San Jose State University Research Foundation San Jose, California, United States
Introduction:: Mechanical heart valves (MHVs) are preferred for long term heart valve implantations due to their excellent durability [1]. However, MHVs suffer from serious thromboembolic complications which lead to patients undergoing lifelong anticoagulant therapy [2]. The lack of a comprehensive testing platform to evaluate MHV thrombogenicity and its connection with the valve design features is a critical barrier to the innovation of coagulation free MHV design. Our novel thrombogenicity tester (TGT) is designed to expose pristine MHVs to blood flow under realistic physiological conditions and assess the level of clot deposition on the valve within a stipulated time. Characterizing clot formation on commercially available bileaflet MHVs may shed light on the role of initial clot deposits on thrombus growth rate. In this study, we wanted to test the hypothesis that platelet count is reduced after circulation in the TGT over a MHV.
Materials and Methods:: Our TGT is a pumpless circulation system, derived from the classic Chandler loop [3], and adapted to replicate physiological blood flow in the presence of a MHV. The loop was rotated at 58 rpm. A motor was used to rotate the loop, alternating clockwise and counterclockwise motion, to replicate systole and diastole (1/3 and 2/3 of one heart cycle, respectively). Opening and closing of the valve leaflets were achieved by change in the rotation direction. We obtained heparinized porcine whole blood from an external laboratory (Lampire Biological Laboratories, Pipersville, PA). Immediately before our tests, we added 200 mg of protamine sulfate to neutralize the Na-heparin anticoagulant and accelerate clotting time. A St. Jude Regent 27-mm bileaflet MHV (Abbott Laboratories) was mounted in a 1-inch-diameter Tygon loop, which was filled with 400 ml of blood, and then incubated at 37oC for at least 2 hours, and no longer than 6 hours. The test was repeated with 9 blood samples. A clamp-on ultrasonic flowmeter (Transonic, Ithaca, NY) was used to non-invasively measure the flow profile inside the loop (Figure 1). Red blood cell (RBC), white blood cell (WBC) and platelet (PLT) counts were measured before and after the experiment using a Element HT5 hematology analyzer. The mass of the MHV was measured before and after incubation. We assumed that the difference is representative of the mass of the blood clots grown on the surfaces of the valve.
Results, Conclusions, and Discussions:: Results and Discussion
In our preliminary tests, we observed the formation of blood clots on the surfaces of the MHV in 8 out of 9 samples. In those 8 samples, the difference in the masses of pre-incubation (3.38g ± 0.02g) and post-incubation valves (11.68g ± 3.1g) was statistically significant (p < 0.05). Figure 2 shows the presence of a large clot from both sides of the valve. Preliminary data appear to indicate a reduction in blood flow in the loop after incubation. The example in Figure 1 shows that after 2 hours of incubation the peak systolic flow rate was reduced by 40-50%. We attributed this decrease to the occlusion of the valve caused by blood clots. Figure 3a shows a decrease in platelet count after incubation. The cell counts are normalized by the average pre-incubation value for each test and shown in Figure 3b. When normalized, significant reduction in the white blood cell count is also observed. Large clots may have trapped white blood cells and agglomeration of platelets which explain their reduction in the post-incubation blood. Conclusion The aim of the present study was to develop a novel in-vitro platform that can replicate blood clotting on MHVs under realistic physiological conditions. Preliminary results are consistent with our hypothesis that platelet count decreases in the blood after incubation, due to clot formation on the leaflets. These initial findings need to be confirmed using in a larger sample size. Future tests with this thrombogenicity tester device will include varying heart rate, varying concentration of protamine sulphate and varying incubation time to investigate their effect on controlling the level of clot deposits. Upon completion of this project, hydrodynamic studies of thrombotic MHVs with different degree of clots placed in mock circulation loops will provide new insights regarding thrombus formation and growth rate in patients with implanted MHVs.
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References (Optional): : < !1. Fiedler, A. G. & Tolis, G. Surgical Treatment of Valvular Heart Disease: Overview of Mechanical and Tissue Prostheses, Advantages, Disadvantages, and Implications for Clinical Use. Curr. Treat. Options Cardiovasc. Med. 20, (2018).
2. Linde, T., Clauser, J., Meuris, B., & Steinseifer, U. (2019). Assessing the thrombogenic potential of heart valve prostheses: an approach for a standardized in-vitro method. Cardiovascular Engineering and Technology, 10, 216-224.
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