The effect of serum protein fractions on liposome-cell interactions in cultured cells and the perfused rat liver (original) (raw)
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Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1987
We compared the metabolic fate of [3H]cholesteryl[ 14C]oleate, [3Hlcholesteryi hexadecylether, t2s l-labeled bovine serum albumin and [3H]inulin as constituents of large immunoglobulin-coupled tmilamellar lipid vesicles following their internalization by rat liver macrophages (Kupffer cells) in monolayer culture. Under serum-free conditions, the cholesteryl oleate that is taken up is hydrolyzed, for the greater part, within 2 h. This occurs in the lysosomal compartment as judged by the inhibitory effect of the lysosomotropic agents monensin and chloroquin. After hydrolysis, the cholesterol moiety is accommodated in the cellular pool of free cholesterol and the oleate is reutilized for the synthesis mainly of phospholipids and, to a lesser extent of triacylglycerols. During incubation in plasma, however, substantial proportions of both the cholesterol and the oleate are shed from the cells, predominantly in the tmesterified form. When the liposomes are labeled with the cholesteryl ester analog [3H]cholesteryl hexadecylether only a very small fraction of the label is released from the cells, even in the presence of plasma. Similar to the label remaining associated with the cells, the released label is identified in that case as unchanged cholesteryl ether. The liposomal aqueous phase marker 1251-1abeled bovine serum albumin is also readily degraded intralysosomally and the radioactive label is rapidly released from the cells in a trichloroacetic acid-soluble form. Also, as much as 20% of the aqueous phase marker [3Hlinulin that becomes cell-associated during a 2-h incubation with inulin-containing liposomes, is released from the cells during a subsequent 4-h incubation period in medium or rat plasma. The usefulness of the various liposomal labels as parameters of liposome uptake and intracellular processing is discussed.
International Journal of Pharmaceutics, 1999
The in vitro binding of the serum proteins human serum albumin (HSA), human serum immunoglobulin (IgG) or human serum high density lipoprotein (HDL) on unilamellar liposomes of different lipid composition was studied. HDL bound on liposomes at higher amounts than IgG and IgG at higher amounts than HSA. The protein binding on liposomes decreased when bovine brain monosialganglioside (GM 1) or poly(ethyleneglycol)-distearoylphosphatidylethanolamine (DSPE-PEG) was included in liposome membrane. With all three proteins, an inverse relationship was found between the amount of protein bound on liposomes after 1 h liposome-protein incubation in vitro and the BLOOD/RES ratio of the same liposomes 2 min after i.v. administration in mice (Panagi et al., Drug Dev. Ind. Pharm., 22 (1996) 217-224). The potential value of this in vitro-in vivo correlation, provided it is extended to additional liposome compositions, is that it may provide an in vitro method to screen liposomes in terms of blood clearance rates.
Interaction of immunoglobulin-coupled liposomes with rat liver macrophages in vitro
Experimental Cell Research, 1987
The interaction between liposomes coated with covalently linked rabbit immunoglobulin (RbIg-liposomes), and rat liver macrophages (Kupffer cells) in monolayer culture was studied biochemically with radioactive tracers andmorphologically by electron microscopy. The attachment of immunoglobulin (Ig) to liposomes caused a five-fold increase in liposome uptake by the Kupffer cells at 37°C in comparison with uncoated liposomes. The uptake was linear with time for at least 4 h and linear with liposome concentration up to a lipid concentration of 0.2 mM. At 4°C uptake, probably representing cell surface-bound liposomes, was reduced to a level of approx. 20 % of the 37°C values. Involvement of the Fc receptor in the uptake process was indicated by the reduction of RbIg-liposome uptake by more than 75% as a result of preincubating the cells with heat-aggregated human or rabbit Ig at concentrations (less than 2 mg/ml) at which bovine serum albumin (BSA) had virtually no effect on uptake. At high concentrations (10-35 mg/ml), however, albumin also reduced liposome uptake significantly (20-30%), which suggests an interaction of the RbIg-liposomes with the Kupffer cells that is partially non-specific. RbIg-liposome uptake was dependent on the amount of RbIg coupled to the liposomes. Maximal uptake values were reached at about 200 ug RbIg/umol liposomal lipid. Electron microscopic observations on cells incubated with horseradish peroxidase-containing RbIg-liposomes demonstrated massive accumulation of peroxidase reaction product in intracellular vacuoles, showing that the uptake observed by label association represents true internalization. 0
Liposome disposition in vivo. III. Dose and vesicle-size effects
Biochimica et biophysica acta, 1981
The effect of lipid dose (4,3-512.8 mumol total lipid/kg body weight), administered intravenously as liposomes encapsulating radioactive inulin, upon the ability of mouse organs to bind and/or take-up the radioactive label has been studied in vivo. Three different liposome diameters were investigated: 0.46 micrometers (L), 0.16 micrometers (M) and 0.058 micrometers(S). All liposomes were negatively charged with lipid composition of phosphatidylcholine/phosphatidic acid/cholesterol/alpha-tocopherol in the molar ration 4 : 1 : 5 : 0.1 or 4 : 1 : 1 : 0.05. Overall radioactive label disposition after 2 h was consistent with localization predominantly in the reticuloendothelial system. A saturation of liver with increasing lipid dose was demonstrated for all three sizes, together with a corresponding increase in blood levels. Spleen radioactivity increased with increasing dose of L- and M-liposomes, but decreased for increasing doses of S-liposomes. Levels in residual carcass exhibited n...
Life Sciences, 1980
Small unilamellar neutral, negatively and positively charged liposomes composed of egg phosphatidylcholine, various amounts of cholesterol and, when appropriate, phosphatidic acid or stearylamine and containing 6-carboxyfluorescein were injected into mice, incubated with mouse whole blood, plasma or serum or stored at 4°C. Liposomal stability, i.e. the extent to which 6-carboxyfluorescein is retained by liposomes, was dependent on their cholesterol content. (1) Cholesterol-rich (egg phosphatidylcholine/ cholesterol, 7: 7 molar ratio) liposomes, regardless of surface charge, remained stable in the blood of intravenously injected animals for up to at least 400 min. In addition, stability of cholesterol-rich liposomes was largely maintained in vitro in the presence of whole blood, plasma or serum for at least 90 min. (2) Cholesterol-poor (egg phosphatidylcholine/cholesterol, 7 :2 molar ratio) or cholesterol-free (egg phosphatidylcholine) liposomes lost very rapidly (at most within 2 min) much of their stability after intravenous injection or upon contact with whole blood, plasma or serum. Whole blood and to some extent plasma were less detrimental to stability than was serum. (3) After intraperitoneal injection, neutral cholesterol-rich liposomes survived in the peritoneal cavity to enter the blood circulation in their intact form. Liposomes injected intramuscularly also entered the circulation, although with somewhat diminished stability. (4) Stability of neutral and negatively charged cholesterol-rich liposomes stored at 4°C was maintained for several days, and by 53 days it had declined only moderately. Stored liposomes retained their unilamellar structure and their ability to remain stable in the blood after intravenous injection.
Uptake of small liposomes by non-reticuloendothelial tissues
Biochimica Et Biophysica Acta - Biomembranes, 1987
The distribution of liposomes within the intravascular space and the extent to which they escape into extravascular space strongly impact on the application of lipid vesicles as a carrier for pharmacologically active agents. The present study investigates how intact small unilamellar vesicles (SUV) may be taken up by different tissues after intravenous injection into mice, using various types of SUV with different entrapped markers, lipid composition, size, doses of liposomal llpids and stability in the blood. Our focus was specifically on sphingomyelin (or distearoyl phosphatidylcholine)/cholesteroi (2:1, mol/mol) SUV, which are known to be stable in the blood circulation. Our results indicated that, in addition to the reticuioendothelial tissues, intact SUV were taken up in several other parts of the body, including intestine, skin, carcass and legs. It appears that the accumulation of SUV in the intestine and the skin increases with time post-injection. Furthermore, from the kinetic data, the process of uptake of SUV by the skin and intestine is compatible with a non-saturable pathway, which follows first-order kinetics. This suggests that the cells involved in the uptake of SUV in the intestine and skin are not phagocytic cells, which are normally saturable.
Uptake and processing of immunoglobulin-coated liposomes by subpopulations of rat liver macrophages
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 1988
In rive uptake an0 processing by liver macruphages (Kupffer cells) of liposome~, covalently coated with rabbit immuno~obulln (|g ilposomes) was studied following intrave~us injection in rats. Rabbit Ig iiposomas were labeled with trace ~uneunts of eho|es~ryq UC]oleate and [3Hiehelesteryl hex~ecyl ether. | h after |n|e~ion of the liposomes, the non-perenchymal cells were iso|atod and subjected to cen~fugal elutriafion with stepwise-lncreasing flow rates; thus, five snb-fr~ons of Kupffer edis were obtalned ranging in size from 9 to 14 pm in dhuneter. The cells were assayod for peroxidese ~tivity and protein content° Rabbit |8 |iposomes were taken up preferentially by Kupffer ceils with diameters is~rger than II pin, which constitute less than 2S% of the total Kupffer cell population. The intralysosomal degradation of the ingos~ iiposomas was monitored by measuring the 3H/uC ratio of the cells. Due to the rapid release from the ceils of the [uC~oleate formed from the ¢ho|esteryi[UC]o|eate and the virtual|y complete retention of the non-metabolizab|e [3H]cho|esteryl hexadecyl ether the 3H/UC ratio of the cells incre~es with proceeding hydrelysls of the liposomes. Thus, we were able to show that~ in v|vo, the Kupffer ceDs of the larger s~e classes, are not only mo~e active in liposome u~eke~ but ere also su~tanfially more active in liposome degradation than smaller ceils. The maintenance of the observed heterogeneity of rat Hver Eupffer e,,ll~ with respect to liposome uptake under in vitro culture conditions, was examined. Subfractions were maintained in mono|ayer culture for 2 days and incubated with rabbit |g lipo~me$. Binding and uptake of Hposomes by the cells was monitored by measuring ceil-assucinted radioactivity at 4°C and 37°C, respectively. In contrast to our in vivo rosults, we observed maximal in vitro fiposome binding and uptake in these subfn~cfions containing small cells (10-H pm diameter), while the fractions containing ceils larger than |2/~m, which were more active in vivo, were substantially less active than the smaller ceils. The maximum we observed was even more pronounced when the llposeme concentration was ineroased. We conclude that liver macrephase subfractions that barely participate in liposome uptake from the bloodstream in vivo, possess the potential to develop the capacity in vitro to phagocy|ose rabbit Ig.coated |iposomes to extents equal to or even higher than the cells belonging to those subfractions containing the phagocyticeHy most active cells under in vivo conditions. Abbreviations: Ig, immunogiobulin; Hopes, 4-(2-hydroxyethyl)-l-piperazineethanesulfomc acid; MPB-PE, maleimido-4-(p.phenylbutyryl)phesphatidy!ethanolamine; PBS, phosphatebuffered safine.