Characterisation of the fibrinolytic system and the Von Willebrand factor-Adamts13 axis in the Chacma baboon
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Date
2021-11
Authors
Joubert, Jaco
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Journal ISSN
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Publisher
University of the Free State
Abstract
Background: The Chacma baboon (Papio ursinus) model of acquired thrombotic thrombocytopenic purpura (aTTP) is ideally suited to investigate novel treatments with potential application in this lethal thrombotic disorder, which hinges on the dysfunction of the VWF–ADAMTS13 axis. One such modality is the thrombolytic drugs, which activate the fibrinolytic system. Our recently published pilot study demonstrated the thrombolytic drug streptokinase as ineffective in resolving aTTP in this model and highlighted the deficits in our knowledge of the Chacma baboon’s fibrinolytic system and VWFADAMTS13 axis. The present study aimed to characterise these components of the Chacma baboon’s haemostatic system to better understand the aTTP model, the effects of thrombolytics in this model, and the implications for other haemostatic disease models in this species. Materials and Methods: Forty baboons were tested using observational and experimental assays. The VWF–ADAMTS13 axis was investigated by determining ADAMTS13 antigen and activity levels, VWF:Ag, VWF:RCo, and VWF:CB levels and VWF multimer patterns. The fibrinolytic system was explored by measuring the concentrations of fibrinogen, plasminogen, tPA, PAI-1, PAP complexes, TAFI, and α2- antiplasmin, and by assessing its in vitro clot lysis ability in a modified clot lysis time assay.The plasminogen activation potentials of streptokinase and tPA were determined using concentration escalation experiments. The effect of tPA-induced plasmin activity on VWF multimer patterns when in the non-globular state was assessed in the presence of the anti-ADAMTS13 mAb 3H9. Thrombin generation, which can influence the aTTP model and its response to thrombolytics, was also evaluated, as well as the effects of sex and ABO blood group. Reference intervals and interindividual variation were calculated and compared with human values. Results and Discussion: ADAMTS13 activities were generally below (but still comparable to) human ranges. VWF:Ag and VWF:CB values tended towards the lower limit of the human reference interval, whereas VWF:RCo activities were higher. All VWF multimer patterns were essentially equivalent to normal human patterns. Fibrinogen concentrations were similar to human values, but tPA, PAP complex, PAI-1 and α2- antiplasmin concentrations all tended toward the lower human ranges. Meaningful results could not be generated for ADAMTS13 antigen, plasminogen, or TAFI, possibly due to structural differences with the human protein. Streptokinase resulted in minimal plasmin activity, but tPA led to concentration-dependent increases. All baboon samples exceeded 100% of human plasmin activity at baseline and had clot lysis times shorter than pooled normal human plasma when activated by tPA. In the absence of ADAMTS13 activity, tPA reduced the non-globular high molecular weight VWF multimers in both human and baboon samples. Baboons had greater overall endogenous thrombin potentials than humans, which was more prominent in females (p=0.0238). Except for lower fibrinogen concentrations (p=0.0134) in male baboons, and PAP complex concentrations which were higher (p=0.0188), no other sex-related differences were apparent. No baboons were typed as ABO group B or AB, and none were Rh(D) negative. Group O baboons had higher fibrinogen concentrations (p=0.0355), but all other parameters were unaffected. Fibrinogen and especially α2-antiplasmin were subject to considerable interindividual biological variation. Conclusion: The central thesis of this research is that the components of the Chacma baboon’s haemostatic system pertinent to the pathogenesis of aTTP and its treatment with thrombolytics are similar enough to their human counterparts to enable continued use of this species as a model of human haemostasis, provided quantitative results are interpreted within the context of the novel reference intervals, and the identified limitations and interspecies differences are considered. Finally, tPA should be explored further in the,Chacma baboon aTTP model in vivo to provide proof-of-concept for the use of thrombolytic drugs in the treatment of aTTP in humans.
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Keywords
Thesis (Ph.D. (Haematology))--University of the Free State, 2021, Thrombotic thrombocytopenic purpura, Chacma baboon, Papio ursinus, Von Willebrand factor–ADAMTS13 axis, Fibrinolytic system, Animal model, Animal model, Streptokinase, Tissue-type plasminogen activator