Liposome Encapsulation of Muramyl Peptides for Activation of Macrophage Cytotoxic Properties

This chapter describes a microcytoxicity assay that measures the tumoricidal activity of macrophages activated by liposome-encapsulated muramyl peptides against tumor target cells prelabeled with [¹²⁵I]iododeoxyuridine ([¹²⁵I]IUdR). It discusses the advantages and disadvantages of hydrophilic and lipophilic muramyl peptide derivatives for the activation of macrophage cytotoxic properties and the techniques employed to generate appropriate phospholipid vesicles for achieving macrophage activation. The procedure to assess macrophage activation is also presented in the chapter. The activation of macrophages by liposomes containing muramyl peptides can be achieved using three different types of liposomes—namely, (1) multilamellar vesicles (MLVs), small unilamellar vesicles (SUVs), and large unilamellar/oligolamellar vesicles produced by reverse-phase evaporation (REVs). The standard method for generating MLVs involves hydrating a dry lipid film on a glass test tube or a lyophilized phospholipid mixture by vortex mixing above the gel-to-liquid crystalline transition temperature of the lipid. An extensive sonication of MLVs results in the formation of smaller unilamellar vesicles (SUVs). The reverse-phase evaporation vesicles (REVs) are formed from water-in-oil emulsions of phospholipids and buffer in an excess organic phase, followed by removal of the organic phase under reduced pressure.