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Arellano‐García LI, Milton‐Laskibar I, Martínez JA, Arán‐González M, Portillo MP. Comparative effects of viable Lactobacillus rhamnosus GG and its heat-inactivated paraprobiotic in the prevention of high-fat high-fructose diet-induced non-alcoholic fatty liver disease in rats. Biofactors 2025; 51:e2116. [PMID: 39135211 PMCID: PMC11680974 DOI: 10.1002/biof.2116] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/04/2024] [Accepted: 07/29/2024] [Indexed: 12/29/2024]
Abstract
Nonalcoholic fatty liver disease (NAFLD) is one of the most prevalent chronic liver alterations worldwide, being gut microbiota dysbiosis one of the contributing factors to its development. The aim of this research is to compare the potential effects of a viable probiotic (Lactobacillus rhamnosus GG) with those exerted by its heat-inactivated paraprobiotic counterpart in a dietary rodent model of NAFLD. The probiotic administration effectively prevented the hepatic lipid accumulation induced by a high-fat high-fructose diet feeding, as demonstrated by chemical (lower TG content) and histological (lower steatosis grade and lobular inflammation) analyses. This effect was mainly mediated by the downregulation of lipid uptake (FATP2 protein expression) and upregulating liver TG release to bloodstream (MTTP activity) in rats receiving the probiotic. By contrast, the effect of the paraprobiotic preventing diet-induced liver lipid accumulation was milder, and mainly derived from the downregulation of hepatic de novo lipogenesis (SREBP-1c protein expression and FAS activity) and TG assembly (DGAT2 and AQP9 protein expression). The obtained results demonstrate that under these experimental conditions, the effects induced by the administration of viable L. rhamnosus GG preventing liver lipid accumulation in rats fed a diet rich in saturated fat and fructose differ from those induced by its heat-inactivated paraprobiotic counterpart.
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Affiliation(s)
- Laura Isabel Arellano‐García
- Nutrition and Obesity Group, Department of Pharmacy and Food Sciences, Faculty of Pharmacy and Lucio Lascaray Research CentreUniversity of the Basque Country (UPV/EHU)Vitoria‐GasteizSpain
- CIBERobn Physiopathology of Obesity and Nutrition, Institute of Health Carlos IIIMadridSpain
| | - Iñaki Milton‐Laskibar
- Nutrition and Obesity Group, Department of Pharmacy and Food Sciences, Faculty of Pharmacy and Lucio Lascaray Research CentreUniversity of the Basque Country (UPV/EHU)Vitoria‐GasteizSpain
- CIBERobn Physiopathology of Obesity and Nutrition, Institute of Health Carlos IIIMadridSpain
- BIOARABA Health Research InstituteVitoria‐GasteizSpain
| | - J. Alfredo Martínez
- CIBERobn Physiopathology of Obesity and Nutrition, Institute of Health Carlos IIIMadridSpain
- Precision Nutrition and Cardiometabolic Health, IMDEA‐Food Institute (Madrid Institute for Advanced Studies), Campus of International Excellence (CEI) UAM+CSIC, Spanish National Research CouncilMadridSpain
| | - Miguel Arán‐González
- Unidad de Gestión Clínica de Anatomía Patológica de GuipúzcoaHospital Universitario DonostiaSan SebastiánSpain
| | - María P. Portillo
- Nutrition and Obesity Group, Department of Pharmacy and Food Sciences, Faculty of Pharmacy and Lucio Lascaray Research CentreUniversity of the Basque Country (UPV/EHU)Vitoria‐GasteizSpain
- CIBERobn Physiopathology of Obesity and Nutrition, Institute of Health Carlos IIIMadridSpain
- BIOARABA Health Research InstituteVitoria‐GasteizSpain
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2
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Thierer JH, Foresti O, Yadav PK, Wilson MH, Moll TOC, Shen MC, Busch-Nentwich EM, Morash M, Mohlke KL, Rawls JF, Malhotra V, Hussain MM, Farber SA. Pla2g12b drives expansion of triglyceride-rich lipoproteins. Nat Commun 2024; 15:2095. [PMID: 38453914 PMCID: PMC10920679 DOI: 10.1038/s41467-024-46102-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2023] [Accepted: 02/14/2024] [Indexed: 03/09/2024] Open
Abstract
Vertebrates transport hydrophobic triglycerides through the circulatory system by packaging them within amphipathic particles called Triglyceride-Rich Lipoproteins. Yet, it remains largely unknown how triglycerides are loaded onto these particles. Mutations in Phospholipase A2 group 12B (PLA2G12B) are known to disrupt lipoprotein homeostasis, but its mechanistic role in this process remains unclear. Here we report that PLA2G12B channels lipids within the lumen of the endoplasmic reticulum into nascent lipoproteins. This activity promotes efficient lipid secretion while preventing excess accumulation of intracellular lipids. We characterize the functional domains, subcellular localization, and interacting partners of PLA2G12B, demonstrating that PLA2G12B is calcium-dependent and tightly associated with the membrane of the endoplasmic reticulum. We also detect profound resistance to atherosclerosis in PLA2G12B mutant mice, suggesting an evolutionary tradeoff between triglyceride transport and cardiovascular disease risk. Here we identify PLA2G12B as a key driver of triglyceride incorporation into vertebrate lipoproteins.
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Affiliation(s)
- James H Thierer
- Department of Embryology, Carnegie Institution for Science, Baltimore, MD, 21218, USA
- Johns Hopkins University in Baltimore, Maryland Department of biology, Baltimore, MD, 21218, USA
| | - Ombretta Foresti
- Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology, Barcelona, 08003, ES, Spain
| | - Pradeep Kumar Yadav
- Department of Foundations of Medicine, NYU Long Island School of Medicine, Mineola, NY, 11501, USA
- Department of Botany, Faculty of Science, University of Allahabad, Prayagraj, India
| | - Meredith H Wilson
- Department of Embryology, Carnegie Institution for Science, Baltimore, MD, 21218, USA
- Johns Hopkins University in Baltimore, Maryland Department of biology, Baltimore, MD, 21218, USA
| | - Tabea O C Moll
- Department of Embryology, Carnegie Institution for Science, Baltimore, MD, 21218, USA
- Johns Hopkins University in Baltimore, Maryland Department of biology, Baltimore, MD, 21218, USA
| | - Meng-Chieh Shen
- Department of Embryology, Carnegie Institution for Science, Baltimore, MD, 21218, USA
| | | | - Margaret Morash
- Department of Molecular Genetics and Microbiology, Duke University, Durham, NC, 27708, USA
| | - Karen L Mohlke
- Department of Genetics, University of North Carolina, Chapel Hill, NC, 27599, USA
| | - John F Rawls
- Department of Molecular Genetics and Microbiology, Duke University, Durham, NC, 27708, USA
| | - Vivek Malhotra
- Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology, Barcelona, 08003, ES, Spain
- Universitat Pompeu Fabra, Barcelona, Spain
- Institució Catalana de Recerca i Estudis Avançats, Barcelona, Spain
| | - M Mahmood Hussain
- Department of Foundations of Medicine, NYU Long Island School of Medicine, Mineola, NY, 11501, USA
| | - Steven A Farber
- Department of Embryology, Carnegie Institution for Science, Baltimore, MD, 21218, USA.
- Johns Hopkins University in Baltimore, Maryland Department of biology, Baltimore, MD, 21218, USA.
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3
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Cai Y, Yang Q, Yu Y, Yang F, Bai R, Fan X. Efficacy and underlying mechanisms of berberine against lipid metabolic diseases: a review. Front Pharmacol 2023; 14:1283784. [PMID: 38034996 PMCID: PMC10684937 DOI: 10.3389/fphar.2023.1283784] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2023] [Accepted: 11/03/2023] [Indexed: 12/02/2023] Open
Abstract
Lipid-lowering therapy is an important tool for the treatment of lipid metabolic diseases, which are increasing in prevalence. However, the failure of conventional lipid-lowering drugs to achieve the desired efficacy in some patients, and the side-effects of these drug regimens, highlight the urgent need for novel lipid-lowering drugs. The liver and intestine are important in the production and removal of endogenous and exogenous lipids, respectively, and have an important impact on circulating lipid levels. Elevated circulating lipids predisposes an individual to lipid deposition in the vascular wall, affecting vascular function. Berberine (BBR) modulates liver lipid production and clearance by regulating cellular targets such as cluster of differentiation 36 (CD36), acetyl-CoA carboxylase (ACC), microsomal triglyceride transfer protein (MTTP), scavenger receptor class B type 1 (SR-BI), low-density lipoprotein receptor (LDLR), and ATP-binding cassette transporter A1 (ABCA1). It influences intestinal lipid synthesis and metabolism by modulating gut microbiota composition and metabolism. Finally, BBR maintains vascular function by targeting proteins such as endothelial nitric oxide synthase (eNOS) and lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). This paper elucidates and summarizes the pharmacological mechanisms of berberine in lipid metabolic diseases from a multi-organ (liver, intestine, and vascular system) and multi-target perspective.
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Affiliation(s)
- Yajie Cai
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Qiaoning Yang
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- National Clinical Research Center for Chinese Medicine Cardiology, Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- NMPA Key Laboratory for Clinical Research and Evaluation of Traditional Chinese Medicine, Beijing, China
| | - Yanqiao Yu
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- Department of Graduate School, Beijing University of Chinese Medicine, Beijing, China
| | - Furong Yang
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Ruina Bai
- Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
- National Clinical Research Center for Chinese Medicine Cardiology, Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing, China
| | - Xiaodi Fan
- Institute of Basic Medical Sciences, Xiyuan Hospital of China Academy of Chinese Medical Sciences, Beijing, China
- Key Laboratory of Pharmacology of Chinese Materia Medica, Beijing, China
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4
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Zhu L, Fu J, Xiao X, Wang F, Jin M, Fang W, Wang Y, Zong X. Faecal microbiota transplantation-mediated jejunal microbiota changes halt high-fat diet-induced obesity in mice via retarding intestinal fat absorption. Microb Biotechnol 2021; 15:337-352. [PMID: 34704376 PMCID: PMC8719817 DOI: 10.1111/1751-7915.13951] [Citation(s) in RCA: 26] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2021] [Revised: 09/13/2021] [Accepted: 10/06/2021] [Indexed: 01/21/2023] Open
Abstract
Faecal Microbiota Transplantation (FMT) is considered as a promising technology to fight against obesity. Wild boar has leanermuscle and less fat in comparison to the domestic pig, which were thought to be related with microbiota. To investigate the function and mechanism of the wild boar microbiota on obesity, we first analysed the wild boar microbiota composition via 16S rDNA sequencing, which showed that Firmicutes and Proteobacteria were the dominant bacteria. Then, we established a high‐fat diet (HFD)‐induced obesity model, and transfer low and high concentrations of wild boar faecal suspension in mice for 9 weeks. The results showed that FMT prevented HFD‐induced obesity and lipid metabolism disorders, and altered the jejunal microbiota composition especially increasing the abundance of the Lactobacillus and Romboutsia, which were negatively correlated with obesity‐related indicators. Moreover, we found that the anti‐obesity effect of wild boar faecal suspension was associated with jejunal N6‐methyladenosine (m6A) levels. Overall, these results suggest that FMT has a mitigating effect on HFD‐induced obesity, which may be due to the impressive effects of FMT on the microbial composition and structure of the jejunum. These changes further alter intestinal lipid metabolism and m6A levels to achieve resistance to obesity.
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Affiliation(s)
- Luoyi Zhu
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Jie Fu
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Xiao Xiao
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Fengqin Wang
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Mingliang Jin
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Weihuan Fang
- Zhejiang University Institute of Preventive Veterinary Medicine, Hangzhou, Zhejiang, 310058, China.,Zhejiang Provincial Key Laboratory of Preventive Veterinary Medicine, Hangzhou, Zhejiang, 310058, China
| | - Yizhen Wang
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
| | - Xin Zong
- National Engineering Laboratory for Feed Safety and Pollution Prevention and Controlling, National Development and Reform Commission, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Molecular Animal Nutrition (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Nutrition and Feed Science (Eastern of China), Ministry of Agriculture and Rural Affairs, Hangzhou, Zhejiang, 310058, China.,Key Laboratory of Animal Feed and Nutrition of Zhejiang Province, Hangzhou, Zhejiang, 310058, China
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5
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Chen P, Li Y, Xiao L. Berberine ameliorates nonalcoholic fatty liver disease by decreasing the liver lipid content via reversing the abnormal expression of MTTP and LDLR. Exp Ther Med 2021; 22:1109. [PMID: 34504563 PMCID: PMC8383777 DOI: 10.3892/etm.2021.10543] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2020] [Accepted: 02/15/2021] [Indexed: 12/14/2022] Open
Abstract
The global incidence of nonalcoholic fatty liver disease (NAFLD) is increasing. The present study explored the effect and mechanism of berberine (BBR) on NAFLD in rats. Thirty-five Sprague-Dawley rats were randomly divided into the control and NAFLD groups, which were fed a normal diet or high-fat diet, respectively, for 8 weeks. Hematoxylin and eosin staining was performed on liver tissues and establishment of the NAFLD model was confirmed by microscopy. NAFLD rats were subsequently randomly subdivided and treated with saline or BBR for 8 weeks. The liver wet weight of rats in each group was measured, the liver tissue structure was observed by microscopy, and alanine aminotransferase (ALT), aspartate aminotransferase (AST), total cholesterol (TC), triglyceride (TG), fasting blood glucose (FBG), low-density lipoprotein (LDL) and high-density lipoprotein (HDL) levels were detected using a semi-automatic biochemical detector. Reverse transcription-quantitative PCR and western blotting were performed to determine the mRNA and protein expression levels of microsomal triglyceride transfer protein (MTTP), apolipoprotein B and low-density lipoprotein receptor (LDLR). Compared with the control group, the liver wet weight of the NAFLD rats was higher; the liver showed obvious fatty degeneration and liver TG levels increased significantly, as did serum levels of ALT, AST, TC, TG, FBG, HDL and LDL, while expression of MTTP and LDLR significantly decreased. Compared with the saline-treated NAFLD rats, the BBR-treated rats had reduced liver wet weight, improved liver steatosis and a significant decrease in liver TG levels, while ALT, AST, TC, TG, and LDL serum levels significantly decreased and MTTP levels were significantly upregulated. In conclusion, BBR treatment ameliorated the fatty liver induced by a high-fat diet in rats. Furthermore, BBR reversed the abnormal expression of MTTP and LDLR in rats with high-fat diet induced-NAFLD. The present findings suggest that fatty liver could be improved by BBR administration, via reversing the abnormal expression of MTTP and LDLR and inhibiting lipid synthesis.
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Affiliation(s)
- Ping Chen
- Department of Pharmacy, Affiliated Hospital of Shandong Medical College, Linyi, Shandong 276000, P.R. China
| | - Yusheng Li
- Department of Pharmacy, Linyi Maternal and Child Health Care Hospital, Linyi, Shandong 276000, P.R. China
| | - Li Xiao
- Department of Pharmacy, Linyi Maternal and Child Health Care Hospital, Linyi, Shandong 276000, P.R. China
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6
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Peng H, Chiu TY, Liang YJ, Lee CJ, Liu CS, Suen CS, Yen JJY, Chen HT, Hwang MJ, Hussain MM, Yang HC, Yang-Yen HF. PRAP1 is a novel lipid-binding protein that promotes lipid absorption by facilitating MTTP-mediated lipid transport. J Biol Chem 2021; 296:100052. [PMID: 33168624 PMCID: PMC7949078 DOI: 10.1074/jbc.ra120.015002] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2020] [Revised: 10/30/2020] [Accepted: 11/09/2020] [Indexed: 01/10/2023] Open
Abstract
Microsomal triglyceride transfer protein (MTTP) is an endoplasmic reticulum resident protein that is essential for the assembly and secretion of triglyceride (TG)-rich, apoB-containing lipoproteins. Although the function and structure of mammalian MTTP have been extensively studied, how exactly MTTP transfers lipids to lipid acceptors and whether there are other biomolecules involved in MTTP-mediated lipid transport remain elusive. Here we identify a role in this process for the poorly characterized protein PRAP1. We report that PRAP1 and MTTP are partially colocalized in the endoplasmic reticulum. We observe that PRAP1 directly binds to TG and facilitates MTTP-mediated lipid transfer. A single amino acid mutation at position 85 (E85V) impairs PRAP1's ability to form a ternary complex with TG and MTTP, as well as impairs its ability to facilitate MTTP-mediated apoB-containing lipoprotein assembly and secretion, suggesting that the ternary complex formation is required for PRAP1 to facilitate MTTP-mediated lipid transport. PRAP1 is detectable in chylomicron/VLDL-rich plasma fractions, suggesting that MTTP recognizes PRAP1-bound TG as a cargo and transfers TG along with PRAP1 to lipid acceptors. Both PRAP1-deficient and E85V knock-in mutant mice fed a chow diet manifested an increase in the length of their small intestines, likely to compensate for challenges in absorbing lipid. Interestingly, both genetically modified mice gained significantly less body weight and fat mass when on high-fat diets compared with littermate controls and were prevented from hepatosteatosis. Together, this study provides evidence that PRAP1 plays an important role in MTTP-mediated lipid transport and lipid absorption.
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Affiliation(s)
- Hubert Peng
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan
| | - Tzu-Yuan Chiu
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan
| | - Yu-Jen Liang
- Institute of Statistical Science, Academia Sinica, Taipei, Taiwan
| | - Chia-Jen Lee
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan
| | - Chih-Syuan Liu
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan
| | - Ching-Shu Suen
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Jeffrey J-Y Yen
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Hung-Ta Chen
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan
| | - Ming-Jing Hwang
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - M Mahmood Hussain
- Foundations of Medicine, NYU Long Island School of Medicine, Mineola, New York, USA
| | - Hsin-Chou Yang
- Institute of Statistical Science, Academia Sinica, Taipei, Taiwan
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7
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Shen Y, Xiao X, Wu K, Wang Y, Yuan Y, Liu J, Sun S, Liu J. Effects and molecular mechanisms of Ninghong black tea extract in nonalcoholic fatty liver disease of rats. J Food Sci 2020; 85:800-807. [PMID: 32090345 DOI: 10.1111/1750-3841.14846] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2019] [Revised: 09/03/2019] [Accepted: 09/16/2019] [Indexed: 12/24/2022]
Abstract
The aim of this study is to observe the effects of Ninghong black tea extract on fat deposition and high-fat diet-induced nonalcoholic fatty liver disease (NAFLD) and to explore the potential mechanisms of these effect. Under 2% Ninghong black tea extract diet feeding in rat model, the results showed that Ninghong black tea extract decreased the body fat ratio and the number of lipid droplets in the liver and significantly alleviated NAFLD in the rat model. The real-time fluorescence quantitative polymerase chain reaction results showed that Ninghong black tea extract significantly upregulated the expression of peroxisome proliferator-activated receptor α (PPARα), which is important in fatty acid β-oxidation, and microsomal triglyceride transfer protein (MTP), which plays an important role in the synthesis of very low density lipoprotein (VLDL). By promoting the expression of PPARα and MTP in liver tissue and thereby promoting fatty acid β-oxidation and VLDL synthesis, Ninghong black tea extract relieves high-fat diet-induced NAFLD.
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Affiliation(s)
- Yu Shen
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Xiaojuan Xiao
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Kunlu Wu
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Yanpeng Wang
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Yijun Yuan
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Jianwei Liu
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Shuming Sun
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
| | - Jing Liu
- Molecular Biology Research Center, School of Life Sciences, Central South Univ., Changsha, 410078, China
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8
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Qiao L, Luo GG. Human apolipoprotein E promotes hepatitis B virus infection and production. PLoS Pathog 2019; 15:e1007874. [PMID: 31393946 PMCID: PMC6687101 DOI: 10.1371/journal.ppat.1007874] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2019] [Accepted: 05/27/2019] [Indexed: 12/16/2022] Open
Abstract
Hepatitis B virus (HBV) is a common cause of liver diseases, including chronic hepatitis, steatosis, fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). HBV chronically infects about 240 million people worldwide, posing a major global health problem. The current standard antiviral therapy effectively inhibits HBV replication but does not eliminate the virus unlike direct-acting antivirals (DAA) for curing hepatitis C. Our previous studies have demonstrated that human apolipoprotein E (apoE) plays important roles in hepatitis C virus infection and morphogenesis. In the present study, we have found that apoE is also associated with HBV and is required for efficient HBV infection. An apoE-specific monoclonal antibody was able to capture HBV similar to anti-HBs. More importantly, apoE monoclonal antibody could effectively block HBV infection, resulting in a greater than 90% reduction of HBV infectivity. Likewise, silencing of apoE expression or knockout of apoE gene by CRISPR/Cas9 resulted in a greater than 90% reduction of HBV infection and more than 80% decrease of HBV production, which could be fully restored by ectopic apoE expression. However, apoE silencing or knockout did not significantly affect HBV DNA replication or the production of nonenveloped (naked) nucleocapsids. These findings demonstrate that human apoE promotes HBV infection and production. We speculate that apoE may also play a role in persistent HBV infection by evading host immune response similar to its role in the HCV life cycle and pathogenesis. Inhibitors interfering with apoE biogenesis, secretion, and/or binding to receptors may serve as antivirals for elimination of chronic HBV infection.
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Affiliation(s)
- Luhua Qiao
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, Alabama, United States of America
| | - Guangxiang George Luo
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, Alabama, United States of America
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9
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Di Filippo M, Collardeau Frachon S, Janin A, Rajan S, Marmontel O, Decourt C, Rubio A, Nony S, Dumont S, Cuerq C, Charrière S, Moulin P, Lachaux A, Hussain MM, Bozon D, Peretti N. Normal serum ApoB48 and red cells vitamin E concentrations after supplementation in a novel compound heterozygous case of abetalipoproteinemia. Atherosclerosis 2019; 284:75-82. [PMID: 30875496 DOI: 10.1016/j.atherosclerosis.2019.02.016] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/14/2018] [Revised: 02/16/2019] [Accepted: 02/19/2019] [Indexed: 02/06/2023]
Abstract
BACKGROUND AND AIMS Abetalipoproteinemia (ABL) is a rare recessive monogenic disease due to MTTP (microsomal triglyceride transfer protein) mutations leading to the absence of plasma apoB-containing lipoproteins. Here we characterize a new ABL case with usual clinical phenotype, hypocholesterolemia, hypotriglyceridemia but normal serum apolipoprotein B48 (apoB48) and red blood cell vitamin E concentrations. METHODS Histology and MTP activity measurements were performed on intestinal biopsies. Mutations in MTTP were identified by Sanger sequencing, quantitative digital droplet and long-range PCR. Functional consequences of the variants were studied in vitro using a minigene splicing assay, measurement of MTP activity and apoB48 secretion. RESULTS Intestinal steatosis and the absence of measurable lipid transfer activity in intestinal protein extract supported the diagnosis of ABL. A novel MTTP c.1868G>T variant inherited from the patient's father was identified. This variant gives rise to three mRNA transcripts: one normally spliced, found at a low frequency in intestinal biopsy, carrying the p.(Arg623Leu) missense variant, producing in vitro 65% of normal MTP activity and apoB48 secretion, and two abnormally spliced transcripts resulting in a non-functional MTP protein. Digital droplet PCR and long-range sequencing revealed a previously described c.1067+1217_1141del allele inherited from the mother, removing exon 10. Thus, the patient is compound heterozygous for two dysfunctional MTTP alleles. The p.(Arg623Leu) variant may maintain residual secretion of apoB48. CONCLUSIONS Complex cases of primary dyslipidemia require the use of a cascade of different methodologies to establish the diagnosis in patients with non-classical biological phenotypes and provide better knowledge on the regulation of lipid metabolism.
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Affiliation(s)
- Mathilde Di Filippo
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France; INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France.
| | - Sophie Collardeau Frachon
- INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France; Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Institut de Pathologie, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Alexandre Janin
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France; Université de Lyon, Université Claude Bernard Lyon 1, Institut NeuroMyoGène, CNRS UMR5310, INSERM U1217, Lyon, F-69622, France.
| | - Sujith Rajan
- NYU Winthrop Hospital, 101 Mineola Blvd, Mineola, USA.
| | - Oriane Marmontel
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France; INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France.
| | - Charlotte Decourt
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Amandine Rubio
- Gastroentérologie et Nutrition Pédiatrique Hôpital Couple Enfant, CHU de Grenoble Alpes, Grenoble, F-38043, France; Laboratoire de Bioénergétique Fondamentale et Appliquée, INSERM U1055, Univ. Grenoble Alpes, F-38000, France.
| | - Séverine Nony
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Sabrina Dumont
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Charlotte Cuerq
- INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France; Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Sud, Service de Biochimie et Biologie Moléculaire, Hospices Civils de Lyon, Pierre, Benite cedex, F-69495, France.
| | - Sybil Charrière
- INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France; Fédération d'endocrinologie, maladies métaboliques, diabète et nutrition, Hôpital Louis Pradel, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Philippe Moulin
- INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France; Fédération d'endocrinologie, maladies métaboliques, diabète et nutrition, Hôpital Louis Pradel, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Alain Lachaux
- Service de Nutrition Pediatrique, Gastroenterologie and Hepatologie, Hôpital Femme Mère Enfants, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | | | - Dominique Bozon
- Laboratoire de Biologie Médicale Multi Sites, Centre de Biologie et de Pathologie Est, Service de Biochimie et Biologie Moléculaire Grand Est, Hospices Civils de Lyon, Bron cedex, F-69677, France.
| | - Noël Peretti
- INSERM U1060, Laboratoire Carmen, Université Lyon 1, INRA U1235, INSA de Lyon, CENS, Centre de Recherche en Nutrition Humaine Rhône Alpes, Villeurbanne F-69621, Oullins cedex, F-69921, France; Service de Nutrition Pediatrique, Gastroenterologie and Hepatologie, Hôpital Femme Mère Enfants, Hospices Civils de Lyon, Bron cedex, F-69677, France.
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10
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Di Filippo M, Varret M, Boehm V, Rabès JP, Ferkdadji L, Abramowitz L, Dumont S, Lenaerts C, Boileau C, Joly F, Schmitz J, Samson-Bouma ME, Bonnefont-Rousselot D. Postprandial lipid absorption in seven heterozygous carriers of deleterious variants of MTTP in two abetalipoproteinemic families. J Clin Lipidol 2018; 13:201-212. [PMID: 30522860 DOI: 10.1016/j.jacl.2018.10.003] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2018] [Revised: 09/25/2018] [Accepted: 10/12/2018] [Indexed: 12/19/2022]
Abstract
BACKGROUND Abetalipoproteinemia, a recessive disease resulting from deleterious variants in MTTP (microsomal triglyceride transfer protein), is characterized by undetectable concentrations of apolipoprotein B, extremely low levels of low-density lipoprotein cholesterol in the plasma, and a total inability to export apolipoprotein B-containing lipoproteins from both the intestine and the liver. OBJECTIVE To study lipid absorption after a fat load and liver function in 7 heterozygous relatives from 2 abetalipoproteinemic families, 1 previously unreported. RESULTS Both patients are compound heterozygotes for p.(Arg540His) and either c.708_709del p.(His236Glnfs*11) or c.1344+3_1344+6del on the MTTP gene. The previously undescribed patient has been followed for 22 years with ultrastructure analyses of both the intestine and the liver. In these 2 families, 5 relatives were heterozygous for p.(Arg540His), 1 for p.(His236Glnfs*11) and 1 for c.1344+3_1344+6del. In 4 heterozygous relatives, the lipid absorption was normal independent of the MTTP variant. In contrast, in 3 of them, the increase in triglyceride levels after fat load was abnormal. These subjects were additionally heterozygous carriers of Asp2213 APOB in-frame deletion, near the cytidine mRNA editing site, which is essential for intestinal apoB48 production. Liver function appeared to be normal in all the heterozygotes except for one who exhibited liver steatosis for unexplained reasons. CONCLUSION Our study suggests that a single copy of the MTTP gene may be sufficient for human normal lipid absorption, except when associated with an additional APOB gene alteration. The hepatic steatosis reported in 1 patient emphasizes the need for liver function tests in all heterozygotes until the level of risk is established.
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Affiliation(s)
- Mathilde Di Filippo
- UF Dyslipidemies, Service de Biochimie et Biologie moléculaire Grand Est, GHE, Hospices Civils de Lyon, Bron Cedex, France; Univ-Lyon, CarMeN laboratory, Inserm U1060, INRA U1397, Université Claude Bernard Lyon 1, INSA Lyon, Villeurbanne, France.
| | - Mathilde Varret
- INSERM U1148, Université Paris Diderot, Hôpital Bichat-Claude Bernard, Paris Cedex 18, France
| | - Vanessa Boehm
- Service de gastroentérologie, MICI et Assistance Nutritive, Hopital Beaujon, Hopital Beaujon (AP-HP), Université Paris VII, Paris, France. INSERM UMR1149, Centre de Recherche sur l'Inflammation Paris Montmartre (CRI), Paris, France
| | - Jean-Pierre Rabès
- INSERM U1148, Université Paris Diderot, Hôpital Bichat-Claude Bernard, Paris Cedex 18, France; AP-HP, HUPIFO, Hôpital Ambroise Paré, Laboratoire de Biochimie et Génétique Moléculaire & UVSQ, UFR des Sciences de la Santé Simone Veil, Montigny-Le-Bretonneux, France
| | - Latifa Ferkdadji
- Service d'anatomie et de cytologie pathologiques, Hôpital Robert Debré, Université Paris 7, Paris, France
| | - Laurent Abramowitz
- Service d'Hépato-Gastroentérologie, Hôpital Bichat-Claude Bernard, Assistance Publique-Hôpitaux de Paris (AP-HP), Paris Cedex 18, France
| | - Sabrina Dumont
- UF Dyslipidemies, Service de Biochimie et Biologie moléculaire Grand Est, GHE, Hospices Civils de Lyon, Bron Cedex, France
| | | | - Catherine Boileau
- INSERM U1148, Université Paris Diderot, Hôpital Bichat-Claude Bernard, Paris Cedex 18, France
| | - Francisca Joly
- Service de gastroentérologie, MICI et Assistance Nutritive, Hopital Beaujon, Hopital Beaujon (AP-HP), Université Paris VII, Paris, France. INSERM UMR1149, Centre de Recherche sur l'Inflammation Paris Montmartre (CRI), Paris, France
| | - Jacques Schmitz
- Département de Gastroentérologie pédiatrique, Hopital Necker-Enfants Malades, Paris, France
| | | | - Dominique Bonnefont-Rousselot
- Service de Biochimie métabolique, Hôpitaux universitaires Pitié-Salpêtrière-Charles Foix (AP-HP), Paris, France; Faculté de Pharmacie de Paris, Université Paris Descartes, Sorbonne Paris Cité, Unité de Technologies Chimiques et Biologiques pour la Santé, U 1022 INSERM, UMR 8258 CNRS, Paris, France
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11
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Sirwi A, Hussain MM. Lipid transfer proteins in the assembly of apoB-containing lipoproteins. J Lipid Res 2018; 59:1094-1102. [PMID: 29650752 DOI: 10.1194/jlr.r083451] [Citation(s) in RCA: 99] [Impact Index Per Article: 14.1] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2018] [Revised: 04/12/2018] [Indexed: 11/20/2022] Open
Abstract
A better understanding of intracellular lipoprotein assembly may help identify proteins with important roles in lipid disorders. apoB-containing lipoproteins (B-lps) are macromolecular lipid and protein micelles that act as specialized transport vehicles for hydrophobic lipids. They are assembled predominantly in enterocytes and hepatocytes to transport dietary and endogenous fat, respectively, to different tissues. Assembly occurs in the endoplasmic reticulum (ER) and is dependent on lipid resynthesis in the ER and on a chaperone, namely, microsomal triglyceride transfer protein (MTTP). Precursors for lipid synthesis are obtained from extracellular sources and from cytoplasmic lipid droplets. MTTP is the major and essential lipid transfer protein that transfers phospholipids and triacylglycerols to nascent apoB for the assembly of lipoproteins. Assembly is aided by cell death-inducing DFF45-like effector B and by phospholipid transfer protein, which may facilitate additional deposition of triacylglycerols and phospholipids, respectively, to apoB. Here, we summarize the current understanding of the different steps in the assembly of B-lps and discuss the role of lipid transfer proteins in these steps to help identify new clinical targets for lipid-associated disorders, such as heart disease.
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Affiliation(s)
- Alaa Sirwi
- School of Graduate Studies, Molecular and Cell Biology Program, State University of New York Downstate Medical Center, Brooklyn, NY
| | - M Mahmood Hussain
- New York University Winthrop Hospital, Mineola, NY and Veterans Affairs New York Harbor Healthcare System, Brooklyn, NY
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12
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Kim K, Goldberg IJ, Graham MJ, Sundaram M, Bertaggia E, Lee SX, Qiang L, Haeusler RA, Metzger D, Chambon P, Yao Z, Ginsberg HN, Pajvani UB. γ-Secretase Inhibition Lowers Plasma Triglyceride-Rich Lipoproteins by Stabilizing the LDL Receptor. Cell Metab 2018; 27:816-827.e4. [PMID: 29576536 PMCID: PMC5884729 DOI: 10.1016/j.cmet.2018.02.010] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/24/2017] [Revised: 12/14/2017] [Accepted: 02/14/2018] [Indexed: 12/31/2022]
Abstract
Excess plasma triglycerides (TGs) are a key component of obesity-induced metabolic syndrome. We have shown that γ-secretase inhibitor (GSI) treatment improves glucose tolerance due to inhibition of hepatic Notch signaling but found additional Notch-independent reduction of plasma TG-rich lipoproteins (TRLs) in GSI-treated, as well as hepatocyte-specific, γ-secretase knockout (L-Ncst) mice, which suggested a primary effect on hepatocyte TRL uptake. Indeed, we found increased VLDL and LDL particle uptake in L-Ncst hepatocytes and Ncst-deficient hepatoma cells, in part through reduced γ-secretase-mediated low-density lipoprotein receptor (LDLR) cleavage and degradation. To exploit this novel finding, we generated a liver-selective Nicastrin ASO, which recapitulated glucose and lipid improvements of L-Ncst mice, with increased levels of hepatocyte LDLR. Collectively, these results identify the role of hepatic γ-secretase to regulate LDLR and suggest that liver-specific GSIs may simultaneously improve multiple aspects of the metabolic syndrome.
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Affiliation(s)
- KyeongJin Kim
- Department of Medicine, Columbia University, New York, NY, USA
| | - Ira J Goldberg
- Department of Medicine, New York University, New York, NY, USA
| | | | - Meenakshi Sundaram
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, ON, Canada
| | - Enrico Bertaggia
- Departments of Pathology and Cell Biology, Columbia University, New York, NY, USA
| | - Samuel X Lee
- Departments of Pathology and Cell Biology, Columbia University, New York, NY, USA
| | - Li Qiang
- Departments of Pathology and Cell Biology, Columbia University, New York, NY, USA
| | - Rebecca A Haeusler
- Departments of Pathology and Cell Biology, Columbia University, New York, NY, USA
| | | | | | - Zemin Yao
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, ON, Canada
| | | | - Utpal B Pajvani
- Department of Medicine, Columbia University, New York, NY, USA.
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13
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Manchekar M, Kapil R, Sun Z, Segrest JP, Dashti N. Relationship between Amphipathic β Structures in the β 1 Domain of Apolipoprotein B and the Properties of the Secreted Lipoprotein Particles in McA-RH7777 Cells. Biochemistry 2017; 56:4084-4094. [PMID: 28702990 DOI: 10.1021/acs.biochem.6b01174] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Our previous studies demonstrated that the first 1000 amino acid residues (the βα1 domain) of human apolipoprotein (apo) B-100, termed apoB:1000, are required for the initiation of lipoprotein assembly and the formation of a monodisperse stable phospholipid (PL)-rich particle. The objectives of this study were (a) to assess the effects on the properties of apoB truncates undergoing sequential inclusion of the amphipathic β strands in the 700 N-terminal residues of the β1 domain of apoB-100 and (b) to identify the subdomain in the β1 domain that is required for the formation of a microsomal triglyceride transfer protein (MTP)-dependent triacylglycerol (TAG)-rich apoB-containing particle. Characterization of particles secreted by stable transformants of McA-RH7777 cells demonstrated the following. (1) The presence of amphipathic β strands in the 200 N-terminal residues of the β1 domain resulted in the secretion of apoB truncates (apoB:1050 to apoB:1200) as both lipidated and lipid-poor particles. (2) Inclusion of residues 300-700 of the β1 domain led to the secretion of apoB:1300, apoB:1400, apoB:1500, and apoB:1700 predominantly as lipidated particles. (3) Particles containing residues 1050-1500 were all rich in PL. (4) There was a marked increase in the lipid loading capacity and TAG content of apoB:1700-containing particles. (5) Only the level of secretion of apoB:1700 was markedly diminished by MTP inhibitor BMS-197636. These results suggest that apoB:1700 marks the threshold for the formation of a TAG-rich particle and support the concept that MTP participates in apoB assembly and secretion at the stage where particles undergo a transition from PL-rich to TAG-rich.
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Affiliation(s)
| | | | | | - Jere P Segrest
- Division of Cardiovascular Medicine, Department of Medicine, Vanderbilt University Medical Center , Nashville, Tennessee 37232, United States
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14
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Liu M, Chung S, Shelness GS, Parks JS. Hepatic ABCA1 deficiency is associated with delayed apolipoprotein B secretory trafficking and augmented VLDL triglyceride secretion. Biochim Biophys Acta Mol Cell Biol Lipids 2017; 1862:1035-1043. [PMID: 28694219 DOI: 10.1016/j.bbalip.2017.07.001] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2017] [Revised: 06/30/2017] [Accepted: 07/05/2017] [Indexed: 11/30/2022]
Abstract
ATP binding cassette transporter A1 (ABCA1) is a membrane transporter that facilitates nascent HDL formation. Tangier disease subjects with complete ABCA1 deficiency have <5% of normal levels of plasma HDL, elevated triglycerides (TGs), and defective vesicular trafficking in fibroblasts and macrophages. Hepatocyte-specific ABCA1 knockout mice (HSKO) have a similar lipid phenotype with 20% of normal plasma HDL levels and a two-fold elevation of plasma TGs due to hepatic overproduction of large, triglyceride-enriched VLDL. We hypothesized that enhanced VLDL TG secretion in the absence of hepatocyte ABCA1 is due to altered intracellular trafficking of apolipoprotein B (apoB), resulting in augmented TG addition to nascent VLDL. We found that trafficking of newly synthesized apoB through the secretory pathway was delayed in ABCA1-silenced rat hepatoma cells and HSKO primary hepatocytes, relative to controls. Endoglycosidase H treatment of cellular apoB revealed a likely delay in apoB trafficking in post-ER compartments. The reduced rate of protein trafficking was also observed for an adenoviral-expressed GPI-linked fluorescent fusion protein, but not albumin, suggesting a selective delay of secretory cargoes in the absence of hepatocyte ABCA1. Our results suggest an important role for hepatic ABCA1 in regulating secretory trafficking and modulating VLDL expansion during the TG accretion phase of hepatic lipoprotein particle assembly.
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Affiliation(s)
- Mingxia Liu
- Department of Internal Medicine-Section on Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA.
| | - Soonkyu Chung
- Department of Internal Medicine-Section on Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA
| | - Gregory S Shelness
- Department of Internal Medicine-Section on Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA
| | - John S Parks
- Department of Internal Medicine-Section on Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA; Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, NC, USA
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15
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Mulligan C, Kondakala S, Yang EJ, Stokes JV, Stewart JA, Kaplan BLF, Howell GE. Exposure to an environmentally relevant mixture of organochlorine compounds and polychlorinated biphenyls Promotes hepatic steatosis in male Ob/Ob mice. ENVIRONMENTAL TOXICOLOGY 2017; 32:1399-1411. [PMID: 27533883 DOI: 10.1002/tox.22334] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/13/2016] [Revised: 07/25/2016] [Accepted: 07/30/2016] [Indexed: 05/19/2023]
Abstract
Hepatic steatosis is recognized as an independent risk factor for the development of cardiovascular disease. While obesity and type 2 diabetes are well-established risk factors in the development of hepatic steatosis, recent studies have revealed exposure to mixtures of persistent organic pollutants (POPs), which are environmental contaminants in various fatty foods, can promote steatosis. Thus, the present study was designed to determine if exposure to a defined mixture of prevalent polychlorinated biphenyls (PCBs) and organochlorine (OC) pesticides or their metabolites promote hepatic steatosis in a genetically induced model of type 2 diabetes, the leptin-deficient ob/ob mouse. Male C57BL/6J wild type (WT) or ob/ob mice were administered an environmentally relevant mixture of PCBs and OCs for 7 weeks via oral gavage. Exposure to POPs did not significantly alter fasting serum glucose or insulin levels. However, POPs exposure significantly increased hepatic triglyceride content in ob/ob animals, while decreasing serum triglyceride levels. This POPs-mediated increase in hepatic triglyceride content did not appear to be associated with significantly increased inflammation in either the liver or adipose. Exposure to POPs significantly induced the expression of cytochrome P450 3a11 in WT animals, yet the expression of this cytochrome was significantly downregulated in ob/ob animals regardless of POPs exposure. Taken together, the present data indicate exposure to an environmentally relevant mixture of both PCBs and OC pesticides in ob/ob mice promotes hepatic steatosis while decreasing hypertriglyceridemia, which demonstrates exposure to a defined mixture of POPs alters systemic lipid metabolism in a genetically induced model of obesity and type 2 diabetes. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1399-1411, 2017.
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Affiliation(s)
- Charlee Mulligan
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - Sandeep Kondakala
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - Eun-Ju Yang
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - John V Stokes
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - James A Stewart
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - Barbara L F Kaplan
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
| | - George E Howell
- Department of Basic Sciences, Mississippi State University, College of Veterinary Medicine, Mississippi State, Mississippi, 39762
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16
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Luo Z, Wei CC, Ye HM, Zhao HP, Song YF, Wu K. Effect of dietary choline levels on growth performance, lipid deposition and metabolism in juvenile yellow catfish Pelteobagrus fulvidraco. Comp Biochem Physiol B Biochem Mol Biol 2016; 202:1-7. [DOI: 10.1016/j.cbpb.2016.07.005] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2016] [Revised: 07/14/2016] [Accepted: 07/22/2016] [Indexed: 01/20/2023]
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17
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Walsh MT, Hussain MM. Targeting microsomal triglyceride transfer protein and lipoprotein assembly to treat homozygous familial hypercholesterolemia. Crit Rev Clin Lab Sci 2016; 54:26-48. [PMID: 27690713 DOI: 10.1080/10408363.2016.1221883] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
Homozygous familial hypercholesterolemia (HoFH) is a polygenic disease arising from defects in the clearance of plasma low-density lipoprotein (LDL), which results in extremely elevated plasma LDL cholesterol (LDL-C) and increased risk of atherosclerosis, coronary heart disease, and premature death. Conventional lipid-lowering therapies, such as statins and ezetimibe, are ineffective at lowering plasma cholesterol to safe levels in these patients. Other therapeutic options, such as LDL apheresis and liver transplantation, are inconvenient, costly, and not readily available. Recently, lomitapide was approved by the Federal Drug Administration as an adjunct therapy for the treatment of HoFH. Lomitapide inhibits microsomal triglyceride transfer protein (MTP), reduces lipoprotein assembly and secretion, and lowers plasma cholesterol levels by over 50%. Here, we explain the steps involved in lipoprotein assembly, summarize the role of MTP in lipoprotein assembly, explore the clinical and molecular basis of HoFH, and review pre-clinical studies and clinical trials with lomitapide and other MTP inhibitors for the treatment of HoFH. In addition, ongoing research and new approaches underway for better treatment modalities are discussed.
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Affiliation(s)
- Meghan T Walsh
- a School of Graduate Studies, Molecular and Cell Biology Program, State University of New York Downstate Medical Center , Brooklyn , NY , USA.,b Department of Cell Biology , State University of New York Downstate Medical Center , Brooklyn , NY , USA
| | - M Mahmood Hussain
- b Department of Cell Biology , State University of New York Downstate Medical Center , Brooklyn , NY , USA.,c Department of Pediatrics , SUNY Downstate Medical Center , Brooklyn , NY , USA.,d VA New York Harbor Healthcare System , Brooklyn , NY , USA , and.,e Winthrop University Hospital , Mineola , NY , USA
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18
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Cheng D, Xu X, Simon T, Boudyguina E, Deng Z, VerHague M, Lee AH, Shelness GS, Weinberg RB, Parks JS. Very Low Density Lipoprotein Assembly Is Required for cAMP-responsive Element-binding Protein H Processing and Hepatic Apolipoprotein A-IV Expression. J Biol Chem 2016; 291:23793-23803. [PMID: 27655915 DOI: 10.1074/jbc.m116.749283] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2016] [Indexed: 12/21/2022] Open
Abstract
Hepatic apolipoprotein A-IV (apoA-IV) expression is correlated with hepatic triglyceride (TG) content in mouse models of chronic hepatosteatosis, and steatosis-induced hepatic apoA-IV gene expression is regulated by nuclear transcription factor cAMP-responsive element-binding protein H (CREBH) processing. To define what aspects of TG homeostasis regulate hepatic CREBH processing and apoA-IV gene expression, several mouse models of attenuated VLDL particle assembly were subjected to acute hepatosteatosis induced by an overnight fast or short term ketogenic diet feeding. Compared with chow-fed C57BL/6 mice, fasted or ketogenic diet-fed mice displayed increased hepatic TG content, which was highly correlated (r2 = 0.95) with apoA-IV gene expression, and secretion of larger, TG-enriched VLDL, despite a lower rate of TG secretion and a similar or reduced rate of apoB100 secretion. When VLDL particle assembly and secretion was inhibited by hepatic shRNA-induced apoB silencing or genetic or pharmacologic reduction in microsomal triglyceride transfer protein (MTP) activity, hepatic TG content increased dramatically; however, CREBH processing and apoA-IV gene expression were attenuated compared with controls. Adenovirus-mediated reconstitution of MTP expression proportionately restored CREBH processing and apoA-IV expression in liver-specific MTP knock-out mice. These results reveal that hepatic TG content, per se, does not regulate CREBH processing. Instead, TG mobilization into the endoplasmic reticulum for nascent VLDL particle assembly activates CREBH processing and enhances apoA-IV gene expression in the setting of acute steatosis. We conclude that VLDL assembly and CREBH activation play key roles in the response to hepatic steatosis by up-regulating apoA-IV and promoting assembly and secretion of larger, more TG-enriched VLDL particles.
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Affiliation(s)
- Dongmei Cheng
- From the Departments of Internal Medicine-Section on Molecular Medicine
| | - Xu Xu
- the Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, New York 10065
| | - Trang Simon
- Internal Medicine-Section on Gastroenterology
| | - Elena Boudyguina
- From the Departments of Internal Medicine-Section on Molecular Medicine
| | | | - Melissa VerHague
- From the Departments of Internal Medicine-Section on Molecular Medicine
| | - Ann-Hwee Lee
- the Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, New York 10065
| | | | | | - John S Parks
- From the Departments of Internal Medicine-Section on Molecular Medicine, .,Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157 and
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19
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Microsomal Triglyceride Transfer Protein (MTP) Associates with Cytosolic Lipid Droplets in 3T3-L1 Adipocytes. PLoS One 2015; 10:e0135598. [PMID: 26267806 PMCID: PMC4534446 DOI: 10.1371/journal.pone.0135598] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2014] [Accepted: 07/16/2015] [Indexed: 12/18/2022] Open
Abstract
Lipid droplets are intracellular energy storage organelles composed of a hydrophobic core of neutral lipid, surrounded by a monolayer of phospholipid and a diverse array of proteins. The function of the vast majority of these proteins with regard to the formation and/or turnover of lipid droplets is unknown. Our laboratory was the first to report that microsomal triglyceride transfer protein (MTP), a lipid transfer protein essential for the assembly of triglyceride-rich lipoproteins, was expressed in adipose tissue of humans and mice. In addition, our studies suggested that MTP was associated with lipid droplets in both brown and white fat. Our observations led us to hypothesize that MTP plays a key role in lipid droplet formation and/or turnover. The objective of these studies was to gain insight into the function of MTP in adipocytes. Using molecular, biochemical, and morphologic approaches we have shown: 1) MTP protein levels increase nearly five-fold as 3T3-L1 cells differentiate into adipocytes. 2) As 3T3-L1 cells undergo differentiation, MTP moves from the juxtanuclear region of the cell to the surface of lipid droplets. MTP and perilipin 2, a major lipid droplet surface protein, are found on the same droplets; however, MTP does not co-localize with perilipin 2. 3) Inhibition of MTP activity has no effect on the movement of triglyceride out of the cell either as a lipid complex or via lipolysis. 4) MTP is found associated with lipid droplets within hepatocytes from human fatty livers, suggesting that association of MTP with lipid droplets is not restricted to adipocytes. In summary, our data demonstrate that MTP is a lipid droplet-associated protein. Its location on the surface of the droplet in adipocytes and hepatocytes, coupled with its known function as a lipid transfer protein and its increased expression during adipocyte differentiation suggest a role in lipid droplet biology.
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González-Reimers E, Quintero-Platt G, Rodríguez-Gaspar M, Alemán-Valls R, Pérez-Hernández O, Santolaria-Fernández F. Liver steatosis in hepatitis C patients. World J Hepatol 2015; 7:1337-1346. [PMID: 26052379 PMCID: PMC4450197 DOI: 10.4254/wjh.v7.i10.1337] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/31/2014] [Revised: 01/31/2015] [Accepted: 03/09/2015] [Indexed: 02/06/2023] Open
Abstract
There is controversy regarding some aspects of hepatitis C virus (HCV) infection-associated liver steatosis, and their relationship with body fat stores. It has classically been found that HCV, especially genotype 3, exerts direct metabolic effects which lead to liver steatosis. This supports the existence of a so called viral steatosis and a metabolic steatosis, which would affect HCV patients who are also obese or diabetics. In fact, several genotypes exert metabolic effects which overlap with some of those observed in the metabolic syndrome. In this review we will analyse the pathogenic pathways involved in the development of steatosis in HCV patients. Several cytokines and adipokines also become activated and are involved in “pure” steatosic effects, in addition to inflammation. They are probably responsible for the evolution of simple steatosis to steatohepatitis, making it difficult to explain why such alterations only affect a proportion of steatosic patients.
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New insights into the pathophysiology of dyslipidemia in type 2 diabetes. Atherosclerosis 2015; 239:483-95. [PMID: 25706066 DOI: 10.1016/j.atherosclerosis.2015.01.039] [Citation(s) in RCA: 278] [Impact Index Per Article: 27.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/30/2014] [Revised: 01/28/2015] [Accepted: 01/30/2015] [Indexed: 02/06/2023]
Abstract
Cardiovascular disease (CVD) remains the leading cause of morbidity and mortality for patients with type 2 diabetes, despite recent significant advances in management strategies to lessen CVD risk factors. A major cause is the atherogenic dyslipidemia, which consists of elevated plasma concentrations of both fasting and postprandial triglyceride-rich lipoproteins (TRLs), small dense low-density lipoprotein (LDL) and low high-density lipoprotein (HDL) cholesterol. The different components of diabetic dyslipidemia are not isolated abnormalities but closely linked to each other metabolically. The underlying disturbances are hepatic overproduction and delayed clearance of TRLs. Recent results have unequivocally shown that triglyceride-rich lipoproteins and their remnants are atherogenic. To develop novel strategies for the prevention and treatment of dyslipidaemia, it is essential to understand the pathophysiology of dyslipoproteinaemia in humans. Here, we review recent advances in our understanding of the pathophysiology of diabetic dyslipidemia.
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Manchekar M, Liu Y, Sun Z, Richardson PE, Dashti N. Phospholipid transfer protein plays a major role in the initiation of apolipoprotein B-containing lipoprotein assembly in mouse primary hepatocytes. J Biol Chem 2015; 290:8196-205. [PMID: 25638820 DOI: 10.1074/jbc.m114.602748] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
In this study, we tested the hypothesis that phospholipid transfer protein (PLTP) is a plausible mediator of phospholipid (PL) transfer to the N-terminal 1000 residues of apoB (apoB:1000) leading to the initiation of apoB-containing lipoprotein assembly. To this end, primary hepatocytes from wild type (WT) and PLTP knock-out (KO) mice were transduced with adenovirus-apoB:1000 with or without co-transduction with adenovirus-PLTP, and the assembly and secretion of apoB:1000-containing lipoproteins were assessed. PLTP deficiency resulted in a 65 and 72% reduction in the protein and lipid content, respectively, of secreted apoB:1000-containing lipoproteins. Particles secreted by WT hepatocytes contained 69% PL, 9% diacylglycerol (DAG), and 23% triacylglycerol (TAG) with a stoichiometry of 46 PL, 6 DAG, and 15 TAG molecules per apoB:1000. PLTP absence drastically altered the lipid composition of apoB:1000 lipoproteins; these particles contained 46% PL, 13% DAG, and 41% TAG with a stoichiometry of 27 PL, 10 DAG, and 23 TAG molecules per apoB:1000. Reintroduction of Pltp gene into PLTP-KO hepatocytes stimulated the lipidation and secretion of apoB:1000-containing lipoproteins by ∼3-fold; the lipid composition and stoichiometry of these particles were identical to those secreted by WT hepatocytes. In contrast to the WT, apoB:1000 in PLTP-KO hepatocytes was susceptible to intracellular degradation predominantly in the post-endoplasmic reticulum, presecretory compartment. Reintroduction of Pltp gene into PLTP-KO hepatocytes restored the stability of apoB:1000. These results provide compelling evidence that in hepatocytes initial recruitment of PL by apoB:1000 leading to the formation of the PL-rich apoB-containing initiation complex is mediated to a large extent by PLTP.
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Affiliation(s)
- Medha Manchekar
- From the Department of Medicine, Division of Gerontology, Geriatrics, and Palliative Care, Basic Sciences Section, University of Alabama, Birmingham, Alabama 35294 and
| | - Yanwen Liu
- From the Department of Medicine, Division of Gerontology, Geriatrics, and Palliative Care, Basic Sciences Section, University of Alabama, Birmingham, Alabama 35294 and
| | - Zhihuan Sun
- From the Department of Medicine, Division of Gerontology, Geriatrics, and Palliative Care, Basic Sciences Section, University of Alabama, Birmingham, Alabama 35294 and
| | - Paul E Richardson
- the Department of Chemistry and Physics, Coastal Carolina University, Conway, South Carolina 29528
| | - Nassrin Dashti
- From the Department of Medicine, Division of Gerontology, Geriatrics, and Palliative Care, Basic Sciences Section, University of Alabama, Birmingham, Alabama 35294 and
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Zhang L, Zhong S, Li Y, Ji G, Sundaram M, Yao Z. Global Inactivation of the Pla2g6 Gene in Mice Does Not Cause Dyslipidemia under Chow or High-fat Diet Conditions. J Cancer Prev 2014; 18:235-48. [PMID: 25337551 PMCID: PMC4189460 DOI: 10.15430/jcp.2013.18.3.235] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2013] [Revised: 09/10/2013] [Accepted: 09/10/2013] [Indexed: 01/30/2023] Open
Abstract
Background: Genome-wide association studies suggest that plasma triacylglyceride (TAG) in humans was associated with variation in the PLA2G6 locus, a gene that encodes calcium-independent phospholipase A2 (iPLA2β). The objective of the present study is to understand the impact of genetic inactivation of iPLA2β on hepatic TAG metabolism in C57BL/6 mice. Methods: Male iPLA2β+/− mice were backcrossed with female iPLA2β−/− mice for up to 10 generations prior to experiments. Lipid and lipoprotein metabolism from plasma, hepatocytes, thigh subcutaneous adipose and thigh skeletal muscle tissues of the mice were determined under various experimental conditions. Results: The iPLA2β−/− mice, either male or female as compared with iPLA2β+/+ littermates, showed no change in fasted or postprandial plasma TAG or total cholesterol at young (12–15 weeks) or old (40–44 weeks) ages under chow diet or high-fat diet (HFD) conditions. However, fractionation of plasma lipoproteins showed that under HFD conditions, there was a significant increase (by 40%) in apoB-100 association with VLDL1 fractions in iPLA2β−/− mice as compared with iPLA2β+/+ littermates. There was no significant difference in triglyceride or cholesterol contents in the liver, muscle, or adipose tissue between iPLA2β−/− and iPLA2β+/+ littermates. Metabolic labeling experiments with cultured primary hepatocytes isolated from iPLA2β−/− mice also showed 2-fold increase in the secretion of [35S]methionine-labeled apoB-100 in VLDL1 fractions as compared with that from iPLA2β+/+ hepatocytes. Likewise, secretion of [3H]palmitate-labeled TAG from the iPLA2β−/− hepatocytes was increased by 2-fold. Conclusions: Although iPLA2β may play a role in TAG-rich VLDL1 production from cultured hepatocytes, there is no evidence that inactivation of iPLA2β would lead to dyslipidemia in mice in vivo.
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Affiliation(s)
- Li Zhang
- Institute of Digestive Disease, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Shumei Zhong
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Canada
| | - Ying Li
- College of Life Science, Chongqing Normal University, Chongqing, China
| | - Guang Ji
- Institute of Digestive Disease, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Meenakshi Sundaram
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Canada
| | - Zemin Yao
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Canada
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Li X, Li Y, Yang W, Xiao C, Fu S, Deng Q, Ding H, Wang Z, Liu G, Li X. SREBP-1c overexpression induces triglycerides accumulation through increasing lipid synthesis and decreasing lipid oxidation and VLDL assembly in bovine hepatocytes. J Steroid Biochem Mol Biol 2014; 143:174-82. [PMID: 24565561 DOI: 10.1016/j.jsbmb.2014.02.009] [Citation(s) in RCA: 70] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/29/2013] [Revised: 01/20/2014] [Accepted: 02/14/2014] [Indexed: 02/06/2023]
Abstract
The natural incidence of fatty liver in ruminants is significantly higher than in monogastric animals. Fatty liver is associated with sterol regulatory element-binding protein 1c (SREBP-1c). The aim of this study was to investigate the regulatory network effects of SREBP-1c on the lipid metabolic genes involved in fatty acid uptake, activation, oxidation, synthesis, and very low-density lipoprotein (VLDL) assembly in bovine hepatocytes. In vitro, bovine hepatocytes were transfected with an adenovirus-mediated SREBP-1c overexpression vector. SREBP-1c overexpression significantly up-regulated the expression and activity of the fatty acid uptake, activation, and synthesis enzymes: liver fatty acid binding protein, fatty acid translocase, acyl-CoA synthetase long-chain 1, acetyl-CoA carboxylase 1, and fatty acid synthase, increasing triglyceride (TG) synthesis and accumulation. SREBP-1c overexpression down-regulated the expression and activity of the lipid oxidation enzymes: carnitine palmitoyltransferase 1 and carnitine palmitoyltransferase 2. Furthermore, the apolipoprotein B100 expression and microsomal triglyceride transfer protein activity were significantly decreased. SREBP-1c overexpression reduced lipid oxidation and VLDL synthesis, thereby decreasing TG disposal and export. Therefore, large amounts of TG accumulated in the bovine hepatocytes. Taken together, these results indicate that SREBP-1c overexpression increases lipid synthesis and decreases lipid oxidation and VLDL export, thereby inducing TG accumulation in bovine hepatocytes.
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Affiliation(s)
- Xinwei Li
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Yu Li
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Wentao Yang
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Chong Xiao
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Shixin Fu
- Institute of Animal Science and Technology, Heilongjiang Bayi Agriculture University, Daqing 163319, Heilongjiang, China
| | - Qinghua Deng
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Hongyan Ding
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Zhe Wang
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China
| | - Guowen Liu
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China.
| | - Xiaobing Li
- Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, Jilin, China.
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Abstract
PURPOSE OF REVIEW To summarize the evidence for the presence of two lipid absorption pathways and their regulation. RECENT FINDINGS Lipid absorption involves hydrolysis of dietary fat in the lumen of the intestine, followed by the uptake of hydrolyzed products by enterocytes. Lipids are resynthesized in the endoplasmic reticulum and are either secreted with chylomicrons and HDLs or stored as cytoplasmic lipid droplets. Lipids in the droplets are hydrolyzed and are secreted at a later time. Secretion of lipids by the chylomicron and HDL pathways are dependent on microsomal triglyceride transfer protein (MTP) and ATP-binding cassette family A protein 1, respectively, and are regulated independently. Gene-ablation studies showed that MTP function and chylomicron assembly is essential for the absorption of triglycerides. Ablation of MTP abolishes triglyceride absorption and results in massive triglyceride accumulation in enterocytes. Although the majority of phospholipid, cholesterol, and vitamin E are absorbed through the chylomicron pathway, a significant amount of these lipids are also absorbed via the HDL pathway. Chylomicron assembly and secretion is increased by the enhanced availability of fatty acids, whereas the HDL pathway is upregulated by liver X receptor agonists. SUMMARY Triglycerides are exclusively transported with chylomicrons and this process is critically dependent on MTP. In addition to chylomicrons, absorption of phospholipids, free cholesterol, retinol, and vitamin E also involves HDLs. These two pathways are complementary and are regulated independently. They may be targeted to lower lipid absorption in order to control hyperlipidemia, obesity, metabolic syndrome, steatosis, insulin resistance, atherosclerosis, and other disorders.
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Affiliation(s)
- M Mahmood Hussain
- aDepartment of Cell Biology bDepartment of Pediatrics, SUNY Downstate Medical Center cVA New York Harbor Healthcare System, Brooklyn, New York, USA
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VerHague MA, Cheng D, Weinberg RB, Shelness GS. Apolipoprotein A-IV Expression in Mouse Liver Enhances Triglyceride Secretion and Reduces Hepatic Lipid Content by Promoting Very Low Density Lipoprotein Particle Expansion. Arterioscler Thromb Vasc Biol 2013; 33:2501-8. [DOI: 10.1161/atvbaha.113.301948] [Citation(s) in RCA: 79] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023]
Affiliation(s)
- Melissa A. VerHague
- From the Department of Pathology (M.A.V., D.C., G.S.S.), Department of Internal Medicine (R.B.W.), and Department of Physiology & Pharmacology (R.B.W.), Wake Forest School of Medicine, Winston-Salem, NC
| | - Dongmei Cheng
- From the Department of Pathology (M.A.V., D.C., G.S.S.), Department of Internal Medicine (R.B.W.), and Department of Physiology & Pharmacology (R.B.W.), Wake Forest School of Medicine, Winston-Salem, NC
| | - Richard B. Weinberg
- From the Department of Pathology (M.A.V., D.C., G.S.S.), Department of Internal Medicine (R.B.W.), and Department of Physiology & Pharmacology (R.B.W.), Wake Forest School of Medicine, Winston-Salem, NC
| | - Gregory S. Shelness
- From the Department of Pathology (M.A.V., D.C., G.S.S.), Department of Internal Medicine (R.B.W.), and Department of Physiology & Pharmacology (R.B.W.), Wake Forest School of Medicine, Winston-Salem, NC
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27
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Borén J, Taskinen MR, Olofsson SO, Levin M. Ectopic lipid storage and insulin resistance: a harmful relationship. J Intern Med 2013; 274:25-40. [PMID: 23551521 DOI: 10.1111/joim.12071] [Citation(s) in RCA: 157] [Impact Index Per Article: 13.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
Obesity increases the risk of metabolic diseases, including insulin resistance and type 2 diabetes, as well as cardiovascular disease. In addition to lipid accumulation in adipose tissue, obesity is associated with increased lipid storage in ectopic tissues, such as skeletal muscle and liver. Furthermore, lipid accumulation in the heart may result in cardiac dysfunction and heart failure. It has recently been demonstrated that intracellular lipid accumulation in ectopic tissues leads to pathological responses and impaired insulin signalling. Here, we will review the current understanding of how lipid storage and lipid droplet physiology affect the risk of developing metabolic diseases.
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Affiliation(s)
- J Borén
- Department of Molecular and Clinical Medicine, University of Gothenburg, Gothenburg, Sweden.
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Endoplasmic reticulum-localized hepatic lipase decreases triacylglycerol storage and VLDL secretion. Biochim Biophys Acta Mol Cell Biol Lipids 2013; 1831:1113-23. [DOI: 10.1016/j.bbalip.2013.01.017] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2012] [Revised: 01/09/2013] [Accepted: 01/23/2013] [Indexed: 01/07/2023]
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Lipoproteins in Drosophila melanogaster--assembly, function, and influence on tissue lipid composition. PLoS Genet 2012; 8:e1002828. [PMID: 22844248 PMCID: PMC3406001 DOI: 10.1371/journal.pgen.1002828] [Citation(s) in RCA: 187] [Impact Index Per Article: 14.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2012] [Accepted: 05/25/2012] [Indexed: 11/19/2022] Open
Abstract
Interorgan lipid transport occurs via lipoproteins, and altered lipoprotein levels correlate with metabolic disease. However, precisely how lipoproteins affect tissue lipid composition has not been comprehensively analyzed. Here, we identify the major lipoproteins of Drosophila melanogaster and use genetics and mass spectrometry to study their assembly, interorgan trafficking, and influence on tissue lipids. The apoB-family lipoprotein Lipophorin (Lpp) is the major hemolymph lipid carrier. It is produced as a phospholipid-rich particle by the fat body, and its secretion requires Microsomal Triglyceride Transfer Protein (MTP). Lpp acquires sterols and most diacylglycerol (DAG) at the gut via Lipid Transfer Particle (LTP), another fat body-derived apoB-family lipoprotein. The gut, like the fat body, is a lipogenic organ, incorporating both de novo-synthesized and dietary fatty acids into DAG for export. We identify distinct requirements for LTP and Lpp-dependent lipid mobilization in contributing to the neutral and polar lipid composition of the brain and wing imaginal disc. These studies define major routes of interorgan lipid transport in Drosophila and uncover surprising tissue-specific differences in lipoprotein lipid utilization.
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Sparks JD, Sparks CE, Adeli K. Selective hepatic insulin resistance, VLDL overproduction, and hypertriglyceridemia. Arterioscler Thromb Vasc Biol 2012; 32:2104-12. [PMID: 22796579 DOI: 10.1161/atvbaha.111.241463] [Citation(s) in RCA: 158] [Impact Index Per Article: 12.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Insulin plays a central role in regulating energy metabolism, including hepatic transport of very low-density lipoprotein (VLDL)-associated triglyceride. Hepatic hypersecretion of VLDL and consequent hypertriglyceridemia leads to lower circulating high-density lipoprotein levels and generation of small dense low-density lipoproteins characteristic of the dyslipidemia commonly observed in metabolic syndrome and type 2 diabetes mellitus. Physiological fluctuations of insulin modulate VLDL secretion, and insulin inhibition of VLDL secretion upon feeding may be the first pathway to become resistant in obesity that leads to VLDL hypersecretion. This review summarizes the role of insulin-related signaling pathways that determine hepatic VLDL production. Disruption in signaling pathways that reduce generation of the second messenger phosphatidylinositide (3,4,5) triphosphate downstream of activated phosphatidylinositide 3-kinase underlies the development of VLDL hypersecretion. As insulin resistance progresses, a number of pathways are altered that further augment VLDL hypersecretion, including hepatic inflammatory pathways. Insulin plays a complex role in regulating glucose metabolism, and it is not surprising that the role of insulin in VLDL and lipid metabolism will prove equally complex.
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Affiliation(s)
- Janet D Sparks
- University of Rochester Medical Center, Department of Pathology and Laboratory Medicine, Rochester, NY, USA
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31
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Ye J. Hepatitis C virus: a new class of virus associated with particles derived from very low-density lipoproteins. Arterioscler Thromb Vasc Biol 2012; 32:1099-103. [PMID: 22517369 DOI: 10.1161/atvbaha.111.241448] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
Hepatitis C virus (HCV) infects 3% of the world population and is the leading cause of liver failure in the United States. A unique feature of HCV is that the viral particles are integral to very low-density lipoprotein (VLDL)-derived lipoprotein particles. The virus is assembled into VLDL in hepatocytes and released out of the cells together with VLDL. The virus then infects more hepatocytes by entering the cells through the low-density lipoprotein receptor, which mediates uptake of majorities of VLDL-derived lipoprotein particles. These observations suggest that HCV may belong to a novel class of viruses that is associated with VLDL. Understanding the relationship between HCV and VLDL metabolism may reveal new strategies to treat HCV infection.
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Affiliation(s)
- Jin Ye
- Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, TX 75390-9046, USA.
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32
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Abstract
Lipid homeostasis is maintained through the coordination of lipid metabolism in various tissues, including adipose tissue and the liver. The disruption of lipid homeostasis often results in the development of metabolic disorders such as obesity, diabetes mellitus, liver steatosis, and cardiovascular diseases. Cell death-inducing DNA fragmentation factor 45-like effector family proteins, including Cidea, Cideb, and Fsp27 (Cidec), are emerging as important regulators of various lipid metabolic pathways and play pivotal roles in the development of metabolic disorders. This review summarizes the latest cell death-inducing DNA fragmentation factor 45-like effector protein discoveries related to the control of lipid metabolism, with emphasis on the role of these proteins in lipid droplet growth in adipocytes and in the regulation of very low-density lipoprotein lipidation and maturation in hepatocytes.
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Affiliation(s)
- Li Xu
- Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China
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Lehner R, Lian J, Quiroga AD. Lumenal lipid metabolism: implications for lipoprotein assembly. Arterioscler Thromb Vasc Biol 2012; 32:1087-93. [PMID: 22517367 DOI: 10.1161/atvbaha.111.241497] [Citation(s) in RCA: 68] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Overproduction of apolipoprotein B (apoB)-containing lipoproteins by the liver and the intestine is 1 of the hallmarks of insulin resistance and type 2 diabetes and a well-established risk factor of cardiovascular disease. The assembly of apoB lipoproteins is regulated by the availability of lipids that form the neutral lipid core (triacylglycerol and cholesteryl ester) and the limiting lipoprotein monolayer (phospholipids and cholesterol). Although tremendous advances have been made over the past decade toward understanding neutral lipid and phospholipid biosynthesis and neutral lipid storage in cytosolic lipid droplets (LDs), little is known about the mechanisms that govern the transfer of lipids to the lumen of the endoplasmic reticulum for apoB lipidation. ApoB-synthesizing organs can deposit synthesized neutral lipids into at least 3 different types of LDs, each decorated with a subset of specific proteins: perilipin-decorated cytosolic LDs, and 2 types of LDs formed in the lumen of the endoplasmic reticulum, the secretion-destined LDs containing apoB, and resident lumenal LDs coated with microsomal triglyceride transfer protein and exchangeable apolipoproteins. This brief review will address the current knowledge of lumenal lipid metabolism in the context of apoB assembly and lipid storage.
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Affiliation(s)
- Richard Lehner
- Department of Pediatrics and Cell Biology, Group on Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada.
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Abstract
PURPOSE OF REVIEW A strong positive correlation between plasma apolipoprotein (apo) C-III and triglyceride concentrations has been invariably observed in human and animal studies. The hypertriglyceridemic effect of apo C-III has been conventionally explained by its extracellular roles in inhibiting lipolysis catalysed by lipoprotein lipase and attenuating triglyceride-rich lipoprotein clearance through receptor-dependent and/or independent mechanisms. However, recent experimental evidence suggests that apo C-III may also play an intracellular role in promoting hepatic triglyceride-rich lipoprotein production. RECENT FINDINGS Kinetic studies with humans and genetically modified mice have shown that apo C-III is linked with increased production of triglyceride-rich lipoproteins, such as very-low-density lipoprotein 1 (VLDL1). Mutational studies on human apo C-III variants (originally identified in humans with hypotriglyceridemia or hyperalphalipoproteinemia) provide the structure-function analysis of human apo C-III, demonstrating that loss-of-function mutations within human apo C-III impair the assembly and secretion of triglyceride-rich VLDL1 under lipid-rich conditions. SUMMARY The current review summarizes recent experimental evidence for an intrahepatic role of human apo C-III in promoting mobilization and utilization of triglyceride during VLDL1 assembly/secretion. Understanding mechanisms by which hepatic apo C-III expression is regulated under insulin resistance and diabetic conditions will lead to better and more rational strategies for the prevention and treatment of diabetic hypertriglyceridemia that is closely related to premature atherosclerosis.
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Affiliation(s)
- Zemin Yao
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Ontario, Canada
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Liu M, Chung S, Shelness GS, Parks JS. Hepatic ABCA1 and VLDL triglyceride production. BIOCHIMICA ET BIOPHYSICA ACTA 2012; 1821:770-7. [PMID: 22001232 PMCID: PMC3272310 DOI: 10.1016/j.bbalip.2011.09.020] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/18/2011] [Revised: 09/23/2011] [Accepted: 09/26/2011] [Indexed: 02/04/2023]
Abstract
Elevated plasma triglyceride (TG) and reduced high density lipoprotein (HDL) concentrations are prominent features of metabolic syndrome (MS) and type 2 diabetes (T2D). Individuals with Tangier disease also have elevated plasma TG concentrations and a near absence of HDL, resulting from mutations in ATP binding cassette transporter A1 (ABCA1), which facilitates the efflux of cellular phospholipid and free cholesterol to assemble with apolipoprotein A-I (apoA-I), forming nascent HDL particles. In this review, we summarize studies focused on the regulation of hepatic very low density lipoprotein (VLDL) TG production, with particular attention on recent evidence connecting hepatic ABCA1 expression to VLDL, LDL, and HDL metabolism. Silencing ABCA1 in McArdle rat hepatoma cells results in diminished assembly of large (>10nm) nascent HDL particles, diminished PI3 kinase activation, and increased secretion of large, TG-enriched VLDL1 particles. Hepatocyte-specific ABCA1 knockout (HSKO) mice have a similar plasma lipid phenotype as Tangier disease subjects, with a two-fold elevation of plasma VLDL TG, 50% lower LDL, and 80% reduction in HDL concentrations. This lipid phenotype arises from increased hepatic secretion of VLDL1 particles, increased hepatic uptake of plasma LDL by the LDL receptor, elimination of nascent HDL particle assembly by the liver, and hypercatabolism of apoA-I by the kidney. These studies highlight a novel role for hepatic ABCA1 in the metabolism of all three major classes of plasma lipoproteins and provide a metabolic link between elevated TG and reduced HDL levels that are a common feature of Tangier disease, MS, and T2D. This article is part of a Special Issue entitled: Triglyceride Metabolism and Disease.
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Affiliation(s)
- Mingxia Liu
- Department of Pathology/Section on Lipid Sciences, Wake Forest School of Medicine, Winston-Salem, NC, USA
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36
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Khatun I, Zeissig S, Iqbal J, Wang M, Curiel D, Shelness GS, Blumberg RS, Hussain MM. Phospholipid transfer activity of microsomal triglyceride transfer protein produces apolipoprotein B and reduces hepatosteatosis while maintaining low plasma lipids in mice. Hepatology 2012; 55:1356-68. [PMID: 22121032 PMCID: PMC3299931 DOI: 10.1002/hep.25504] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/23/2011] [Accepted: 11/13/2011] [Indexed: 12/12/2022]
Abstract
UNLABELLED Microsomal triglyceride transfer protein (MTP), essential for apolipoprotein B (apoB) biosynthesis, evolved as a phospholipid transfer protein and acquired triglyceride transfer activity during a transition from invertebrates to vertebrates. But it is unknown whether MTP directly transfers lipids onto apoB in vivo and, if it does, whether both neutral and polar lipid transfer activities of MTP are critical for lipoprotein assembly. The molecular bases for differences in lipid transfer activities with respect to distinct domains in Drosophila MTP (dMTP) and human MTP (hMTP) are not obvious because both proteins have very similar primary, secondary, and tertiary structures. We used an in vivo approach to delineate physiological significance of these distinct lipid transfer activities by expressing dMTP (transfers phospholipids) and hMTP (transfers phospholipids and triglycerides) orthologs using adenoviruses in liver-specific MTP-deficient (L-MTP(-/-)) mice that have low plasma and high hepatic lipids. Both orthologs improved plasma lipids but plasma triglycerides were lower in dMTP mice due to lower hepatic triglyceride and apoB production. Hepatosteatosis in L-MTP(-/-) mice was ameliorated to similar levels by both. Attenuation of hepatosteatosis upon dMTP expression pertained to enhanced β-oxidation with no changes in lipogenesis. Phospholipid transfer activity of MTP promoted biogenesis of both apoB48 and apoB100-containing very low density lipoprotein in addition to a phospholipid-rich apoB48-containing high-density lipoprotein particle. Triglyceride transfer activity augmented the biosynthesis of triglyceride-rich lipoproteins by increasing the formation of these particles in the lumen of the endoplasmic reticulum. CONCLUSION Based on these findings, we posit that the selective inhibition of MTP triglyceride transfer activity might reduce hyperlipidemia while protecting liver from excess lipid accumulation.
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Affiliation(s)
- Irani Khatun
- School of Graduate Studies, Molecular and Cellular Biology Program, Brooklyn, NY,Departments of Cell Biology and Pediatrics, SUNY Downstate Medical Center, Brooklyn, NY
| | - Sebastian Zeissig
- Division of Gastroenterology, Hepatology, and Endoscopy, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA
| | - Jahangir Iqbal
- Departments of Cell Biology and Pediatrics, SUNY Downstate Medical Center, Brooklyn, NY
| | - Minghui Wang
- University of Alabama, Birmingham, Winston-Salem, NC
| | - David Curiel
- University of Alabama, Birmingham, Winston-Salem, NC
| | | | - Richard S. Blumberg
- Division of Gastroenterology, Hepatology, and Endoscopy, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA
| | - M. Mahmood Hussain
- Departments of Cell Biology and Pediatrics, SUNY Downstate Medical Center, Brooklyn, NY
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37
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Weinberg RB, Gallagher JW, Fabritius MA, Shelness GS. ApoA-IV modulates the secretory trafficking of apoB and the size of triglyceride-rich lipoproteins. J Lipid Res 2012; 53:736-43. [PMID: 22257482 DOI: 10.1194/jlr.m019992] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Although the evidence linking apoA-IV expression and triglyceride (TG)-rich lipoprotein assembly and secretion is compelling, the intracellular mechanisms by which apoA-IV could modulate these processes remain poorly understood. We therefore examined the functional impact of apoA-IV expression on endogenous apoB, TG, and VLDL secretion in stably transfected McA-RH7777 rat hepatoma cells. Expression of apoA-IV modified with the endoplasmic reticulum (ER) retention signal KDEL (apoA-IV-KDEL) dramatically decreased both the rate and efficiency of endogenous apoB secretion, suggesting a presecretory interaction between apoA-IV-KDEL and apoB or apoB-containing lipoproteins. Expression of native apoA-IV using either a constitutive or tetracycline-inducible promoter delayed the initial rate of apoB secretion and reduced the final secretion efficiency by ∼40%. However, whereas apoA-IV-KDEL reduced TG secretion by 75%, expression of native apoA-IV caused a 20-35% increase in TG secretion, accompanied by a ∼55% increase in VLDL-associated apoB, an increase in the TG:phospholipid ratio of secreted d < 1.006 lipoproteins, and a 10.1 nm increase in peak VLDL(1) particle diameter. Native apoA-IV expression had a negligible impact on expression of the MTP gene. These data suggest that by interacting with apoB in the secretory pathway, apoA-IV alters the trafficking kinetics of apoB-containing TG-rich lipoproteins through cellular lipidation compartments, which in turn, enhances particle expansion and increases TG secretion.
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Affiliation(s)
- Richard B Weinberg
- Department of Internal Medicine, Wake Forest School of Medicine, Winston-Salem, NC 27157, USA
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38
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Zhou H, Wang F, Wang Y, Ning Z, Hou W, Wright TG, Sundaram M, Zhong S, Yao Z, Figeys D. Improved recovery and identification of membrane proteins from rat hepatic cells using a centrifugal proteomic reactor. Mol Cell Proteomics 2011; 10:O111.008425. [PMID: 21749988 DOI: 10.1074/mcp.o111.008425] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Despite their importance in many biological processes, membrane proteins are underrepresented in proteomic analysis because of their poor solubility (hydrophobicity) and often low abundance. We describe a novel approach for the identification of plasma membrane proteins and intracellular microsomal proteins that combines membrane fractionation, a centrifugal proteomic reactor for streamlined protein extraction, protein digestion and fractionation by centrifugation, and high performance liquid chromatography-electrospray ionization-tandem MS. The performance of this approach was illustrated for the study of the proteome of ER and Golgi microsomal membranes in rat hepatic cells. The centrifugal proteomic reactor identified 945 plasma membrane proteins and 955 microsomal membrane proteins, of which 63 and 47% were predicted as bona fide membrane proteins, respectively. Among these proteins, >800 proteins were undetectable by the conventional in-gel digestion approach. The majority of the membrane proteins only identified by the centrifugal proteomic reactor were proteins with ≥ 2 transmembrane segments or proteins with high molecular mass (e.g. >150 kDa) and hydrophobicity. The improved proteomic reactor allowed the detection of a group of endocytic and/or signaling receptor proteins on the plasma membrane, as well as apolipoproteins and glycerolipid synthesis enzymes that play a role in the assembly and secretion of apolipoprotein B100-containing very low density lipoproteins. Thus, the centrifugal proteomic reactor offers a new analytical tool for structure and function studies of membrane proteins involved in lipid and lipoprotein metabolism.
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Affiliation(s)
- Hu Zhou
- Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, 451 Smyth Road, Ottawa, Canada
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39
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Qin W, Sundaram M, Wang Y, Zhou H, Zhong S, Chang CC, Manhas S, Yao EF, Parks RJ, McFie PJ, Stone SJ, Jiang ZG, Wang C, Figeys D, Jia W, Yao Z. Missense mutation in APOC3 within the C-terminal lipid binding domain of human ApoC-III results in impaired assembly and secretion of triacylglycerol-rich very low density lipoproteins: evidence that ApoC-III plays a major role in the formation of lipid precursors within the microsomal lumen. J Biol Chem 2011; 286:27769-80. [PMID: 21676879 DOI: 10.1074/jbc.m110.203679] [Citation(s) in RCA: 89] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Hepatic assembly of triacylglycerol (TAG)-rich very low density lipoproteins (VLDL) is achieved through recruitment of bulk TAG (presumably in the form of lipid droplets within the microsomal lumen) into VLDL precursor containing apolipoprotein (apo) B-100. We determined protein/lipid components of lumenal lipid droplets (LLD) in cells expressing recombinant human apoC-III (C3wt) or a mutant form (K58E, C3KE) initially identified in humans that displayed hypotriglyceridemia. Although expression of C3wt markedly stimulated secretion of TAG and apoB-100 as VLDL(1), the K58E mutation (located at the C-terminal lipid binding domain) abolished the effect in transfected McA-RH7777 cells and in apoc3-null mice. Metabolic labeling studies revealed that accumulation of TAG in LLD was decreased (by 50%) in cells expressing C3KE. A Fat Western lipid protein overlay assay showed drastically reduced lipid binding of the mutant protein. Substituting Lys(58) with Arg demonstrated that the positive charge at position 58 is crucial for apoC-III binding to lipid and for promoting TAG secretion. On the other hand, substituting both Lys(58) and Lys(60) with Glu resulted in almost entire elimination of lipid binding and loss of function in promoting TAG secretion. Thus, the lipid binding domain of apoC-III plays a key role in the formation of LLD for hepatic VLDL assembly and secretion.
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Affiliation(s)
- Wen Qin
- Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai Diabetes Institute, Shanghai Clinical Center of Diabetes, Shanghai 200233, China
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40
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Blade AM, Fabritius MA, Hou L, Weinberg RB, Shelness GS. Biogenesis of apolipoprotein A-V and its impact on VLDL triglyceride secretion. J Lipid Res 2011; 52:237-44. [PMID: 21115968 PMCID: PMC3023543 DOI: 10.1194/jlr.m010793] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2010] [Revised: 11/09/2010] [Indexed: 12/12/2022] Open
Abstract
Apolipoprotein A-V (apoA-V) is a potent regulator of intravascular triglyceride (TG) metabolism, yet its plasma concentration is very low compared with that of other apolipoproteins. To examine the basis for its low plasma concentration, the secretion efficiency of apoA-V was measured in stably transfected McA-RH7777 rat hepatoma cells. Pulse-chase experiments revealed that only ∼20% of newly synthesized apoA-V is secreted into culture medium within 3 h postsynthesis and that ∼65% undergoes presecretory turnover; similar results were obtained with transfected nonhepatic Chinese hamster ovary cells. ApoA-V secreted by McA-RH7777 cells was not associated with cell surface heparin-competable binding sites. When stably transfected McA-RH7777 cells were treated with oleic acid, the resulting increase in TG synthesis caused a reduction in apoA-V secretion, a reciprocal increase in cell-associated apoA-V, and movement of apoA-V onto cytosolic lipid droplets. In a stably transfected doxycycline-inducible McA-RH7777 cell line, apoA-V expression inhibited TG secretion by ∼50%, increased cellular TG, and reduced Z-average VLDL(1) particle diameter from 81 to 67 nm; however, no impact on apoB secretion was observed. These data demonstrate that apoA-V inefficiently traffics within the secretory pathway, that its intracellular itinerary can be regulated by changes in cellular TG accumulation, and that apoA-V synthesis can modulate VLDL TG mobilization and secretion.
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Affiliation(s)
- Anna M. Blade
- Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
| | - Melissa A. Fabritius
- Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
| | - Li Hou
- Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
| | - Richard B. Weinberg
- Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
- Physiology and Pharmacology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
| | - Gregory S. Shelness
- Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, NC 27157
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41
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Bamji-Mirza M, Sundaram M, Zhong S, Yao EF, Parks RJ, Yao Z. Secretion of triacylglycerol-poor VLDL particles from McA-RH7777 cells expressing human hepatic lipase. J Lipid Res 2010; 52:540-8. [PMID: 21189265 DOI: 10.1194/jlr.m012476] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Hepatic lipase (HL) plays a role in the catabolism of apolipoprotein (apo)B-containing lipoproteins through its lipolytic and ligand-binding properties. We describe a potential intracellular role of HL in the assembly and secretion of VLDL. Transient or stable expression of HL in McA-RH7777 cells resulted in decreased (by 40%) incorporation of [(3)H]glycerol into cell-associated and secreted triacylglycerol (TAG) relative to control cells. However, incorporation of [(35)S]methionine/cysteine into cell and medium apoB-100 was not decreased by HL expression. The decreased (3)H-TAG synthesis/secretion in HL expressing cells was not attributable to decreased expression of genes involved in lipogenesis. Fractionation of medium revealed that the decreased [(3)H]TAG from HL expressing cells was mainly attributable to decreased VLDL. Expression of catalytically-inactive HL (HL(SG)) (Ser-145 at the catalytic site was substituted with Gly) in the cells also resulted in decreased secretion of VLDL-[(3)H]TAG. Examination of lumenal contents of microsomes showed a 40% decrease in [(3)H]TAG associated with lumenal lipid droplets in HL or HL(SG) expressing cells as compared with control. The microsomal membrane-associated [(3)H]TAG was decreased by 50% in HL expressing cells but not in HL(SG) expressing cells. Thus, expression of HL, irrespective of its lipolytic function, impairs formation of VLDL precursor [(3)H]TAG in the form of lumenal lipid droplets. These results suggest that HL expression in McA-RH7777 cells result in secretion of [(3)H]TAG-poor VLDL.
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Affiliation(s)
- Michelle Bamji-Mirza
- Department of Biochemistry, Microbiology & Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Canada, K1H 8M5
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42
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Chung S, Gebre AK, Seo J, Shelness GS, Parks JS. A novel role for ABCA1-generated large pre-beta migrating nascent HDL in the regulation of hepatic VLDL triglyceride secretion. J Lipid Res 2010; 51:729-42. [PMID: 20215580 DOI: 10.1194/jlr.m900083] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
In Tangier disease, absence of ATP binding cassette transporter A1 (ABCA1) results in reduced plasma HDL and elevated triglyceride (TG) levels. We hypothesized that hepatocyte ABCA1 regulates VLDL TG secretion through nascent HDL production. Silencing of ABCA1 expression in oleate-stimulated rat hepatoma cells resulted in: 1) decreased large nascent HDL (>10 nm diameter) and increased small nascent HDL (<10 nm) formation, 2) increased large buoyant VLDL1 particle secretion, and 3) decreased phosphatidylinositol-3 (PI3) kinase activation. Nascent HDL-containing conditioned medium from rat hepatoma cells or HEK293 cells transfected with ABCA1 was effective in increasing PI3 kinase activation and reducing VLDL TG secretion in ABCA1-silenced hepatoma cells. Addition of isolated large nascent HDL particles to ABCA1-silenced hepatoma cells inhibited VLDL TG secretion to a greater extent than small nascent HDL. Similarly, addition of recombinant HDL, but not human plasma HDL, was effective in attenuating TG secretion and increasing PI3 kinase activation in ABCA1-silenced cells. Collectively, these data suggest that large nascent HDL particles, assembled by hepatic ABCA1, generate a PI3 kinase-mediated autocrine signal that attenuates VLDL maturation and TG secretion. This pathway may explain the elevated plasma TG concentration that occurs in most Tangier subjects and may also account, in part, for the inverse relationship between plasma HDL and TG concentrations in individuals with compromised ABCA1 function.
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Affiliation(s)
- Soonkyu Chung
- Department of Pathology/Section on Lipid Sciences, Wake Forest University School of Medicine, Winston-Salem, NC 27157, USA
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43
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Soriguer F, García-Serrano S, Garrido-Sánchez L, Gutierrez-Repiso C, Rojo-Martínez G, Garcia-Escobar E, García-Arnés J, Gallego-Perales JL, Delgado V, García-Fuentes E. Jejunal wall triglyceride concentration of morbidly obese persons is lower in those with type 2 diabetes mellitus. J Lipid Res 2010; 51:3516-23. [PMID: 20855567 DOI: 10.1194/jlr.m007815] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The overproduction of intestinal lipoproteins may contribute to the dyslipidemia found in diabetes. We studied the influence of diabetes on the fasting jejunal lipid content and its association with plasma lipids and the expression of genes involved in the synthesis and secretion of these lipoproteins. The study was undertaken in 27 morbidly obese persons, 12 of whom had type 2 diabetes mellitus (T2DM). The morbidly obese persons with diabetes had higher levels of chylomicron (CM) triglycerides (P < 0.001) and apolipoprotein (apo)B48 (P = 0.012). The jejunum samples obtained from the subjects with diabetes had a lower jejunal triglyceride content (P = 0.012) and angiopoietin-like protein 4 (ANGPTL4) mRNA expression (P = 0.043). However, the apoA-IV mRNA expression was significantly greater (P = 0.036). The jejunal triglyceride content correlated negatively with apoA-IV mRNA expression (r = -0.587, P = 0.027). The variables that explained the jejunal triglyceride content in a multiple linear regression model were the insulin resistance state and the apoA-IV mRNA expression. Our results show that the morbidly obese subjects with diabetes had lower jejunal lipid content and that this correlated negatively with apoA-IV mRNA expression. These findings show that the jejunum appears to play an active role in lipid homeostasis in the fasting state.
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Affiliation(s)
- F Soriguer
- Servicios de Endocrinología y Nutrición y Cirugía General, Málaga, Spain
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Sundaram M, Yao Z. Recent progress in understanding protein and lipid factors affecting hepatic VLDL assembly and secretion. Nutr Metab (Lond) 2010; 7:35. [PMID: 20423497 PMCID: PMC2873297 DOI: 10.1186/1743-7075-7-35] [Citation(s) in RCA: 118] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2009] [Accepted: 04/27/2010] [Indexed: 02/06/2023] Open
Abstract
Excess lipid induced metabolic disorders are one of the major existing challenges for the society. Among many different causes of lipid disorders, overproduction and compromised catabolism of triacylglycerol-rich very low density lipoproteins (VLDL) have become increasingly prevalent leading to hyperlipidemia worldwide. This review provides the latest understanding in different aspects of VLDL assembly process, including structure-function relationships within apoB, mutations in APOB causing hypobetalipoproteinemia, significance of modulating microsomal triglyceride-transfer protein activity in VLDL assembly, alterations of VLDL assembly by different fatty acid species, and hepatic proteins involved in vesicular trafficking, and cytosolic lipid droplet metabolism that contribute to VLDL assembly. The role of lipoprotein receptors and exchangeable apolipoproteins that promote or diminish VLDL assembly and secretion is discussed. New understanding on dysregulated insulin signaling as a consequence of excessive triacylglycerol-rich VLDL in the plasma is also presented. It is hoped that a comprehensive view of protein and lipid factors that contribute to molecular and cellular events associated with VLDL assembly and secretion will assist in the identification of pharmaceutical targets to reduce disease complications related to hyperlipidemia.
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Affiliation(s)
- Meenakshi Sundaram
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5, Canada
| | - Zemin Yao
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5, Canada
- Department of Pathology and Laboratory Medicine, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5, Canada
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45
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Chung S, Gebre AK, Seo J, Shelness GS, Parks JS. A novel role for ABCA1-generated large pre-β migrating nascent HDL in the regulation of hepatic VLDL triglyceride secretion. J Lipid Res 2010. [DOI: 10.1194/jlr.m900083-jlr200] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
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46
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Sundaram M, Zhong S, Bou Khalil M, Links PH, Zhao Y, Iqbal J, Hussain MM, Parks RJ, Wang Y, Yao Z. Expression of apolipoprotein C-III in McA-RH7777 cells enhances VLDL assembly and secretion under lipid-rich conditions. J Lipid Res 2010. [PMID: 19622837 DOI: 10.1194/jlr.m900346-jlr200] [Citation(s) in RCA: 89] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Apolipoprotein (apo) C-III plays a regulatory role in VLDL lipolysis and clearance. In this study, we determined a potential intracellular role of apoC-III in hepatic VLDL assembly and secretion. Stable expression of recombinant apoC-III in McA-RH7777 cells resulted in increased secretion efficiency of VLDL-associated triacylglycerol (TAG) and apoB-100 in a gene-dosage-dependent manner. The stimulatory effect of apoC-III on TAG secretion was manifested only when cells were cultured under lipid-rich (i.e., media supplemented with exogenous oleate) but not lipid-poor conditions. The stimulated TAG secretion was accompanied by increased secretion of apoB-100 and apoB-48 as VLDL(1). Expression of apoC-III also increased mRNA and activity of microsomal triglyceride transfer protein (MTP). Pulse-chase experiments showed that apoC-III expression promoted VLDL(1) secretion even under conditions where the MTP activity was inhibited immediately after the formation of lipid-poor apoB-100 particles, suggesting an involvement of apoC-III in the second-step VLDL assembly process. Consistent with this notion, the newly synthesized apoC-III was predominantly associated with TAG within the microsomal lumen that resembled lipid precursors of VLDL. Introducing an Ala23-to-Thr mutation into apoC-III, a naturally occurring mutation originally identified in two Mayan Indian subjects with hypotriglyceridemia, abolished the ability of apoC-III to stimulate VLDL secretion from transfected cells. Thus, expression of apoC-III in McA-RH7777 cells enhances hepatic TAG-rich VLDL assembly and secretion under lipid-rich conditions.
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Affiliation(s)
- Meenakshi Sundaram
- Department of Biochemistry, Microbiology and Immunology, Department of Pathology and Laboratory Medicine, Ottawa Institute of Systems Biology, University of Ottawa Canada K1H 8M5
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47
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Sundaram M, Zhong S, Bou Khalil M, Links PH, Zhao Y, Iqbal J, Hussain MM, Parks RJ, Wang Y, Yao Z. Expression of apolipoprotein C-III in McA-RH7777 cells enhances VLDL assembly and secretion under lipid-rich conditions. J Lipid Res 2010; 51:150-61. [PMID: 19622837 DOI: 10.1194/m900346-jlr200] [Citation(s) in RCA: 93] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023] Open
Abstract
Apolipoprotein (apo) C-III plays a regulatory role in VLDL lipolysis and clearance. In this study, we determined a potential intracellular role of apoC-III in hepatic VLDL assembly and secretion. Stable expression of recombinant apoC-III in McA-RH7777 cells resulted in increased secretion efficiency of VLDL-associated triacylglycerol (TAG) and apoB-100 in a gene-dosage-dependent manner. The stimulatory effect of apoC-III on TAG secretion was manifested only when cells were cultured under lipid-rich (i.e., media supplemented with exogenous oleate) but not lipid-poor conditions. The stimulated TAG secretion was accompanied by increased secretion of apoB-100 and apoB-48 as VLDL(1). Expression of apoC-III also increased mRNA and activity of microsomal triglyceride transfer protein (MTP). Pulse-chase experiments showed that apoC-III expression promoted VLDL(1) secretion even under conditions where the MTP activity was inhibited immediately after the formation of lipid-poor apoB-100 particles, suggesting an involvement of apoC-III in the second-step VLDL assembly process. Consistent with this notion, the newly synthesized apoC-III was predominantly associated with TAG within the microsomal lumen that resembled lipid precursors of VLDL. Introducing an Ala23-to-Thr mutation into apoC-III, a naturally occurring mutation originally identified in two Mayan Indian subjects with hypotriglyceridemia, abolished the ability of apoC-III to stimulate VLDL secretion from transfected cells. Thus, expression of apoC-III in McA-RH7777 cells enhances hepatic TAG-rich VLDL assembly and secretion under lipid-rich conditions.
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Affiliation(s)
- Meenakshi Sundaram
- Department of Biochemistry, Microbiology and Immunology, Department of Pathology and Laboratory Medicine, Ottawa Institute of Systems Biology, University of Ottawa Canada K1H 8M5
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48
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Liu Y, Manchekar M, Sun Z, Richardson PE, Dashti N. Apolipoprotein B-containing lipoprotein assembly in microsomal triglyceride transfer protein-deficient McA-RH7777 cells. J Lipid Res 2010; 51:2253-64. [PMID: 20181985 DOI: 10.1194/jlr.m005371] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023] Open
Abstract
Microsomal triglyceride transfer protein (MTP) is required for the assembly and secretion of apolipoprotein (apo) B-containing lipoproteins. Previously, we demonstrated that the N-terminal 1,000 residues of apoB (apoB:1000) are necessary for the initiation of apoB-containing lipoprotein assembly in rat hepatoma McA-RH7777 cells and that these particles are phospholipid (PL) rich. To determine if the PL transfer activity of MTP is sufficient for the assembly and secretion of primordial apoB:1000-containing lipoproteins, we employed microRNA-based short hairpin RNAs (miR-shRNAs) to silence Mttp gene expression in parental and apoB:1000-expressing McA-RH7777 cells. This approach led to 98% reduction in MTP protein levels in both cell types. Metabolic labeling studies demonstrated a drastic 90-95% decrease in the secretion of rat endogenous apoB100-containing lipoproteins in MTP-deficient McA-RH7777 cells compared with cells transfected with negative control miR-shRNA. A similar reduction was observed in the secretion of rat endogenous apoB48 under the experimental conditions employed. In contrast, MTP absence had no significant effect on the synthesis, lipidation, and secretion of human apoB:1000-containing particles. These results provide strong evidence in support of the concept that in McA-RH7777 cells, acquisition of PL by apoB:1000 and initiation of apoB-containing lipoprotein assembly, a process distinct from the conventional first-step assembly of HDL-sized apoB-containing particles, do not require MTP. This study indicates that, in hepatocytes, a factor(s) other than MTP mediates the formation of the PL-rich primordial apoB:1000-containing initiation complex.
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Affiliation(s)
- Yanwen Liu
- Department of Medicine, Basic Sciences Section, Atherosclerosis Research Unit, University of Alabama at Birmingham Medical Center, Birmingham, AL 35294, USA
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Sundaram M, Zhong S, Bou Khalil M, Zhou H, Jiang ZG, Zhao Y, Iqbal J, Hussain MM, Figeys D, Wang Y, Yao Z. Functional analysis of the missense APOC3 mutation Ala23Thr associated with human hypotriglyceridemia. J Lipid Res 2010; 51:1524-34. [PMID: 20097930 DOI: 10.1194/jlr.m005108] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
We have shown that expression of apolipoprotein (apo) C-III promotes VLDL secretion from transfected McA-RH7777 cells under lipid-rich conditions. To determine structural elements within apoC-III that confer to this function, we contrasted wild-type apoC-III with a mutant Ala23Thr originally identified in hypotriglyceridemia subjects. Although synthesis of [(3)H]glycerol-labeled TAG was comparable between cells expressing wild-type apoC-III (C3wt cells) or Ala23Thr mutant (C3AT cells), secretion of [(3)H]TAG from C3AT cells was markedly decreased. The lowered [(3)H]TAG secretion was associated with an inability of C3AT cells to assemble VLDL(1). Moreover, [(3)H]TAG within the microsomal lumen in C3AT cells was 60% higher than that in C3wt cells, yet the activity of microsomal triglyceride-transfer protein in C3AT cells was not elevated. The accumulated [(3)H]TAG in C3AT microsomal lumen was mainly associated with lumenal IDL/LDL-like lipoproteins. Phenotypically, this [(3)H]TAG fractionation profiling resembled what was observed in cells treated with brefeldin A, which at low dose specifically blocked the second-step VLDL(1) maturation. Furthermore, lumenal [(35)S]Ala23Thr protein accumulated in IDL/LDL fractions and was absent in VLDL fractions in C3AT cells. These results suggest that the presence of Ala23Thr protein in lumenal IDL/LDL particles might prevent effective fusion between lipid droplets and VLDL precursors. Thus, the current study reveals an important structural element residing within the N-terminal region of apoC-III that governs the second step VLDL(1) maturation.
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Affiliation(s)
- Meenakshi Sundaram
- Department of Biochemistry, Ottawa Institute of Systems Biology, University of Ottawa, Canada
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50
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Zhong S, Magnolo AL, Sundaram M, Zhou H, Yao EF, Di Leo E, Loria P, Wang S, Bamji-Mirza M, Wang L, McKnight CJ, Figeys D, Wang Y, Tarugi P, Yao Z. Nonsynonymous mutations within APOB in human familial hypobetalipoproteinemia: evidence for feedback inhibition of lipogenesis and postendoplasmic reticulum degradation of apolipoprotein B. J Biol Chem 2009; 285:6453-64. [PMID: 20032471 DOI: 10.1074/jbc.m109.060467] [Citation(s) in RCA: 56] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Five nontruncating missense APOB mutations, namely A31P, G275S, L324M, G912D, and G945S, were identified in heterozygous carriers of familial hypobetalipoproteinemia (FHBL) in the Italian population. To test that the FHBL phenotype was a result of impaired hepatic secretion of mutant apoB proteins, we performed transfection studies using McA-RH7777 cells stably expressing wild type or mutant forms of human apolipoprotein B-48 (apoB-48). All mutant proteins displayed varied impairment in secretion, with G912D the least affected and A31P barely secreted. Although some A31P was degraded by proteasomes, a significant proportion of it (although inappropriately glycosylated) escaped endoplasmic reticulum (ER) quality control and presented in the Golgi compartment. Degradation of the post-ER A31P was achieved by autophagy. Expression of A31P also decreased secretion of endogenous apoB and triglycerides, yet the impaired lipoprotein secretion did not lead to lipid accumulation in the cells or ER stress. Rather, expression of genes involved in lipogenesis was down-regulated, including liver X receptor alpha, sterol regulator element-binding protein 1c, fatty acid synthase, acetyl-CoA carboxylase 1, stearoyl-CoA desaturase 1, and lipin-1. These results suggest that feedback inhibition of hepatic lipogenesis in conjunction with post-ER degradation of misfolded apoB proteins can contribute to reduce fat accumulation in the FHBL liver.
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Affiliation(s)
- Shumei Zhong
- Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada
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