FUNCTIONAL-CHARACTERISTICS OF THE RAT-LIVER MACROPHAGE POPULATION AFTER A SINGLE INTRAVENOUS-INJECTION OF LIPOSOME-ENCAPSULATED MURAMYL PEPTIDES

In this study, we investigated different functional characteristics of the rat liver macrophage population after a single i.v. injection of liposome-encapsulated muramyl dipeptide (MDP) or its lipophilic derivative muramyl tripeptide-phosphatidylethanolamine (MTP-PE). The in situ induced tumoricidal activity of the liver macrophage population was determined in vitro against C26 colon adenocarcinoma cells. For investigating which cells are responsible for the observed cytotoxic effects, subfractions of the liver macrophage population, differing in cell size, were isolated at different intervals after injection of the liposomal muramyl peptides. From these subfractions, the number of cells, degree of cytotoxicity, and the response to an additional activation with free MDP in vitro were determined. Maximal induction of tumoricidal activity of the liver macrophage population was reached between 12 and 24 hours after injection of liposomal MDP, while no significant differences between the subfractions were observed. Heterogeneity of tumor cytolytic capacity was observed in subfractions of macrophages isolated at 2 and 48 hours after injection. At these time points, highest cytolytic activity was observed for the small to intermediate-size macrophages. No significant cytotoxicity was detectable in any subfraction 72 hours after injection of liposomal MDP. An identical pattern of macrophage tumoricidal activity was observed after injection of liposomal MTP-PE, although slightly lower cytotoxicity levels were found. When isolated during the first 12 hours after injection of liposomal MDP, the macrophage population was unable to respond to a subsequent in vitro exposure to MDP, with respect to tumor cytotoxicity. Twenty-four and 48 hours after injection, the smallest cells could be slightly reactivated, whereas the larger cells still remained unresponsive. Seventy-two hours after injection, reactivating potential was only slightly decreased, compared with macrophages isolated from control animals. We observed an approximately two-fold increase in the number of isolated cells from 2 up to 24 hours after injection of liposomal MDP, mainly as a result of an increase in the number of cells in the small-cell fractions. At 2 hours after injection, this increase is mainly due to an increase in the number of granulocytes in these fractions, which coincides with an increase in the percentage of granulocytes in the blood at this time point. At 12 and 24 hours after injection, no granulocytes were observed in the different subfractions. The increase in the number of small-size cells at 12 and 24 hours after injection is accompanied by an increase in the percentage of monocytes in the blood at these time points. We conclude that it is likely that these cells play an important role in the cytotoxic potential of the liver, induced by liposomal muramyl peptides.