Effect of the polar headgroup of phospholipids on their interaction with actin.

It is generally admitted that actin filaments are anchored to a membrane by membranar actin-binding-proteins. However, we found that actin may also interact directly with membrane phospholipids. The actin-phospholipid complex has been investigated at the air-water interface using a film balance technique. In order to probe the effect of the phospholipid headgroup on the actin-phospholipid interaction, we focus mainly on phospholipids that have the same acyl chain length but different headgroups. For all the phospholipids, the apparent area per molecule (the total surface divided by the number of lipid molecules) increases after the injection of the protein into the subphase, which suggests an intercalation of actin between the phospholipid molecules. This effect seems to be more important for DMPE and DMPS than for DMPG, suggesting that the headgroup plays an important role in this intercalation. The critical surface pressure associated to the liquid expanded-liquid condensed (LE-LC) phospholipid transition increases with the concentration of G-actin and thus suggests that G-actin acts as an impurity, simply competing as a surfactant at the air-water interface. On the other hand, F-actin affects the LE to LC transition of phospholipids differently. In this case, the LE to LC transition is broader and F-actin slightly decreases the critical surface pressure, which suggests that electrostatic interactions are involved.

Optimisation of a silicon/silicon dioxide substrate for a fluorescence DNA microarray.

This paper presents a comprehensive theory and experimental characterisation of the modulation of the fluorescence intensity by the construction of optical interferences on oxidised silicon substrates used for DNA microarrays. The model predicts a 90-fold variation of the fluorescence signal depending on the oxide thickness. For a Cy3 dye, the signal is maximal for a 90 nm oxide thickness corresponding to a 7.5-fold enhancement with respect to a standard glass substrate. For experimental validation of the model, we have prepared Si/SiO2 substrates with different parallel steps of decreasing oxide thicknesses on the same sample using a buffered oxide etch (BOE) etching process after thermal oxidation. The SiO2 surface has been functionalized by a silane monolayer before in situ synthesis of L185 oligonucleotide probes. After hybridisation with complementary targets, the variations of the fluorescence intensity versus oxide thickness are in very good accordance with the theoretical model. The experimental comparison against a glass substrate shows a 10-fold enhancement of the detection sensitivity. Our results demonstrate that a Si/SiO2 substrate is an attractive alternative to standard glass slides for the realisation of fluorescence DNA microarrays whenever detection sensitivity is an important issue.

Adsorption of actin at the air-water interface: a monolayer study.

The intrinsic surface activity of the contractile protein actin has been determined from surface tension measurements using the Wilhelmy hanging-plate method. Actin, a very soluble protein, moves from the subphase to the air-water interface to make a film. In the absence of magnesium, actin is monomeric and is known as G-actin. During the compression the monomers change their conformation or orientation at the interface and they are then pushed reversibly into the subphase upon further compression. No collapse occurs. Actin monomers in the presence of magnesium become activated; at concentrations greater than some critical value, actin polymerizes to form filaments of F-actin. The actin filaments have a higher surface activity than the actin monomers either because they are more hydrophobic or because F-actin, a rigid polymer, is much more efficient at creating excluded volume. The actin filaments then form a rigid film at the interface that collapses when the surface area is decreased. At less than the critical concentration, the actin monomers are present in the subphase in their activated form. However, their concentration increases at the interface during film compression until the critical concentration is reached. The surface pressure isotherm in this case has the characteristics of a G-actin film at the beginning of the compression and of an F-actin film at the end of the compression process.

Surface film pressure of actin: interactions with lipids in mixed monolayers.

The interactions of actin with neutral lipid films made from DLPC, and with positively charged films built from DLPC and stearylamine (SA), have been characterized by the monolayer technique. Injection of actin underneath an expanded lipid film produces an increase in the surface pressure that is consistent with a penetration of the lipid molecules by actin. This adsorption of actin to the lipid is more pronounced either with positively charged films or with Mg(2+) present in the sub-phase, suggesting that the mechanism involves an electrostatic attraction. During compression, the actin molecules are squeezed out into the sub-phase, carrying along some lipid molecules; this suggests a strong affinity of the lipids for actin. An analysis of the dilational modulus shows that when actin is found as monomers at the interface, the mixed actin-lipid film undergoes three phase changes upon compression. On the other hand, when actin is polymerized at the interface, the actin and the lipid form a rigid film for which the compressibility is mostly dominated by actin.

Interaction between G-actin and various types of liposomes: A 19F, 31P, and 2H nuclear magnetic resonance study.

We have investigated in the present study the interaction between G-actin and various types of liposomes, zwitterionic, positively charged, and negatively charged. To investigate at the molecular level the conformation of actin in the presence of lipids, we have selectively attached a fluorinated probe, 3-bromo-1,1,1-trifluoropropanone, to the actin cysteine residues 10, 285, and 374 and used high-resolution 19F nuclear magnetic resonance spectroscopy to investigate the probe resonances. The results indicate a change in the mobility of the 19F labels when G-actin is in the presence of positively charged liposomes made of DMPC and stearylamine and in the presence of DMPG, a negatively charged lipid. No conformational change was observed in the actin molecule in the presence of neutral liposomes. Electron micrographs of these systems reveal the formation of paracrystalline arrays of actin filaments at the surface of the positively charged liposomes, while no evidence of actin polymerization or paracrystallization was observed in the presence of DMPG. The interaction between actin and the lipid polar headgroup has also been investigated using solid-state phosphorus and deuterium NMR. The results indicate no evidence of interaction between actin and zwitterionic liposomes but show an interaction between the positively charged liposomes and a negative charge on the actin molecules. Interestingly, the negatively charged liposomes interact with a positive charge, which is most likely associated with the three residues (His-Arg-Lys) preceding the cysteine 374 residue in the protein.

Cardiac arrhythmias during the combined use of intravenous aminophylline and terbutaline in status asthmaticus.

The purpose of this study was to evaluate prospectively the occurrence of cardiac arrhythmias during the combined therapy with intravenous aminophylline and terbutaline in 29 consecutive patients with status asthmaticus. The 24-hour Holter recordings were performed during continuous intravenous infusions of aminophylline (0.56 +/- 0.20 mg/kg/h) and terbutaline (0.034 +/- 0.014 microgram/kg/min). Serum theophylline concentration was 12.1 +/- 3.8 micrograms/ml and never reached the toxic level (greater than 20 micrograms/ml). Premature ventricular contractions (PVCs) were absent in 35 percent of patients and 48 percent had rare unifocal PVCs (less than 10/h). Only 17 percent of patients (five of 29) exhibited severe ventricular arrhythmias: PVCs greater than 10/h (n = 3), multifocal PVCs (n = 1); and a short run of ventricular tachycardia (n = 1). Serious supraventricular arrhythmias occurred in only 7 percent of patients (two of 29) who developed sustained runs of atrial tachycardia. These arrhythmias were always clinically well tolerated and spontaneously resolved without any antiarrhythmic treatment. We conclude that severe arrhythmias are rarely observed during combined therapy with aminophylline and terbutaline in status asthmaticus.

[Acute myocarditis disclosing leptospirosis]. / Myocardite aiguë révélant une leptospirose.

A 44 year old patient presents with acute myocarditis and cardiogenic shock. The evolution is progressively favorable at the price of a residual involvement of the left ventricular function, evolving to a dilated cardiopathy, within three years. The responsibility of an advanced ictero-hemorrhagic leptospirosis is established. The severity of this myocarditis and the revealing characteristics of the leptospirosis are peculiar to this observation which is discussed in terms of data from the literature.

Les hemoglobines des amphibiens: Separation et caracterisation preliminaire des chaines d'une hemoglobine du crapaud bufo bufo.

A toad (Bufo bufo) hemoglobin has been purified and two types of chain have been separated by counter-current distribution. The alpha-chain has a N-terminal sequence Ac-Ala-Leu and a C-terminal sequence Tyr-Arg. The beta-chain has a N-terminal sequence Val-His-Leu and a C-terminal sequence Tyr-His.