Assessment of mitochondrial physiology of murine white adipose tissue by mechanical permeabilization and lipid depletion

Abstract

White adipose tissue (WAT) represents a major site of triacylglycerol energy storage and is directly associated with metabolic disorders. Mitochondria regulate cellular energy expenditure and are active in WAT. Although isolated mitochondria have been classically used to assess their functions, several artifacts can be introduced by this approach. Furthermore, important limitations exist in the available methods to determine mitochondrial physiology in permeabilized WAT. Here, we established and validated a method for functional evaluation of mice mesenteric WAT (mWAT) mitochondria by using MEchanical Permeabilization and LIpid DEpletion (MEPLIDE) coupled to high-resolution respirometry. We observed that mild stirring of mWAT for 20 min at room temperature with 4% fatty acid-free albumin (FAF-BSA) followed by 50 min without FAF-BSA selectively permeabilized white adipocytes plasma membrane. In these conditions, mWAT mitochondria were intact, exhibiting succinate-induced respiratory rates that were sensitive to classical oxidative phosphorylation modulators. Finally, the respiratory capacity of mWAT in female mice was significantly higher than in males, an observation that agrees with reported data. Therefore, the functional assessment of mWAT mitochondria through MEPLIDE coupled to high resolution respirometry proposed here will contribute to a better understanding of WAT biology in several pathophysiological contexts.