The effect of block co-polymers on the uptake of model polystyrene microspheres by Kupffer cells--in vitro and in vivo studies.

A major problem in targeting of intravenously injected colloidal particles in the form of microspheres, emulsions and liposomes to sites of inflammation and tumours is their rapid recognition and sequestration by Kupffer cells of the liver and macrophages of the spleen [I]. We have shown previously that the uptake of small polystyrene microspheres (less than 100 nm in diameter) by liver and spleen can be dramatically reduced by means of coating microspheres with polyoxypropylene-polyoxyethylene block copolymers comprising poloxamine and poloxamer series [2,3]. In this communication we describe the effect of poloxamine block copolymers, selected from a range of molecular weight and hydrophobic/hydrophilic properties, on the uptake of polystyrene microspheres by liver non-parenchymal cells both in vifro and in vivo. Polystyrene microspheres (60 nm in diameter) were obtained from Polysciences (U.K.) and sbrface labelled with 125Nal as described previously [2,3]. Coating of microspheres was acheived by preincubation in 1 .O %w/v solutions of selected poloxamines (BASF, U S A ) at room temperature overnight. For in vitro studies, freshly isolated liver non-parenchymal cells were incubated at 37°C for Ihr with microspheres (0.125mg) both in the absence and presence of fresh rat serum [4]. Our in vifro results supports the earlier studies of Van Oss [5] and suggests that in the absence of serum, hydrophobic forces are probably involved on interaction of uncoated latex particles with liver cells. Diminishing the hydrophobicity of particles by coating them with poloxamines. such as poloxamine 908, dramatically reduced their interaction with liver cells in the absence of serum (Table 1 ) . On the other hand, interaction of both uncoated and coated microspheres with liver cells was significantly reduced in the presence of serum when compared to corresponding control incubations (Table 1 ) . In this case, however, serum suppressed the interaction of coated particles by 2-fold in comparison to incubations containing uncoated particles. Centrifugation of serum in Amicon centricon-30 and -100 membrane cones suggested that at least two serum components may be responsible for exerting the dysopsonic effect on uncoated polystyrene particles. A component of molecular weight below 30000. and a second component, more potent in its dysopsonic effect, of molecular weight greater than 100 000. The latter component may probably exert its dysopsonic effect on poloxamine coated microspheres (Table 1 ) . The dysopsonic effect of serum was previously demonstrated on the uptake of polystyrene particles and a number of different bacteria by lung and peritoneal macrophages [6.7,8] and more recently on the uptake of cholesterol-rich liposomes and vesicles composed of saturated phospholipids by Kupffer cells [5,9]. It has previously been suggested that the presence of such inhibitors in serum could prevent macrophages from being killed by excessive binding and ingestion of particles and lethal microorganisms IS]. For in vivo studies, groups of three male Wistar rats (150 f 10 g) were injected intravenously with 10l3 of either uncoated or poloxamine coated’ 251-labelled polystyrene microspheres via the tail vein. Particle biodistribution in organs of the reticuloendothelial system was determined 1 hr following administration [I 01. The hepatic uptake of all particles coated with poloxamines were dramatically reduced in comparison to uncoated particles. For example the hepatic uptake of plain particles (uncoated), poloxamine 904, 908 and 1508 coated particles was 38.6 f 7.1%, 5.5 f 1.8%. 4.9 f 4.2% and 2.3 f 0.1% of injected dose respectively. The corresponding circulatory activities were 2.0 f 0.5%. 22.9 f 6.4%, 35.8 ? 2.3% and 48.9 f 0.3% of injected dose respectively. Preinjection of animals with free polymers 15 min prior to injection of particles had no effect on their biodistribution as compared to control animals, thus, suggesting the inability of the polymers to block the phagocytic function of the Kupffer cells [3]. Table 1 The effect of serum fractions on interaction of polystyrene microspheres with liver non-parenchymal cells in suspension