title: Increased intracellular triglyceride in C(2)C(12) muscle cells transfected with human lipoprotein lipase.
abstract: Much of the knowledge about the cell biology of lipoprotein lipase (LPL) in vitro has been gained from adipose tissue model systems. However, the importance of skeletal muscle lipoprotein lipase (SMLPL) to both lipoprotein and muscle metabolism remains unclear. Although the production of LPL in cultured myocytes has been documented, the amount of enzyme activity produced is small. To develop a more suitable tissue culture model for SMLPL, mouse C(2)C(12) myoblasts were stably transduced with a retroviral vector encoding the full-length human LPL (hLPL) cDNA. Control cells were transduced with a vector encoding beta-galactosidase. LPL expression was assayed as a function of cell growth by measuring LPL activity on days 3, 7, 9, 11, and 14 after subculture. The hLPL-transduced myoblasts increasingly overexpressed both heparin-releasable (HR) and intracellular (IN) LPL activity compared to nontransduced myoblasts (P < 0.001 at Day 11) and myoblasts transduced with the control vector (P < 0.001 at Day 11). This increase occurred while LPL mRNA levels remained stable between days 3 and 14. As expected, IN LPL activity was also increased in the transduced cells. High levels of LPL activity were also obtained after differentiating the C(2)C(12) cells into myotubes by serum deprivation. Additionally, throughout the time course, C(2)/LPL cells had greater amounts of intracellular triglyceride than both the C(2)C(12) and the C(2)/beta-GEO cells (P = 0.005 and P < 0.001, respectively) with the largest differences seen on day 14 of the time course (P = 0.001, C(2)/LPL vs C(2)C(12) (r) or C(2)/beta-GEO cells). Thus, C(2)C(12) myoblasts stably transduced with hLPL markedly overexpressed both HR and IN LPL activity compared to control cells which, in turn, was associated with increases in intracellular triglyceride content. Because LPL regulation in tissues is mostly posttranslational, this new in vitro model will permit the in-depth study of the posttranslational regulation of SMLPL and provide new insights into the fate of lipoprotein-derived fatty acids in muscle.
authors.affiliation: Division of Endocrinology, Metabolism and Diabetes, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
authors.forename: P
authors.initials: P
authors.lastname: Poirier
authors.forename: T
authors.initials: T
authors.lastname: Marcell
authors.forename: P U
authors.initials: PU
authors.lastname: Huey
authors.forename: I R
authors.initials: IR
authors.lastname: Schlaepfer
authors.forename: G C
authors.initials: GC
authors.lastname: Owens
authors.forename: D R
authors.initials: DR
authors.lastname: Jensen
authors.forename: R H
authors.initials: RH
authors.lastname: Eckel
mesh_terms: D000818:Animals; D002460:Cell Line; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008071:Lipoprotein Lipase / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014280:Triglycerides / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D014280:Triglycerides; D006493:Heparin; D008071:Lipoprotein Lipase