Exogenous surfactant as a drug delivery agent.

Pulmonary surfactant is a complex mixture of lipids and several specific surfactant proteins, which together render it with unique spreading properties and a dynamic surface tension behavior. These characteristics are heralded as ideal for a carrier of choice to instil therapeutic agents into the lung, because this combination enables high local therapeutic levels while minimizing systemic side-effects of the instilled agent. This review outlines the rationale to use exogenous surfactant in lung injury, including opening-up inaccessible regions of the lung to other therapeutic agents. Especially the combination of anti-microbial agents and surfactant offers an alternative for critically ill patients with pneumonia. Some caution is also indicated in combining surfactant with antibiotics without proper evaluation of possible interactions. Some other applications for surfactant as a carrier are discussed. Overall, the benefits of surfactant as a carrier warrants clinical trials and promises additional therapeutic tools for the clinician.

Ventilator-induced lung injury leads to loss of alveolar and systemic compartmentalization of tumor necrosis factor-alpha.

OBJECTIVES: To determine the effect on compartmentalization of the tumor necrosis factor (TNF)-alpha response in the lung and systemically after ventilation with high peak inspiratory pressure with and without positive end-expiratory pressure (PEEP). DESIGN AND SETTING: Prospective, randomized, animal study in an experimental laboratory of a university. SUBJECTS AND INTERVENTIONS: 85 male Sprague-Dawley rats. Lipopolysaccharide was given intratracheally or intraperitoneally to stimulate TNF-alpha production; control animals received a similar amount of saline. Animals were subsequently ventilated for 20 min in a pressure control mode with peak inspiratory pressure/PEEP ratio of either 45/0 or 45/10 (frequency 30 bpm, I/E ratio 1:2, FIO2 = 1). MEASUREMENTS AND RESULTS: Blood gas tension and arterial pressures were recorded at 1, 10, and 20 min after start of mechanical ventilation. After killing of the animals pressure-volume curves were recorded, and bronchoalveolar lavage (BAL) was performed for assessment of protein content and the small/large surfactant aggregate ratio. TNF-alpha was determined in serum and BAL. TNF-alpha levels were significantly increased after lipopolysaccharide stimulation; furthermore ventilation without PEEP resulted in a significant shift of TNF-alpha to the nonstimulated compartment as opposed to ventilation with a PEEP level of 10 cmH2O. CONCLUSIONS: Ventilation strategies which are known to induce ventilation-induced lung injury (VILI) disturb the compartmentalization of the early cytokines response in the lung and systemically. Furthermore, the loss of compartmentalization is a two-way disturbance, with cytokines shifting from the vascular side to the alveolar side and vice versa. A ventilation strategy (PEEP level of 10 cmH2O) which prevents VILI significantly diminished this shift in cytokines.

Synthesis and antiproliferative activity of cytidine-5'-alkylphosphonophosphates and structurally related compounds.

The chemical synthesis of cytidine-5'-alkyl- and cytidine-5'-alkyl (acyl)deoxyglycerophosphonophosphates is reported. The compounds obtained represent a novel class of cytostatically active agents based on phospholipids, which inhibit the growth of various tumor cell lines in vitro. They are phosphono analogs of the cytidine-5'-diphosphate-diacylglycerol (CDP-DAG) possessing a structurally modified lipid moiety and a phospholipase C-resistant P-C bond. The antiproliferative efficacy of the cytidine-5'-alkylphosphonophosphates strongly depends on the alkyl chain length. The cytidine-5'-hexadecylphosphonophosphate was found to be the most effective compound tested in this study. Its cytostatic effect was distinctly higher than that of the alkyl(acyl) deoxyglycero derivatives and of the corresponding diphosphates. The structure of the new compounds were confirmed by fast atom bombardment mass spectrometry (FAB). The FAB fragmentation pattern is discussed.

Quantitative reconstitution of isolated influenza haemagglutinin into liposomes by the detergent method and the immunogenicity of haemagglutinin liposomes.

The reconstitution of influenza virus haemagglutinin into liposomes from lipid/protein/detergent mixtures by detergent removal provides vesicles that are similar in structure to viral particles. The dissociation properties of haemagglutinin aggregates and the molar ratio of lipid to protein in the starting mixture are the key factors for the individual and total yield of protein incorporation into liposomes. Structural properties of the detergent used as well as special reconstitution conditions are of minor importance for the formation of haemagglutinin liposomes. As determined by radial immunodiffusion-, haemolysis- and fusion experiments, specific properties of haemagglutinin were maintained to a large extent on liposomal incorporation, but its immunogenicity is increased, if the antigen is incorporated into the lipid bilayer of liposomes.

[Binding and entrapment of insulin by lecithin-phosphatidic acid liposomes in acid solutions]. / Bindung und Einschluss von Insulin durch Lecithin-Phosphatidsäure-Liposomen in saurer Lösung.

Microvesicles made of mixtures of lecithin and phosphatidic acid were precipitated by insulin in acid solution. Insulin was bound by the precipitates up to saturation, that was governed by the portion of phosphatidic acid in the liposomal matrix. The amount of bound insulin was described by a binding isotherm for a bivalent ligand. In neutral solutions the precipitates were repeptised thereby releasing bound insulin. An increase of the particle size, observed by gel filtration analysis, was explained by liposome fusion. If the liposomes were generated in acid solutions of insulin by sonication, up to 50% of the insulin remained entrapped even in the neutralized solution. Especially with cholesterol as a lipid component the entrapped insulin was highly protected from the proteolytic attack by pronase.

Preparation and properties of GnRH-loaded multilamellar liposomes.

Binding of the decapeptide gonadotropin releasing hormone (GnRH) to multilamellar liposomes was investigated with variation of the lipid matrix and of the medium conditions. It is demonstrated, that binding capacity of liposomes may be varied within wide limits if negatively charged phosphatidic acid is made a constituent part of the liposomes. Binding to liposomes is protecting GnRH against enzymatic hydrolysis by proteases, if structural integrity of liposomes is maintained. These properties are in favour of the application of liposomes as a drug carrier system for GnRH. The determination of the binding capacity and of the association constant for a definite system GnRH/lipid and, additionally, comparison of these data with analogous results concerning substance P [Pharmazie 39, 765 (1984)] furnish some new information with respect to the interaction of oppositely charged peptides and lipids.

Substance P: binding to unilamellar and multilamellar liposomes made from mixtures of phosphatidylcholine and phosphatidic acid.

Binding of the undecapeptide substance P to unilamellar and multilamellar lipid vesicles, made from mixtures of phosphatidylcholine and phosphatidic acid, was investigated, because liposomes may be supposed to become a useful pharmaceutical tool for the delivery of low molecular mass peptide hormones. It is demonstrated that binding is largely electrostatic in nature, and that the amount of bound peptide may be varied within rather broad limits by changing the lipid matrix and/or the aqueous medium conditions. Concerning the distribution of the peptide between inner and outer surfaces of the liposomes, large differences may be realised too, as revealed by enzymatic hydrolysis of the outside bound portions of substance P. But additionally, electrostatic binding is demonstrated also to bring about peculiar, unexpected properties of the system. These imply special relevance to a potential pharmaceutical application of substance P liposomes as well as some general relevance to in vivo application of liposomes as drug carriers.

[The image of the public health office in the public consciousness of a small city]. / Das Image des Gesundheitsamtes im Bewusstsein der Bevölkerung einer Kleinstadt.

130 inhabitants of a small town (12,500 inhabitants) in Baden-Württemberg, Germany, were asked by health office employees about their opinion on the public health office and their knowledge of its tasks. The results of the poll show that there is some knowledge regarding the tasks of the public health office in the population; however, this knowledge is confined to a few points. As a result of the poll, the population has a neutral or rather positive point of view of the public health office. An intensification of the public relations several scopes of duties e.g. the scope of the pollution control, seems necessary to achieve the proper position in the image of the population.

Radioiodination of S-type lipopolysaccharide. Possibilities and limitations of labelling of the carbohydrate chain by periodate oxidation.

S-type Lipopolysaccharide (LPS) from E. coli O 139:K 82 (B) was radiolabelled by coupling 125I-tyramine to the carbohydrate chain of LPS after oxidation of sugar residues carrying vicinal hydroxy groups, with sodium metaperiodate. The specific activity amounted to 1.3 kBq/microgram LPS. As LPS is a mixture of molecules with different carbohydrate chain lengths that are associated to large micelles of high kinetic stability, the preparation and the properties of the tracer exhibit some peculiarities. Most important, partial cleavage of the carbohydrate chains of LPS is caused on radioiodination by the procedure described. As a consequence, the tracer differs markedly from the starting LPS with respect to carbohydrate chain length distribution. Another feature of special interest is the variation of the specific activity (radioactivity/unit mass) for LPS molecules of different carbohydrate chain lengths. Because of the kinetic stability of LPS micelles, the equilibration of radioiodinated and non-radioactive LPS aggregates requires their solubilization by a detergent and its subsequent removal by dialysis. It could be demonstrated that short-chain LPS molecules predominantly associate to larger micelles than long-chain molecules. Regardless of these restrictions, the 125I-LPS proved to be useful for certain analytical purposes.

Incorporation of the cytochrome P-450 monooxygenase system into large unilamellar liposomes using octylglucoside, especially for measurements of protein diffusion in membranes.

Cytochrome P-450 and NADPH cytochrome P-450 reductase were incorporated into large unilamellar lipid vesicles (200-300 nm in diameter) removing octylglucoside from mixed micelles by dialysis. The large size of the protein-containing liposomes guarantees a negligibly small vesicle tumbling. Such large vesicles are better suited for studies of protein rotation in reconstituted membranes than vesicles prepared by use of bile salts. At present the octyl-glucoside reconstituted monooxygenase system seems to be the most appropriate model for studying protein-protein and protein-lipid interactions in liver microsomes due to the similarity with respect to the main structural and functional properties, including size.

Preparation and properties of large octylglucoside dialysis/adsorption liposomes.

The suitability and capacity of the polystyrene resin Wofatit EP 60 for the adsorption of octylglucoside, Triton X-100, Cholate, and CHAPS was studied. Optimal detergent/bead ratios and the maximum capacity of Wofatit EP 60 for the four detergents were determined as prerequisites for optimal application of the beads in liposome preparations. It is shown that large unilamellar liposomes can be prepared easily, quickly and cost-effectively using a combined dialysis/adsorption method with octylglucoside as detergent and Wofatit EP 60 and adsorbing polystyrene beads. Structure, composition, size, homogeneity, lamellarity, stability, internal volume, and residual octylglucoside concentration were studied by gel chromatography, radioactive assay, dynamic light scattering and electron microscopy. Vesicle size and homogeneity depend on lipid concentration, lipid composition, cholesterol content, and the rate of octylglucoside removal, but not on the detergent/lipid ratio. The reliability of the method and the properties of the vesicles are compared with those of other methods and researchers.