Download PDF
Thromb Haemost 2009; 102(06): 1204-1211
DOI: 10.1160/TH09-03-0194
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Expression of thrombin-activatable fibrinolysis inhibitor (TAFI) is up-regulated by increase in intracellular cyclic AMP levels in cultured HepG2 cells

Hidemi Ishii
1   Laboratory of Molecular and Cellular Pathophysiology, Showa Pharmaceutical University, Tokyo, Japan
2   High Technology Research Center, Showa Pharmaceutical University, Tokyo, Japan
,
Yuka Kojima
1   Laboratory of Molecular and Cellular Pathophysiology, Showa Pharmaceutical University, Tokyo, Japan
,
Yutaka Masuda
1   Laboratory of Molecular and Cellular Pathophysiology, Showa Pharmaceutical University, Tokyo, Japan
2   High Technology Research Center, Showa Pharmaceutical University, Tokyo, Japan
,
Kimihiko Takada
1   Laboratory of Molecular and Cellular Pathophysiology, Showa Pharmaceutical University, Tokyo, Japan
,
Katsuyoshi Sugimoto
2   High Technology Research Center, Showa Pharmaceutical University, Tokyo, Japan
,
Tomohiro Ikeda
3   Department of Ophthalmology, Hyogo College of Medicine, Hyogo, Japan
› Author AffiliationsFinancial support: This work was supported by Grant-in-Aid for High Technology Research Centre Project (19–8) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
Further Information

Publication History

Received: 26 March 2009

Accepted after minor revision: 07 July 2009

Publication Date:
28 November 2017 (online)

Also available at
    Permissions and Reprints

Summary

Thrombin-activatable fibrinolysis inhibitor (TAFI), a carboxypeptidase B-like proenzyme, is predominantly biosynthesised in the liver and released into circulating plasma. Activated TAFI has a role in maintaining the balance between blood coagulation and fibrinolysis. We investigated the regulation of TAFI expression in cultured human hepatoma HepG2 cells. Stimulation of the cells with forskolin and dibutyryl cyclic AMP (DBcAMP) increased TAFI antigen levels in the cells in parallel with TAFI mRNA levels and antigen release from the cells into the conditioned medium. The elevated TAFI expression was abolished by pretreatment of the cells with KT5720, a protein kinase A (PKA) inhibitor. The promoter activity of the TAFI gene and the half-life of the TAFI transcript in DBcAMP-stimulated HepG2 cells increased to 1.5-fold and 2.0-fold, respectively, of those in the control cells. The increased promoter activity and the prolonged half-life were abolished by pretreatment of the cells with KT5720.These results suggest that an increase in intracellular cAMP levels upregulates TAFI expression in the cells in accompaniment with an elevation of TAFI mRNA levels, and that the elevated mRNA levels are derived from both transcriptional and post-transcriptional regulations of the TAFI gene mediated by activation of the AMP/PKA signaling pathway.

Keywords

Thrombin-activatable fibrinolysis inhibitor (TAFI) - TAFI mRNA stability - cyclic AMP - protein kinase A - TAFI promoter activity
 

  • References

  • 1 Bouma BN, Marx PF, Mosnier LO. et al. Thrombin-activatable fibrinolysis inhibitor (TAFI, plasma procarboxypeptidase B, procarboxypeptidase R, procarboxypeptidase U). Thromb Res 2001; 101: 329-354.
    CrossrefPubMedGoogle Scholar
  • 2 Leurs J, Hendriks D. Carboxypeptidase U (TAFIa): a metallocarboxypeptidase with a distinct role in haemostasis and a possible risk factor for thrombotic disease. Thromb Haemost 2005; 94: 471-487.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 3 Redlitz A, Tan AK, Eaton DL, Plow EF. Plasma carboxypeptidases as regulators of the plasminogen system. J Clin Invest 1995; 96: 2534-2538.
    CrossrefPubMedGoogle Scholar
  • 4 Wu C, Dong N, da Cunha V, Martin-McNulty B. et al. Activated thrombin-activatable fibrinolysis inhibitor attenuates spontaneous fibrinolysis of batroxobin-induced fibrin deposition in rat lungs. Thromb Haemost 2003; 90: 414-421.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 5 Bouma BN, Meijers JC. Thrombin-activatable fibrinolysis inhibitor (TAFI, plasma procarboxypeptidase B, procarboxypeptidase R, procarboxypeptidase U). J Thromb Haemost 2003; 01: 1566-1574.
    CrossrefPubMedGoogle Scholar
  • 6 Muto Y, Suzuki K, Sato E. et al. Carboxypeptidase B inhibitors reduce tissue factor-induced renal microthrombi in rats. Eur J Pharmacol 2003; 461: 181-189.
    CrossrefPubMedGoogle Scholar
  • 7 Suzuki K, Muto Y, Fushihara K. et al. Enhancement of fibrinolysis by EF6265 [(S)-7-amino-2-[[[(R)-2-methyl-1-(3-phenylpropanoylamino)propyl]hydroxypho sphinoyl] methyl]heptanoic acid], a specific inhibitor of plasma carboxypeptidase B. J Pharmacol Exp Ther 2004; 309: 607-615.
    CrossrefPubMedGoogle Scholar
  • 8 Eichinger S, Schonauer V, Weltermann A. et al. Thrombin-activatable fibrinolysis inhibitor and the risk for recurrent venous thromboembolism. Blood 2004; 103: 3773-3776.
    CrossrefPubMedGoogle Scholar
  • 9 Montaner J, Ribo M, Monasterio J. et al. Thrombin-activable fibrinolysis inhibitor levels in the acute phase of ischemic stroke. Stroke 2003; 34: 1038-1040.
    CrossrefPubMedGoogle Scholar
  • 10 Santamaria A, Oliver A, Borrell M. et al. Risk of ischemic stroke associated with functional thrombin-activatable fibrinolysis inhibitor plasma levels. Stroke 2003; 34: 2387-2391.
    CrossrefPubMedGoogle Scholar
  • 11 Leebeek FW, Goor MP, Guimaraes AH. et al. High functional levels of thrombin-activatable fibrinolysis inhibitor are associated with an increased risk of first ischemic stroke. J Thromb Haemost 2005; 03: 2211-2218.
    CrossrefPubMedGoogle Scholar
  • 12 Ladenvall C, Gils A, Jood K. et al. Thrombin activatable fibrinolysis inhibitor activation peptide shows association with all major subtypes of ischemic stroke and with TAFI gene variation. Arterioscler Thromb Vasc Biol 2007; 27: 955-962.
    CrossrefPubMedGoogle Scholar
  • 13 Campbell W, Okada N, Okada H. Carboxypeptidase R is an inactivator of complement-derived inflammatory peptides and an inhibitor of fibrinolysis. Immunol Rev 2001; 180: 162-167.
    CrossrefPubMedGoogle Scholar
  • 14 Tan AK, Eaton DL. Activation and characterization of procarboxypeptidase B from human plasma. Biochemistry 1995; 34: 5811-5816.
    CrossrefPubMedGoogle Scholar
  • 15 Boffa MB, Hamill JD, Bastajian N. et al. A role for CCAAT/enhancer-binding protein in hepatic expression of thrombin-activable fibrinolysis inhibitor. J Biol Chem 2002; 277: 25329-25336.
    CrossrefPubMedGoogle Scholar
  • 16 Boffa MB, Hamill JD, Maret D. et al. Acute phase mediators modulate thrombin-activable fibrinolysis inhibitor (TAFI) gene expression in HepG2 cells. J Biol Chem 2003; 278: 9250-9257.
    CrossrefPubMedGoogle Scholar
  • 17 Mullarky IK, Szaba FM, Winchel CG. et al. In situ assays demonstrate that interferon-gamma suppresses infection-stimulated hepatic fibrin deposition by promoting fibrinolysis. J Thromb Haemost 2006; 04: 1580-1587.
    CrossrefPubMedGoogle Scholar
  • 18 Maret D, Boffa MB, Brien DF. et al. Role of mRNA transcript stability in modulation of expression of the gene encoding thrombin activable fibrinolysis inhibitor. J Thromb Haemost 2004; 02: 1969-1979.
    CrossrefPubMedGoogle Scholar
  • 19 Hori Y, Nakatani K, Morioka K. et al. Insulin enhanced thrombin-activable fibrinolysis inhibitor expression through PI3 kinase/Akt pathway. Int J Mol Med 2005; 15: 265-268.
    PubMedGoogle Scholar
  • 20 Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 1983; 65: 55-63.
    CrossrefPubMedGoogle Scholar
  • 21 Eaton DL, Malloy BE, Tsai SP. et al. Isolation, molecular cloning, and partial characterization of a novel carboxypeptidase B from human plasma. J Biol Chem 1991; 266: 21833-21838.
    PubMedGoogle Scholar
  • 22 Ishii H, Horie S, Kizaki K. et al. Retinoic acid counteracts both the downregulation of thrombomodulin and the induction of tissue factor in cultured human endothelial cells exposed to tumor necrosis factor. Blood 1992; 80: 2556-2562.
    PubMedGoogle Scholar
  • 23 Wilson GM, Deeley RG. An episomal expression vector system for monitoring sequence-specific effects on mRNA stability in human cell lines. Plasmid 1995; 33: 198-207.
    CrossrefPubMedGoogle Scholar
  • 24 Dani C, Piechaczyk M, Audigier Y. et al. Characterization of the transcription products of glyceraldehyde 3-phosphate-dehydrogenase gene in HeLa cells. Eur J Biochem 1984; 145: 299-304.
    CrossrefPubMedGoogle Scholar
  • 25 Hori Y, Gabazza EC, Yano Y. et al. Insulin resistance is associated with increased circulating level of thrombin-activatable fibrinolysis inhibitor in type 2 diabetic patients. J Clin Endocrinol Metab 2002; 87: 660-665.
    CrossrefPubMedGoogle Scholar
  • 26 Kolacz E, Lewandowski K, Zawilska K. The role of thrombin activatable fibrinolysis inhibitor in disturbances of fibrinolytic system in obesity. Pol Arch Med Wewn 2005; 113: 424-430.
    PubMedGoogle Scholar
  • 27 Aubert H, Frere C, Aillaud MF. et al. Weak and nonindependent association between plasma TAFI antigen levels and the insulin resistance syndrome. J Thromb Haemost 2003; 01: 791-797.
    CrossrefPubMedGoogle Scholar
  • 28 Garand M, Bastajian N, Nesheim ME. et al. Molecular analysis of the human thrombin-activatable fibrinolysis inhibitor gene promoter. Br J Haematol 2007; 138: 231-244.
    CrossrefPubMedGoogle Scholar
  • 29 Cluck MW, Murphy LO, Olson J. et al. Amylin gene expression mediated by cAMP/PKA and transcription factors HNF-1 and NFY. Mol Cell Endocrinol 2003; 210: 63-75.
    CrossrefPubMedGoogle Scholar
  • 30 Massaad C, Houard N, Lombes M. et al. Modulation of human mineralocorticoid receptor function by protein kinase A. Mol Endocrinol 1999; 13: 57-65.
    CrossrefPubMedGoogle Scholar
  • 31 Tebo J, Der S, Frevel M. et al. Heterogeneity in control of mRNA stability by AU-rich elements. J Biol Chem 2003; 278: 12085-12093.
    CrossrefPubMedGoogle Scholar
  • 32 Adams DJ, Beveridge DJ, van der Weyden L. et al. HADHB, HuR, and CP1 bind to the distal 3’-untranslated region of human renin mRNA and differentially modulate renin expression. J Biol Chem 2003; 278: 44894-44903.
    CrossrefPubMedGoogle Scholar
  • 33 Jungmann RA, Kiryukhina O. Cyclic AMP and AKAP-mediated targeting of protein kinase A regulates lactate dehydrogenase subunit A mRNA stability. J Biol Chem 2005; 280: 25170-25177.
    CrossrefPubMedGoogle Scholar