Decreased Platelet Reactivity and Function in a Mouse Model of Human Pancreatic Cancer

Tomohiro Kawano; Yohei Hisada; Steven P. Grover; Wyatt J. Schug; David S. Paul; Wolfgang Bergmeier; Nigel Mackman

doi:10.1055/s-0043-1761419

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 2023; 123(05): 501-509
DOI: 10.1055/s-0043-1761419

Cellular Haemostasis and Platelets

Decreased Platelet Reactivity and Function in a Mouse Model of Human Pancreatic Cancer

Autor*innen

Tomohiro Kawano

¹Division of Hematology, Department of Medicine, UNC Blood Research Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Yohei Hisada

¹Division of Hematology, Department of Medicine, UNC Blood Research Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Steven P. Grover

¹Division of Hematology, Department of Medicine, UNC Blood Research Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Wyatt J. Schug

²Department of Biochemistry and Biophysics, UNC Blood Research Center, University of North Carolina at Chapel Hill, North Carolina, United States
David S. Paul

²Department of Biochemistry and Biophysics, UNC Blood Research Center, University of North Carolina at Chapel Hill, North Carolina, United States
Wolfgang Bergmeier

²Department of Biochemistry and Biophysics, UNC Blood Research Center, University of North Carolina at Chapel Hill, North Carolina, United States
Nigel Mackman

¹Division of Hematology, Department of Medicine, UNC Blood Research Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States

Abstract

Cancer patients have increased thrombosis and bleeding compared with the general population. Cancer is associated with activation of both platelets and coagulation. Mouse models have been used to study the dysregulation of platelets and coagulation in cancer. We established a mouse model of pancreatic cancer in which tissue factor-expressing human pancreatic tumors (BxPC-3) are grown in nude mice. Tumor-bearing mice have an activated coagulation system and increased venous thrombosis compared to control mice. We also showed that tumor-derived, tissue factor-positive extracellular vesicles activated platelets ex vivo and in vivo. In this study, we determined the effect of tumors on a platelet-dependent arterial thrombosis model. Unexpectedly, we observed significantly reduced carotid artery thrombosis in tumor-bearing mice compared to controls. In addition, we observed significantly increased tail bleeding in tumor-bearing mice compared to controls. These results suggested that the presence of the tumor affected platelets. Indeed, tumor-bearing mice exhibited a significant decrease in platelet count and an increase in mean platelet volume and percentage of reticulated platelets, findings that are consistent with increased platelet turnover. Levels of the platelet activation marker platelet factor 4 were also increased in tumor-bearing mice. We also observed decreased platelet receptor expression in tumor-bearing mice and reduced levels of active α_IIb/β₃ integrin in response to PAR4 agonist peptide and convulxin in platelets from tumor-bearing mice compared with platelets from control mice. In summary, our study suggests that in tumor-bearing mice there is chronic platelet activation, leading to thrombocytopenia, decreased receptor expression, and impaired platelet adhesive function.

Keywords

bleeding - cancer - extracellular vesicles - platelet - thrombosis

Volltext

Referenzen

References
1 Khorana AA, Dalal M, Lin J, Connolly GC. Incidence and predictors of venous thromboembolism (VTE) among ambulatory high-risk cancer patients undergoing chemotherapy in the United States. Cancer 2013; 119 (03) 648-655
2 Navi BB, Reiner AS, Kamel H. et al. Risk of arterial thromboembolism in patients with cancer. J Am Coll Cardiol 2017; 70 (08) 926-938
3 Timp JF, Braekkan SK, Versteeg HH, Cannegieter SC. Epidemiology of cancer-associated venous thrombosis. Blood 2013; 122 (10) 1712-1723
4 Hsu C, Patell R, Zwicker JI. The prevalence of thrombocytopenia in patients with acute cancer-associated thrombosis. Blood Adv 2022; (e-pub ahead of print).
5 Prandoni P, Lensing AW, Piccioli A. et al. Recurrent venous thromboembolism and bleeding complications during anticoagulant treatment in patients with cancer and venous thrombosis. Blood 2002; 100 (10) 3484-3488
6 Weitz JI, Haas S, Ageno W. et al; GARFIELD-VTE investigators. Cancer associated thrombosis in everyday practice: perspectives from GARFIELD-VTE. J Thromb Thrombolysis 2020; 50 (02) 267-277
7 Riedl J, Pabinger I, Ay C. Platelets in cancer and thrombosis. Hamostaseologie 2014; 34 (01) 54-62
8 Riedl J, Hell L, Kaider A. et al. Association of platelet activation markers with cancer-associated venous thromboembolism. Platelets 2016; 27 (01) 80-85
9 Ay C, Simanek R, Vormittag R. et al. High plasma levels of soluble P-selectin are predictive of venous thromboembolism in cancer patients: results from the Vienna Cancer and Thrombosis Study (CATS). Blood 2008; 112 (07) 2703-2708
10 Poruk KE, Firpo MA, Huerter LM. et al. Serum platelet factor 4 is an independent predictor of survival and venous thromboembolism in patients with pancreatic adenocarcinoma. Cancer Epidemiol Biomarkers Prev 2010; 19 (10) 2605-2610
11 Riedl J, Kaider A, Marosi C. et al. Decreased platelet reactivity in patients with cancer is associated with high risk of venous thromboembolism and poor prognosis. Thromb Haemost 2017; 117 (01) 90-98
12 Levi M. Disseminated intravascular coagulation in cancer: an update. Semin Thromb Hemost 2019; 45 (04) 342-347
13 Sallah S, Wan JY, Nguyen NP, Hanrahan LR, Sigounas G. Disseminated intravascular coagulation in solid tumors: clinical and pathologic study. Thromb Haemost 2001; 86 (03) 828-833
14 Dvorak HF, Quay SC, Orenstein NS. et al. Tumor shedding and coagulation. Science 1981; 212 (4497): 923-924
15 Yu JL, Rak JW. Shedding of tissue factor (TF)-containing microparticles rather than alternatively spliced TF is the main source of TF activity released from human cancer cells. J Thromb Haemost 2004; 2 (11) 2065-2067
16 Geddings JE, Mackman N. Tumor-derived tissue factor-positive microparticles and venous thrombosis in cancer patients. Blood 2013; 122 (11) 1873-1880
17 Mir Seyed Nazari P, Riedl J, Pabinger I, Ay C. The role of podoplanin in cancer-associated thrombosis. Thromb Res 2018; 164 (Suppl 1): S34-S39
18 Geddings JE, Hisada Y, Boulaftali Y. et al. Tissue factor-positive tumor microvesicles activate platelets and enhance thrombosis in mice. J Thromb Haemost 2016; 14 (01) 153-166
19 Hisada Y, Geddings JE, Ay C, Mackman N. Venous thrombosis and cancer: from mouse models to clinical trials. J Thromb Haemost 2015; 13 (08) 1372-1382
20 Davila M, Amirkhosravi A, Coll E. et al. Tissue factor-bearing microparticles derived from tumor cells: impact on coagulation activation. J Thromb Haemost 2008; 6 (09) 1517-1524
21 Wang JG, Geddings JE, Aleman MM. et al. Tumor-derived tissue factor activates coagulation and enhances thrombosis in a mouse xenograft model of human pancreatic cancer. Blood 2012; 119 (23) 5543-5552
22 Hisada Y, Ay C, Auriemma AC, Cooley BC, Mackman N. Human pancreatic tumors grown in mice release tissue factor-positive microvesicles that increase venous clot size. J Thromb Haemost 2017; 15 (11) 2208-2217
23 Thomas GM, Panicot-Dubois L, Lacroix R, Dignat-George F, Lombardo D, Dubois C. Cancer cell-derived microparticles bearing P-selectin glycoprotein ligand 1 accelerate thrombus formation in vivo. J Exp Med 2009; 206 (09) 1913-1927
24 Thomas GM, Brill A, Mezouar S. et al. Tissue factor expressed by circulating cancer cell-derived microparticles drastically increases the incidence of deep vein thrombosis in mice. J Thromb Haemost 2015; 13 (07) 1310-1319
25 Sasano T, Cho MS, Rodriguez-Aguayo C. et al. Role of tissue-factor bearing extracellular vesicles released from ovarian cancer cells in platelet aggregation in vitro and venous thrombosis in mice. Thrombosis Update 2021; 2: 100020
26 Stark K, Schubert I, Joshi U. et al. Distinct pathogenesis of pancreatic cancer microvesicle-associated venous thrombosis identifies new antithrombotic targets in vivo. Arterioscler Thromb Vasc Biol 2018; 38 (04) 772-786
27 Mezouar S, Darbousset R, Dignat-George F, Panicot-Dubois L, Dubois C. Inhibition of platelet activation prevents the P-selectin and integrin-dependent accumulation of cancer cell microparticles and reduces tumor growth and metastasis in vivo. Int J Cancer 2015; 136 (02) 462-475
28 Hisada Y, Grover SP, Maqsood A. et al. Neutrophils and neutrophil extracellular traps enhance venous thrombosis in mice bearing human pancreatic tumors. Haematologica 2020; 105 (01) 218-225
29 Owens III AP, Lu Y, Whinna HC, Gachet C, Fay WP, Mackman N. Towards a standardization of the murine ferric chloride-induced carotid arterial thrombosis model. J Thromb Haemost 2011; 9 (09) 1862-1863
30 Schiviz A, Magirr D, Leidenmühler P, Schuster M, Muchitsch EM, Höllriegl W. Subcommittee on Animal Models of the Scientific and Standardization Committee of the International Society on Thrombosis and Hemostasis. Influence of genetic background on bleeding phenotype in the tail-tip bleeding model and recommendations for standardization: communication from the SSC of the ISTH. J Thromb Haemost 2014; 12 (11) 1940-1942
31 Sommeijer DW, van Oerle R, Reitsma PH. et al. Analysis of blood coagulation in mice: pre-analytical conditions and evaluation of a home-made assay for thrombin-antithrombin complexes. Thromb J 2005; 3: 12
32 Stefanini L, Paul DS, Robledo RF. et al. RASA3 is a critical inhibitor of RAP1-dependent platelet activation. J Clin Invest 2015; 125 (04) 1419-1432
33 Grover SP, Mackman N. How useful are ferric chloride models of arterial thrombosis?. Platelets 2020; 31 (04) 432-438
34 Palacios-Acedo AL, Mezouar S, Mège D, Crescence L, Dubois C, Panicot-Dubois L. P2RY12-inhibitors reduce cancer-associated thrombosis and tumor growth in pancreatic cancers. Front Oncol 2021; 11: 704945
35 Morowski M, Vögtle T, Kraft P, Kleinschnitz C, Stoll G, Nieswandt B. Only severe thrombocytopenia results in bleeding and defective thrombus formation in mice. Blood 2013; 121 (24) 4938-4947
36 Liu Y, Jennings NL, Dart AM, Du XJ. Standardizing a simpler, more sensitive and accurate tail bleeding assay in mice. World J Exp Med 2012; 2 (02) 30-36
37 Bergmeier W, Rackebrandt K, Schröder W, Zirngibl H, Nieswandt B. Structural and functional characterization of the mouse von Willebrand factor receptor GPIb-IX with novel monoclonal antibodies. Blood 2000; 95 (03) 886-893
38 Montague SJ, Andrews RK, Gardiner EE. Mechanisms of receptor shedding in platelets. Blood 2018; 132 (24) 2535-2545
39 Elvers M, Stegner D, Hagedorn I. et al. Impaired alpha(IIb)beta(3) integrin activation and shear-dependent thrombus formation in mice lacking phospholipase D1. Sci Signal 2010; 3 (103) ra1
40 Hisada Y, Mackman N. Mouse models of cancer-associated thrombosis. Thromb Res 2018; 164 (Suppl 1): S48-S53
41 Mege D, Mezouar S, Dignat-George F, Panicot-Dubois L, Dubois C. Microparticles and cancer thrombosis in animal models. Thromb Res 2016; 140 (Suppl 1): S21-S26
42 Mezouar S, Frère C, Darbousset R. et al. Role of platelets in cancer and cancer-associated thrombosis: experimental and clinical evidences. Thromb Res 2016; 139: 65-76
43 Mwiza JMN, Lee RH, Paul DS. et al. Both G protein-coupled and immunoreceptor tyrosine-based activation motif receptors mediate venous thrombosis in mice. Blood 2022; 139 (21) 3194-3203