ROLES OF BLOOD FLOW IN PLATELET ADHESION AND AGGREGATION*
Eric F. Grabowski, Ph.D. Department of Laboratory Medicine, Mayo Clinic Rochester, }1N ABSTRACT The investigations described in this review concern platelet adhesion and aggregation as blood flow-dependent phenomena. The experimental approach utilizes a chamber in which are controlled both blood flow and, in aggregation studies, the rate at which ADP enters flowing blood by localized diffusion through a Cuprophan membrane. Videomicroscopy and densitometry of video recordings allow quantification of the in situ growth of surface-adherent platelet aggregates. Platelet adhesion is studied separately using phase contrast microscopy. As a result, with the present method one can distinguish adhesion from aggregation, utilize whole blood, simulate arterial (or extracorporeal) shear rate conditions, independently vary ADP entry rate, and quantify aggregate growth in situ continuously. One finding with this experimental system is that human platelet adhesion to Cuprophan membrane (and to at least two other surfaces) exposed to flowing blood for 10 min to 3 hr is negligible in comparison to dog platelet adhesion to these same surfaces. The species difference is present in both citrated and heparinized blood and at surface shear rates comparable to those in mammalian arteries, i.e., 99 to 986 sec-I. Tests for the thromboresistance of biomaterials and for platelet adhesion under controlled flow conditions must take into better account variations between humans and other species. More generally, initial rates of platelet ad*Supported by NIH Grants HL-17443, HL-17430, and the Minnesota Heart Association. 73
H. J. Day et al. (eds.), Thrombosis © Springer Science+Business Media New York 1978
74
E.F. GRABOWSKI
hesion to Cuprophan for eight species are either "diffusion-limited" (dog, rabbit) or "surface-reaction limited" (human and five other species). In either case there operates a macroscopic transport mechanism for platelets which involves blood flow. With citrated dog blood and ADP in vmolal concentrations at the membrane-blood surface, another finding is that aggregate rate of growth prior to embolization passes through a maximum (between 394 and 635 sec-I) with respect to surface shear rate. This result can be explained by competition between platelet convection and diminution of blood levels of ADP with increased flow and suggests the existence of flow regimens favoring or inhibiting aggregate growth. Furthermore, aggregate growth seems to depend upon degree of platelet adhesion prior to the onset of ADP entry and, consequently, species and the particular synthetic surface in question. Shear flow, but not the presence of erythrocytes, is essential to the aggregation process. For shear rates up to 394 sec- l , the above results suggest a role for platelets in thrombosis which increases as shear rate increases. Such a role is compatible with the greater importance of platelets in arterial thrombosis (shear rates above about 100 sec-I) than in venous thrombosis (shear rates below about 100 sec-I). Arterial thrombi, characteristically more whitish in color, are in large part composed of platelets and fibrin, while the more reddish venous thrombi are principally red blood cells entrapped in a fibrin mesh. As a result, anticoagulants are less effective in prevention of arterial than venous thrombosis; antiplatelet agents are being evaluated in the management of arterial thrombosis, particularly treatment of coronary artery disease and cerebrovascular disease (1,2). INTRODUCTION Platelets, 5 ' -adenosine diphosphate (ADP) released from platelets or other formed elements, and certain clotting proteins have been regarded for some time as important in the pathogenesis of arterial thrombosis and also perhaps in venous thrombosis. Now it is also recognized that these elements undergo convection by flow and diffusion and are therefore subject to the laws of momentum and mass transport. The degree of platelet adhesion to synthetic surfaces exposed to flowing blood (3-6) and subendothelium (7-9) depends upon blood flow rate and exposure time. A layer of singly adherent platelets forms more rapidly at higher rates of flow. The degree or rate of platelet aggregation shows divergent responses. The rate of platelet aggregation on synthetic surfaces
BLOOD FLOW AND PLATELET ADHESION AND AGGREGATION
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exposed to f1ow1ng blood (10), subendothe1ium (8), or endothelium with ADP applied just outside a venule wall by iontophoresis (11) increases with increasing blood flow rate at low rates of flow. An example is Begent and Born's finding (11) that this applies at mean flow velocities of
Eric F. Grabowski, Ph.D. Department of Laboratory Medicine, Mayo Clinic Rochester, }1N ABSTRACT The investigations described in this review concern platelet adhesion and aggregation as blood flow-dependent phenomena. The experimental approach utilizes a chamber in which are controlled both blood flow and, in aggregation studies, the rate at which ADP enters flowing blood by localized diffusion through a Cuprophan membrane. Videomicroscopy and densitometry of video recordings allow quantification of the in situ growth of surface-adherent platelet aggregates. Platelet adhesion is studied separately using phase contrast microscopy. As a result, with the present method one can distinguish adhesion from aggregation, utilize whole blood, simulate arterial (or extracorporeal) shear rate conditions, independently vary ADP entry rate, and quantify aggregate growth in situ continuously. One finding with this experimental system is that human platelet adhesion to Cuprophan membrane (and to at least two other surfaces) exposed to flowing blood for 10 min to 3 hr is negligible in comparison to dog platelet adhesion to these same surfaces. The species difference is present in both citrated and heparinized blood and at surface shear rates comparable to those in mammalian arteries, i.e., 99 to 986 sec-I. Tests for the thromboresistance of biomaterials and for platelet adhesion under controlled flow conditions must take into better account variations between humans and other species. More generally, initial rates of platelet ad*Supported by NIH Grants HL-17443, HL-17430, and the Minnesota Heart Association. 73
H. J. Day et al. (eds.), Thrombosis © Springer Science+Business Media New York 1978
74
E.F. GRABOWSKI
hesion to Cuprophan for eight species are either "diffusion-limited" (dog, rabbit) or "surface-reaction limited" (human and five other species). In either case there operates a macroscopic transport mechanism for platelets which involves blood flow. With citrated dog blood and ADP in vmolal concentrations at the membrane-blood surface, another finding is that aggregate rate of growth prior to embolization passes through a maximum (between 394 and 635 sec-I) with respect to surface shear rate. This result can be explained by competition between platelet convection and diminution of blood levels of ADP with increased flow and suggests the existence of flow regimens favoring or inhibiting aggregate growth. Furthermore, aggregate growth seems to depend upon degree of platelet adhesion prior to the onset of ADP entry and, consequently, species and the particular synthetic surface in question. Shear flow, but not the presence of erythrocytes, is essential to the aggregation process. For shear rates up to 394 sec- l , the above results suggest a role for platelets in thrombosis which increases as shear rate increases. Such a role is compatible with the greater importance of platelets in arterial thrombosis (shear rates above about 100 sec-I) than in venous thrombosis (shear rates below about 100 sec-I). Arterial thrombi, characteristically more whitish in color, are in large part composed of platelets and fibrin, while the more reddish venous thrombi are principally red blood cells entrapped in a fibrin mesh. As a result, anticoagulants are less effective in prevention of arterial than venous thrombosis; antiplatelet agents are being evaluated in the management of arterial thrombosis, particularly treatment of coronary artery disease and cerebrovascular disease (1,2). INTRODUCTION Platelets, 5 ' -adenosine diphosphate (ADP) released from platelets or other formed elements, and certain clotting proteins have been regarded for some time as important in the pathogenesis of arterial thrombosis and also perhaps in venous thrombosis. Now it is also recognized that these elements undergo convection by flow and diffusion and are therefore subject to the laws of momentum and mass transport. The degree of platelet adhesion to synthetic surfaces exposed to flowing blood (3-6) and subendothelium (7-9) depends upon blood flow rate and exposure time. A layer of singly adherent platelets forms more rapidly at higher rates of flow. The degree or rate of platelet aggregation shows divergent responses. The rate of platelet aggregation on synthetic surfaces
BLOOD FLOW AND PLATELET ADHESION AND AGGREGATION
75
exposed to f1ow1ng blood (10), subendothe1ium (8), or endothelium with ADP applied just outside a venule wall by iontophoresis (11) increases with increasing blood flow rate at low rates of flow. An example is Begent and Born's finding (11) that this applies at mean flow velocities of
