Isolation and Phospholipid Enrichment of Muscle Mitochondria and Mitoplasts

Abstract

The efficient ATP production in mitochondria relies on the highly specific organization of its double membrane. Notably, the inner mitochondrial membrane (IMM) displays a massive surface extension through its folding into cristae, along which concentrate respiratory complexes and oligomers of the ATP synthase. Evidence has accumulated to highlight the importance of a specific phospholipid composition of the IMM to support mitochondrial oxidative phosphorylation. Contribution of specific phospholipids to mitochondrial ATP production is classically studied by modulating the activity of enzymes involved in their synthesis, but the interconnection of phospholipid synthesis pathways often impedes the determination of the precise role of each phospholipid. Here, we describe a protocol to specifically enrich mitochondrial membranes with cardiolipin or phosphatidylcholine, as well as a fluorescence-based method to quantify phospholipid enrichment. This method, based on the fusion of lipid vesicles with isolated mitochondria, may further allow a precise evaluation of phospholipid contribution to mitochondrial functions.

Cardiolipin Alterations during Obesity: Exploring Therapeutic Opportunities

“…Alteration of mitochondrial membranes composition is associated with a reduction of mitochondrial coupling, which drives an increase of energy expenditure that protects mice against HFD-induced obesity and its associated deleterious consequences. Importantly, enrichment of mitochondria isolated from Hacd1 -KO mice with cardiolipin rescued mitochondrial coupling, in vitro demonstrating a key role of cardiolipin in mitochondrial coupling efficiency in muscle [ 46 , 153 ]. Targeting of other genes involved in VLCFA synthesis confirmed the interest of its modulation in metabolic syndrome, although mitochondrial efficiency and cardiolipin content were not investigated in these studies [ 154 , 155 , 156 , 157 , 158 ].…”