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Electron microscope studies were done on mouse liver, from 5 min to 8 wk after an intravenous injection of liposomes containing ethylenediaminetetraacetic acid (EDTA). Livers of mice receiving an injection of liposomes containing KCL instead of EDTA or an injection of a solution of EDTA were also examined. Liposomes were shown to be phagocytized by hepatocytes as well as by Kupffer cells within minutes after the injection. Initially, there was a close contact between the liposomal membrane and the cellular membrane, followed by an invagination of the latter and the formation of a distinct vesicle surrounding a single liposome or a cluster of several liposomes. No fusion between the liposomal membrane and the cell membrane was observed. Between 15 min and 6 h after liposome injection, the Kupffer cells were found to have an increased number of lysosomes and autophagic vacuoles. Within the latter, morphologically intact liposomes or remnants of liposomes could be seen. At 12 h after injection, a striking increase in macrophages was observed in the liver sinusoids of EDTA-liposome-injected mice, but not in those of KCl-liposome-injected mice. Within the macrophages, remnants of liposomes occasionally could be observed. However, the origin and the physiological role of these cells are unknown. In the hepatocytes, morphological changes were first observed 24 h after injection; there were large numbers of autophagic vacuoles, and some cells showed extensive areas of focal cytoplasmic degeneration. The morphology of the liver cells returned to normal about 7 days after injection. No morphological changes were observed in livers of mice receiving EDTA solution without liposomes. A possible mechanism by which the liposome-encapsulated chelating agents can successfully remove intracellular toxic metals is discussed. The use of liposomes as carriers seems to be a useful tool for intracellular delivery of chelating agents or drugs in general.

Liposomes containing chelating agents. Cellular penetration and a possible mechanism of metal removal.