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In the present study, the effects of initial collagen fiber orientation on the medium-term (up to 50 Â 10 6 cycles) fatigue response of heart valve soft tissue biomaterials was investigated. Glutaraldehyde treated bovine pericardium (GLBP), preselected for uniform structure and collagen fiber orientation, was used as the representative heart valve biomaterial. Using specialized instrumentation, GLBP specimens were subjected to cyclic tensile loading to maximum stress levels of 500 6 50 kPa at a frequency of 22 Hz. Two sample groups were examined, one with the preferred collagen fiber direction parallel (PD) and perpendicular (XD) to the direction of applied strain. The primary findings indicated that GLBP fatigue response was highly sensitive to the direction of loading with respect to fiber orientation. Specifically, when loading perpendicular to the preferred collagen fiber orientation, fiber reorientation is the dominant mechanism. In contrast, when loaded parallel to the preferred fiber direction a reduction in both collagen fiber crimp and fiber reorientation occurred. Moreover, alterations in the degree and direction of mechanical anisotropy can be inducted by cyclic loading when specimens are loaded perpendicular to the preferred fiber direction. Fourier Transform Infrared Spectroscopy (FT-IR) results indicate that molecular-level damage to collagen occurs in both groups after only 20 Â 10 6 cycles. Taken as a whole, the results of this study suggest that initial collagen orientation plays a critical role in bioprosthetic heart valve biomaterial fatigue response.

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Platelet activation is the initiating step to thromboembolic complications in blood-contacting medical devices. Currently, there are no widely accepted testing protocols or relevant metrics to assess platelet activation during the in vitro evaluation of new medical devices. In this article, two commonly used platelet activation marker antibodies, CD62P (platelet surface P-selectin) and PAC1 (activated GP IIb/IIIa), were evaluated using flow cytometry. Anticoagulant citrate dextrose solution A (ACDA) and heparin anticoagulated human blood from healthy donors were separately exposed to shear stresses of 0, 10, 15, and 20 Pa for 120 s using a cone-plate rheometer model, and immediately mixed with the platelet marker antibodies for analysis. To monitor for changes in platelet reactivity between donors and over time, blood samples were also evaluated after exposure to 0, 2, and 20 µM of adenosine diphosphate (ADP). Following ADP stimulation, the percentage of both CD62P and PAC1 positive platelets increased in a dose dependent fashion, even 8 h after the blood was collected. After shear stress stimulation, both CD62P and PAC1 positive platelets increased significantly at shear stress levels of 15 and 20 Pa when ACDA was used as the anticoagulant. However, for heparinized blood, the PAC1 positive platelets decreased with increasing shear stress, while the CD62P positive platelets increased. Besides the anticoagulant effect, the platelet staining buffer also impacted PAC1 response, but had little effect on CD62P positive platelets. These data suggest that CD62P is a more reliable marker compared with PAC1 for measuring shear-dependent platelet activation and it has the potential for use during in vitro medical device testing.

As platelet activation plays a critical role in physiological hemostasis and pathological thrombosis, it is important in the overall hemocompatibility evaluation of new medical devices and biomaterials to assess their effects on platelet function. However, there are currently no widely accepted in vitro test methods to perform this assessment. In an effort to develop effective platelet tests for potential use in medical device evaluation, this study compared the sensitivity of platelet responses to shear stress stimulation of human and bovine blood using multiple platelet activation markers. Fresh whole blood samples anticoagulated with heparin or anticoagulant citrate dextrose, solution A (ACDA) were exposed to shear stresses up to 40 Pa for 2 min using a cone-and-plate rheometer model. Platelet activation was characterized by platelet counts, platelet surface P-selectin expression, and serotonin release into blood plasma. The results indicated that exposure to shear stresses above 20 Pa caused significant changes in all three of the platelet markers for human blood and that the changes were usually greater with ACDA anticoagulation than with heparin. In contrast, for bovine blood, the markers did not change with shear stress stimulation except for plasma serotonin in heparin anticoagulated blood. The differences observed between human and bovine platelet responses suggest that the value of using bovine blood for in vitro platelet testing to evaluate devices may be limited.

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A reliable in vitro dynamic test method to evaluate device thrombogenicity is very important for the improvement of the design and safety of blood-contacting medical devices, while reducing the use of animal studies. In this study, a recirculating flow loop system was developed for thrombogenicity testing, using donor sheep blood anticoagulated with Anticoagulant Citrate Dextrose Solution A (ACDA) and used within 24–36 hr postdraw. Immediately before testing, the blood was recalcified and heparinized to a donor-specific target concentration. The heparinization level was based on a static pretest, in which latex tubes were incubated at room temperature for 30 min in blood with a series of heparin concentrations and evaluated for thrombus deposition. For dynamic testing, blood was recirculated at room temperature through a polyvinyl chloride (PVC) tubing loop containing a test material for 1 hr at 200 ml/min using a roller pump. Nine materials were investigated: a negative control (polytetrafluoroethylene [PTFE]), a positive control (latex), and seven commonly used biomaterials including PVC, two silicones with different formulations (Q-Sil and V-Sil), nylon, polyurethane (PU), high-density polyethylene (HDPE), and polyether block amide (PEBAX). The results showed that latex was significantly more thrombogenic than all the other materials (p < 0.05), PVC and Q-Sil exhibited intermediate thrombogenicity with significantly more thrombus surface coverage and thrombus weight than PTFE (p < 0.05), whereas PTFE and the rest of the biomaterials had little to no thrombus deposition. In summary, the test loop system was able to effectively differentiate materials with different thrombogenic potentials.

Nanocrystalline diamond (NCD) coatings have been investigated for improved wear resistance and enhanced hemocompatibility of cardiovascular devices. The goal of this study was to evaluate the effects of NCD surface nanotopography on in vitro hemocompatibility. NCD coatings with small (NCD-S) and large (NCD-L) grain sizes were deposited using microwave plasma chemical vapor deposition and characterized using scanning electron microscopy, atomic force microscopy, contact angle testing, and Raman spectroscopy. NCD-S coatings exhibited average grain sizes of 50-80 nm (RMS 5.8 nm), while NCD-L coatings exhibited average grain sizes of 200-280 nm (RMS 23.1 nm). In vitro hemocompatibility testing using human blood included protein adsorption, hemolysis, nonactivated partial thromboplastin time, platelet adhesion, and platelet activation. Both NCD coatings demonstrated low protein adsorption, a nonhemolytic response, and minimal activation of the plasma coagulation cascade. Furthermore, the NCD coatings exhibited low thrombogenicity with minimal platelet adhesion and aggregation, and similar morphological changes to surface-bound platelets (i.e., activation) in comparison to the HDPE negative control material. For all assays, there were no significant differences in the blood-material interactions of NCD-S versus NCD-L. The two tested NCD coatings, regardless of nanotopography, had similar hemocompatibility profiles compared to the negative control material (HDPE) and should be further evaluated for use in blood-contacting medical devices. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 253-264, 2017.