Inflammatory and Procoagulant Mediator Interactions in an Experimental Baboon Model of Venous Thrombosis

Summary

Theoretic and in vitro evidence suggests that thrombosis and inflammation are interrelated. The purpose of the present study was to define the relationship between inflammation and deep venous thrombosis (DVT) in an in vivo model. Initiation of DVT was accomplished by administration of antibody to protein C (HPC₄, 2 mg/kg) and tumor necrosis factor (TNF, 150 μg/kg); stasis; and subtle venous catheter injury. Thrombosis was assessed by thrombin-antithrombin assay (TAT), ¹²⁵I-fibrinogen scanning (scan) over both the proximal and distal iliac veins, and ascending venography. Cytokines TNF, interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), and interleukin-8 (IL-8) were measured along with differential white blood cell counts, platelet counts, fibrinogen (FIB), and erythrocyte sedimentation rates (ESR). Baboon pairs were sacrificed on day 3 (T + 3d), T + 6d, and T + 9d and veins removed. All animals developed inferior vena cava and left iliofemoral DVT by venography; no right DVT was found. TAT was elevated by T + 1hr and peaked at T + 3hrs. Left iliofemoral DVT was found at T + 1hr by scan and reached a 20% uptake difference between the affected left and nonaffected right side at T + 3hrs. TNF peaked at T + 1hr; MCP-1 peaked at T + 6hrs; IL-8 and IL-6 peaked on T + 2d; all cytokines declined to baseline. TNF and TAT elevations were found to correlate with all cytokines; elevations in IL-8 were correlated with elevations in MCP-1 and IL-6 (p <0.05). Correlation between cytokines and scan revealed a significant (p <0.05) relationship only between elevations in IL-6 and distal iliac fibrin accumulation; no significant correlation was found between IL-8 and MCP-1 and scan. Increased mature polymorphonuclear leukocytes were found by T + 2d; immature forms were prominent at T + 3hrs, T + 6hrs, and T + 2d. Increased monocytes were noted by T + 4d; increased lymphocytes and platelets by T + 8d. ESR and FIB were elevated by T + 3d. Histopathologic study revealed venous inflammation at T + 3d, with beginning thrombus organization by T + 6d. MCP-1 localized to areas of thrombus and phlebitis. The development of DVT in this model involves inflammatory as well as coagulant activity. We conclude that this model allows studies on the role of inflammatory mediators in the development and natural history of DVT.

• References

• 1 Esmon NL, Esmon CT. Protein C and the endothelium. Semin Thromb Hemostas 1988; 14: 210-215
• 2 Nawroth PP, Stern DM. Modulation of endothelial cell hemostatic properties by tumor necrosis factor. J Exp Med 1986; 163: 740-745
• 3 Bevilacqua MP, Pober JS, Majeau GR, Fiers W, Cotran RS, Gimbrone MA Jr. Recombinant tumor necrosis factor induces procoagulant activity in cultured human vascular endothelium: characterization and comparison with the actions of interleukin 1. Proc Natl Acad Sci USA 1986; 83: 4533-4537
• 4 Conway EM, Bach R, Rosenberg RD, Konigsberg WH. Tumor necrosis factor enhances expression of tissue factor mRNA in endothelial cells. Thromb Res 1989; 53: 231-241
• 5 Schleef RR, Bevilacqua MP, Sawdey M, Gimbrone Jr MA, Loskutoff DJ. Cytokine activation of vascular endothelium. Effects on tissue-type plasminogen activator and type 1 plasminogen activator inhibitor. J Biol Chem 1988; 263: 5797-5803
• 6 van Hinsbergh VW, Kooistra T, van den Berg EA, Princen HM, Fiers W, Emeis JJ. Tumor necrosis factor increases production of plasminogen activator inhibitor in human endothelial cells in vitro and in rats in vivo. Blood 1988; 72: 1467-1473
• 7 Medina R, Socher SH, Han JH, Friedman PA. Interleukin-1, endotoxin, or tumor necrosis factor/cachectin enhance the level of plasminogen activator inhibitor messenger RNA in bovine aortic endothelial cells. Thromb Res 1989; 54: 41-52
• 8 Van der Poll T, Levi M, Büller HR, van Deventer SJH, de Boer JP, Hack CE, ten Cate JW. Fibrinolytic response to tumor necrosis factor in healthy subjects. J Exp Med 1991; 174: 729-732
• 9 Wakefield TW, Wrobleski SK, Sarpa MS, Taylor FB Jr, Esmon CT, Chang A, Greenfield LJ. Deep venous thrombosis in the baboon: an experimental model. J Vase Surg 1991; 14: 588-598
• 10 Taylor FB, Esmon CT, Chang A. Primate model of deep vein thrombosis induced by inflammatory mediators [Abstract]. Circulation 1990; 82 (03) 770
• 11 Strieter RM, Remick DG, Ham JM, Colletti LM, Lynch III JP, Kunkel SL. Tumor necrosis factor-alpha gene expression in human whole blood. J Leukocyte Biol 1990; 47: 366-370
• 12 Standiford TJ, Kunkel SL, Basha MA, Chensue SW, Lynch III JP, Toews GB, Westwick J, Strieter RM. Interleukin-8 gene expression by a pulmonary epithelial cell line. A model for cytokine networks in the lung. J Clin Invest 1990; 86: 1945-1953
• 13 Van Oers MH, Van der Heyden AA, Aarden LA. Interleukin 6 (IL-6) in serum and urine of renal transplant recipients. Clin Exp Immunol 1988; 71: 314-319
• 14 Evanoff HL, Burdick MD, Moore SA, Kunkel SL, Strieter RM. A sensitive ELISA for the detection of human monocyte chemoattrac-tant protein-1 (MCP-1). Immunol Invest 1992; 21: 39-45
• 15 Taylor Jr FB, Chang AC, Esmon CT, Hinshaw LB. Baboon model of Escherichia coli sepsis: description of its four stages and the role of tumor necrosis factor, tissue factors, and the protein C system in septic shock. Curr Stud Hematol Blood Transfus 1991; 58: 8-14
• 16 Voss BL, DeBault LE, Blick KE, Chang AC, Stiers DL, Hinshaw LB, Taylor FB Jr. Sequential renal alterations in septic shock in the primate. Circ Shock 1991; 33: 142-155
• 17 Hinshaw LB, Archer LT, Beller-Todd BK, Coalson JJ, Flournoy DJ, Passey R, Benjamin B, White GL. Survival of primates in LD100 septic shock following steroid/antibiotic therapy. J Surg Res 1980; 28: 151-170
• 18 Creasey AA, Stevens P, Kenney J, Allison AC, Warren K, Catlett R, Hinshaw LB, Taylor FB Jr. Endotoxin and cytokine profile in plasma of baboons challenged with lethal and sublethal Escherichia coli. Circ Shock 1991; 33: 84-91
• 19 Taylor Jr FB, Chang A, Ruf W, Morrissey JH, Hinshaw L, Catlett R, Blick K, Edginton TS. Lethal E. coli septic shock is prevented by blocking tissue factor with monoclonal antibody. Circ Shock 1991; 33: 127-134
• 20 Day KC, Hoffman LC, Palmier MO, Kretzmer KK, Huang MD, Pyla EY, Spokas E, Broze GJ Jr, Warren TG, Wun TC. Recombinant lipoprotein-associated coagulation inhibitor inhibits tissue thromboplastin-induced intravascular coagulation in the rabbit. Blood 1990; 76: 1538-1545
• 21 Taylor Jr FB, Chang AC, Peer GT, Mather T, Blick K, Catlett R, Lockhart MS, Esmon CT. DEGR-factor Xa blocks disseminated intravascular coagulation initiated by Escherichia coli without preventing shock or organ damage. Blood 1991; 78: 364-368
• 22 Taylor F, Chang A, Ferrell G, Mather T, Catlett R, Blick K, Esmon CT. C4b-binding protein exacerbates the host response to Escherichia coli. Blood 1991; 78: 357-363
• 23 Taylor Jr FB, Chang A, Esmon CT, D'Angelo A, Vigano-D'ngelo S, Blick KE. Protein C prevents the coagulopathic and lethal effects of Escherichia coli infusion in the baboon. J Clin Invest 1987; 79: 918-925
• 24 Taylor FB Jr, Emerson Jr TE, Jordan R, Chang AC, Blick KE. Antithrombin III prevents the lethal effects of E. coli infusion in baboons. Circ Shock 1988; 26: 227-235
• 25 Kunkel SL, Standiford T, Metinko AP, Strieter RM. Endothelial cell-derived novel chemotactic cytokines. In: Lung Vascular Injury: Molecular and Cellular Response. Lefant C. (ed). Marcel Dekker, Inc.; New York: 1991
• 26 Hebert CA, Luscinskas FW, Kiely JM, Luis EA, Darbonne WC, Bennett GL, Liu CC, Obin MS, Gibrone Jr MA, Baker JB. Endothelial and leukocyte forms of IL-8. Conversion by thrombin and interactions with neutrophils. J Immunol 1990; 145: 3033-3040
• 27 Wang JM, Sica A, Peri G, Walter S, Padura IM, Libby P, Ceska M, Lindley I, Colotta F, Mantovani A. Expression of monocyte chemotactic protein and interleukin-8 by cytokine-activated human vascular smooth muscle cells. Arterioscler Thromb 1991; 11: 1166-1174
• 28 Strieter RM, Wiggins R, Phan SH, Wharram BL, Showell HJ, Remick DG, Chensue SW, Kunkel SL. Monocyte chemotactic protein gene expression by cytokine-treated human fibroblasts and endothelial cells. Biochem Biophys Res Commun 1989; 162: 694-700
• 29 Sica A, Wang JM, Colotta F, Dejana E, Mantovani A, Oppenheim JJ, Larsen CG, Zachariae CO, Matsushima K. Monocyte chemotactic and activating factor gene expression induced in endothelial cells by IL-1 and tumor necrosis factor. J Immunol 1990; 144: 3034-3038
• 30 Schaub RG, Simmons CA, Koets MH, Romano PJ, Stewart GJ. Early events in the formation of a venous thrombus following local trauma and stasis. Lab Invest 1984; 51: 218-224
• 31 Simmons CA, Burdick MD, Schaub RG. Heparin inhibits fibrin, but not leukocytes, in a model of deep-vein thrombosis. J Surg Res 1987; 43: 468-475
• 32 Loppnow H, Libby P. Adult human vascular endothelial cells express the IL-6 gene differentially in response to LPS or IL-1. Cell Immunol 1989; 122: 493-503
• 33 Loppnow H, Libby P. Proliferating or interleukin 1-activated human vascular smooth muscle cells secrete copious interleukin 6. J Clin Invest 1990; 85: 731-738
• 34 Hirano T, Akira S, Taga T, Kishimoto T. Biological and clinical aspects of interleukin 6. Immunol Today 1990; 11: 443-449
• 35 Van der Poll T, van Deventer SJH, Hack CE, Wolbink GJ, Aarden LA, Buller HR, ten Cate JW. Effects on leukocytes after injection of tumor necrosis factor into healthy humans. Blood 1992; 79: 693-698
• 36 Hoek JA, Sturk A, ten Cate JW, Lamping RJ, Berenda F, Borm JJ. Laboratory and clinical evaluation of an assay of thrombin-antithrombin III complexes in plasma. Clin Chem 1988; 34: 2058-2062
• 37 Loppnow H, Libby P. Functional significance of human vascular smooth muscle cell-derived interleukin 1 in paracrine and autocrine regulation pathways. Exp Cell Res 1992; 198: 283-290
• 38 Mileski W, Borgstrom D, Lightfoot E, Rothlein R, Faanes R, Lipsky P, Baxter C. Inhibition of leukocyte-endothelial adherence following thermal injury. J Surg Res 1992; 52: 334-339
• 39 Van der Poll T, Büller HR, ten Cate H, Wortel CH, Bauer KA, van Deventer SJ, Hack CE, Sauerwein HP, Rosenberg RD, ten Cate JW. Activation of coagulation after administration of tumor necrosis factor to normal subjects. N Engl J Med 1990; 322: 1622-1627