Human Platelet Factor 4 and Its C-Terminal Peptides: Heparin Binding and Clearance from the Circulation

Summary

Human platelet factor 4 (PF4), a high affinity heparin binding protein, is released from stimulated platelets and stored at vascular sites, predominantly in liver, from where it can be brought back into circulation by heparin. We attempted to define structural requirements for PF4 binding to heparin and for the pattern of its clearance from the circulation. Intact PF4 bound strongly to heparin agarose and was eluted at 1.4 M NaCl, while reduced PF4 and PF4 C-terminal peptides PF4 (47-70) and PF4 (58-70) bound weakly and were eluted at 0.2-0.5 M NaCl. ¹²⁵I-radiolabeled intact PF4, reduced PF4 and C-terminal PF4 peptides injected into rabbits were cleared from the circulation in a biphasic pattern with components having half-life time of 1-2 min and 20-140 min. Heparin eliminated the fast component of PF4 clearance, but it did not affect clearance of reduced PF4 or C-terminal PF4 peptides. In contrast to reduced PF4 and PF4 (47-70), intact PF4 that accumulated in the liver and spleen, was displaced by heparin into circulating blood. In conclusion, specific binding sites and native conformation of the molecule are critical for high affinity PF4 binding to insolubilized heparin and for a pattern of PF4 clearance from the circulation in the presence of heparin.

• References

• 1 Niewiarowski S, Thomas DP. Platelet factor 4 and adenosine diphosphate release during human platelet aggregation. Nature (London) 1969; 222: 1269-70
• 2 Barber AJ, Kaser-Glanzmann R, Jakabova M, Luscher EF. Characterization of chondroitin sulfate proteoglycan carrier for heparin-neutralizing activity (PF4) released from human blood platelets. Biochim Biophys Acta 1972; 286: 312-29
• 3 Moore S, Pepper DS, Cash JD. Platelet antiheparin activity. The isolation and characterization of platelet factor 4 released from thrombin-aggregated washed human platelets and its dissociation into subunits and the isolation of membrane-bound antiheparin activity. Biochim Biophys Acta 1975; 379: 370-84
• 1 Deuel TF, Keim PS, Farmer M, Heinrikson RL, Platelet factor. Complete amino acid sequence. Proc Natl Acad Sci USA 1977; 74: 2256-8
• 5 Hermodson M, Schmer G, Kurachi K. Isolation, characterization and primary amino acid sequence of human platelet factor 4. J Biol Chem 1977; 252: 6276-9
• 6 Walz DA, Wu VY, DeLamo R, Dene H, McCoy LE. Primary structure of human platelet factor 4. Thromb Res 1977; 11: 892-8
• 7 Handin RI, Cohen JH. Purification and binding properties of human platelet factor 4. J Biol Chem 1976; 251: 4273-82
• 8 Loscalzo J, Melnick B, Handin RI. The interaction of platelet factor 4 and glycosaminoglycans. Arch Biochem Biophys 1985; 240: 446-55
• 9 Moore S, Pepper DS. Identification and characterization of a platelet specific release product: β-thromboglobulin. In: Platelets in Biology and Pathology Gordon JL. ed. Elsevier/North-Holland Biomedical Press; Amsterdam: 1976: 293-311
• 10 Rucinski B, Niewiarowski S, James P, Walz DA, Budzynski AZ. Antiheparin proteins secreted by human platelets. Purification, characterization and immunoassay. Blood 1979 53. 47-62
• 11 Begg GS, Pepper DS, Chesterman CN, Morgan FJ. Complete covalent structure of human β-thromboglobulin. Biochemistry 1978; 17: 1739-44
• 12 Cowan S, Bakshi EN, Machim KJ, Isaacs NW. Binding of heparin to human platelet factor 4. Biochem J 1986; 234: 485-8
• 13 St Charles R, Walz DA, Edwards BF P. The three-dimensional structure of bovine platelet factor 4 at 3.0. - A resolution. J Biol Chem 1989; 264: 2092-9
• 14 Mayo KH, Chen M-J. Human platelet factor 4 monomer-dimer-trimer equilibria investigated by 1H NMR spectroscopy. Biochemistry 1989; 28: 9469-78
• 15 Laterra J, Norton EK, Izzard CS, Culp LA. Contact formation by fibroblasts adhering to heparan sulfate-binding substrata (fibronectin or platelet factor 4). Exp Cell Res 1983; 146: 15-27
• 16 Deuel TF, Senior RM, Chang D, Griffin GL, Heinrikson RL. Platelet factor 4 is chemotactic for neutrophils and monocytes. Proc Natl Acad Sci USA 1981; 78: 4854-7
• 17 Katz IR, Thorbecke GJ, Bell MK, Yin J-Z, Clarke D, Zucker MB. Protease-induced immunoregulatory activity of platelet factor 4. Proc Natl Acad Sci USA 1986; 83: 3491-5
• 18 Gewirtz AM, Calabretta B, Rucinski B, Niewiarowski S, Xu W-Y. Inhibition of human megakaryocytopoiesis in vitro by platelet factor 4 (PF4) and a synthetic C-terminal PF4 peptide. J Clin Invest 1989; 83: 1477-86
• 19 Osterman DG, Griffin GI, Senior RM, Kaiser ET, Deuel TF. The carboxy-terminal tridecapeptide of platelet factor 4 is a potent chemotactic agent for monocytes. Biochem Biophys Res Comm 1982; 107: 130-5
• 20 Zucker MB, Katz IR, Thorbecke GJ, Milot DC, Holt JC. Immunoregulatory activity of peptides related to platelet factor 4. Proc Natl Acad Sci USA 1989; 86: 7571-4
• 21 Busch C, Dawes J, Pepper DW, Wasteson A. Binding of platelet factor 4 to cultured human umbilical vein endothelial cells. Thromb Res 1980; 19: 129-38
• 22 Dawes JC W, Pumphrey KM, McLaren KM, Prowse CV, Pepper DS. The in vivo release of human platelet factor 4 by heparin. Thromb Res 1982; 27: 65-76
• 23 Rao AK, Niewiarowski S, James P, Holt JC, Harris M, Elfenbein B, Bastl C. Effect of heparin on the in vivo release and clearance of human platelet factor 4. Blood 1983; 61: 1208-14
• 24 Rybak ME, Gimbrone Jr MA, Davies PF, Handin RI. Interaction of platelet factor 4 with cultured vascular endothelial cells. Blood 1989; 73: 1534-9
• 25 Rucinski B, Knight LC, Niewiarowski S. Clearance of human platelet factor 4 by liver and kidney: its alteration by heparin. Amcr J Physiol 1986; 251: H800
• 26 Rucinski B, Stewart GJ, DeFeo PA, Boden G, Niewiarowski S. Uptake and processing of human platelet factor 4 by hepatocytes. Proc Soc Exp Biol Med 1987; 186: 361-7
• 27 Busch C, Owen WG. Identification in vitro of an endothelial cell surface cofactor for antithrombin III: parallel studies with isolated perfused rat hearts and microcarriers culture of bovine endothelium. J Clin Invest 1982; 69: 726-9
• 28 Marcum JA, McKenney JB, Rosenberg RD. Acceleration of thrombin-antithrombin complex formation in rat hindquarters via heparin-like molecules bound to the endothelium. J Clin Invest 1984; 74: 341-50
• 29 Bastl CP, Musial J, Kloczewiak M, Guzzo J, Berman I, Niewiarowski S. Role of kidney in the catabolic clearance of human platelet antiheparin proteins from rat circulation. Blood 1981; 57: 233-8
• 30 Kloczewiak M, Timmons S, Lukas TJ, Hawiger J. Platelet receptor recognition site on human fibrinogen. Synthesis and structure-function relationship of peptides corresponding to the carboxyterminal segment of the gamma chain. Biochemistry 1984 23. 1767-74
• 31 Huang T-F, Holt JC, Lukasiewicz H, Niewiarowski S. Trigramin - A low molecular weight peptide inhibiting fibrinogen interaction with platelet receptors expressed on glycoprotein Ilb/IIIa complex. J Biol Chem 1987; 262: 16157-63
• 32 Rucinski B, Niewiarowski S, Strzyzewski M, Holt JC, Mayo KH. Structural requirements for the expression of heparin binding activity of platelet factor 4 and its clearance from the circulation. Blood 1988; 72: 1255