Large liposomes containing ganglioside GM1 accumulate effectively in spleen

Large liposomes, with a composition of egg phosphatidylcholine, cholesterol and ganglioside GM1, prepared by an extrusion method, were injected intravenously into mice. After 24 h, up to 50% of injected dose was accumulated in spleen compared with about 15% in spleen for liposomes containing no GM1. The effect of GM1 on spleen accumulation of liposomes was liposome size dependent. Only relatively large liposomes (d greater than 300 nm) showed high accumulation; smaller liposomes were progressively less accumulated. The spleen accumulation increased with increasing injection dose of the liposomes. It was noted that the enhanced uptake by spleen was accompanied by a decrease in the liver uptake, but the total uptake of liposomes by liver and spleen was not dependent on the diameter of liposome or the presence of the ganglioside GM1. Autoradiographs of fixed and sectioned spleen using 125I-labeled tyraminylinulin as a content marker for the liposomes, showed that liposomes localized at the reticular meshwork of the red pulp. These results suggest that larger liposomes containing GM1 are filtered by the spleen during the circulation in blood. The smaller ones with a mean diameter of less than 100 nm are not retained by the filter. The function of GM1 is to prevent liposomes from a rapid uptake by the liver so that liposomes may circulate through the spleen and be filtered. These results, together with the observation that the liposome-entrapped proteins were degraded by the spleen, suggest the potential use of these liposomes for specific drug delivery to the spleen.     

Modulation of the in vitro activity of lysosomal phospholipase A1 by membrane lipids

Lysosomal phospholipases play a critical role for degradation of cellular membranes after their lysosomal segregation. We investigated the regulation of lysosomal phospholipase A1 by cholesterol, phosphatidylethanolamine, and negatively-charged lipids in correlation with changes of biophysical properties of the membranes induced by these lipids. Lysosomal phospholipase A1 activity was determined towards phosphatidylcholine included in liposomes of variable composition using a whole-soluble lysosomal fraction of rat liver as enzymatic source. Phospholipase A1 activity was then related to membrane fluidity, lipid phase organization and membrane potential as determined by fluorescence depolarization of DPH, 31P NMR and capillary electrophoresis. Phospholipase A1 activity was markedly enhanced when the amount of negatively-charged lipids included in the vesicles was increased from 10 to around 30% of total phospholipids and the intensity of this effect depended on the nature of the acidic lipids used (ganglioside GM1相似文献   

Inhibitory effect of cholesterol on the uptake of liposomes by liver and spleen

The effect of cholesterol content of small unilamellar (SUV) and reverse phase (REV) liposomes on blood clearance and tissue distribution has been studied. [¹⁴C]Inulin has been used as an aqueous marker of liposomes to represent the uptake of intact liposomes in tissues. The blood clearance of the intravenously-injected SUV and REV liposomes depends on the cholesterol content of liposomes. The cholesterol-free (0 mol%) liposomes are cleared more readily from the circulation than the cholesterol-poor liposomes (20 mol%) and the cholesterol-poor are cleared more rapidly than the cholesterol-rich (46.6 mol%) liposomes. This clearance pattern of liposomes from the circulation is not attributed to the change of size of liposomes due to the increase in cholesterol content of liposomes. However, poor stability of cholesterol-free or cholesterol-poor liposomes in the circulation is partly responsible, but the predominant factor responsible for the observed blood clearance pattern is the inhibitory effect of cholesterol on the uptake of liposomes by reticuloendothelial-rich tissues liver and spleen. Uptake of liposomes by these organs is decreased with increasing cholesterol content of vesicles. It is suggested that to produce liposome preparations with a long circulating half life in vivo it is necessary to inhibit their uptake by liver and spleen.     

Receptor-mediated uptake of asialoganglioside liposomes: sub-cellular distribution of the liposomal marker in isolated liver cell types

The use of asialo GM1-containing small unilamellar liposome preparations in vivo caused a 2.8-fold increase in the uptake by the liver as compared with the control (neutral) preparations (without asialo GM1). The uptake of negatively charged dicetylphosphate and dipalmitoyl phosphatidic acid-containing small unilamellar liposomes was found to be 1.6-and 1.8-fold respectively higher than that of the neutral preparations. In studies with isolated liver cell types, inhibition of the galactosylated liposome uptake by asialofetuin indicated a possible involvement of hepatic galactose receptors in the recognition of asialo GM1 liposomes by the hepatic parenchymal cells, which in turn were found to be mainly responsible for the enhanced incorporation of these liposomes in the liver. Sub-cellular distribution studies with isolated liver cell types indicated lysosomal localization of the liposomes both in parenchymal and nonparenchymal cells, and it has been proposed that the asialo GM1 liposomes are cointernalized with asialofetuin through a common lysosomal route of ligand internalization.     

Effect of the lipid composition of liposomes on their clearance from the circulation and accumulation in the liver of mice

The effect of phospholipid and ganglioside liposome composition on the liposome clearance from the circulation and accumulation in the mouse liver has been studied. It has been shown that liposomes constituting of liver lipids are quicker removed from the circulation and accumulated in the liver. Liver gangliosides increased liposome uptake by the mouse liver and their clearance from the circulation.     

The effect of blood on the uptake of liposomal lipid by perfused rat liver

Liposomes have been prepared from dipalmitoylphosphatidylcholine (DPPC) and its mixtures with phosphatidylinositol (PI) and stearylamine. The absorption of the liposomes by perfused rat liver has been studied as a function of blood level (0-7% haematocrit). It has been found that the rate constant for uptake of liposomes (perfusion constant, kp) is markedly reduced by addition of blood to the perfusate particularly in the haematocrit range 0-3%. The perfusion constant is dependent on the liposome composition and decreases with incorporation of PI and increases with incorporation of stearylamine into DPPC liposomes, but is independent of the initial size of the liposomes in the range of weight-average diameter from 40-400 nm. The possible effects of blood components on the liposomes and their subsequent absorption by the liver are discussed.     

Phosphatidylinositol promotes cholesterol transport and excretion

Administration of phosphatidylinositol (PI) to New Zealand White rabbits increases HDL negative charge and stimulates reverse cholesterol transport. Intravenously administered PI (10 mg/kg) associated almost exclusively with the HDL fraction in rabbits. PI promoted an increase in the hepatic uptake of plasma free cholesterol (FC) and a 21-fold increase in the biliary secretion of plasma-derived cholesterol. PI also increased cholesterol excretion into the feces by 2.5-fold. PI directly affects cellular cholesterol metabolism. In cholesterol-loaded macrophages, PI stimulated cholesterol mass efflux to lipid-poor reconstituted HDL. PI was about half as effective as cAMP at stimulating efflux, and the effects of cAMP and PI were additive. In cultured HepG2 cells, PI-enriched HDL also enhanced FC uptake from HDL by 3-fold and decreased cellular cholesterol synthesis and esterification. PI enrichment had no effect on the selective uptake of cholesterol esters or on the internalization of HDL particles. PI-dependent metabolic events were efficiently blocked by inhibitors of protein kinase C and the inositol signaling cascade.The data suggest that HDL-PI acts via cell surface ATP binding cassette transporters and signaling pathways to regulate both cellular and intravascular cholesterol homeostasis.     

The role of beta2-glycoprotein I in liposome-hepatocyte interaction

Adsorption of serum proteins to the liposomal surface plays a critical role in liposome clearance from the blood. The aim of this study was to investigate the role of liposome-adsorbed serum proteins in the interaction of liposomes with hepatocytes. We analyzed the serum proteins adsorbing to the surface of differently composed small unilamellar liposomes during incubation with human or rat serum, and found that one protein, with a molecular weight of around 55 kDa, adsorbed in a large amount to negatively charged liposomes containing phosphatidylserine (PS) or phosphatidylglycerol (PG). The binding was dependent on the liposomal charge density. The approximately 55-kDa protein was identified as beta2-glycoprotein I (beta2GPI) by Western blotting. Despite the high affinity of beta2GPI for strongly negatively charged liposomes, in vitro uptake and binding experiments with isolated rat hepatocytes, Kupffer cells or liver endothelial cells, and with HepG2 cells showed no enhancing effect of this protein on the association of negatively charged liposomes with any of these cells. On the contrary, an inhibitory effect was observed. We conclude that despite abundant adsorption to negatively charged liposomes, beta2GP1 inhibits, rather than enhances, liposome uptake by liver cells.     

Uptake and biodistribution of free and liposomally incorporated lipopolysaccharide of aeromonas salmonicida administered via different routes to rainbow trout: (Oncorhynchus mykiss)

Abstract

The effects on uptake and biodistribution of radiolabelled lipopolysaccharide (LPS) due to changing routes of administration, encapsulation of LPS within liposomes and altering liposomal surface charge were examined in rainbow trout (Oncorhynchus mykiss). ³H-labelled LPS, positively- and negatively-charged (¹⁴C-labelled) liposomes or ¹⁴C-labelled liposomes containing ³H-LPS were administered to trout via intravenous, intraperitoneal, intramuscular, or oral routes. Twenty-four hours following administration, relative uptake of LPS and multilamellar vesicles (MLV) based on detection of ³H and ^1AC, respectively, was determined in samples taken from the kidney, spleen, liver, plasma, blood cells and skeletal muscle. In general, regardless of the route of administration, ³H-LPS, ^1AC-MLV and liposomally encapsulated LPS were recovered primarily in the kidney and spleen. Intravenous administration resulted in the greatest uptake of radiolabel by the kidney and spleen, followed by the intraperitoneal and intramuscular routes. Although oral administration yielded the lowest overall uptake of labelled material, detection of ³H and ¹⁴C in the liver was enhanced when compared with the other routes. Negatively-charged MLV were delivered more efficiently to the kidney and spleen than positively-charged MLV; but negatively- and positively-charged MLV containing LPS demonstrated the opposite relationship between charge and distribution among the kidney and spleen. These results suggest that liposomal encapsulation (particularly within positively-charged MLV) enhances delivery of LPS to the primary hemopoietic organs in rainbow trout.     

Effect of Macrophage Elimination Using Liposome-Encapsulated Dichloromethylene Diphosphonate on Tissue Distribution of Liposomes

Abstract

Effect of macrophage elimination using liposomal dichloromethylene diphosphonate (C1₂MDP)1 on tissue distribution of different types of liposomes was examined in mice. Intravenously administration into mice with CI2MDP encapsulated in liposomes composed of phosphatidylcholine, cholesterol and phosphatidylserine exhibits a temporary blockade of liver and spleen function for liposome uptake. At a low dose of 90 (ig/mouse, the liposome uptake by the liver was significantly decreased. Such decrease was accompanied by an increase in liposome accumulation in either spleen or blood depending on liposome composition and size. Direct correlation between the administration dose of liposomal CI2MDP and the liposome circulation time in blood was also obtained even for liposomes with an average diameter of more than 500 nm. These results indicate that temporary elimination of macrophages of the liver and spleen using liposomal CI2MDP may prove to be useful to enhance the drug delivery efficiency of liposomes.     

Phosphatidylinositol promotes cholesterol transport in vivo

To examine the role that lipoprotein charge plays in cholesterol metabolism in vivo, we characterized the effects of an intravenous injection of 40 micromol of an uncharged phospholipid (phosphatidylcholine, PC) or an anionic phospholipid (phosphatidylinositol, PI) into fasted rabbits. PC injection had a negligible effect on lipoprotein charge and composition, similar to that observed in a saline-injected animal. In contrast, PI injection caused a significant increase in the net negative surface charge of all lipoproteins after only 10 min, followed by a gradual return to normal by 24 h. Lipoprotein compositional analysis showed that PI caused a significant increase of cholesteryl ester (CE) and cholesterol (FC) in the VLDL pool by 3 h, with no changes in VLDL-triglyceride content. While the bulk of the plasma CE was located in the HDL pool in the PC-injected animals, in the PI animals, VLDL became the major CE storage compartment. No major changes in the levels or composition of HDL or LDL were evident over the 24-h turnover period. Co-injection of [(3)H]FC revealed a 30-fold greater rate of clearance of the labeled cholesterol from the PI-injected rabbit plasma. In addition, the rate of cholesterol esterification by lecithin:cholesterol acyltransferase was almost completely inhibited in the PI animals. In summary, a bolus injection of PI into rabbits appears to enhance the mobilization of cellular sterol and promote a rapid clearance of both FC and CE from the plasma compartment. The data show that lipoprotein charge can affect cholesterol transport and that this process can be selectively manipulated.     

Lactosylceramide-induced stimulation of liposome uptake by Kupffer cells in vivo

Incorporation of 8 mol percent lactosylceramide into small unilamellar vesicles consisting of cholesterol and sphingomyelin in an equimolar ratio and containing [³H]inulin as a marker resulted in an increase in total liver uptake and a drastic change in intrahepatic distribution of the liposomes after intravenous injection into rats. The control vesicles without glycolipid accumulated predominantly in the hepatocytes, but incorporation of the glycolipid resulted in a larger stimulation of Kupffer-cell uptake (3.2-fold) than of hepatocyte uptake (1.2-fold). Liposome preparations both with and without lactosylceramide in which part of the sphingomyelin was replaced by phosphatidylserine, resulting in a net negative charge of the vesicles, were cleared much more rapidly from the blood and taken up by the liver to higher extents. The negative charge had, however, no influence on the intrahepatic distributions. The fast hepatic uptake of the negatively charged liposomes allowed competition experiments with substrates for the galactose receptors on liver cells. Inhibition of blood clearance and liver uptake of lactosylceramide-containing liposomes by $\text{N-}\text{acetyl-}\text{d}\text{-galactosamine}$ indicated the involvement of specific recognition sites for the liposomal galactose residues. This inhibitory effect of $\text{N-}\text{acetyl-}\text{d}\text{-galactosamine}$ was shown to be mainly the result of a decreased liposome uptake by the Kupffer cells, compatible with the reported presence of a galactose specific receptor on this cell type (Kolb-Bachofen et al. (1982) Cell 29, 859–866). The difference between the results on sphingomyelin-based liposomes as described in this paper and those on phosphatidylcholine-based liposomes as published previously (Spanjer and Scherphof (1983) Biochim. Biophys. Acta 734, 40–47) are discussed.     

The effect of polyphosphoinositides and phosphatidic acid on the phosphatidylinositol transfer protein from bovine brain: a kinetic study

The phosphatidylinositol transfer protein from bovine brain (PI-TP) has lipid transfer characteristics which make it well suited to maintain phosphatidylinositol (PI) levels in intracellular membranes (Van Paridon, P.A., Gadella, Jr., T.W.J., Somerharju, P.J. and Wirtz, K.W.A. (1987) Biochim. Biophys. Acta 903, 68-77). Using a continuous fluorimetric transfer assay we have investigated in what way phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidic acid (PA) affect the transfer activity of this protein in model systems. The effects were analysed by application of a kinetic model which yielded the association constant (K) and dissociation rate constant (k-) for the PI-TP/vesicle complex. Incorporation of PA, PIP and PIP2 into the phosphatidylcholine-containing vesicles increased the association constant solely by diminishing the dissociation rate constant. This effect could be completely accounted for by changes in the membrane surface charge density. In contrast to the inhibitory effect of PA, the inhibition caused by PIP2 was completely abolished by the addition of neomycin, in agreement with the observed preferential binding of this polyamine antibiotic to PIP2. A rise in pH from 5.5 to 8 drastically reduced the association constant for vesicles containing 16 mol% PA (e.g., from 38 to 2 mM-1), without affecting the Vmax. This effect could be mainly attributed to an increase in the negative charge on PI-TP (isoelectric point 5.5), resulting in an enhanced repulsion. Increasing the negative membrane surface charge at pH 7.4 had the opposite effect. This is interpreted to indicate that the membrane interaction site on PI-TP must be positively charged, overcoming the repulsive forces between PI-TP and the vesicle. Addition of PIP2 micelles as a third component in the transfer assay strongly inhibited PI-TP transfer activity. The extent of inhibition suggests a very high affinity of PI-TP for this lipid.     

Tissue distribution of EDTA encapsulated within liposomes containing glycolipids or brain phospholipids.

Multilameller liposomes were prepared with various asialoglycolipids, gangliosides, sialic acid, or brain phospholipids in the liposome membrane and with ethylenediaminetetraacetic acid (EDTA) encapsulated in the aqueous compartments. The liposomes containing glycolipids or sialic acid were prepared from a mixture of phosphatidylcholine, cholesterol, and one of the following test substances: galactocerebroside, glucocerebroside, galactocerebroside sulfate, mixed gangliosides, monosialoganglioside GM1, monosialoganglioside GM2, monosialoganglioside GM3, disialoganglioside GD1a, or sialic acid. The liposomes containing brain phospholipids were mixtures of either sphingomyelin and cholesterol or a brain total phospholipid extract and cholesterol. Distributions of 14C-labeled EDTA were determined in mouse tissues from 15 min to 6 h or 12 h after a single injection of liposome preparation. Liver uptake up encapsulated EDTA was lowest from all liposome preparations containing sialic acid or sialogangliosides, regardless of the amount of sialic acid moiety present or the identity of the particular ganglioside; highest uptake of encapsulated EDTA by liver was from liposomes containing galactocerebroside or brain phospholipids. Lungs and brain took up the largest amounts of EDTA from liposomes containing sphingomyelin and lesser amounts from liposomes containing GD1a. Use of mouse brain phospholipid extract to prepare liposomes did not increase uptake of encapsulated EDTA by the brain. EDTA in liposomes containing monosialogangliosides, brain phospholipids, galactocerebroside, or sialic acid was taken up well by spleen and marrow. Highest thymus uptake of encapsulated EDTA was from liposomes containing GD1a. These results demonstrate that inclusion of sialogangliosides in liposome membranes decreases uptake of liposomes by liver, thus making direction of encapsulated drugs to other organs more feasible. Liposomes containing glycolipids also have potential uses as probes of cell surface receptors.     

The absorption of phospholipid vesicles by perfused rat liver depends on vesicle surface charge

Two types of sonicated vesicle have been prepared from dipalmitoylphosphatidylcholine (DPPC) by incorporation of phosphatidylinositol (PI) to give negatively charged vesicles and stearylamine to give positively charged vesicles. The absorption of the vesicles by rat liver has been investigated by perfusion techniques. A steady state of vesicle absorption is rapidly established in approx. 2 min and the initial rates of absorption decrease with PI content of the vesicles and increase with stearylamine content. In the steady state, the uptake of vesicles by the liver is similarly dependent on vesicle charge, being inhibited by PI and enhanced by incorporation of stearylamine in the vesicles. Fractionation of the liver into subcellular fractions following perfusion showed that most of the vesicular lipid could be found associated with a nuclear (plus plasma membrane) fraction. The suppression of vesicle absorption by PI may be of value as a means of bypassing the liver in relation to the use of vesicles as a delivery system.     

Localization of sterically stabilized liposomes in Klebsiella pneumoniae-infected rat lung tissue: influence of liposome characteristics.

Sterically stabilized liposomes are able to localize at sites of infection and could serve as carriers of antimicrobial agents. For a rational optimization of liposome localization, the blood clearance kinetics and biodistribution of liposomes differing in poly(ethylene glycol) (PEG) density, particle size, bilayer fluidity or surface charge were studied in a rat model of a unilateral pneumonia caused by Klebsiella pneumoniae. It is shown that all liposome preparations studied localize preferentially in the infected lung compared to the contralateral non-infected lung. A reduction of the PEG density or rise in particle size resulted in a higher uptake by the mononuclear phagocyte system, lower blood circulation time and lower infected lung localization. Differences in bilayer fluidity did not affect blood clearance kinetics or localization in the infected lung. Increasing the amount of negatively charged phospholipids in the liposome bilayer did not affect blood clearance kinetics, but did reduce localization of this liposome preparation at the site of lung infection. In conclusion, the degree of localization at the infected site is remarkably independent of the physicochemical characteristics of the PEG liposomes. Substantial selective liposome localization can be achieved provided that certain criteria regarding PEG density, size and inclusion of charged phospholipids are met. These properties seem to be a direct consequence of the presence of the polymer coating operating as a repulsive steric barrier opposing interactions with biological components.     

Interaction of cells from murine organs with liposomes of various composition

The distribution of liposomes prepared from total mouse liver lipids and containing (3H)-labelled platelet activation factor in mouse organs was studied. It was shown that the majority of intraperitoneally injected liposomes prepared from total mouse liver lipids were transported to mouse liver and spleen. The interaction of liposomes with spleen cells in vitro revealed that the affinity of liposomes prepared from total spleen macrophage or total spleen lymphocyte lipids for mouse spleen cells was much higher than that of liposomes prepared from a model lipid mixture. The liposome binding to isolated spleen macrophages or lymphocytes was much higher than the liposome uptake by these cells in the total population of mouse spleen cells.     

Germacrone improves liver fibrosis by regulating the PI3K/AKT/mTOR signalling pathway

Liver fibrosis is a primary threat to public health, owing to limited therapeutic options. Germacrone (GM) has been shown to exert various curative effects against human diseases, including liver injury. The aim of this study was to investigate the pharmacological effects of GM in the pathophysiology of hepatic fibrosis and determine its potential mechanisms of action. A liver fibrosis rat model was established via carbon tetrachloride (CCl₄) treatment, and LX-2 cells were stimulated with TGF-β1. The effects of GM on liver fibrosis and its relationship with the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signalling pathway were investigated. In the CCl₄ fibrosis-induced rat model, GM improved histological damage, inhibited the activity of hepatic α-smooth muscle actin and improved serum alanine aminotransferase and aspartate aminotransferase levels in a dose-dependent manner. GM potently inhibited hepatic stellate cells (HSCs) growth and epithelial–mesenchymal transition (EMT) progression, as reflected by the altered expression of proliferative (Ki-67, PCNA and cleaved caspase-3) and EMT-related (E-cadherin and vimentin) proteins. In TGF-β1-stimulated LX-2 cells, GM significantly inhibited the survival and activation of HSCs and induced cell apoptosis. GM also suppressed the migration ability and reversed the EMT process in HSCs. Following GM treatment, the phosphorylation of the PI3K, AKT and mTOR proteins was reduced in the liver of CCl₄-treated rats and TGF-β1-stimulated LX-2 cells, indicating that GM may attenuate hepatic fibrosis via the PI3K/AKT/mTOR signalling pathway. These outcomes highlight the anti-fibrotic effects of GM and suggest that it is a potential therapeutic agent for the treatment of liver fibrosis.     

Parameters affecting the liposome-mediated insertion of RNA into eucaryotic cells in vitro

The effect of various parameters on the liposome-mediated insertion of RNA into eucaryotic cells in vitro has been studied. Maximization of the insertion of liposome-encapsulated RNA into cells was approached at three levels: (1) alteration of liposome membrane composition, (2) alteration of the recipient cell membrane, and (3) manipulation of the conditions of liposome-cell cocultivation. (1) Changes in liposome membrane composition failed to affect the amount of RNA sequestered within liposomes but did alter the efficiency and mode of liposome uptake by human epithelial carcinoma cells, rabbit spleen lymphocytes, and carrot protoplasts. Addition of lysolecithin to the liposome membrane enhanced the cellular uptake of liposome-sequestered RNA by a “fusion” mechanism (uptake in the presence of cytochalasin B), while addition of cholesterol was inhibitory. (2) Uptake of liposome-sequestered RNA was enhanced when (a) cells were in the mitotic phase of the cell cycle; (b) cells were pretreated with cholesterol-free liposomes; and (c) cells were treated with Piracetam (2-oxo-1-pyrrolidine acetamide). The increased cellular uptake of liposomes appeared in most cases to be due to enhanced cell membrane fluidity. (3) Liposome uptake by cells was directly proportional to the time of liposome-cell cocultivation and to cell number. Increasing doses of liposomes resulted in a reduction of the percentage of RNA uptake, possibly due to a saturation phenomenon. When several of the investigated parameters were simultaneously maximized, as high as 20% of the liposome-sequestered RNA was inserted into human epithelial carcinoma cells.     

The role of β2-glycoprotein I in liposome-hepatocyte interaction

Adsorption of serum proteins to the liposomal surface plays a critical role in liposome clearance from the blood. The aim of this study was to investigate the role of liposome-adsorbed serum proteins in the interaction of liposomes with hepatocytes. We analyzed the serum proteins adsorbing to the surface of differently composed small unilamellar liposomes during incubation with human or rat serum, and found that one protein, with a molecular weight of around 55 kDa, adsorbed in a large amount to negatively charged liposomes containing phosphatidylserine (PS) or phosphatidylglycerol (PG). The binding was dependent on the liposomal charge density. The ∼55-kDa protein was identified as β₂-glycoprotein I (β₂GPI) by Western blotting. Despite the high affinity of β₂GPI for strongly negatively charged liposomes, in vitro uptake and binding experiments with isolated rat hepatocytes, Kupffer cells or liver endothelial cells, and with HepG2 cells showed no enhancing effect of this protein on the association of negatively charged liposomes with any of these cells. On the contrary, an inhibitory effect was observed. We conclude that despite abundant adsorption to negatively charged liposomes, β₂GP1 inhibits, rather than enhances, liposome uptake by liver cells.