Background: Endosomal cholesterol storage in Niemann-Pick type C1 deficiency is associated with elevated mitochondrial cholesterol and alterations in energy metabolism. Results: Blocking endosomal cholesterol transport to mitochondria prevented the metabolic alterations in NPC1-deficient cells. Conclusion: Mitochondrial cholesterol accumulation increases oxidative stress and alters energy metabolism. Significance: Mitochondrial cholesterol is a regulator of energy metabolism and mitochondrial function.

The mitochondrial Ca2+ uniporter is a highly Ca2+-selective protein complex that consists of the pore-forming mitochondrial Ca2+ uniporter protein (MCU), the scaffolding essential MCU regulator (EMRE), and mitochondrial calcium uptake 1 and 2 (MICU1/2), which negatively regulate mitochondrial Ca2+ uptake. We have previously reported that uncoupling proteins 2 and 3 (UCP2/3) are also engaged in the activity of mitochondrial Ca2+ uptake under certain conditions, while the mechanism by which UCP2/3 facilitates mitochondrial Ca2+ uniport remains elusive. This work was designed to investigate the impact of UCP2 on the three distinct mitochondrial Ca2+ currents found in mitoplasts isolated from HeLa cells, the intermediate- (i-), burst- (b-) and extra-large (xl-) mitochondrial/mitoplast Ca2+ currents (MCC). Using the patch clamp technique on mitoplasts from cells with reduced MCU and EMRE unveiled a very high affinity of MCU for xl-MCC that succeeds that for i-MCC, indicating the coexistence of at least two MCU/EMRE-dependent Ca2+ currents. The manipulation of the expression level of UCP2 by either siRNA-mediated knockdown or overexpression changed exclusively the open probability (NPo) of xl-MCC by approx. 38 % decrease or nearly a 3-fold increase, respectively. These findings confirm a regulatory role of UCP2 in mitochondrial Ca2+ uptake and identify UCP2 as a selective modulator of just one distinct MCU/EMRE-dependent mitochondrial Ca2+ inward current.

Cellular TG stores are efficiently hydrolyzed by adipose TG lipase (ATGL). Its coactivator comparative gene identification-58 (CGI-58) strongly increases ATGL-mediated TG catabolism in cell culture experiments. To investigate the consequences of CGI-58 deficiency in murine macrophages, we generated mice with a targeted deletion of CGI-58 in myeloid cells (macCGI-58−/− mice). CGI-58−/− macrophages accumulate intracellular TG-rich lipid droplets and have decreased phagocytic capacity, comparable to ATGL−/− macrophages. In contrast to ATGL−/− macrophages, however, CGI-58−/− macrophages have intact mitochondria and show no indications of mitochondrial apoptosis and endoplasmic reticulum stress, suggesting that TG accumulation per se lacks a significant role in processes leading to mitochondrial dysfunction. Another notable difference is the fact that CGI-58−/− macrophages adopt an M1-like phenotype in vitro. Finally, we investigated atherosclerosis susceptibility in macCGI-58/ApoE-double KO (DKO) animals. In response to high-fat/high-cholesterol diet feeding, DKO animals showed comparable plaque formation as observed in ApoE−/− mice. In agreement, antisense oligonucleotide-mediated knockdown of CGI-58 in LDL receptor−/− mice did not alter atherosclerosis burden in the aortic root. These results suggest that macrophage function and atherosclerosis susceptibility differ fundamentally in these two animal models with disturbed TG catabolism, showing a more severe phenotype by ATGL deficiency.

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