<p>The first step in hemostasis and thrombosis is the adherence of platelets to the damaged vessel wall. The factors involved in platelet adherence are not well understood, mainly because of the lack of a suitable method to measure quantitatively platelet adherence to surfaces. The aim of the present studies was to develop a quantitative method to measure platelet adherence to collagen containing surfaces. The following results were obtained with this method: (1) In this quantitative method for measuring platelet adherence to collagen-coated glass surfaces and to subendothelium the platelelets are labeled with ⁵¹Cr and adherence can be measured over a large area without sampling bias. Platelets are suspended at pH 7.35, at 37°C in an artificial medium which can be modified as required. The medium contains physiological concentrations of calcium and magnesium and platelet aggregation is prevented by the inclusion of apyrase to degrade any ADP released. There is no plasma present and thus no thrombin generation or fibrin formation occurs. Adherence is measured using a rotating probe device under controlled flow conditions and various surfaces can be tested (collagen-coated glass, everted aorta damaged to expose the subendothelium). (2) Platelet adherence to collagen-coated glass or to subendothelium is greatly reduced in the absence of divalent cations. Thus, methods which measure platelet adherence in the presence of chelating agents are difficult to interpret. (3) Increasing the albumin concentration of the medium decreases platelet adherence, but there is less variation among replicates. (4) Increasing the hematocrit increases the number of platelets adhering to the surface. (5) Platelets adhere tightly to collagen or to subendothelium and are not dislodged by agents which readily deaggregate platelets such as EDTA, EGTA, or PGE₁. (6) Exposure of undamaged endothelium to thrombin increases the number of platelets adherent to the surface and this is blocked by heparin. (7) This method of measuring platelet adherence to collagen can be used to screen drugs which may be useful as antithrombotic agents. The best system is to test the effect of a drug in the presence of 4% albumin and 40% hematocrit, however, for initial screening a simpler system can be used (0.35% albumin, zero hematocrit). (9) Several treatments which modify the platelet surface have been tested for their effect on adherence to collagen. Removal of sialic acid with neuraminidase do not affect platelet adherence whereas treatment with sodium periodate and the proteolytic enzymes thrombin, plasmin and chymotrypsin decrease adherence. (10) UDP and UDPG do not have an effect on platelet adherence to collagen and this observation does not support the collagen: glucosyltransferase theory of platelet adhesion to collagen. (11) Clq, a subcomponent of the first component of complement specifically inhibits platelet adhesion to collagen and may compete with the collagen receptor on the platelet membrane. (12) A wide variety of agents inhibits platelet adherence to collagen-coated surfaces and to subendothelium. Among the inhibitors which decrease platelet adherence in a system containing 4% albumin and 40% hematocrit are agents which chelate divalent cations (EGTA, EDTA, citrate), indomethacin, agents which increase cAMP levels (prostaglandin E₁, dipyridamole, RA 433), methylprednisolone, penicillin G and cephalothin. (13) Aspirin inhibits platelet adherence to collagen-coated surface or to subendothelium, but its inhibitory effect is not evident in the presence of 40% hematocrit or citrated plasma indicating that the conditions of the experiments are important in determining the effect of drugs. (14) Modifications of platelelets by treatment with thrombin, penicillin G or cephalothin which inhibit platelet adherence to collagen or subendothelium do not affect platelet survival. Modification of the platelet surface sialic acid by neuraminidase or periodate is followed by rapid clearance of platelets from the circulation. Thus there is no correlation between platelet adherence and platelet survival. (15) Treatments which decrease platelet to collagen and to the subendothelium also reduce the platelets in hemostasis.</p> / Doctor of Philosophy (PhD)
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/7932 |
| Date | 05 1900 |
| Creators | Cazenave, Jean-Pierre |
| Contributors | Mustard, J.F., Medical Sciences |
| Source Sets | McMaster University |
| Detected Language | English |
| Type | thesis |
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