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. 1988 Feb;81(2):398–406. doi: 10.1172/JCI113332

Regulation of lipoprotein lipase immunoreactive mass in isolated human adipocytes.

P A Kern 1, J M Ong 1, J W Goers 1, M E Pedersen 1
PMCID: PMC329582  PMID: 3276727

Abstract

Previous studies of human adipose tissue lipoprotein lipase (LPL) have focused on enzyme catalytic activity, and have not measured the LPL protein directly. To study the regulation of the LPL protein, an antibody against purified bovine LPL was used. To demonstrate the specificity of the antiserum, adipose homogenates were Western blotted, and adipocytes were radiolabeled and the cell homogenates immunoprecipitated, yielding a single specific band at 53 kD. Breakdown products of LPL were demonstrated at 35 and 20 kD by Western blotting. An ELISA for human adipose LPL was established, in which LPL was sandwiched between affinity-purified antibody and biotinylated affinity-purified antibody. The standard curves for bovine LPL and human adipose LPL were parallel, and LPL activity correlated strongly with LPL immunoreactive mass. Thus, the bovine LPL standard curve was used to estimate LPL immunoreactive mass from human adipose tissue. The regulation of LPL activity and immunoreactive mass were compared in cultured adipocytes in the presence an absence of insulinlike growth factor-I/somatomedin C (IGF-I), insulin, and fetal bovine serum. IGF-I and a high insulin concentration (70 nM) stimulated only the heparin-releasable (HR) component of LPL activity and immunoreactive mass, and neither IGF-I nor insulin affected LPL specific activity. In contrast, 10% fetal bovine serum stimulated HR activity, HR mass, and cellular extractable (EXT) immunoreactive mass, with no effect on EXT activity. This resulted in a decrease in EXT specific activity in response to serum. The effects of the locally produced nucleosides adenosine and inosine were studied in a similar manner. As with serum, adenosine stimulated HR activity, HR mass, and EXT immunoreactive mass, resulting in a decrease in EXT specific activity. Inosine stimulated an increase in HR activity and HR mass, but had no effect on EXT, and thus did not change LPL specific activity. Thus, a sensitive ELISA for adipose tissue LPL has been developed using a specific, well-characterized antibody. Regulation of human LPL immunoreactive mass was demonstrated in vitro by IGF-I, serum, high concentrations of insulin, adenosine, and inosine. This method will permit further investigations into the regulation of the LPL protein.

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Selected References

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  1. Amri E. Z., Vannier C., Etienne J., Ailhaud G. Maturation and secretion of lipoprotein lipase in cultured adipose cells. II. Effects of tunicamycin on activation and secretion of the enzyme. Biochim Biophys Acta. 1986 Feb 12;875(2):334–343. doi: 10.1016/0005-2760(86)90184-0. [DOI] [PubMed] [Google Scholar]
  2. Arch J. R., Newsholme E. A. The control of the metabolism and the hormonal role of adenosine. Essays Biochem. 1978;14:82–123. [PubMed] [Google Scholar]
  3. Belfrage P., Vaughan M. Simple liquid-liquid partition system for isolation of labeled oleic acid from mixtures with glycerides. J Lipid Res. 1969 May;10(3):341–344. [PubMed] [Google Scholar]
  4. Bourdeaux A. M., Nordmann R., Giudicelli Y. Stimulatory effects of adenosine, adenosine analogs and insulin on adipose tissue lipoprotein lipase activity and their prevention by phosphodiesterase inhibitors. FEBS Lett. 1984 Dec 3;178(1):132–136. doi: 10.1016/0014-5793(84)81256-9. [DOI] [PubMed] [Google Scholar]
  5. Bourdeaux A. M., Rebourcet M. C., Nordmann J., Nordmann R., Giudicelli Y. Rat adipocyte lipoprotein lipase activity is inhibited by theophylline and stimulated by inosine through adenosine-independent mechanisms. Biochem Biophys Res Commun. 1982 Jul 16;107(1):59–67. doi: 10.1016/0006-291x(82)91669-2. [DOI] [PubMed] [Google Scholar]
  6. Brunzell J. D., Schwartz R. S., Eckel R. H., Goldberg A. P. Insulin and adipose tissue lipoprotein lipase activity in humans. Int J Obes. 1981;5(6):685–694. [PubMed] [Google Scholar]
  7. Chajek-Shaul T., Friedman G., Knobler H., Stein O., Etienne J., Stein Y. Importance of the different steps of glycosylation for the activity and secretion of lipoprotein lipase in rat preadipocytes studied with monensin and tunicamycin. Biochim Biophys Acta. 1985 Nov 14;837(2):123–134. doi: 10.1016/0005-2760(85)90235-8. [DOI] [PubMed] [Google Scholar]
  8. Chamberlain J. P. Fluorographic detection of radioactivity in polyacrylamide gels with the water-soluble fluor, sodium salicylate. Anal Biochem. 1979 Sep 15;98(1):132–135. doi: 10.1016/0003-2697(79)90716-4. [DOI] [PubMed] [Google Scholar]
  9. Cheng C. F., Oosta G. M., Bensadoun A., Rosenberg R. D. Binding of lipoprotein lipase to endothelial cells in culture. J Biol Chem. 1981 Dec 25;256(24):12893–12898. [PubMed] [Google Scholar]
  10. Cheung A. H., Bensadoun A., Cheng C. F. Direct solid phase radioimmunoassay for chicken lipoprotein lipase. Anal Biochem. 1979 Apr 15;94(2):346–357. doi: 10.1016/0003-2697(79)90371-3. [DOI] [PubMed] [Google Scholar]
  11. Eckel R. H., Prasad J. E., Kern P. A., Marshall S. Insulin regulation of lipoprotein lipase in cultured isolated rat adipocytes. Endocrinology. 1984 May;114(5):1665–1671. doi: 10.1210/endo-114-5-1665. [DOI] [PubMed] [Google Scholar]
  12. Elkeles R. S. Lipoprotein lipase in human adipose tissue. Clin Sci Mol Med. 1974 Jun;46(6):753–762. doi: 10.1042/cs0460753. [DOI] [PubMed] [Google Scholar]
  13. Etienne J., Noé L., Rossignol M., Arnaud C., Vydelingum N., Kissebah A. H. Antibody against rat adipose tissue lipoprotein lipase. Biochim Biophys Acta. 1985 Mar 27;834(1):95–102. doi: 10.1016/0005-2760(85)90180-8. [DOI] [PubMed] [Google Scholar]
  14. Fredholm B. B. Adenosine and lipolysis. Int J Obes. 1981;5(6):643–649. [PubMed] [Google Scholar]
  15. Goers J. W., Pedersen M. E., Kern P. A., Ong J., Schotz M. C. An enzyme-linked immunoassay for lipoprotein lipase. Anal Biochem. 1987 Oct;166(1):27–35. doi: 10.1016/0003-2697(87)90541-0. [DOI] [PubMed] [Google Scholar]
  16. Goldberg I. J., Blaner W. S., Goodman D. S. Immunologic and enzymatic comparisons between human and bovine lipoprotein lipase. Arch Biochem Biophys. 1986 Feb 1;244(2):580–584. doi: 10.1016/0003-9861(86)90626-0. [DOI] [PubMed] [Google Scholar]
  17. Goldberg I. J., Paterniti J. R., Jr, France D. S., Martinelli G., Cornicelli J. A. Production and use of an inhibitory monoclonal antibody to human lipoprotein lipase. Biochim Biophys Acta. 1986 Sep 12;878(2):168–176. doi: 10.1016/0005-2760(86)90143-8. [DOI] [PubMed] [Google Scholar]
  18. Hernell O., Egelrud T., Olivecrona T. Serum-stimulated lipases (lipoprotein lipases). Immunological crossreaction between the bovine and the human enzymes. Biochim Biophys Acta. 1975 Feb 13;381(2):233–241. [PubMed] [Google Scholar]
  19. Iverius P. H., Ostlund-Lindqvist A. M. Preparation, characterization, and measurement of lipoprotein lipase. Methods Enzymol. 1986;129:691–704. doi: 10.1016/0076-6879(86)29099-0. [DOI] [PubMed] [Google Scholar]
  20. Jonasson L., Hansson G. K., Bondjers G., Bengtsson G., Olivecrona T. Immunohistochemical localization of lipoprotein lipase in human adipose tissue. Atherosclerosis. 1984 May-Jun;51(2-3):313–326. doi: 10.1016/0021-9150(84)90179-5. [DOI] [PubMed] [Google Scholar]
  21. Kern P. A., Mandic A., Eckel R. H. Regulation of lipoprotein lipase by glucose in primary cultures of isolated human adipocytes. Relevance to hypertriglyceridemia of diabetes. Diabetes. 1987 Nov;36(11):1238–1245. doi: 10.2337/diab.36.11.1238. [DOI] [PubMed] [Google Scholar]
  22. Kern P. A., Marshall S., Eckel R. H. Regulation of lipoprotein lipase in primary cultures of isolated human adipocytes. J Clin Invest. 1985 Jan;75(1):199–208. doi: 10.1172/JCI111675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Londos C., Wolff J. Two distinct adenosine-sensitive sites on adenylate cyclase. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5482–5486. doi: 10.1073/pnas.74.12.5482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nilsson-Ehle P., Carlström S., Belfrage P. Rapid effect on lipoprotein lipase activity in adipose tissue of humans after carbohydrate and lipid intake. Scand J Clin Lab Invest. 1975 Jul;35(4):373–378. [PubMed] [Google Scholar]
  25. Nilsson-Ehle P., Garfinkel A. S., Schotz M. C. Lipolytic enzymes and plasma lipoprotein metabolism. Annu Rev Biochem. 1980;49:667–693. doi: 10.1146/annurev.bi.49.070180.003315. [DOI] [PubMed] [Google Scholar]
  26. Nissley S. P., Rechler M. M. Somatomedin/insulin-like growth factor tissue receptors. Clin Endocrinol Metab. 1984 Mar;13(1):43–67. doi: 10.1016/s0300-595x(84)80008-0. [DOI] [PubMed] [Google Scholar]
  27. Ohisalo J. J., Strandberg H., Kostiainen E., Kuusi T., Ehnholm C. Stimulation of lipoprotein lipase activity of rat adipose tissue and post-heparin plasma by N6-(phenylisopropyl)adenosine. FEBS Lett. 1981 Sep 14;132(1):121–123. doi: 10.1016/0014-5793(81)80442-5. [DOI] [PubMed] [Google Scholar]
  28. Olivecrona T., Bengtsson G. Immunochemical properties of lipoprotein lipase. Development of an immunoassay applicable to several mammalian species. Biochim Biophys Acta. 1983 Jun 16;752(1):38–45. doi: 10.1016/0005-2760(83)90230-8. [DOI] [PubMed] [Google Scholar]
  29. Parkin S. M., Walker K., Ashby P., Robinson D. S. Effects of glucose and insulin on the activation of lipoprotin lipase and on protein-synthesis in rat adipose tissue. Biochem J. 1980 Apr 15;188(1):193–199. doi: 10.1042/bj1880193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Patten R. L. The reciprocal regulation of lipoprotein lipase activity and hormone-sensitive lipase activity in rat adipocytes. J Biol Chem. 1970 Nov 10;245(21):5577–5584. [PubMed] [Google Scholar]
  31. Persson B., Hood B., Angervall G. Effects of prolonged fast on lipoprotein lipase activity eluted from human adipose tissue. Acta Med Scand. 1970 Sep;188(3):225–229. doi: 10.1111/j.0954-6820.1970.tb08030.x. [DOI] [PubMed] [Google Scholar]
  32. Pykälistö O. J., Smith P. H., Brunzell J. D. Determinants of human adipose tissue lipoprotein lipase. Effect of diabetes and obesity on basal- and diet-induced activity. J Clin Invest. 1975 Nov;56(5):1108–1117. doi: 10.1172/JCI108185. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sadur C. N., Eckel R. H. Insulin stimulation of adipose tissue lipoprotein lipase. Use of the euglycemic clamp technique. J Clin Invest. 1982 May;69(5):1119–1125. doi: 10.1172/JCI110547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Schwabe U., Schönhöfer P. S., Ebert R. Facilitation by adenosine of the action of insulin on the accumulation of adenosine 3':5'-monophosphate, lipolysis, and glucose oxidation in isolated fat cells. Eur J Biochem. 1974 Aug 1;46(3):537–545. doi: 10.1111/j.1432-1033.1974.tb03647.x. [DOI] [PubMed] [Google Scholar]
  35. Shirai K., Wisner D. A., Johnson J. D., Srivastava L. S., Jackson R. L. Immunological studies on bovine milk lipoprotein lipase. Effects of Fab fragments on enzyme activity. Biochim Biophys Acta. 1982 Jul 20;712(1):10–20. doi: 10.1016/0005-2760(82)90078-9. [DOI] [PubMed] [Google Scholar]
  36. Socorro L., Jackson R. L. Monoclonal antibodies to bovine milk lipoprotein lipase. Evidence for proteolytic degradation of the native enzyme. J Biol Chem. 1985 May 25;260(10):6324–6328. [PubMed] [Google Scholar]
  37. Taskinen M. R., Nikkilä E. A. Lipoprotein lipase activity of adipose tissue and skeletal muscle in insulin-deficient human diabetes. Relation to high-density and very-low-density lipoproteins and response to treatment. Diabetologia. 1979 Dec;17(6):351–356. doi: 10.1007/BF01236268. [DOI] [PubMed] [Google Scholar]
  38. Taskinen M. R., Nikkilä E. A. Lipoprotein lipase activity of adipose tissue and skeletal muscle in insulin-deficient human diabetes. Relation to high-density and very-low-density lipoproteins and response to treatment. Diabetologia. 1979 Dec;17(6):351–356. doi: 10.1007/BF01236268. [DOI] [PubMed] [Google Scholar]
  39. Vannier C., Amri E. Z., Etienne J., Négrel R., Ailhaud G. Maturation and secretion of lipoprotein lipase in cultured adipose cells. I. Intracellular activation of the enzyme. J Biol Chem. 1985 Apr 10;260(7):4424–4431. [PubMed] [Google Scholar]
  40. Vydelingum N. A., AlQuadan F., Kissebah A. H., Etienne J. Characterization of rat adipose tissue lipoprotein lipase using a monospecific antibody. Biochim Biophys Acta. 1986 May 21;876(3):399–412. doi: 10.1016/0005-2760(86)90026-3. [DOI] [PubMed] [Google Scholar]
  41. Wion K. L., Kirchgessner T. G., Lusis A. J., Schotz M. C., Lawn R. M. Human lipoprotein lipase complementary DNA sequence. Science. 1987 Mar 27;235(4796):1638–1641. doi: 10.1126/science.3823907. [DOI] [PubMed] [Google Scholar]
  42. Yki-Järvinen H., Taskinen M. R., Koivisto V. A., Nikkilä E. A. Response of adipose tissue lipoprotein lipase activity and serum lipoproteins to acute hyperinsulinaemia in man. Diabetologia. 1984 Sep;27(3):364–369. doi: 10.1007/BF00304851. [DOI] [PubMed] [Google Scholar]

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