Fast atom bombardment tandem mass spectrometric identification of diacyl, alkylacyl, and alk-1-enylacyl molecular species of glycerophosphoethanolamine in human polymorphonuclear leukocytes.

Fast atom bombardment ionization with tandem mass spectrometry of both positive and negative ions is a useful technique for the identification of intact glycerophosphoethanolamine (GPE) phospholipids, providing information as to polar head group and fatty acyl substituents. In the identification of GPE molecular species, positive ion neutral loss scanning for 141 units was attempted to confirm the presence of the phosphoethanolamine polar head group. This scan was found to discriminate against the abundant subclass of phospholipids having an 1-O-alk-1'-enyl linkage, termed plasmalogens, as well as 1-O-alkyl ether species. The neutral loss process is suggested to involve attack of a carbonyl oxygen from either sn-1 or sn-2 on the sn-3 methylene carbon with loss of neutral phosphoethanolamine. Using FAB/MS/MS alone, it is not possible to differentiate between plasmalogens and other 1-O-alkyl ether molecular species having the same molecular weight. The combination of mild acid hydrolysis, which selectively hydrolyzes the labile 1-O-alk-1'-enyl bond, with subsequent FAB/MS/MS distinguished species of these distinct subclasses. Using these techniques and precursor ion scans for the arachidonoyl carboxylate anion, m/z 303, the arachidonic acid containing glycerophosphoethanolamine molecular species were identified and the relative abundance of arachidonoyl plasmalogen, alkylacyl, and 1,2-diacyl GPE molecular species in the human polymorphonuclear leukocyte (neutrophil) was determined to be 75.4%, 12.1%, and 12.5%, respectively. These values were not significantly different from that reported in the literature using conventional methodology.     

The distribution of brominated long-chain fatty acids in sponge and symbiont cell types from the tropical marine spongeAmphimedon terpenensis

The tropical marine spongeAmphimedon terpenensis (family Niphatidae, order Haplosclerida) has previously been shown to possess unusual lipids, including unusual fatty acids. The biosynthetic origin of these fatty acids is of interest as the sponge supports a significant population of eubacterial and cyanobacterial symbionts. The total fatty acid composition of the sponge was analyzed by gas chromatography/mass spectrometry of the methyl esters. Among the most abundant of the fatty acids in intact tissue were 16∶0, 18∶0 and 3,7,11,15-tetramethylhexadecanoic (phytanic) acid. In addition, three brominated fatty acids, (5E,9Z)-6-bromo-5,9-tetracosadienoic acid (24∶2Br), (5E,9Z)-6-bromo-5,9-pentacosadienoic acid (25∶2Br) and (5E,9Z)-6-bromo-5,9-hexacosadienoic acid (26∶2Br) were also present. The three brominated fatty acids, together with phytanic acid, were isolated from both ectosomoal (superficial) and choanosomal (internal) regions of the sponge. Analysis of extracts prepared from sponge/symbiont cells, partitioned by density gradient centrifugation on Ficoll, indicated that phytanic acid and the three brominated fatty acids were associated with sponge cells only. Further, a fatty acid methyl ester sample from intact tissue ofA. terpenensis was partitioned according to phospholipid class, and the brominated fatty acids were shown to be associated with the phosphatidylserine and phosphatidylethanolamine fractions that are commonly present in marine sponge lipids. The phosphatidylcholine and phosphatidylglycerol fractions were rich in the relatively shorter chain fatty acids (16∶0 and 18∶0). The association of brominated long-chain fatty acids (LCFA) with sponge cells has been confirmed. The findings allow comment on the use of fatty acid profiles in chemotaxonomy and permit further interpretation of LCFA biosynthetic pathways in sponges. The assignment of the sponge studied, which is currently placed asA. terpenensis, is being supported to some extent, but the species is unusual in having C₂₅ fatty acids as the major constituent in this group. Other factors, such as season or microenvironmental conditions, may influence observed fatty acid composition which tends to reduce the usefulness of fatty acid profiles as markers in sponge chemotaxonomy.     

Finite element computation of electromagnetic fields [hyperthermiatreatment]

A three dimensional finite element solution scheme is developed for numerically computing electromagnetically induced power depositions. The solution method is applicable to those problems for which it can be reasonably assumed that the magnetic permeability is homogeneous. The method employs an incident field/scattered field approach where the incident field is precalculated and used as the forcing function for the computation of the scattered field. A physically logical condition is used for the numerical boundary conditions to overcome the fact that electromagnetic problems are generally unbounded (i.e., the boundary condition is applied at infinity) but numerical models must have a boundary condition applied to some finite location. At that numerical boundary, an outgoing spherical wave is simulated. Finally, an alternate to a direct solution scheme is described. This alternate method, a preconditioned conjugate gradient solver, provides both a storage and CPU time advantage over direct solution methods. For example, a one-thousand fold decrease in CPU time was achieved for simple test cases. Unlike most iterative methods, the preconditioned conjugate gradient technique used has the important property of guaranteed convergence. Solutions obtained from this finite element method are compared to analytic solutions demonstrating that the solution method is second-order accurate     

Residues essential for the function of SecE, a membrane component of the Escherichia coli secretion apparatus, are located in a conserved cytoplasmic region

Protein export in Escherichia coli is absolutely dependent on two integral membrane proteins, SecY and SecE. Previous deletion mutagenesis of the secE gene showed that only the third of three membrane-spanning segments and a portion of the second cytoplasmic region are necessary for its function in protein export. Here we further define the residues important for SecE function. Alignment of the SecE homologues of various eubacteria reveals that they all contain one membrane-spanning segment, compared with three in E. coli SecE, and that the most conserved region among them lies in their putative cytoplasmic amino termini; little homology exists in their membrane-spanning segments. The SecE homologue of the extreme thermophilic bacterium Thermotoga maritima was cloned and found to complement a deletion of secE in E. coli. Deletion or replacement of the cytoplasmic region of E. coli SecE eliminated SecE function, indicating that this sequence is essential for a functional secretion machinery. Mutant analysis suggests that the most important function of the third membrane-spanning segment is to maintain the proper topological arrangement of the conserved cytoplasmic domain.     

Differential contractile response of cultured microvascular pericytes to vasoactive agents

OBJECTIVE: Microvascular pericytes may contract in two different ways: In the first, a circumferential or radial mechanical force applied at right angles to the long axis of the vessel may constrict the underlying vessel affecting blood flow and transmural pressure. Retraction and elongation of pericyte processes may also occur tangentially and at right angles to the vessel axis and alter microvessel permeability by changing the amount of ablumenal surface covered or the openness of interendothelial junctions. In this study, cultured pericytes were utilized as a model experimental system to determine if vasoactive stimulation changes their shape in a manner consistent with this hypothesis. METHODS: Pericytes cultured from isolated rate capillaries were subjected to angiotensin II and histamine. Their response was monitored by measuring the area of non-yielding substrate covered by the pericytes and the manner in which their shape changed. Shape changes were quantified by calculating the surface area: perimeter perimeter ratios. RESULTS: Histamine significantly reduced surface area covered and the surface area:perimeter ratio. The pericyte processes retracted, resulting in elongated, spindle-shaped cells. These effects were nullified by the H1 blocker diphenhydramine suggesting a receptor-specific response. Angiotensin II also elicited contraction and reduced surface area, but the cells contracted laterally and longitudinally. The surface area: perimeter ratios also decreased. CONCLUSIONS: These results indicate that pericytes are capable of two types of contractile responses in culture, depending on the specific vasoactive stimulus.     

Reduction and uptake of methylene blue from rat air spaces

The use of methylene blue (MB) to estimate dilution of epithelial lining fluid, which occurs during bronchoalveolar lavage (BAL), is complicated by loss of this redox dye from the air spaces. The rate of MB uptake from the air spaces of isolated rat lungs and the effects of oxidation and reduction on this process were investigated in this study. Movement of MB from the air spaces to perfusate was compared with the corresponding transport of 125I-labeled albumin, [14C]-dextran, 99mTc-labeled diethylenetriaminepentaacetate, [3H]-sucrose, and 3H2O. By the end of 2 min, MB concentrations in the BAL had fallen by 58 +/- 4% (SE; n = 11) and 3H2O by 78 +/- 2% (n = 13), whereas concentrations of the other indicators decreased by approximately 6%. All but 10% of the 3H2O lost from the air spaces was found in the perfusate, whereas 19% of the lost MB was not recovered in the perfusate, suggesting retention of MB in the pulmonary tissues. Absorption of MB from the air spaces was slowed by 20% when the lungs were left unperfused, and absorption was accelerated threefold by reduction of MB to leukomethylene blue with Na2S2O4. In contrast, MB losses from the air space were slowed by the oxidizing agent K3Fe(CN)6 and by addition of superoxide dismutase or ascorbic oxidase. It is therefore possible that ascorbic acid and O2- entering the air spaces reduce MB to the uncharged leuko form. Lowering the pH of the BAL fluid to 3.5 also slowed MB reabsorption. This suggests that acid aspiration may stimulate release of oxidants into the air spaces.     

Clinical trial of a new lightwand device (Trachlight) to intubate the trachea

BACKGROUND: Transillumination of the soft tissue of the neck using a lighted stylet (lightwand) is an effective and safe intubating technique. A newly designed lightwand (Trachlight) incorporates modifications to improve the brightness of the light source as well as flexibility. The goal of this study was to determine the effectiveness and safety of this device in intubating the trachea of elective surgical patients. METHODS: Healthy surgical patients were studied. Patients with known or potential problems with intubation were excluded. During general anesthesia, the tracheas were intubated randomly using either the Trachlight or the laryngoscope. Failure to intubate was defined as lack of successful intubation after three attempts. The duration of each attempt was recorded as the time from insertion of the device into the oropharynx to the time of its removal. The total time to intubation (TTI), an overall measure of the ease of intubation, was defined as the sum of the durations of all (as many as three) intubation attempts. Complications, such as mucosal bleeding, lacerations, dental injury, and sore throat, were recorded. RESULTS: Nine hundred fifty patients (479 in the Trachlight group and 471 in the laryngoscope group) were studied. There was a 1% failure rate with the Trachlight, and 92% of intubations were successful on the first attempt, compared with a 3% failure rate and an 89% success rate on the first attempt with the laryngoscope (P not significant). All failures were followed by successful intubation using the alternate device. The TTI was significantly less with the Trachlight compared with the laryngoscope (15.7 +/- 10.8 vs. 19.6 +/- 23.7 s). For laryngoscopic intubation, the TTI was longer for patients with limited mandibular protrusion and mentohyoid distance, with a larger circumference of the neck, and with a high classification according to Mallampatti et al. However, there was no relation between the TTI and any of the airway parameters for Trachlight. There were significantly fewer traumatic events in the Trachlight group than in the laryngoscope group (10 vs. 37). More patients complained of sore throat in the laryngoscope group than in the Trachlight group (25.3% vs. 17.1%). CONCLUSIONS: In contrast to laryngoscopy, the ease of intubation using the Trachlight does not appear to be influenced by anatomic variations of the upper airway. Intubation occasionally failed with the Trachlight but in all cases was resolved with direct laryngoscopy. The failures of direct laryngoscopy were resolved with Trachlight. Thus the combined technique was 100% successful in intubating the tracheas of all patients.     

One-step synthesis of functional Co nanoparticles for surface-initiated polymerization

Much effort is conducted to construct artificial objects that are capable of converting chemical or electromagnetic energy into a specific, predetermined motion on the nanoscale. We present results on the synthesis of core–shell nanoparticles capable to be set in rotation by the application of electromagnetic fields. The nanorotors implied in the study are based on the cobalt nanospheres decorated with a stabilizing brush shell composed of poly(?-caprolactone) that is attached by surface-initiated polymerization. The functional cores used as macroinitiators can be fabricated alternatively by a two-step or a one-step process.