Thromb Haemost 2022; 122(05): 755-766
DOI: 10.1055/s-0041-1735531
Cellular Haemostasis and Platelets

Characteristics of the Thrombus Formation in Transgenic Mice with Platelet-Targeted Factor VIII Expression

Yun Wang*
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Jianhua Mao*
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Li Li*
2   Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
,
Bing Xiao
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
3   Institute for Developmental and Regenerative Cardiovascular Medicine, Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Zheng Ruan
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Yichen Liu
4   Department of Hematology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, China
,
Guowei Zhang
5   Key Laboratory of Aging and Cancer Biology of Zhejiang Province, Department of Basic Medical Sciences, Hangzhou Normal University School of Medicine, Hangzhou, Zhejiang, China
,
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Jian-Qing Mi
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Chao Fang
2   Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
,
Xiaodong Xi
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
,
Xiaofeng Shi
4   Department of Hematology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu, China
6   Department of Hematology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, China
,
Jin Wang
1   State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, National Research Center for Translational Medicine at Shanghai, Pôle Sino-Français des Sciences du Vivant et Genomique, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
› Author Affiliations
Funding This work was supported by grants from the National Natural Science Foundation of China (81970112, 81700130, 81670127, 81570178, 81270594), Shanghai Municipal Health Commission in China (201940342), and Novo Nordisk Haemophilia Research Fund in China.

Abstract

Platelet-targeted FVIII gene therapy can efficiently recover bleeding phenotype for hemophilia A (HA), yet characteristics of thrombus formation with this ectopic expression of factor VIII (FVIII) in platelets remain unclear. Here, we generated 2bF8trans mice restrictively expressing human B-domain–deleted FVIII (hBDD FVIII) in platelets on a hemophilic (FVIIInull) mice background. The results showed no statistical difference in clot strength and stability between wild-type (WT) and 2bF8trans mice, but with a prolonged reaction time (R-time), by thromboelastography. Fluid dynamics analysis showed that at the shear rates of 500 to 1,500 s−1, where physiological hemostasis often develops, the thrombi formed in 2bF8trans mice were more stable than those in FVIIInull mice, while at high pathological shear rates (2,500 s−1), mimicking atherosclerosis, thrombus size and fibrin deposition in 2bF8trans mice were less than those in WT mice. Thrombus morphology analysis showed that there was a locally concentrated deposition of fibrin in thrombus at the injured site and fibrin co-localized with activated platelets in 2bF8trans mice. Moreover, a higher ratio of fibrin to platelets was found in thrombus from 2bF8trans mice following laser-induced injury in cremaster arterioles, which might be the underlying mechanism of thrombus stability in 2bF8trans mice at physiological arterial circumstance. These observations suggest that specific morphological features of the thrombi might contribute to the efficacy and safety of platelet-targeted FVIII gene therapy for HA.

* These authors contributed equally to this article.


Supplementary Material



Publication History

Received: 26 March 2021

Accepted: 31 July 2021

Article published online:
29 September 2021

© 2021. Thieme. All rights reserved.

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

 
  • References

  • 1 Srivastava A, Brewer AK, Mauser-Bunschoten EP. et al; Treatment Guidelines Working Group on Behalf of The World Federation Of Hemophilia. Guidelines for the management of hemophilia. Haemophilia 2013; 19 (01) e1-e47
  • 2 Chen SL. Economic costs of hemophilia and the impact of prophylactic treatment on patient management. Am J Manag Care 2016; 22 (5, Suppl): s126-s133
  • 3 Thorat T, Neumann PJ, Chambers JD. Hemophilia burden of disease: a systematic review of the cost-utility literature for hemophilia. J Manag Care Spec Pharm 2018; 24 (07) 632-642
  • 4 Hermans C, Altisent C, Batorova A. et al; European Haemophilia Therapy Standardisation Board. Replacement therapy for invasive procedures in patients with haemophilia: literature review, European survey and recommendations. Haemophilia 2009; 15 (03) 639-658
  • 5 Iorio A, Halimeh S, Holzhauer S. et al. Rate of inhibitor development in previously untreated hemophilia A patients treated with plasma-derived or recombinant factor VIII concentrates: a systematic review. J Thromb Haemost 2010; 8 (06) 1256-1265
  • 6 Mancuso ME, Mannucci PM, Rocino A, Garagiola I, Tagliaferri A, Santagostino E. Source and purity of factor VIII products as risk factors for inhibitor development in patients with hemophilia A. J Thromb Haemost 2012; 10 (05) 781-790
  • 7 Franchini M, Coppola A, Rocino A. et al; Italian Association of Hemophilia Centers (AICE) Working Group. Systematic review of the role of FVIII concentrates in inhibitor development in previously untreated patients with severe hemophilia a: a 2013 update. Semin Thromb Hemost 2013; 39 (07) 752-766
  • 8 Nathwani AC. Gene therapy for hemophilia. Hematology (Am Soc Hematol Educ Program) 2019; 2019 (01) 1-8
  • 9 Shi Q. Platelet-targeted gene therapy for hemophilia. Mol Ther Methods Clin Dev 2018; 9: 100-108
  • 10 Shi Q, Wilcox DA, Fahs SA. et al. Factor VIII ectopically targeted to platelets is therapeutic in hemophilia A with high-titer inhibitory antibodies. J Clin Invest 2006; 116 (07) 1974-1982
  • 11 Shi Q, Fahs SA, Kuether EL, Cooley BC, Weiler H, Montgomery RR. Targeting FVIII expression to endothelial cells regenerates a releasable pool of FVIII and restores hemostasis in a mouse model of hemophilia A. Blood 2010; 116 (16) 3049-3057
  • 12 Baumgartner CK, Mattson JG, Weiler H, Shi Q, Montgomery RR. Targeting factor VIII expression to platelets for hemophilia A gene therapy does not induce an apparent thrombotic risk in mice. J Thromb Haemost 2017; 15 (01) 98-109
  • 13 Chavin SI. Factor VIII: structure and function in blood clotting. Am J Hematol 1984; 16 (03) 297-306
  • 14 Shi Q, Wilcox DA, Fahs SA, Kroner PA, Montgomery RR. Expression of human factor VIII under control of the platelet-specific alphaIIb promoter in megakaryocytic cell line as well as storage together with VWF. Mol Genet Metab 2003; 79 (01) 25-33
  • 15 Mosesson MW. The roles of fibrinogen and fibrin in hemostasis and thrombosis. Semin Hematol 1992; 29 (03) 177-188
  • 16 Mosesson MW. Fibrinogen and fibrin structure and functions. J Thromb Haemost 2005; 3 (08) 1894-1904
  • 17 Thachil J. Deep vein thrombosis. Hematology 2014; 19 (05) 309-310
  • 18 Shi X, Yang J, Huang J. et al. Effects of different shear rates on the attachment and detachment of platelet thrombi. Mol Med Rep 2016; 13 (03) 2447-2456
  • 19 Stalker TJ, Traxler EA, Wu J. et al. Hierarchical organization in the hemostatic response and its relationship to the platelet-signaling network. Blood 2013; 121 (10) 1875-1885
  • 20 Maxwell MJ, Westein E, Nesbitt WS, Giuliano S, Dopheide SM, Jackson SP. Identification of a 2-stage platelet aggregation process mediating shear-dependent thrombus formation. Blood 2007; 109 (02) 566-576
  • 21 Baumgartner CK, Zhang G, Kuether EL, Weiler H, Shi Q, Montgomery RR. Comparison of platelet-derived and plasma factor VIII efficacy using a novel native whole blood thrombin generation assay. J Thromb Haemost 2015; 13 (12) 2210-2219
  • 22 Chen J, Schroeder JA, Luo X, Montgomery RR, Shi Q. The impact of GPIbα on platelet-targeted FVIII gene therapy in hemophilia A mice with pre-existing anti-FVIII immunity. J Thromb Haemost 2019; 17 (03) 449-459
  • 23 Gao C, Schroeder JA, Xue F. et al. Nongenotoxic antibody-drug conjugate conditioning enables safe and effective platelet gene therapy of hemophilia A mice. Blood Adv 2019; 3 (18) 2700-2711
  • 24 Ramiz S, Hartmann J, Young G, Escobar MA, Chitlur M. Clinical utility of viscoelastic testing (TEG and ROTEM analyzers) in the management of old and new therapies for hemophilia. Am J Hematol 2019; 94 (02) 249-256
  • 25 Castellino FJ, Donahue DL, Navari RM, Ploplis VA, Walsh M. An accompanying genetic severe deficiency of tissue factor protects mice with a protein C deficiency from lethal endotoxemia. Blood 2011; 117 (01) 283-289
  • 26 Fressinaud E, Sakariassen KS, Rothschild C, Baumgartner HR, Meyer D. Shear rate-dependent impairment of thrombus growth on collagen in nonanticoagulated blood from patients with von Willebrand disease and hemophilia A. Blood 1992; 80 (04) 988-994
  • 27 Tomaiuolo M, Matzko CN, Poventud-Fuentes I, Weisel JW, Brass LF, Stalker TJ. Interrelationships between structure and function during the hemostatic response to injury. Proc Natl Acad Sci U S A 2019; 116 (06) 2243-2252
  • 28 Yarovoi H, Nurden AT, Montgomery RR, Nurden P, Poncz M. Intracellular interaction of von Willebrand factor and factor VIII depends on cellular context: lessons from platelet-expressed factor VIII. Blood 2005; 105 (12) 4674-4676
  • 29 Welsh JD, Stalker TJ, Voronov R. et al. A systems approach to hemostasis: 1. The interdependence of thrombus architecture and agonist movements in the gaps between platelets. Blood 2014; 124 (11) 1808-1815
  • 30 Receveur N, Nechipurenko D, Knapp Y. et al. Shear rate gradients promote a bi-phasic thrombus formation on weak adhesive proteins, such as fibrinogen in a VWF-dependent manner. Haematologica 2020; 105 (10) 2471-2483
  • 31 Casa LDC, Ku DN. Thrombus formation at high shear rates. Annu Rev Biomed Eng 2017; 19: 415-433
  • 32 Casa LD, Deaton DH, Ku DN. Role of high shear rate in thrombosis. J Vasc Surg 2015; 61 (04) 1068-1080
  • 33 Li M, Hotaling NA, Ku DN, Forest CR. Microfluidic thrombosis under multiple shear rates and antiplatelet therapy doses. PLoS One 2014; 9 (01) e82493
  • 34 Roka-Moiia Y, Walk R, Palomares DE. et al. Platelet activation via shear stress exposure induces a differing pattern of biomarkers of activation versus biochemical agonists. Thromb Haemost 2020; 120 (05) 776-792
  • 35 Kroll MH, Hellums JD, McIntire LV, Schafer AI, Moake JL. Platelets and shear stress. Blood 1996; 88 (05) 1525-1541
  • 36 Rumbaut RE, Randhawa JK, Smith CW, Burns AR. Mouse cremaster venules are predisposed to light/dye-induced thrombosis independent of wall shear rate, CD18, ICAM-1, or P-selectin. Microcirculation 2004; 11 (03) 239-247
  • 37 Li W, Nieman M, Sen Gupta A. Ferric chloride-induced murine thrombosis models. J Vis Exp 2016; 5 (115) e54479
  • 38 Kaneva VN, Dunster JL, Volpert V, Ataullahanov F, Panteleev MA, Nechipurenko DY. Modeling thrombus shell: linking adhesion receptor properties and macroscopic dynamics. Biophys J 2021; 120 (02) 334-351