Properties of a hormone-stimulated lipase from rat adipose cells

Abstract

A well defined, easily reproducible system for the study of hormone-stimulated lipolysis in broken cell preparations from rat adipocytes has been described. Addition of hormone prior to cellular rupture results in a homogenate which liberates free fatty acids (FFA) in a linear fashion for at least 1 hr. The quantity of FFA liberated from nonhormone-treated control homogenates is much smaller but is also linear for at least 1 hr. The activation process occurring after addition of hormone appears to be complete within 1 min. Preincubation of the cells prior to addition of the hormone does not increase the magnitude of the response to the hormone. Broken cell preparations made from both control and hormone-treated cells show a pH optimum in the range of 6.5-1.5, and both preparations show a necessity for the presence of bovine serum albumin in the assay medium. The centrifugation of an activated homogenate at 0° and subsequent recombination of the resulting fractions (fat cake, infranatant and pellet) produced a preparation showing lipolysis. When the infranatant from an untreated homogenate was recombined with the fat cake from a hormone-treated homogenate, hormone-stimulated lipolysis was still seen; the reverse combination showed no such stimulation of lipolytic activity. When the infranatant fluid was removed entirely and replaced with warm (37°) Krebs-Ringer phosphate buffer (KRP), lipolysis was greater in hormone-treated than in control preparations, and the specific activity was increased 5-fold. It is tentatively concluded, therefore, that the “active” lipolytic properties are associated with the fat cake; the infranatant fluid appears to inhibit the lipolytic process.