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Thromb Haemost 1995; 73(02): 268-274
DOI: 10.1055/s-0038-1653763
Original Article
Fibrinolysis
Schattauer GmbH Stuttgart

Up-Regulated Expression of Plasminogen Activator Inhibitor-1 in Hep G2 Cells: Interrelationship between Insulin and Insulin-Like Growth Factor 1

F Anfosso
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
,
M C Alessi
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
,
G Nalbone
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
,
N Chomiki
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
,
M Henry
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
,
I Juhan-Vague
The Laboratoire d'Hématologie, INSERM CJF 93/12, Faculté de Médecine, Marseille, France
› Author Affiliations
Further Information

Publication History

Received 29 April 1994

Accepted after resubmission 19 October 1994

Publication Date:
09 July 2018 (online)

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Summary

Insulin resistance represents a situation with a high risk of athero-thrombosis and is accompanied by increased plasma plasminogen activator inhibitor-1 (PAI-1) levels. Fasting insulin level is highly correlated with PAI-1 levels in plasma. It has been shown that insulin increases PAI-1 synthesis by the human hepatoma cell line Hep G2. Moreover when Hep G2 cells expressing a down-regulation of insulin receptors by incubation with 10-7 M insulin, were stimulated by 10-9 M insulin, an overexpression of PAI-1 synthesis was observed despite a reduced number of insulin receptors. Insulin-like growth factor 1 (IGF-1) shares many properties with insulin. The aim of the present study was to evaluate the effect of IGF-1 on PAI-1 synthesis by Hep G2 cells down-regulated either by insulin or IGF-1.

Incubation of Hep G2 cells with increasing doses from 10-9 to 10-7 M IGF-1 induced a dose-dependent stimulation of PAI-1 synthesis up to 4.5-fold the control level. When cells were first pre-incubated with 10-7M IGF-1 for 18 h, acid washed, and then stimulated with 10-9 M IGF-1, the expression of IGF-1 receptors was greatly reduced (up to 70%). In contrast PAI-1 secretion was increased 3.4-fold the level of control cells and by 1.9-fold the level of cells first stimulated with 10-9M IGF-1. Both transcripts of PAI-1 mRNA were also increased. The overexpression of PAI-1 synthesis was observed irrespective of the hormone used in the down-regulation step (i.e. 10-9 M insulin or IGF-1) or in the stimulation step (i. e. 10-9 M insulin or IGF-1). The results showed an interrelationship between insulin and IGF-1 on PAI-1 synthesis in down-regulated Hep G2 cells. They also suggest that in the insulin resistant state, IGF-1 would be able to participate in the increase in PAI-1 plasma levels by stimulating down-regulated insulin target cells.

 

  • References

  • 1 Bachmann FW. Fibrinolysis. In: Thrombosis Haemostasis. Verstraete M, Vermylen J, Lijnen HR, Amout J. eds Leuven University Press, Leuven; Belgium: 1987. pp 227-265
    Google Scholar
  • 2 Emeis J, Van Hinsbergh VW, Verheijen J, Wijngaards G. Inhibitor of the tissue-type plasminogen activator by conditioned medium from cultured human and porcine vascular endothelial cells. Biochem Biophys Res Commun 1983; 110: 392-397
    CrossrefPubMedGoogle Scholar
  • 3 Sprengers E, Princen HM, Kooistra T, Van Hinsbergh VW. Inhibition of plasminogen activators by conditioned medium of human hepatocytes and hepatoma cell line Hep G2. Lab Clin Med 1985; 10: 751-756
    PubMedGoogle Scholar
  • 4 Juhan-Vague I, Alessi MC. Plasminogen activator inhibitor 1 and athero- thrombosis. Thromb Haemost 1993; 70: 138-143
    Article in Thieme ConnectPubMedGoogle Scholar
  • 5 Hamsten A, De FaireU, Walldius G, Dahlen G, Szamosi A, Landov D, Blämback M, Wiman B. Plasminogen activator inhibitor in plasma: risk factor for recurrent myocardial infarction. Lancet 1987; 2: 2-3
    PubMedGoogle Scholar
  • 6 Fontbonne A, Eschwege E. Diabetes, hyperglycaemia, hyperinsulinemia and atherosclerosis: epidemiological data. Diab Metab 1987; 13: 350-353
    PubMedGoogle Scholar
  • 7 Reaven G. The Banting lecture. Role of insulin resistance in human disease Diabetes 1988; 37: 1595-1607
    CrossrefPubMedGoogle Scholar
  • 8 Juhan-Vague I, Alessi MC, Vague P. Increased plasma plasminogen activator inhibitor 1 levelsA possible link between insulin resistance and athero- thrombosis. Diabetologia 1991; 34: 457-462
    CrossrefPubMedGoogle Scholar
  • 9 Alessi MC, Juhan-Vague I, Kooistra T, Declerck PJ, Collen D. Insulin stimulates the synthesis of plasminogen activator inhibitor 1 by the hepatocellular cell line Hep G2. Thromb Haemost 1988; 60: 491-495
    Article in Thieme ConnectPubMedGoogle Scholar
  • 10 Kooistra T, Bosma P, Täns H, Van denBerg, Meyer P, Princen H. Plasminogen activator inhibitor 1: biosynthesis and mRNA level are increased by insulin in cultured human hepatocytes. Thromb Haemost 1989; 62: 723-728
    Article in Thieme ConnectPubMedGoogle Scholar
  • 11 Grant PJ, Riiegg M, Medcalf RL. Basal expression and insulin-mediated induction of PAI-1 mRNA in Hep G2 cells. Fibrinolysis 1991; 5: 81-86
    PubMedGoogle Scholar
  • 12 Caro JF, Dohm L, Sinha M. Cellular alterations in liver, skeletal muscle and adipose tissue responsible for insulin resistance in obesity and type II diabetes. Diabetes Metab Rev 1989; 5: 665-681
    CrossrefPubMedGoogle Scholar
  • 13 Blake AD, Hayes NS, Slater EE, Strader CD. Insulin receptor desensitization correlates with attenuation of tyrosine kinase activity but not of receptor endocytosis. Biochem J 1987; 245: 257-264
    CrossrefPubMedGoogle Scholar
  • 14 Anfosso F, Chomiki N, Alessi MC, Vague P, Juhan-Vague I. Plasminogen activator inhibitor-1 synthesis in the human hepatoma cell line Hep G2 Metformin inhibits the stimulating effect of insulin. J Clin Invest 1993; 91: 2185-2193
    CrossrefPubMedGoogle Scholar
  • 15 Froesch E, Schmid C, Schwander J. Action of insulin-like growth factors. Am Rev Physiol 1985; 47: 443-467
    CrossrefPubMedGoogle Scholar
  • 16 Sara V, Hall K. Insulin-like growth factors and their binding proteins. Physiological Reviews 1990; 70: 591-614
    CrossrefPubMedGoogle Scholar
  • 17 Verspohl EJ, Roth RA, Vigneri R, Goldftne D. Dual regulation of glycogen metabolism by insulin and insulin-like growth factors in human's hepatoma cell (Hep G2). Analysis with an anti-receptor monoclonal antibody J Clin Invest 1984; 74: 1436-1443
    CrossrefPubMedGoogle Scholar
  • 18 Soos MA, Siddle KImmunological. Evidence for structural heterogeneity of insulin-like growth factor I receptors involving hybrids with insulin receptors. Biochem J 1989; 263: 553-563
    CrossrefPubMedGoogle Scholar
  • 19 Guler HP, Zapf J, Froesch E. Short-term metabolic effects of recombinant human insulin-like growth factor I in healthy adults. N Engl J Med 1987 317: 137-140
    PubMedGoogle Scholar
  • 20 Jacob R, Barrett E, Plewe G, Fagin K, Sherwin R. Acute effects of insulin-like growth factor I on glucose and amino-acid metabolism in the awake-fasted rat. Comparison with insulin. J Clin Invest 1989; 83: 1717-1723
    CrossrefPubMedGoogle Scholar
  • 21 Verspohl E, Maddrux B, Goldfine D. Insulin and insulin-like growth factor I regulate the same biological functions in Hep G2 cells via their own specific receptor. J Clin Endocrinol Metab 1988; 67: 169-174
    CrossrefPubMedGoogle Scholar
  • 22 Anfosso F, Alessi MC, Latron Y, Juhan-Vague I. IGF-1 increases PAI-1 synthesis by human hepatoma cell line Hep G2. Fibrinolysis 1990; 4 Suppl (Suppl. 03) 175 (abstr.)
    PubMedGoogle Scholar
  • 23 Schneider DJ, Sobel BE. Augmentation of synthesis of plasminogen activator inhibitor type I by insulin and insulin-like growth factor type I: implica tions for vascular disease in hyperinsulinemic states. Proc Natl Acad Sci USA 1991; 88: 9959-9963
    CrossrefPubMedGoogle Scholar
  • 24 Padayatti SJ, Orme S, Zenobi PD, Stickland MH, Belchetz PE, Grant PJ. The effects of insulin-like growth factor-1 on plasminogen activator inhibitor-1 synthesis and secretion: results from in vitro and in vivo studies. Thromb Haemost 1993; 70: 1009-1013
    Article in Thieme ConnectPubMedGoogle Scholar
  • 25 Jaffe E, Nachman R, Becker C, Minick C. Culture of human endothelial cells obtained from umbilical veins Identification by morphologic and immunological criteria. J Clin Invest 1973; 52: 2745-2756
    CrossrefPubMedGoogle Scholar
  • 26 Declerck PJ, Alessi MC, Verstreken M, Kruithof EK O, Juhan-Vague I, Collen D. Measurement of plasminogen activator inhibitor 1 in biologic fluids with a murine monoclonal antibody based enzyme linked immunosorbent assay. Blood 1988; 71: 220-225
    PubMedGoogle Scholar
  • 27 Garvey T, Olefsky JM, Marshall S. Insulin induces progressive insulin resistance in cultured rat adipocytes. Sequential effects at receptor and multiple post receptor sites Diabetes 1986; 35: 258-267
    CrossrefPubMedGoogle Scholar
  • 28 Melton DA, Krieg PA, Rebagliati MR, Maniatis T, Zinn K, Green MR. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. . Nucl Acid Res 1984; 12: 7035-7056
    CrossrefPubMedGoogle Scholar
  • 29 Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidin thiocyanate-phenol chloroform extraction. Anal Biochem 1987; 162: 156-159
    PubMedGoogle Scholar
  • 30 Medcalf RL, Van den Berg EA, Schleuning WD. Glucocorticoid modulates gene expression of tissue and urinary-type plasminogen activators and plasminogen activator inhibitor 1 and 2. J Cell Biol 1988; 106: 971-978
    CrossrefPubMedGoogle Scholar
  • 31 Stout RW, Vallance-Owen J. Insulin and atheroma, 20 yr perspective. Diabetes Care 1990; 13: 631-654
    CrossrefPubMedGoogle Scholar
  • 32 Loskutoff D, Linders M, Keijer J, Veerman H, van HeerikhuizenH, Panne- koekH. Structure of the human plasminogen activator inhibitor 1 gene: nonrandom distribution of introns. Biochemistry 1987; 26: 3763-3768
    CrossrefPubMedGoogle Scholar
  • 33 Schneider DJ, Nordt TK, Sobel BE. Stimulation by proinsulin of expression of plasminogen activator inhibitor type-1 in endothelial cells. Diabetes 1992; 41: 890-895
    CrossrefPubMedGoogle Scholar
  • 34 Jialal I, Crettaz M, Hachiya H, Kahn C, Moses A, Buzney S, King G. Characterization of the receptors for insulin and the insulin-like growth factors on micro- and macro-vascular tissues. Endocrinology 1985; 117: 1222-1229
    CrossrefPubMedGoogle Scholar
  • 35 Bar R, Boes M, Yorek M. Processing of insulin-like growth factors I and II by capillary and large vessel endothelial cells. Endocrinology 1986; 118: 1072-1080
    CrossrefPubMedGoogle Scholar
  • 36 Rosenfeld RG, Dollar LA. Characterization of a specific receptor for somatomedin C (SM-C) on cultured human lymphocytes: evidence that SM-C modulates homologous receptor concentrations. Endocrinology 1980; 107: 1841-1848
    CrossrefPubMedGoogle Scholar
  • 37 Zick Y, Sakasi N, Rees-Jones RW, Grungenberg G, Nissley SP, Rechler MM. Insulin-like growth factor-1 (IGF-1) stimulates tyrosine kinase activity in purified receptors from a rat liver cell line. Biochem Biophys Res Commun 1984; 119: 6-13
    CrossrefPubMedGoogle Scholar
  • 38 Moxham CP, Duronio V, Jacobs S. Insulin-like growth factor I receptor β-subunit heterogeneity. Evidence for hybrid tetramers composed of insulin-like growth factor I and insulin receptor heterodimers J Biol Chem 1989; 264: 13238-13244
    PubMedGoogle Scholar
  • 39 Maegawa H, Olefsky JM, Thies S, Boyd D, Ullrich A, McClain DA. Insulin receptors with defective tyrosine kinase inhibit normal receptor function at the level of substrate phosphorylation. J Biol Chem 1988; 263: 12629-12637
    PubMedGoogle Scholar
  • 40 Treadway JL, Morrison BD, Soos MA, Siddle K, Olefsky JM, Ullrich A, McClain DA, Pessin JE. Transdominant inhibition of tyrosine kinase activity in mutant insulin/insulin-like growth factor I hybrid receptors. Proc Natl Acad Sci USA 1991; 88: 214-218
    CrossrefPubMedGoogle Scholar
  • 41 Frattali AL, Treadway JL, Pessin JE. Insulin/IGF-1 hybrid receptors. Implications for the dominant-negative phenotype in syndrome of insulin resistance J Cell Biochem 1992; 48: 43-50
    CrossrefPubMedGoogle Scholar
  • 42 Williams J, Olefsky JM. Defective insulin receptor function in down-regulated Hep G2 cells. Endocrinology 1990; 127: 1706-1717
    CrossrefPubMedGoogle Scholar
  • 43 Sung CK. Insulin receptor signaling through non-tyrosine kinase pathways: evidence from anti-receptor antibodies and insulin receptor mutants. J Cell Biochem 1992; 48: 26-32
    CrossrefPubMedGoogle Scholar
  • 44 Lin YJ, Harada S, Loten EG, Smith RM, Jarrett L. Direct stimulation of immediate-early genes by intranuclear insulin in trypsin-treated H35 hepatoma cells. Proc Natl Acad Sci USA 1992; 89: 9691-9694
    CrossrefPubMedGoogle Scholar
  • 45 Kolaczynski JW, Caro JF. Insulin-like growth factor-1 therapy in diabetes: physiologic basis, clinical benefits, and risks. Ann Intern Med 1994; 120: 47-55
    CrossrefPubMedGoogle Scholar
  • 46 Hochberg Z, Hertz P, Colin V, Ish-Shalom S, Yeshurun D, Youdim M, Amit T. The distal axis of growth hormone (GH) in nutritional disorders: GH-binding protein, insulin-like growth factor I (IGF-1) and IGF-1 receptors in obesity and anorexia nervosa. Metabolism 1992; 41: 106-12
    CrossrefPubMedGoogle Scholar
  • 47 Loskutoff DJ. Regulation of PAI-1 gene expression. Fibrinolysis 1991; 5: 197-206
    PubMedGoogle Scholar
  • 48 Caro JF, Poulos J, Ittoop O, Pories WJ, Flickinger EG, Sinha MK. Insulin-like growth factor I binding in hepatocytes from human liver, human hepatoma, regenerating, and fetal rat liver. J Clin Invest 1988; 81: 976-981
    CrossrefPubMedGoogle Scholar
  • 49 Schneiderman J, Sawdey J, Craig H, Thinnes T, Bordin G, Loskutoff DJ. Type I plasminogen activator inhibitor gene expression following partial hepatectomy. Am J Pathol 1993; 143: 753-762
    PubMedGoogle Scholar