The objective of this study was to determine the cellular uptake of stavudine (an approved drug for AIDS treatment) by human monocyte/macrophages (U 937). The effect of lipid used, cholesterol concentration and the presence of charge on the liposome bilayer, on the cellular uptake by monocyte/macrophages was investigated. Liposomes employed in the study were prepared by reverse phase evaporation. The lipids egg PC, DMPC, DPPC, DSPC, DMPG and sphingomyelin were employed in this study. The effect of cholesterol on cellular uptake was studied by using liposomes containing a constant amount of lipid and varying amounts of cholesterol. Stearylamine or dicetylphosphate (10 mol%) was used to induce positive or negative charge on the bilayer. The cells were separated from liposomes by centrifugation in membrane filters and the amount of stavudine taken up by macrophages was estimated using tritium labeled drug as a marker. Stavudine uptake was found to be the maximum (approximately 950 ng/million cells) in liposomes containing dipalmitoyl phosphatidylcholine (DPPC). The presence of sphingomyelin, which increases bilayer rigidity decreased cellular uptake of stavudine and the presence of negative charge on the bilayer, enhanced the uptake of stavudine compared to positive charge. There is no apparent difference in uptake when varying amounts of cholesterol was added to liposomal formulations. The present study shows that the sensitivity of macrophages to different charge and lipid type can be used to either decrease or increase cellular uptake as desired.
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The aim of present work was to elucidate the interaction of solid lipid nanoparticles (SLNs) with cellular plasma-membrane to gain insight of intracellular drug delivery. To this aim we followed the uptake of coumarin-6 (a drug model) either free in the extracellular medium or loaded on SLN (c-SLN). Alveolar epithelial cells were exposed to a biocompatible concentration of c-SLN (0.01 mg/ml of tripalmitin) prepared by warm microemulsion whose lipid matrix was constituted by low melting point molecules (fatty acids, triglycerides). Intracellular fluorescence and preferential accumulation in the perinuclear region were increased by 54.8% on comparing c-SLN to the same amount of free coumarin-6 in the medium. Lowering temperature from 37° to 4°C decreased the intracellular signal intensity by about 48% equally for the free as well as for loaded drug, thus suggesting the inhibition of a similar non-endocytotic entrance pathway. No specific co-localization of the fluorescence with intracellular organelles was found. The c-SLN calorimetric profile obtained with differential scanning calorimetry (DSC), revealing transition within the range 58-62°C, altered remarkably upon incubation with cells, suggesting a change in SLN structure after association with cells membranes. We propose that the uptake of the model drug loaded on SLN is only partly related to the endocytotic pathway; it occurs despite the loss of integrity of the original SLN structure and it appears to be more efficient when the drug is vehicled rather than being free in the culture medium.
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