Measurement of phosphoinositide 3-kinase and its products to study adipogenic signal transduction

Abstract

Adipogenesis is an important component of adipose tissue development and growth. Alterations in adipogenesis may promote adipose tissue insulin resistance and inflammation. The ability of preadipocytes to differentiate into mature adipocytes depends on the activation of phosphoinositide 3-kinase (PI3K). This chapter describes the methodology used to measure the cellular accumulation of phosphoinositide products of PI3K. This approach involves labeling the cells with myo-[2-3H]-inositol, extraction and deacylation of the phosphoinositides, and HPLC separation of the deacylated derivatives. The assay of PI3K activity itself is also described in detail. The ability to analyze PI3K and its phosphoinositide products is a useful tool for ongoing endeavours to understand adipogenesis and adipose tissue dysfunction. © 2008 Humana Press, a part of Springer Science+Business Media, LLC.