Cardioprotection by the TSPO ligand 4′-chlorodiazepam is associated with inhibition of mitochondrial accumulation of cholesterol at reperfusion

Abstract

The translocator protein (TSPO) is located on the outer mitochondrial membrane where it is responsible for the uptake of cholesterol into mitochondria of steroidogenic organs. TSPO is also present in the heart where its role remains uncertain. We recently showed that TSPO ligands reduced infarct size and improved mitochondrial functions after ischaemia-reperfusion. This study, thus, sought to determine whether cholesterol could play a role in the cardioprotective effect of TSPO ligands. In a model of 30 min coronary occlusion/15 min reperfusion in Wistar rat, we showed that reperfusion induced lipid peroxidation as demonstrated by the increase in conjugated diene and thiobarbituric acid reactive substance formation and altered mitochondrial function (decrease in oxidative phosphorylation and increase in the sensitivity of mitochondrial permeability transition pore opening) in ex-vivo isolated mitochondria. This was associated with an increase in mitochondrial cholesterol uptake (89.5 ± 12.2 vs. 39.9 ± 3.51 nmol/mg protein in controls, P < 0.01) and a subsequent strong generation of auto-oxidized oxysterols, i.e. 7α- and 7β-hydroxycholesterol, 7-ketocholesterol, cholesterol-5α,6α-epoxide, and 5β,6β-epoxide (+173, +149, +165, +165, and +193%vs. controls, respectively; P < 0.01). Administration of the selective TSPO ligand 4'-chlorodiazepam inhibited oxidative stress, improved mitochondrial function, and abolished both mitochondrial cholesterol accumulation and oxysterol production. This was also observed with the new TSPO ligand TRO40303. These data suggest that 4'-chlorodiazepam inhibits oxidative stress and oxysterol formation by reducing the accumulation of cholesterol in the mitochondrial matrix at reperfusion and prevents mitochondrial injury. This new and original mechanism may contribute to the cardioprotective properties of TSPO ligands.