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C R Alving

Publications and source records attributed to C R Alving.

At least 127 records · Page 7Linked to original sources
PubMed1983

Interactions of lipid a and liposome-associated lipid A with Limulus polyphemus amoebocytes.

Lipid A or lipid A fractions and liposomes containing lipid A were tested for the ability to gel Limulus amoebocyte lysates and for effects on intact Limulus amoebocytes. Liposomes having a relatively low concentration of lipid A did not produce coagulation of lysate and were designated as Limulus-negative, but liposomes having a high concentration of lipid A were Limulus-positive. Limulus-negative liposomes had no effect on intact amoebocytes. Limulus-positive liposomes caused a striking transformation in the appearance of amoebocytes in that the cells sent out long filamentous extensions that formed a tangled network of processes between cells. The filamentous projections were similar to those that have been previously observed in the presence of gram-negative bacteria. We conclude that amoebocytes have the ability to recognize Limulus-positive liposomes, but the lack of activation of Limulus lysate or the absence of amoebocyte recognition does not prove the absence of liposomal lipid A. We also found that individual lipid A fractions were heterogeneous in their ability to gel lysate. Of eight fractions tested, one (fraction 1) had no detectable activity above the background, and the seven others had activity that ranged from 10-fold to 10,000-fold above the background. The heterogeneity of lipid A fractions detected in assays with amoebocyte lysate was consistent with the finding of heterogeneity in other functional assays of lipid A fractions.

Animals↗
PubMed1983

Antibodies reactive with liposomal phospholipids are produced during experimental Trypanosoma rhodesiense infections in rabbits.

Antibodies against phospholipids appeared spontaneously during the course of experimental Trypanosoma rhodesiense infections in rabbits. These antibodies were observed in rabbits infected either with a lethal strain or with a strain newly discovered to give a spontaneous self-cure. Serum antibodies reacting with liposomes containing dimyristoyl phosphatidylcholine (DMPC), phosphatidylinositol (Pl), phosphatidylinositol phosphate (PlP), or cardiolipin were detected at 3 to 4 wk by complement-mediated release of trapped marker from liposomes. Antibodies were also detected against a trypanosomal lipid fraction (TrF2) that contained Pl as a major constituent. The antibody activities against DMPC, Pl, or TrF2 all reacted (or cross-reacted) with DMPC, and were removed from the serum by adsorbing with liposomes containing DMPC as the only phospholipid. Phosphocholine inhibited the antibodies reactive with liposomes containing either DMPC or DMPC and Pl as phospholipids. Antibodies against PlP, however, reacted only with liposomes containing PlP and were not removed by adsorbing with liposomes lacking PlP. We conclude that anti-phospholipid antibodies appear during the course of trypanosomal infections that either undergo apparent self-cure or are lethal, and at least two anti-phospholipid antibody specificities can be detected.

Animals↗
PubMed1983

The malaria merozoite surface: a 140,000 m.w. protein antigenically unrelated to other surface components on Plasmodium knowlesi merozoites.

We previously identified three proteins on the surface of merozoites (140,000, 105,000 and 75,000 m.w.). To determine if 140,000 m.w. protein was related to other surface proteins, we immunized mice with liposomes containing merozoite proteins from the 140,000 m.w. region of the polyacrylamide gel. The immune sera reacted with the surface of viable merozoites and acetone-fixed schizonts by immunofluorescence. The sera immunoprecipitated only the 140,000 m.w. protein from surface-labeled merozoites. We demonstrated that monoclonal antibody 13C11 immunoprecipitated a 250,000 m.w. protein from metabolically labeled schizonts and bound to the merozoite surface. This monoclonal antibody immunoprecipitated the 75,000 and lower m.w. proteins from surface-labeled merozoites but did not bring down the 140,000 m.w. protein. Because the mouse immune sera did not immunoprecipitate the 250,000 m.w. protein from metabolically labeled schizonts or proteins other than the 140,000 m.w. protein from surface-labeled merozoites, we conclude that the 140,000 m.w. protein is unrelated to other merozoite surface antigens identified to date. The mouse immune sera against the 140,000 m.w. protein on the merozoite surface did not immunoprecipitate a 140,000 m.w. protein from metabolically labeled schizonts. Instead, the major protein immunoprecipitated had a m.w. of 144,000. By analogy to the 250,000 m.w. protein and its cleavage products, we propose that the 140,000 m.w. protein on the merozoite surface is a cleavage product of the higher m.w. protein.

Animals↗
PubMed1982

Membrane lipid composition modulates the binding specificity of a monoclonal antibody against liposomes.

A hybridoma secreting a monoclonal IgM 'anti-liposome' antibody was produced after injecting a mouse with liposomes containing dipalmitoylphosphatidylcholine, cholesterol, dicetyl phosphate, and lipid A. The antibody was selected by assaying for complement-dependent damage to liposomes lacking lipid A. The monoclonal antibody reacted best with liposomes containing the original immunizing mixture of lipids. Deletion of individual lipid constituents from liposomes diminished the ability of the liposomes to bind (adsorb) the antibody. Binding of the antibody was enhanced by including lipid A or galactosylceramide in the lipid bilayer, or by substituting egg phosphatidylcholine for dimyristoyl- (or dipalmitoyl-) phosphatidylcholine. Sphingomyelin could be substituted for dimyristoylphosphatidylcholine without altering the adsorption of antibody. Although the monoclonal anti-liposome antibody was completely inhibited by phosphocholine, it was probably not a conventional anti-phosphocholine antibody. The antibody apparently had a partial specificity for phosphate, and was inhibited by glycerophosphocholine, glycerophosphate, sodium phosphate, sodium sulfate, and inositol hexaphosphate, but not by choline or inositol.

Animals↗
PubMed1981

Interaction of C-reactive protein with liposomes. III. Membrane requirements for binding.

We have previously presented a model for CRP-membrane interactions using liposomes composed of dimyristoyl phosphatidylcholine (DMPC), cholesterol (CHOL), stearylamine (SA), and galactosyl ceramide. We have shown that the interaction of cRP with these liposomes in the presence of human serum results in consumption of hemolytic C and membrane lysis. In the present paper we have directly examined the binding reaction between CRP and liposomes using radiolabeled CRP. We have found that this binding is more characteristic of CRP interactions with polycations than CRP interactions with phosphocholine- (PC) containing molecules. CRP binding to liposomes was dependent on the presence of SA in the membrane and could occur with dimyristoyl phosphatidylethanolamine in place of DMPC. The binding was not inhibited by ethylenediaminetetraacetate (EDTA) but could be inhibited by CaCl2, whereas CRP binding to PC-Sepharose was inhibited by EDTA and required CaCl2. We have further examined the effects of changes in membrane composition on CRP binding to liposomes. In liposomes with a limiting density of SA, we found increased CRP binding with changes in composition that would increase membrane fluidity. In most cases, the amount of CRP binding correlated with the amount of C activation observed previously. However, increasing the amount of CHOL in the membrane was found to increase C activation while decreasing CRP binding. These findings indicate that CRP binding to membranes and subsequent C activation can occur through cationic molecules as well as phospholipids.

Amines↗
PubMed1980

Influence of temperature on complement-dependent immune damage to liposomes.

Maximal release of trapped liposomal glucose, in the presence of saturating amounts of liposomal antigen (galactocerebroside), antiserum (anti-galactocerebroside), and complement, was dependent on temperature. At lower temperatures (20--25 degrees C), maximal glucose release was inversely related to liposomal phospholipid fatty acyl chain length (dimyristoyl phosphatidylcholine > dipalmitoyl phosphatidylcholine > distearoyl phosphatidylcholine > sphingomyelin). At higher temperatures (32--35 degrees C) a limiting plateau of glucose release, at approx. 60%, was reached, or approached, by all preparations. Sphingomyelin liposomes still released less glucose than those prepared from other phospholipids, even at 35 degrees C. The titers of antiserum and complement (ABL50/ml and CL50/ml) were dependent on temperature, and differences based on liposomal phospholipid fatty acyl chain length were observed. Analysis of antiserum and complement-dependence on temperature, and on phospholipid type, revealed that although antibody binding to galactocerebroside undoubtedly was subject to steric hindrance due to interference by surrounding phospholipids at 20--25 degrees C, steric hindrance did not play a major role in blocking antibody binding above 32 degrees C.

Antibodies↗
PubMed1980

Binding of diphtheria toxin to phospholipids in liposomes.

Diphtheria toxin bound to the phosphate portion of some, but not all, phospholipids in liposomes. Liposomes consisting of dimyristoyl phosphatidylcholine and cholesterol did not bind toxin. Addition of 20 mol% (compared to dimyristoyl phosphatidylcholine) of dipalmitoyl phosphatidic acid, dicetyl phosphate, phosphatidylinositol phosphate, cardiolipin, or phosphatidylserine in the liposomes resulted in substantial binding of toxin. Inclusion of phosphatidylinositol in dimyristol phosphatidylcholine/cholesterol liposomes did not result in toxin binding. The calcium salt of dipalmitoyl phosphatidic acid was more effective than the sodium salt, and the highest level of binding occurred with liposomes consisting only of dipalmitoyl phosphatidic acid (calcium salt) and cholesterol. Binding of toxin to liposomes was dependent on pH, and the pattern of pH dependence varied with liposomes having different compositions. Incubation of diphtheria toxin with liposomes containing dicetyl phosphate resulted in maximal binding at pH 3.6, whereas binding to liposomes containing phosphatidylinositol phosphate was maximal above pH 7. Toxin did not bind to liposomes containing 20 mol% of a free fatty acid (palmitic acid) or a sulfated lipid (3-sulfogalactosylceramide). Toxin binding to dicetyl phosphate or phosphatidylinositol phosphate was inhibited by UTP, ATP, phosphocholine, or p-nitrophenyl phosphate, but not by uracil. We conclude that (a) diphtheria toxin binds specifically to the phosphate portion of certain phospholipids, (b) binding to phospholipids in liposomes is dependent on pH, but is not due only to electrostatic interaction, and (c) binding may be strongly influenced by the composition of adjacent phospholipids that do not bind toxin. We propose that a minor membrane phospholipid (such as phosphatidylinositol phosphate or phosphatidic acid), or that some other phosphorylated membrane molecule (such as a phosphoprotein) may be important in the initial binding of diphtheria toxin to cells.

Cardiolipins↗
PubMed1980

Liposomes as vehicles for vaccines.

Lipid A from Shigella flexneri LPS, or acylated derivatives of muramyl dipeptide (MDP), were incorporated into the lipid bilayer of liposomes to enhance the adjuvanticity of the liposomes in rabbits. Liposomes containing lipid A induced antibodies against lipid A, phosphocholine, phosphatidylcholine, and sphingomyelin. Lipid A was resolved into eight fractions, some of which did, and others of which did not, induce antibodies against liposomes. Anti-liposome antibodies also were induced (in the absence of liposomes) by complete Freund's adjuvant, and by acid treated bacterial cells coated with lipid A, but were not induced either by lipid A or liposomes alone, or by liposomes containing acylated MDP. We tested the liposomes for the ability to enhance the immunogenicity of a protein antigen. The antigen consisted of liposomes having acylated MDP and ganglioside GM1 (the receptor for cholera toxin, or CT) in the lipid membrane, and CT bound to the outer surface of the liposomes. The liposomal antigen (having surface-bound CT) produced a much greater anti-CT titer than that obtained by injection of CT alone. We conclude that liposomes containing only phosphatidylcholine, cholesterol, and dicetyl phosphate are poorly immunogenic, but that antibodies against them can be induced by inclusion of lipid A. Liposomes that are appropriately formulated can strongly enhance the immunogenicity of a liposome-bound protein antigen.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1980

Effects of lipid A and liposomes containing lipid A on platelet and fibrinogen production in rabbits.

The effect of the lipid A moiety of endotoxin on platelet and fibrinogen production was studied in rabbits. Lipid A was infused intravenously in doses ranging from 1 to 100 micrograms/kg body mass; 18 hr later, selenomethionine-75Se was injected intravenously and its incorporation into fibrinogen and platelets determined. Lipid A in saline stimulated fibrinogen and platelet production, but the dose required was 50--100 times that required for an intact endotoxin. Although lipid A solubilized in triethylamine (TEA) was at least 60 times more active in the Limulus amebocyte lysate assay than was lipid A suspended in saline, the sensitivity of platelet and fibrinogen production to solubilized lipid A was increased only twofold. Incorporation of lipid A into liposomes had no effect on its Limulus activity. Lipid A in liposomes continued to stimulate platelet, but not fibrinogen, production. Leukopenia that was induced by lipid A in TEA did not occur when rabbits received the same dose of lipid A in liposomes. Lipid A, like intact endotoxin, can stimulate platelet and fibrinogen production and induce leukopenia but the doses required are high. The low solubility of lipid A in aqueous solutions may be only one factor that determines its biologic activity.

Animals↗
PubMed1979

Sporozoite-induced malaria: therapeutic effects of glycolipids in liposomes.

Liposomes containing neutral glycolipids with a terminal glucose or galactose, when injected intravenously, prevented the appearance of erythrocytic forms of malaria (Plasmodium berghei) in mice previously injected with sporozoites. Inhibitory glycolipids included glucosyl, galactosyl, or lactosyl ceramide. Inhibition was not observed with liposomes containing ceramide, phosphocholine ceramide, sulfogalactosyl ceramide (sulfatide), or ganglioside GM1. Liposomes containing glycolipids did not inhibit infection transmitted by injecting blood containing erythrocytic stages of malaria. These results may have therapeutic implications in the treatment of malaria. Analysis of the mechanism of interference with the life cycle of malaria by liposomal glycolipids may yield information about the interactions of parasites with cellular membranes.

Animals↗
PubMed1979

Cholesterol-dependent tetanolysin damage to liposomes.

Tetanolysin caused membrane damage, resulting in release of trapped glucose from liposomes containing cholesterol. Maximum glucose release occurred from liposomes that contained 50 mol% cholesterol. At higher or lower levels of cholesterol, glucose release was reduced and glucose release did not occur at all below 40 mol% cholesterol. The apparent activity of tetanolysin was not influenced by temperature (24 degrees C compared to 32 degrees C) or by liposomal phospholipid fatty acyl chain length. We conclude that tetanolysin caused cholesterol-dependent lysin-mediated damage to liposomes, possibly by means of a pore consisting of a complex of toxin and cholesterol.

Bacterial Toxins↗
PubMed1979

Choleragen (cholera toxin): a bacterial lectin.

Choleragen (cholera toxin) agglutinated erythrocytes and liposomes containing the toxin receptor, galactosyl-N-acetylgalactosaminyl-(N-acetylneuraminyl)-galactosylglucosylceramide (ganglioside GM1). Cells that had been exposed to GM1 were agglutinated, but agglutination was not observed when cells had been exposed to other gangliosides (GM2, GM3, GD1a, GD1b). Choleragen-dependent agglutination of liposomes was slightly less specific, because liposomes containing either GM1 or GD1b, but neither GM2, GD1a, nor GM3 were agglutinated. The oligosaccharide isolated from GM1 inhibited both the agglutination of cells and liposomes containing GM1 and the binding of choleragen to liposomes containing GM1. Galactose and sialic acid were less effective inhibitors of liposomal agglutination and did not inhibit cellular agglutination or binding of choleragen to liposomes. Liposomal agglutination was dependent on choleragen concentration and occurred with the B but not the A protomer of choleragen. These results suggest that choleragen, through its binding to the oligosaccharide portion of a glycolipid, exhibits lectinlike activity, which results in agglutination of liposomes and erythrocytes.

Agglutination↗
PubMed1979

Lipid A from endotoxin: antigenic activities of purified fractions in liposomes.

Isolation of lipid A by acid hydrolysis of Shigella flexneri lipopolysaccharide resulted in a product that consisted of a heterogeneous mixture of bands when visualized by thin layer chromatography. Differential extraction with ethyl acetate and chloroform, or extraction with EDTA, followed by chloroform-methanol-water (Bligh-Dyer extraction), or a combination of both extraction schemes, resulted in partial purification of immunologically active lipid A. Eight fractions were purified further by preparative thin layer chromatography, and each of the fractions had phosphate, carbohydrate, and esterified fatty acids. Upon incorporation into liposomes, five of the eight purified fractions reacted with anti-lipid A serum, but the three fractions with the most number of esterified fatty acids failed to react with anti-lipid A serum. At least one fraction that originally was unreactive with anti-lipid A serum became reactive as a hapten inhibitor upon removal of esterified fatty acids by alkaline hydrolysis. Alkali-treated fractions from "unreactive" and "reactive" lipid A had similar activities as hapten inhibitors. Our data suggest that lipid A can exist in multiple forms that differ by the number and placement, and possibly by the type, of fatty acids linked to the carbohydrate of lipid A. Highly acylated forms of lipid A do not react with antiserum against the unpurified lipid A mixture, but removal of fatty acids does expose immunoreactive groups.

Animals↗