Caveolin-1 regulates mitochondrial cholesterol and hepatocellular susceptibility to TNF/FAS

Abstract

Caveolin-1 has been shown to function predominantly as a scaffolding protein involved in organizing the activity of multiple signalling molecules in caveolae. In addition, caveolin-1 exhibits binding properties towards fatty acids and cholesterol, suggesting it could play a role in the intracellular transport of lipids, including cholesterol. We have previously shown that caveolin-1 deficiency impairs liver regeneration, which is restored upon glucose supplementation, implying inefficient mitochondrialderived energy production. Caveolins can act independently of caveolae and are found in non-caveolar regions of caveolaecontaining cells including ER, Golgi and possibly mitochondria. Mitochondria can be closely apposed to plasma membrane caveolae and caveolin-1 has been detected in liver mitochondria. Since mitochondrial cholesterol modulates hepatocellular susceptibility to TNF/Fas, our aim was to examine the role of caveolin-1 in the regulation of mitochondrial cholesterol and fate of hepatocytes following TNF/Fas challenge. [Methods]: Primary hepatocytes and isolated mitochondria were prepared from wild-type and caveolin-1 knockout mice (KO). Oxygen consumption, GSH, free cholesterol levels, mitochondrial membrane permeabilization and subsequent release of cytochrome c and Smac/Diablo were analyzed in isolated mitochondria, while susceptibility to TNF/Fas-mediated apoptosis was examined in primary hepatocytes. [Results]: Impaired state 3 respiration and decreased acceptor control ratio were observed in mitochondria from caveolin-1 KO mice compared to wild-type mice. tBid+Bax-induced release of cytochrome c and Smac/Diablo was greater in isolated mitochondria from caveolin-1 KO mice than from wild-type mice. Moreover, these findings were accompanied by higher mitochondrial free cholesterol levels (30–50%) and GSH depletion (40–60%). Consistent with the latter results, hepatoyctes from caveolin-1 KO mice were more susceptible to Fas-mediated cytochrome c release and apoptosis with respect to wild-type mice, an outcome that was prevented upon GSH ethyl ester pretreatment. Finally, mitochondrial cholesterol extraction by methylcyclodextrin improved mitochondrial respiration and prevented tBid+Bax-mediated cytochrome c release. [Conclusions]: These findings indicate a novel role of caveolin-1 in the hepatocellular susceptibility to TNF/Fas-mediated apoptosis by regulating mitochondrial cholesterol trafficking and GSH homeostasis, implying that caveolin-1 could play a role in liver diseases, such as alcoholic and non-alcoholic steatohepatitis.