Elsevier

Molecular Metabolism

Volume 3, Issue 5, August 2014, Pages 544-553
Molecular Metabolism

Original article
ER calcium release promotes mitochondrial dysfunction and hepatic cell lipotoxicity in response to palmitate overload

https://doi.org/10.1016/j.molmet.2014.05.004Get rights and content
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open access

Abstract

Palmitate overload induces hepatic cell dysfunction characterized by enhanced apoptosis and altered citric acid cycle (CAC) metabolism; however, the mechanism of how this occurs is incompletely understood. We hypothesize that elevated doses of palmitate disrupt intracellular calcium homeostasis resulting in a net flux of calcium from the ER to mitochondria, activating aberrant oxidative metabolism. We treated primary hepatocytes and H4IIEC3 cells with palmitate and calcium chelators to identify the roles of intracellular calcium flux in lipotoxicity. We then applied 13C metabolic flux analysis (MFA) to determine the impact of calcium in promoting palmitate-stimulated mitochondrial alterations. Co-treatment with the calcium-specific chelator BAPTA resulted in a suppression of markers for apoptosis and oxygen consumption. Additionally, 13C MFA revealed that BAPTA co-treated cells had reduced CAC fluxes compared to cells treated with palmitate alone. Our results demonstrate that palmitate-induced lipoapoptosis is dependent on calcium-stimulated mitochondrial activation, which induces oxidative stress.

Keywords

Metabolic flux analysis
Lipotoxicity
Oxidative stress
ER stress
Fatty liver

Abbreviations

APE
atom percent enrichment
BSA
bovine serum albumin
CAC
citric acid cycle
FFA
free fatty acid
GC–MS
gas chromatography–mass spectrometry
H2DCFDA
2′,7′-dichlorodihydrofluorescein diacetate
MFA
metabolic flux analysis
MUFA
monounsaturated fatty acid
NAFLD
non-alcoholic fatty liver disease
NASH
non-alcoholic steatohepatitis
OA
oleate
PA
palmitate
PI
propidium iodide
ROS
reactive oxygen species
SERCA
sarcoplasmic-endoplasmic reticulum calcium ATPase
SFA
saturated fatty acid

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