoand three-intersecting straight line models were examined for fit. The interactions of six hybrids viz. MH 110, MH 113, MH 114, MH 115, MH 120 and MBH 110 were explained by the linear regression model. The response of the remaining nine hybrids was largely non-linear. The two and three-intersecting straight line models fit better than the quadratic and cubic models and explained non-linearity of response. The two-intersecting straight line models fit for 6 hybrids MH 106, MH 107, MH 112, MH 116, MH 117 and BJ 104. The response of MH 109 was best explained by a three-intersecting straight line model, but there still existed a significant remainder variation. The truncation of environmental range by assuming moving division points was more efficient than the fixed division points for the segmental regression models. The stability of hybrid varieties on the best fitting model has been discussed.     

Molecular evolution among someDrosophila species groups as indicated by two-dimensional electrophoresis

Summary The evolutionary and phylogenetic relationships of sevenDrosophila species groups (represented byD. melanogaster, D. mulleri, D. mercatorum, D. robusta, D. virilis, D. immigrans, D. funebris, andD. melanica) were investigated by the use of two-dimensional electrophoresis. The resulting phylogeny is congruent with the current views of evolution among these groups based on morphological characters and immunological distances. Previous studies indicated that the ability of one-dimensional electrophoresis to resolve relationships between distantly related taxa extended to about the Miocene [25 million years (Myr) ago], but the present study demonstrates that two-dimensional electrophoresis is a useful indicator of phylogeny even back to the Paleocene (65 Myr ago). In addition, two-dimensional electrophoresis is shown to be a useful technique for detecting slowly evolving structural proteins such as actins and tropomyosins.     

Sexual dimorphism in the early Miocene species ofProconsul from the Kisingiri Formation of east Africa: A morphometric examination using multivariate statistics

This paper examines the pattern(s) of sexual dimorphism within the upper dentition ofProconsul specimens from the early Miocene of east Africa. These fossils are compared against the corresponding dentition ofPan troglodytes andGorilla gorilla using principal components and cluster analyses. This paper demonstrates that both sexes ofPan andGorilla are characterized by their own distinctive shape patterns. It is also demonstrated that someProconsul specimens examined here display a pattern that is dissimilar from otherProconsul specimens also examined. This suggests that at least two species ofProconsul may have to be recognized as having lived in this region during the early Miocene. The identification of distinct patterns withinProconsul also suggests that their overall shape and size range are more similar toPan than toGorilla.     

Gramicidin in chromatophores of Rhodobacter sphaeroides

Chromatophores of Rhodobacter sphaeroides were excited with light flashes to generate a transmembrane electrical potential difference. The electric relaxation was measured by electrochromic absorption changes as a function of added gramicidin. At low gramicidin/bacteriochlorophyll (BChl) molar ratios the decay of the electrochromic absorption changes showed a biphasic behaviour, with a fast phase relaxing at some s, and a slow phase relaxing at more than 100 ms. This was attributable to a mixture of vesicles containing gramicidin dimers with others containing none. The concentration dependence of this effect was linear. This implied full dimerization of gramicidin. The data were interpreted to yield an average bacteriochlorophyll content per chromatophore of 770(±150) and the conductance of a single gramicidin dimer in the chromatophore membrane of 15(±4) pS (in about 115 mM KCl).Abbreviations BChl Bacteriochlorphyll - tricine N-Tris[hydroxymethyllmethylglycine Offprint requests to: W. Junge     

Montane plant communities of Fynbos Biome

The vegetation of the mountains of the Fynbos Biome is classified and described, mostly on the basis of vegetation structure and higher taxa. Various gradients can be recognized. A gradient in soil conditions runs from coarse textured, nutrient-poor soils in non-grassy fynbos (Mountain Fynbos) through grassy fynbos and grassy non-fynbos (Eastern Fynbos and Grasslands & Grassy Shrubland) to finer textured and less nutrient-poor soils in the non-fynbos Karroid & Renoster Shrubland. Another gradient of significance can be likened to the tropical gradient running from the dry conditions of hot semi-deserts to savannas or grasslands to woodlands. At the dry extreme an open shrubland occurs (Karroid & Renoster Shrubland, Asteraceous Fynbos), at intermediate positions a herbland occurs (Restioid Fynbos) and at the wet extreme another type of shrubland occurs (Ericaceous Fynbos). This kind of moisture gradient occurs on individual mountains with the xeric end being at the base of the north slopes, and the mesic end being on the upper south aspects. Soil characteristics are closely linked to this gradient; the better developed soils being found on the south aspects. Another moisture gradient is apparent in the vegetation. This gradient runs from the mesic southern coastal mountains to the north west where summer droughts are extremely severe although total rainfall may be similar.Nomenclature follows that used in the Government Herbarium, Stellenbosch.I thank William Bond, Richard Cowling, John Rushworth, Eddy van der Maarel and Marinus Werger for critical comments and discussion.     

Electron microprobe analysis of intracellular electrolytes in resting and isoproterenol-stimulated exocrine glands of frog skin

Summary In the intact, in vitro frog skin, isoproterenol (ISO) stimulates and amiloride-insensitive increase in short-circuit current (SCC) that can be localized to the exocrine glands and is associated with secretion of chloride. To determine which cells in the glands respond to stimulation we measured the intracellular electrolyte concentrations of the various cell types of the mucous and seromucous glands of the skin using freeze-dried cryosections and electron microprobe analysis. In the resting state, the various cell types of the glands have intracellular electrolyte concentrations similar to the epithelial cells of the skin. Exposure to amiloride (10^–4 m) has little effect on the concentration of Na and Cl in the cells of the glands. The effect of isoproterenol has two distinct phases. Analysis of glands in tissues frozen at the peak of the SCC response (13 min after addition of isoproterenol) shows that the only significant change is an increase in Na and Ca in a group of cells at the ductal pole of the acini of both gland types. These are termed gland cells. The duct cells and cells that secrete macromolecules did not show any significant changes at this timepoint. In the gland cells, after a one-hour exposure to isoproterenol the Na concentration is at prestimulation levels while Cl drops. There is also a smaller drop in Cl in the duct and skin epithelial cells. Ouabain, which can completely block the isoproterenol SCC response, has little short-term effect on Na and Cl in the control gland but accentuates the gain of Na and drop in Cl in the isoproterenol-treated condition. Bumetanide and, to a lesser extent, furosemide, also blocks the isoproterenol SCC response and causes a further drop in Cl. The results provide indirect evidence that a major portion of the ionic component of the gland secretion is produced by a distinct group of cells separate from those producing the macromolecular component and that the mechanism of secretion involves a Na:Cl coupled transport system linked to the activity of the basolateral Na pump.     

Characterisation of the magnesium status of soils by means of the EUF-Mg fractions

Summary On the basis of pot experiments carried out on soils with different Mg contents using ryegrass, it could be established that Mg uptake by ryegrass is closely related to Mg in the EUF extracts. The Mg contents in the EUF extracts are therefore suitable indices of actually and potentially available Mg in soils.The ratio between Mg in the EUF extracts and Mg deposited on the electrode + filter varies. The absolute amount of Mg in the EUF extracts increases with increasing Mg content of the soils. The higher, however, the total desorbable Mg the lower are the Mg contents in the EUF extracts in comparison with the Mg contents deposited on electrode + filter. Besides total desorbable Mg the EUF-Mg fractions are influenced as well by the organic and inorganic CEC and by soil pH. The amount of EUF filtrate flowing through filter and electrode also has an influence on the amount of Mg in the EUF filtrate.Consequently for a proper evaluation of available soil Mg total EUF-Mg (Mg in the EUF filtrate + Mg deposited on electrode + filter) should be used.     

A highly purified allergen from excretory and secretory products of Dirofilaria immitis

Preceding data revealed that the allergen concentrated mainly in excretory and secretory (ES) products exhausted by adult Dirofilaria immitis. The present paper reported that a highly purified allergen was obtainable from ES products more easily and effectively. An allergen in ES products was purified by a combination of DEAE-Sephadex A-50 chromatography, Sephadex G-200 and Sephadex G-100 gel filtration. The purified preparation was proved to be one protein by sodium dodecyl sulfate (SDS) gel electrophoresis and to be exact the same allergen with the one obtained from the crude extract of adult Dirofilaria worms. The molecular weight of the purified allergen was estimated to be 15,000, and the allergen was inclined to aggregate in the buffered solution.     

Theoretical conformational analysis of phospholipids II. Role of hydration in the gel to liquid crystal transition of phospholipids

To obtain a satisfactory agreement between computed transition temperatures and those determined experimentally, we introduce explicitly water molecules which hydrate the polar headgroup of dipalmitoylphosphatidylethanolamine molecules. The calculated free energy curves as a function of the intermolecular interchain distance and the degree of hydration of the polar groups permit the determination of the transition of the phospholipid system from the gel to the liquid crystalline phase. The detailed structure of the hydration shell is defined using the supermolecular approach.