Culture of bone marrow-derived mast cells: A model for studying oxidative metabolism of arachidonic acid and synthesis of other molecules derived from membrane phospholipids

The chapter presents a discussion on a model for studying oxidative metabolism of arachidonic acid and synthesis of other molecules derived from membrane phospholipids. This chapter provides information regarding the techniques of growing bone marrow-derived cultured mouse and human mast cells (BMMC), their activation by various stimuli, and ways of activating separately either arachidonic acid metabolism or exocytosis in these cells. These culture systems can be used as a model of homogeneous cell population responding to a biologically relevant activating secretion stimulus, with the release of lipid mediators originating from membrane phospholipids. The chapter describes several procedures, including exocytosis of preformed and newly generated mediators, phosphatidylinositol (PI) turnover, production of lipid metabolites in activated BMMC, and so on. The chapter mentions on distinguishing the processes of degranulation and 5- lipoxygenase activation in mouse BMMC using the following three methods: (1) Drug intervention; (2) selective stimuli, and (3) oxygen depletion. It is shown that thrombin, a highly specific procoagulant enzyme that has numerous affects on platelets, endothelial, and smooth muscle cells could activate mouse BMMC to degranulate.