Bile acid signaling is a critical regulator of glucose and energy metabolism, mainly through the nuclear receptor FXR and the G protein-coupled receptor TGR. The purpose of the present study was to investigate whether dual activation of FXR and TGR5 plays a significant role in the prevention of atherosclerosis progression. To evaluate the effects of bile acid signaling in atherogenesis, ApoE−/− mice and LDLR−/− mice were treated with an FXR/TGR5 dual agonist (INT-767). INT-767 treatment drastically reduced serum cholesterol levels. INT-767 treatment significantly reduced atherosclerotic plaque formation in both ApoE−/− and LDLR−/− mice. INT-767 decreased the expression of pro-inflammatory cytokines and chemokines in the aortas of ApoE−/− mice through the inactivation of NF-κB. In addition, J774 macrophages treated with INT-767 had significantly lower levels of active NF-κB, resulting in cytokine production in response to LPS through a PKA dependent mechanism. This study demonstrates that concurrent activation of FXR and TGR5 attenuates atherosclerosis by reducing both circulating lipids and inflammation.

R e s e a R c h a R t i c l e 4 5 4 4jci.org Volume 125 Number 12 December 2015 IntroductionVascular calcification is a major complication in patients who are aging, have diabetes, or have chronic kidney disease (CKD) and is an active process that differentiates vascular smooth muscle cells (VSMCs) into osteoblast-like cells (1, 2). This process is highly regulated by transcription factors involved in osteogenic differentiation, such as RUNX2, MSX2, and ATF4 (3-5). Several in vitro and in vivo studies have shown lipids to play a causative role in the pathogenesis of vascular calcification in addition to inorganic phosphate, inflammatory cytokines, and oxidative stress (6-13). Treatment with unsaturated fatty acids (UFAs) inhibits vascular mineralization and osteogenic differentiation (14, 15), whereas oxidized lipids, such as oxysterols and oxidized phospholipids, elicit procalcific effects in vascular cells (9,16,17). In addition to this evidence, we also previously reported that saturated fatty acids (SFAs) and calcium simultaneously accumulate during osteogenic differentiation of vascular cells (18,19). Ectopic accumulation of excess lipids, called lipotoxicity, plays a central role in the pathogenesis of cardio-metabolic diseases, including diabetes, obesity, atherosclerosis, and vascular calcification (20-24). However, evidence from a number of experimental systems is emerging, stating that SFAs and UFAs have distinct contributions to lipotoxicity (25,26). SFAs such as palmitic acid (16:0) and stearic acid (18:0) induce apoptosis, oxidative stress, and ER stress in a variety of mammalian cell lines (including hepatocytes, macrophages, and VSMCs), whereas UFAs such as oleic acid have no or minimal lipotoxic properties (5,(25)(26)(27)(28)(29)(30). In addition, cotreatment with UFAs blocks SFA-mediated lipotoxic effects (25,29). However, the specific metabolite of SFAs that induces lipotoxicity, the mechanism underlying SFA-mediated lipotoxicity, and how UFAs block SFA-mediated lipotoxicity are largely unknown.Proper intracellular SFA and UFA balance is controlled by a lipogenic enzyme called stearoyl-CoA desaturase (SCD) (31,32). SCD catalyzes the conversion of SFAs to monounsaturated FAs, mainly 16:0 into palmitoleate (16:1n-7), and 18:0 into oleate (18:1n-9) (33,34). This introduction of a double bond markedly impacts several chemical properties, including a decrease in melting point and an increase in solubility. The activation of SCD therefore neutralizes SFA-mediated lipotoxicity (5, 25). The expression of SCD is highly regulated by a number of hormonal and dietary factors (33,(35)(36)(37). Recently, we found that the accumulation of SFAs by either supplementation with exogenous SFAs or inhibition of SCD induces mineralization of VSMCs (5,19). In addition, SFA-mediated lipotoxicity and vascular calcification were completely blocked by an acyl-CoA synthetase inhibitor and were attenuated by the shRNA-mediated inhibition of fatty acid elongase-6 (Elovl6) (5), suggesting that stearoyl-CoA (18:0-CoA) or its ...

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BackgroundCardiovascular diseases such as atherosclerosis and vascular calcification are a major cause of death in patients with chronic kidney disease (CKD). Recently, the long‐awaited results of the Study of Heart and Renal Protection trial were reported. This large randomized clinical trial found that an extensive cholesterol‐lowering therapy through the combination of simvastatin and ezetimibe significantly reduced cardiovascular diseases in a wide range of patients with CKD. However, the mechanism by which this cholesterol‐lowering therapy reduces CKD‐dependent vascular diseases remains elusive. The objective of the present study was to determine the contribution of the oxysterol‐induced pro‐apoptotic transcription factor CCAAT/enhancer‐binding protein homologous protein (CHOP) on the pathogenesis of CKD‐dependent cardiovascular diseases through endoplasmic reticulum stress signaling.Methods and ResultsCKD increased levels of serum oxysterols such as 7‐ketocholesterol in human patients and ApoE−/− mice. Treatment with simvastatin plus ezetimibe strongly reduced levels of serum oxysterols and attenuated CKD‐dependent atherosclerosis, vascular cell death, vascular calcification, and cardiac dysfunction. This therapy also reduced aortic endoplasmic reticulum stress induced by CKD. The short hairpin RNA‐mediated knockdown of CHOP and activating transcription factor‐4 in vascular smooth muscle cells attenuated oxysterol‐induced mineralization, osteogenic differentiation, and endoplasmic reticulum stress. In addition, CHOP deficiency protected ApoE−/− mice from CKD‐dependent vascular calcification, cardiac dysfunction, and vascular cell death.ConclusionsThese data reveal that the cholesterol‐lowering therapy of simvastatin plus ezetimibe attenuates CKD‐dependent vascular diseases through a reduction of oxysterol‐mediated endoplasmic reticulum stress. CHOP plays a crucial role in the pathogenesis of CKD‐dependent vascular calcification.

Autophagy is a conserved system that adapts to nutrient starvation, after which proteins and organelles are degraded to recycle amino acids in response to starvation. Recently, the ER was added to the list of targets of autophagic degradation. Autophagic degradation pathways of bulk ER and the specific proteins sorted through the ER are considered key mechanisms in maintaining ER homeostasis. Four ER-resident proteins (FAM134B, CCPG1, SEC62, and RTN3) have been identified as ER-resident cargo receptors, which contain LC3-interacting regions. In this study, we identified an N-terminal–truncated isoform of FAM134B (FAM134B-2) that contributes to starvation-induced ER-related autophagy. Hepatic FAM134B-2 but not full-length FAM134B (FAM134B-1) is expressed in a fed state. Starvation drastically induces FAM134B-2 but no other ER-resident cargo receptors through transcriptional activation by C/EBPβ. C/EBPβ overexpression increases FAM134B-2 recruitment into autophagosomes and lysosomal degradation. FAM134B-2 regulates lysosomal degradation of ER-retained secretory proteins such as ApoCIII. This study demonstrates that the C/EBPβ-FAM134B-2 axis regulates starvation-induced selective ER-phagy.