Inhibition of apolipoprotein B synthesis stimulates endoplasmic reticulum autophagy that prevents steatosis
Donna M. Conlon, … , Jing Liu, Henry N. Ginsberg
Donna M. Conlon, … , Jing Liu, Henry N. Ginsberg
Published September 6, 2016
Citation Information: J Clin Invest. 2016;126(10):3852-3867. https://doi.org/10.1172/JCI86028.
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Research Article Metabolism

Inhibition of apolipoprotein B synthesis stimulates endoplasmic reticulum autophagy that prevents steatosis

Abstract

Inhibition of VLDL secretion reduces plasma levels of atherogenic apolipoprotein B (apoB) lipoproteins but can also cause hepatic steatosis. Approaches targeting apoB synthesis, which lies upstream of VLDL secretion, have potential to effectively reduce dyslipidemia but can also lead to hepatic accumulation of unsecreted triglycerides (TG). Here, we found that treating mice with apoB antisense oligonucleotides (ASOs) for 6 weeks decreased VLDL secretion and plasma cholesterol without causing steatosis. The absence of steatosis was linked to an increase in ER stress in the first 3 weeks of ASO treatment, followed by development of ER autophagy at the end of 6 weeks of treatment. The latter resulted in increased fatty acid (FA) oxidation that was inhibited by both chloroquine and 3-methyl adenine, consistent with trafficking of ER TG through the autophagic pathway before oxidation. These findings support the concept that inhibition of apoB synthesis traps lipids that have been transferred to the ER by microsomal TG transfer protein (MTP), inducing ER stress. ER stress then triggers ER autophagy and subsequent lysosomal lipolysis of TG, followed by mitochondrial oxidation of released FA, leading to prevention of steatosis. The identification of this pathway indicates that inhibition of VLDL secretion remains a viable target for therapies aiming to reduce circulating levels of atherogenic apoB lipoproteins.

Authors

Donna M. Conlon, Tiffany Thomas, Tatyana Fedotova, Antonio Hernandez-Ono, Gilbert Di Paolo, Robin B. Chan, Kelly Ruggles, Sarah Gibeley, Jing Liu, Henry N. Ginsberg

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Figure 1

MTP ASO and apoB ASO reduce secretion of TG and newly synthesized apoB similarly, but only MTP ASO increases hepatic TG levels.

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MTP ASO and apoB ASO reduce secretion of TG and newly synthesized apoB s...
(A) Measurement of Apob and Mttp mRNA in the liver of apobec-1–KO mice treated with ASO directed against MTP, apoB, or control for 6 weeks by real-time quantitative (qRT-PCR). Data were normalized to actin and are expressed relative to the ASO control group. N = 4–5 per group. (B) apobec-1–KO mice were injected i.v. with Triton WR1339 and 35S-methionine. Blood samples were obtained over the next 120 minutes. TG levels in plasma were measured enzymatically. N = 4–5 per group. (C) apoB was isolated by 4% SDS-PAGE from the 120-minute plasma samples in the Triton studies described in B, bands were cut, and radioactivity was quantitated by scintillation counting. Results are displayed as cpm in plasma relative to control ASO. The quantity of plasma loaded was adjusted for TCA-precipitable cpm. N = 5–6 per group. *P < 0.05 versus control ASO–treated group, by ANOVA. (D) TG and cholesterol levels were measured enzymatically in plasma from mice bled after a 4-hour fast. N = 12–15 per group. TC, total cholesterol. (E) Liver TG was extracted from mice treated with control ASO, MTP ASO, or apoB ASO for 6 weeks. Liver TG levels were measured enzymatically and normalized to liver protein. N = 12–15 per group. Groups with different letters above the bars are significantly different from the others at P < 0.05, by ANOVA. (F) Representative images from 5-μm liver sections that were stained for neutral lipid using oil red O. Original magnification, ×400. All values represent the mean ± SD. (A and D) *P < 0.05 versus control ASO– and MTP ASO–treated groups, by ANOVA; **P < 0.01 versus control ASO– and apoB ASO–treated groups, by ANOVA.