Journal of Lipid Research, Vol 37, 1468-1480, Copyright © 1996 by Lipid Research, Inc.

ARTICLES

Inhibition of apolipoprotein B and triglyceride secretion in human hepatoma cells (HepG2)

M Haghpassand, D Wilder and JB Moberly
Department of Cardiovascular and Metabolic Diseases, Pfizer, Inc., Groton, CT 06340, USA.

Apolipoprotein B (apoB), the major protein component of triglyceride- rich lipoproteins, is assembled into a lipoprotein particle via a complex, multistep process. Recent studies indicate that triglyceride- rich lipoprotein assembly requires the activity of the heterodimeric protein, microsomal triglyceride transfer protein (MTP). We identified a novel inhibitor of apolipoprotein B secretion using the human hepatoma cell line, HepG2. CP-10447, a derivative of the hypnotic drug methaqualone (Quaalude), inhibited apoB secretion from HepG2 cells with an IC50 of approximately 5 microM. CP-10447 also inhibited apoB secretion from Caco-2 cells, a model of intestinal lipoprotein production. In experiments using [3H]glycerol as a precursor for triglyceride synthesis, CP-10447 (20 microM) inhibited radiolabeled triglyceride secretion by approximately 83% (P < 0.0001) in HepG2 cells and 76% (P < 0.05) in Caco-2 cells with no effect on radiolabel incorporation into cellular triglyceride, indicating that CP-10447 inhibited triglyceride secretion without affecting triglyceride synthesis. RNA solution hybridization assay indicated that CP-10447 did not affect apoB or apoA-I mRNA levels. Pulse-chase experiments in HepG2 cells confirmed that CP-10447 inhibited the secretion of apoB (not its synthesis) without affecting secretion of total proteins or albumin and suggested that CP-10447 stimulates the early intracellular degradation of apoB in the endoplasmic reticulum (ER). Further studies demonstrated that CP-10447 is a potent inhibitor of human liver microsomal triglyceride transfer activity (IC50 approximately 1.7 microM) in an in vitro assay containing artificial liposomes and partially purified human MTP. These data suggest that CP-10447 may inhibit apoB and triglyceride secretion by inhibiting MTP activity and stimulating the early ER degradation of apoB. CP-10447 should provide a useful tool for further study of the mechanisms of apoB secretion and triglyceride-rich lipoprotein assembly.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				C. Ameen, U. Edvardsson, A. Ljungberg, L. Asp, P. Akerblad, A. Tuneld, S.-O. Olofsson, D. Linden, and J. Oscarsson Activation of Peroxisome Proliferator-activated Receptor {alpha} Increases the Expression and Activity of Microsomal Triglyceride Transfer Protein in the Liver J. Biol. Chem., January 14, 2005; 280(2): 1224 - 1229. [Abstract] [Full Text] [PDF]

				H. Athar, J. Iqbal, X.-C. Jiang, and M. M. Hussain A simple, rapid, and sensitive fluorescence assay for microsomal triglyceride transfer protein J. Lipid Res., April 1, 2004; 45(4): 764 - 772. [Abstract] [Full Text] [PDF]

				L. L. Swift, M.-Y. Zhu, B. Kakkad, A. Jovanovska, M. D. Neely, K. Valyi-Nagy, R. L. Roberts, D. E. Ong, and W. G. Jerome Subcellular localization of microsomal triglyceride transfer protein J. Lipid Res., October 1, 2003; 44(10): 1841 - 1849. [Abstract] [Full Text] [PDF]

				C. E. Chandler, D. E. Wilder, J. L. Pettini, Y. E. Savoy, S. F. Petras, G. Chang, J. Vincent, and H. J. Harwood Jr. CP-346086: an MTP inhibitor that lowers plasma cholesterol and triglycerides in experimental animals and in humans J. Lipid Res., October 1, 2003; 44(10): 1887 - 1901. [Abstract] [Full Text] [PDF]

				H. J. Harwood Jr., S. F. Petras, L. D. Shelly, L. M. Zaccaro, D. A. Perry, M. R. Makowski, D. M. Hargrove, K. A. Martin, W. R. Tracey, J. G. Chapman, et al. Isozyme-nonselective N-Substituted Bipiperidylcarboxamide Acetyl-CoA Carboxylase Inhibitors Reduce Tissue Malonyl-CoA Concentrations, Inhibit Fatty Acid Synthesis, and Increase Fatty Acid Oxidation in Cultured Cells and in Experimental Animals J. Biol. Chem., September 26, 2003; 278(39): 37099 - 37111. [Abstract] [Full Text] [PDF]

				M. M. Hussain, J. Shi, and P. Dreizen Microsomal triglyceride transfer protein and its role in apoB-lipoprotein assembly J. Lipid Res., January 1, 2003; 44(1): 22 - 32. [Abstract] [Full Text] [PDF]

				N. Sakata, T. E. Phillips, and J. L. Dixon Distribution, transport, and degradation of apolipoprotein B-100 in HepG2 cells J. Lipid Res., December 1, 2001; 42(12): 1947 - 1958. [Abstract] [Full Text] [PDF]

				J. A. Sellers and G. S. Shelness Lipoprotein assembly capacity of the mammary tumor-derived cell line C127 is due to the expression of functional microsomal triglyceride transfer protein J. Lipid Res., November 1, 2001; 42(11): 1897 - 1904. [Abstract] [Full Text] [PDF]

				S.-H. H. Juo, Z. Han, J. D. Smith, L. Colangelo, and K. Liu Common Polymorphism in Promoter of Microsomal Triglyceride Transfer Protein Gene Influences Cholesterol, ApoB, and Triglyceride Levels in Young African American Men : Results From the Coronary Artery Risk Development in Young Adults (CARDIA) Study Arterioscler. Thromb. Vasc. Biol., May 1, 2000; 20(5): 1316 - 1322. [Abstract] [Full Text] [PDF]

				G. K. Leung, M. M. Veniant, S. K. Kim, C. H. Zlot, M. Raabe, J. Bjorkegren, R. A. Neese, M. K. Hellerstein, and S. G. Young A Deficiency of Microsomal Triglyceride Transfer Protein Reduces Apolipoprotein B Secretion J. Biol. Chem., March 10, 2000; 275(11): 7515 - 7520. [Abstract] [Full Text] [PDF]

				U. J.F. Tietge, A. Bakillah, C. Maugeais, K. Tsukamoto, M. Hussain, and D. J. Rader Hepatic overexpression of microsomal triglyceride transfer protein (MTP) results in increased in vivo secretion of VLDL triglycerides and apolipoprotein B J. Lipid Res., November 1, 1999; 40(11): 2134 - 2139. [Abstract] [Full Text]

				N. S. Kumar and C. M. Mansbach II Prechylomicron transport vesicle: isolation and partial characterization Am J Physiol Gastrointest Liver Physiol, February 1, 1999; 276(2): G378 - G386. [Abstract] [Full Text] [PDF]

				E. Nicodeme, F. Benoist, R. McLeod, Z. Yao, J. Scott, C. C. Shoulders, and T. Grand-Perret Identification of Domains in Apolipoprotein B100 That Confer a High Requirement for the Microsomal Triglyceride Transfer Protein J. Biol. Chem., January 22, 1999; 274(4): 1986 - 1993. [Abstract] [Full Text] [PDF]

				J.-s. Liang, X. Wu, H. Jiang, M. Zhou, H. Yang, P. Angkeow, L.-S. Huang, S. L. Sturley, and H. Ginsberg Translocation Efficiency, Susceptibility to Proteasomal Degradation, and Lipid Responsiveness of Apolipoprotein B Are Determined by the Presence of beta  Sheet Domains J. Biol. Chem., December 25, 1998; 273(52): 35216 - 35221. [Abstract] [Full Text] [PDF]

				D. M. Mitchell, M. Zhou, R. Pariyarath, H. Wang, J. D. Aitchison, H. N. Ginsberg, and E. A. Fisher Apoprotein B100 has a prolonged interaction with the translocon during which its lipidation and translocation change from dependence on the microsomal triglyceride transfer protein to independence PNAS, December 8, 1998; 95(25): 14733 - 14738. [Abstract] [Full Text] [PDF]

				M. M. Hussain, A. Bakillah, N. Nayak, and G. S. Shelness Amino Acids 430-570 in Apolipoprotein B Are Critical for Its Binding to Microsomal Triglyceride Transfer Protein J. Biol. Chem., October 2, 1998; 273(40): 25612 - 25615. [Abstract] [Full Text] [PDF]

				M. Zhou, E. A. Fisher, and H. N. Ginsberg Regulated Co-translational Ubiquitination of Apolipoprotein B100. A NEW PARADIGM FOR PROTEASOMAL DEGRADATION OF A SECRETORY PROTEIN J. Biol. Chem., September 18, 1998; 273(38): 24649 - 24653. [Abstract] [Full Text] [PDF]

				M. Raabe, L. M. Flynn, C. H. Zlot, J. S. Wong, M. M. Veniant, R. L. Hamilton, and S. G. Young Knockout of the abetalipoproteinemia gene in mice: Reduced lipoprotein secretion in heterozygotes and embryonic lethality in homozygotes PNAS, July 21, 1998; 95(15): 8686 - 8691. [Abstract] [Full Text] [PDF]

				H. Jamil, C.-H. Chu, J. K . Dickson , Jr., Y. Chen, M. Yan, S. A. Biller, R. E. Gregg, J. R. Wetterau, and D. A. Gordon Evidence that microsomal triglyceride transfer protein is limiting in the production of apolipoprotein B-containing lipoproteins in hepatic cells J. Lipid Res., July 1, 1998; 39(7): 1448 - 1454. [Abstract] [Full Text]

				F. Karpe, B. Lundahl, E. Ehrenborg, P. Eriksson, and A. Hamsten A Common Functional Polymorphism in the Promoter Region of the Microsomal Triglyceride Transfer Protein Gene Influences Plasma LDL Levels Arterioscler. Thromb. Vasc. Biol., May 1, 1998; 18(5): 756 - 761. [Abstract] [Full Text] [PDF]

				M. M. J. van Greevenbroek, M. G. Robertus-Teunissen, D. W. Erkelens, and T. W. A. de Bruin Participation of the microsomal triglyceride transfer protein in lipoprotein assembly in Caco-2 cells: interaction with saturated and unsaturated dietary fatty acids J. Lipid Res., January 1, 1998; 39(1): 173 - 185. [Abstract] [Full Text]

				J. Luchoomun, Z. Zhou, A. Bakillah, H. Jamil, and M. M. Hussain Assembly and Secretion of VLDL in Nondifferentiated Caco-2 Cells Stably Transfected With Human Recombinant ApoB48 cDNA Arterioscler. Thromb. Vasc. Biol., November 1, 1997; 17(11): 2955 - 2963. [Abstract] [Full Text]

				F. Benoist and T. Grand-Perret Co-translational Degradation of Apolipoprotein B100 by the Proteasome Is Prevented by Microsomal Triglyceride Transfer Protein. SYNCHRONIZED TRANSLATION STUDIES ON HepG2 CELLS TREATED WITH AN INHIBITOR OF MICROSOMAL TRIGLYCERIDE TRANSFER PROTEIN J. Biol. Chem., August 15, 1997; 272(33): 20435 - 20442. [Abstract] [Full Text] [PDF]

				Y. Wang, R. S. McLeod, and Z. Yao Normal Activity of Microsomal Triglyceride Transfer Protein Is Required for the Oleate-induced Secretion of Very Low Density Lipoproteins Containing Apolipoprotein B from McA-RH7777 Cells J. Biol. Chem., May 9, 1997; 272(19): 12272 - 12278. [Abstract] [Full Text] [PDF]

				M. Pan, J.-s. Liang, E. A. Fisher, and H. N. Ginsberg Inhibition of Translocation of Nascent Apolipoprotein B across the Endoplasmic Reticulum Membrane Is Associated with Selective Inhibition of the Synthesis of Apolipoprotein B J. Biol. Chem., August 25, 2000; 275(35): 27399 - 27405. [Abstract] [Full Text] [PDF]

				J.-s. Liang and H. N. Ginsberg Microsomal Triglyceride Transfer Protein Binding and Lipid Transfer Activities Are Independent of Each Other, but Both Are Required for Secretion of Apolipoprotein B Lipoproteins from Liver Cells J. Biol. Chem., July 20, 2001; 276(30): 28606 - 28612. [Abstract] [Full Text] [PDF]

				A. Bakillah and M. M. Hussain Binding of Microsomal Triglyceride Transfer Protein to Lipids Results in Increased Affinity for Apolipoprotein B. EVIDENCE FOR STABLE MICROSOMAL MTP-LIPID COMPLEXES J. Biol. Chem., August 10, 2001; 276(33): 31466 - 31473. [Abstract] [Full Text] [PDF]

				M. Pan, J.-s. Liang, E. A. Fisher, and H. N. Ginsberg The Late Addition of Core Lipids to Nascent Apolipoprotein B100, Resulting in the Assembly and Secretion of Triglyceride-rich Lipoproteins, Is Independent of Both Microsomal Triglyceride Transfer Protein Activity and New Triglyceride Synthesis J. Biol. Chem., February 1, 2002; 277(6): 4413 - 4421. [Abstract] [Full Text] [PDF]