Preparation of asymmetric,cerebroside sulfate-containing phospholipid vesicles

A procedure is described which inserts asymmetrically cerebroside sulfate (‘sulfatide’) into the outer leaflet of bilayered phospholipid vesicles. Cerebroside sulfate is adsorbed onto a cellulose, filter-paper support and, when incubated with phosphatidylcholine vesicles is transferred to and inserted into the outer leaflet of these vesicles. This transfer occurs at, or above the transition temperature of the phospholipid and follows a similar pattern with small or larger (‘fused’) unilamellar vesicles. The transfer is linear with time for 1–2 h and is maximal after about 6 h, when the sulfatide content reaches about 6 mol% of the total quantity of phospholipid, corresponding to about 10 mol% of the phospholipids present in the outer layer. Initial rates of sulfatide transfer were somewhat increased when the vesicles contained a positively charged lipid (e.g. stearylamine) and decreased when this lipid was negatively charged (e.g. dicetyl phosphate) or hydrophobic (e.g. cholesterol). Divalent ions markedly inhibited sulfatide transfer and monovalent ions did so to a lesser degree. Once incorporated into the outer leaflet of the vesicle, the sulfatide could not be removed by washing with buffer, 1 M NaCl or 1 M urea.     

Comparative titration of arginyl residues in purified D-β-hydroxybutyrate apodehydrogenase and in the reconstituted phospholipid-enzyme complex

In order to titrate and understand the role of arginyl residues of D-β-hydroxybutyrate dehydrogenase, arginyl specific reagents: butanedione, 1,2-cyclohexanedione and phenylglyoxal were incubated with three different forms of the enzyme; native enzyme (inner mitochondrial membrane bound), purified apoenzyme (phospholipid -free) and phospholipid-enzyme complex (reconstituted active form).After complete inactivation of the enzyme by [¹⁴C]-phenylglyoxal, the number of modified arginyl residues was different: one with the lipid-free apoenzyme and three with the phospholipid-enzyme complex, suggesting a conformational change of the enzyme triggered by the presence of phospholipids.After exhaustive chemical modification either of the apoenzyme or of the phospholipid-enzyme complex with [¹⁴C]-phenylglyoxal, four arginyl residues were titrated indicating that these residues are located in the hydrophilic part of the enzyme, not interacting with phospholipids.Reconstituted enzyme inactivated by butanedione could no longer bind a pseudosubstrate (succinate) which indicates that an arginyl residue is involved in the enzyme-substrate complex formation.The values of second order rate constants of D-β-hydroxybutyrate dehydrogenase inactivation by butanedione and 1,2-cyclohexanedione were unchanged with the three enzyme forms, suggesting that phospholipids are not involved in the substrate binding mechanism.     

Die Rezeptoren an den ersten Antennen vonLeptestheria dahalacensis Rüppel (Crustacea,Conchostraca)

Zusammenfassung Bei dem ConchostracenLeptestheria dahalacensis kommen auf den ersten Antennen etwa 600 gleich aussehende Sinneshaare vor, die in Gruppen von jeweils 25–30 zusammengefaßt sind. Diese Sinneshaare sind in zwei Teile gegliedert, die durch das lichtmikroskopisch gut sichtbare Basalstück (basal bead) voneinander getrennt sind. Dieses bildet die Basis des Haares, dessen Wand im wesentlichen aus Epicuticula besteht. Apikal wird das Haar durch das Endkügelchen (terminal pellet) abgeschlossen. Das Basalstück wird von der untersten Lage der Epicuticula gebildet. Die 4–10 Receptorcilien, die jeweils einzeln ebensovielen Dendriten aufsitzen, ziehen aus dem inneren Teil des Rezeptors, der von insgesamt 5 Hüllzellen umgeben wird, durch das Basalstück, in dem sie stark eingeengt werden und verzweigen sich dann im äußeren Teil des Rezeptors. Sie ziehen bis zum Endkügelchen, in das sie durch einen Porus, den man als Häutungsporus ansprechen kann, eintreten. In der Häutungsvorbereitung wird der Haarbalg von der Hüllzelle 5, das Basalstück von der Hüllzelle 4, der Haarschaft dagegen von der Hüllzelle 3 gebildet. Dabei spaltet sich die Hüllzelle 3 ringspaltförmig auf, so daß in diesem Spalt der neuangelegte Haarschaft handschuhfingerförmig eingestülpt liegt. Die Hüllzelle 2 formt die Spitze des neuen Haares, während die Dendritenscheide von der Hüllzelle 1 abgegeben wird.

---

The receptors on the first antennae ofLeptestheria dahalacensis Rüppel (Crustacea, Conchostraca)

Summary On the antennulae ofLeptestheria dahalacensis (Conchostraca) nearly 600 sensory setae of one type are found. They are gathered in groups of 25–30. The single sensory seta is divided into two parts by the basal bead which is easily visible in the light microscope. The basal bead is the socket of the seta, whose wall is mainly built up by the epicuticle. The terminal pellet closes the tip of the seta. The basal bead is derived from the innermost layer of the epicuticle. 4–10 dendrites each with one receptorcilium innervate the receptor. The receptorcilia stretch through the interior part of the receptor and the basal bead into the exterior part, where they branch. They enter the terminal pellet in a porus, which seems to be a moulting porus. The interior part of the receptor is surrounded by 5 sheath cells. During the premoult it becomes obvious, that the socket of the seta is built by the sheath cell 5, the basal bead by the sheath cell 4 and the shaft by the sheath cell 3. For this the sheath cell 3 is divided into two parts. Between this two parts the newly formed cuticle is invaginated. The sheath cell 2 formes the tip and the sheath cell 1 the cuticular sheath of the new bristle.

     

Organophosphate poisoning in the developed world – A single centre experience from here to the millennium

Organophosphate (OP) poisoning is still associated with high morbidity and mortality rates, both in resource-poor settings and in well-developed countries. Despite numerous publications dealing with this particular poison, detailed clinical data on more severe overdoses with these agents are relatively sparsely reported. A retrospective study was consequently conducted on 33 patients with OP poisoning admitted to our intensive care unit (ICU) to provide additional data on clinical features. We included moderate to severe poisonings between 2000 and 2012 who required admission to ICU.     

Transmembrane myosin chitin synthase involved in mollusc shell formation produced in Dictyostelium is active

Several mollusc shells contain chitin, which is formed by a transmembrane myosin motor enzyme. This protein could be involved in sensing mechanical and structural changes of the forming, mineralizing extracellular matrix. Here we report the heterologous expression of the transmembrane myosin chitin synthase Ar-CS1 of the bivalve mollusc Atrina rigida (2286 amino acid residues, M.W. 264 kDa/monomer) in Dictyostelium discoideum, a model organism for myosin motor proteins. Confocal laser scanning immunofluorescence microscopy (CLSM), chitin binding GFP detection of chitin on cells and released to the cell culture medium, and a radiochemical activity assay of membrane extracts revealed expression and enzymatic activity of the mollusc chitin synthase in transgenic slime mold cells. First high-resolution atomic force microscopy (AFM) images of Ar-CS1 transformed cellulose synthase deficient D. discoideumdcsA⁻ cell lines are shown.     

Surfactant protein D increases phagocytosis and aggregation of pollen-allergen starch granules

Recent studies have shown that surfactant components, in particular the collectins surfactant protein (SP)-A and -D, modulate the phagocytosis of various pathogens by alveolar macrophages. This interaction might be important not only for the elimination of pathogens but also for the elimination of inhaled allergens and might explain anti-inflammatory effects of SP-A and SP-D in allergic airway inflammation. We investigated the effect of surfactant components on the phagocytosis of allergen-containing pollen starch granules (PSG) by alveolar macrophages. PSG were isolated from Dactylis glomerata or Phleum pratense, two common grass pollen allergens, and incubated with either rat or human alveolar macrophages in the presence of recombinant human SP-A, SP-A purified from patients suffering from alveolar proteinosis, a recombinant fragment of human SP-D, dodecameric recombinant rat SP-D, or the commercially available surfactant preparations Curosurf and Alveofact. Dodecameric rat recombinant SP-D enhanced binding and phagocytosis of the PSG by alveolar macrophages, whereas the recombinant fragment of human SP-D, SP-A, or the surfactant lipid preparations had no effect. In addition, recombinant rat SP-D bound to the surface of the PSG and induced aggregation. Binding, aggregation, and enhancement of phagocytosis by recombinant rat SP-D was completely blocked by EDTA and inhibited by d-maltose and to a lesser extent by d-galactose, indicating the involvement of the carbohydrate recognition domain of SP-D in these functions. The modulation of allergen phagocytosis by SP-D might play an important role in allergen clearance from the lung and thereby modulate the allergic inflammation of asthma.     

The role of cutaneous feedback for anticipatory grip force adjustments during object movements and externally imposed variation of the direction of gravity

Grip force adjustments to changes of object loading induced by external changes of the direction of gravity during discrete arm movements with a grasped object were analyzed during normal and anesthetized finger sensibility. Two subjects were seated upright in a rotatable chair and rotated backwards into a horizontal position during discrete movements with a hand-held instrumented object. The movement direction varied from vertical to horizontal inducing corresponding changes in the direction of gravity, but the orientation of the movement in relation to the body remained unaffected. During discrete vertical movements a maximum of load force occurs early in upward and late in downward movements; during horizontal movements two load force peaks result from both acceleratory and deceleratory phases of the movement. During performance with normal finger sensibility grip force was modulated in parallel with fluctuations of load force during vertical and horizontal movements. The grip force profile adopted to the varying load force profile during the transition from the vertical to the horizontal position. The maximum grip force occurred at the same time of maximum load force irrespective of the movement plane. During both subjects' first experience of digital anesthesia the object slipped from the grasp during rotation to the horizontal plane. During the following trials with anesthetized fingers subjects substantially increased their grip forces, resulting in elevated force ratios between maximum grip and load force. However, grip force was still modulated with the movement-induced load fluctuations and maximum grip force coincided with maximum load force during vertical and horizontal movements. This implies that the elevated force ratio between maximum grip and load force does not alter the feedforward system of grip force control. Cutaneous afferent information from the grasping digits seems to be important for the economic scaling of the grip force magnitude according to the actual loading conditions and for reactive grip force adjustments in response to load perturbations. However, it plays a subordinate role for the precise anticipatory temporal coupling between grip and load forces during voluntary object manipulation.