阿司匹林与氯吡格雷抗血小板聚集作用效果观察

目的 探讨阿司匹林与氯吡格雷抑制血小板聚集的临床效果。方法 将107例急性脑梗死患者分为阿司匹林组34例、氯吡格雷组19例、联合组54例。阿司匹林组口服阿司匹林100 mg/d;氯吡格雷组口服氯吡格雷75 mg/d;联合组同时服用以上两种药物（剂量同前）。三组均连续服用7 d后采用血栓弹力图检测花生四烯酸（AA）途径血小板抑制率（AA%）及二磷酸腺苷（ADP）途径血小板抑制率（ADP%）。结果 阿司匹林组血小板抑制敏感率明显高于氯吡格雷组,P〈0.01;联合组AA%明显高于阿司匹林组,ADP%明显高于氯吡格雷组,P均〈0.01;联合组血小板抑制敏感率AA为90.7%（49/54）,ADP为70.4%（38/54）,较其余两组明显增高（P均〈0.01）。结论 阿司匹林与氯吡格雷联合应用可协同抑制血小板聚集。     

速效救心丸对大鼠血小板聚集率及环磷腺苷水平的影响

目的探讨速效救心丸对血小板聚集的作用及机制。方法采用比浊法测定速效救心丸、阿司匹林、阿魏酸哌嗪对二磷酸腺苷(ADP)、胶原和花生四烯酸(AA)诱导的大鼠血小板聚集率和环磷腺苷(cAMP)水平的影响。结果与空白组比较,速效救心丸中、高剂量组可显著抑制ADP、胶原、AA诱导大鼠的血小板聚集(P0.01);速效救心丸高剂量组对AA诱导的大鼠血小板聚集抑制作用较阿司匹林组好;速效救心丸可提高大鼠血小板内cAMP的含量(P0.05)。结论速效救心丸能够抑制大鼠ADP、胶原和AA诱导的血小板聚集,其机制可能通过增加cAMP的含量而发挥作用。     

男性素食者血小板MPV及血小板聚集变化的观察

目的比较男性素食者与对照组MPV及血小板聚集能力有无差异。方法检测37例男性素食者和26例对照组的MPV及血小板聚集率,包括ADP0.5μmol/L、ADP2.0μmol/L和Adr 5.56μmol/L诱导的最大聚集率,比较两组有无差异以及分析MPV与血小板聚集率的相关性。结果素食者MPV明显高于对照组,ADP 0.5μmol/L、ADP2.0μmol/L、Adr 5.56μmol/L诱导的血小板最大聚集率均高于对照组,差异均有统计学意义(P均0.05)。相关分析结果显示:MPV与ADP0.5μmol/L所测得最大聚集率呈正相关(r=0.264,P0.05)。结论男性素食者MPV增高,且血小板聚集能力也增加,结果显示饮食方式对血小板活化有重要影响。     

三氧化二砷抑制血小板聚集功能的研究

目的:分析三氧化二砷(ATO)对血小板聚集功能的影响。方法:用不同浓度的ATO(0、0.25、0.5、1、1.5、2、2.5、5和10μmol/L)孵育富血小板血浆(PRP)0、3、15、30、45和60 min,在2 mg/L胶原或2μmol/L腺苷二磷酸(ADP)刺激下,检测血小板聚集功能。结果:在刺激剂为2 mg/L胶原时,ATO(≥5μmol/L)孵育PRP 60 min,可显著抑制血小板聚集,在相应的时间梯度实验中,ATO浓度从5μmol/L上升到10μmol/L,抑制血小板聚集所需要的孵育时间从45 min下降到30 min,ATO对胶原诱导的血小板聚集具有浓度(r=-0.902,P=0.001)和时间(r=-0.964,P=0.002;r=-0.910,P=0.032)依赖性;在刺激剂为2μmol/L ADP时,ATO(≥2μmol/L)孵育PRP 60 min,可显著抑制血小板聚集,在相应的时间梯度实验中,ATO浓度从5μmol/L上升到10μmol/L,抑制血小板聚集所需要的孵育时间从45 min下降到15 min,ATO对ADP诱导的血小板聚集亦具有浓度(r=-0.815,P=0.007)和时间(r=-0.921,P=0.009;r=-0.963,P=0.009)依赖性。结论:低浓度的ATO对血小板聚集功能没有明显的作用,但是高浓度ATO(≥2μmol/L)可抑制血小板的聚集功能,并且该抑制作用具有浓度和时间依赖性。     

三氧化二砷抑制血小板聚集功能的研究

目的:分析三氧化二砷(ATO)对血小板聚集功能的影响。方法:用不同浓度的ATO(0、0.25、0.5、1、1.5、2、2.5、5和10μmol/L)孵育富血小板血浆(PRP)0、3、15、30、45和60 min,在2 mg/L胶原或2μmol/L腺苷二磷酸(ADP)刺激下,检测血小板聚集功能。结果:在刺激剂为2 mg/L胶原时,ATO(≥5μmol/L)孵育PRP 60 min,可显著抑制血小板聚集,在相应的时间梯度实验中,ATO浓度从5μmol/L上升到10μmol/L,抑制血小板聚集所需要的孵育时间从45 min下降到30 min,ATO对胶原诱导的血小板聚集具有浓度(r=-0.902,P=0.001)和时间(r=-0.964,P=0.002;r=-0.910,P=0.032)依赖性;在刺激剂为2μmol/L ADP时,ATO(≥2μmol/L)孵育PRP 60 min,可显著抑制血小板聚集,在相应的时间梯度实验中,ATO浓度从5μmol/L上升到10μmol/L,抑制血小板聚集所需要的孵育时间从45 min下降到15 min,ATO对ADP诱导的血小板聚集亦具有浓度(r=-0.815,P=0.007)和时间(r=-0.921,P=0.009;r=-0.963,P=0.009)依赖性。结论:低浓度的ATO对血小板聚集功能没有明显的作用,但是高浓度ATO(≥2μmol/L)可抑制血小板的聚集功能,并且该抑制作用具有浓度和时间依赖性。     

不同负荷剂量氯吡格雷对血小板聚集率的影响

目的比较两种剂量氯吡格雷的起效时间及安全性,为急性冠状动脉（冠脉）综合征患者用药方案提供依据。方法60例急性冠脉综合征使用不同负荷剂量氯吡格雷患者随机分为A组（300mg）和B组（600mg）,均予氯吡格雷75mg／d后续治疗。以腺苷二磷酸（ADP）5μmol／L及20μmol／L作为诱导剂检测服药前及服药后2h和6h的血小板聚集率,并检测服药前及服药后第3天的血自细胞及血小板计数。结果在ADP20μmoL／L诱导的血小板聚集检测中,两组均显示服药后6h比服药后2h达到更高的血小板聚集抑制水平[A组（29．75±12．11）％比（43．63±14．31）％,P〈0．05;B组（28．86±10．24）％比（34．86±10．84）％,P〈0．05]。B组与A组相比,在服药后2h即起到更加明显的血小板聚集抑制作用[（34．86±10．84）％比（43．63±14．31）％,P〈0．05]。服药后3d内所有人选患者均无出血、自细胞减少及血小板减少等事件发生。结论氯吡格雷600mg作为负荷剂量较之300mg可以更快地达到较高水平的血小板抑制作用,且两者安全性相似。     

松胞菌素D对血小板聚集和血小板—中性粒细胞间相互作用的影响

目的探讨松胞菌素D对血小板聚集及血小板—中性粒细胞间相互作用的影响。方法应用玫瑰花结试验和Born方法观察松胞菌素D对血小板—中性粒细胞之间相互作用及体内外对血小板聚集功能的影响。结果松胞菌素D在体内外对花生四烯酸（AA）、血小板活化因子（PAF）诱导的血小板聚集无明显抑制作用,而对腺苷二磷酸（ADP）诱导的血小板聚集则有明显的抑制作用;松胞菌素D可以阻抑血小板与中性粒细胞间的黏附作用;松胞菌素D明显抑制PAF（1μmol/L）激活的中性粒细胞悬液引起的洗涤血小板聚集。结论松胞菌素D在体内外均具有选择性抑制ADP诱导的血小板聚集作用,且明显阻抑血小板与中性粒细胞间的相互作用。     

血红蛋白在三氧化二砷抑制血小板聚集中的功能研究

目的:检测血红蛋白与砷的结合,分析血红蛋白在三氧化二砷(ATO)抑制血小板聚集功能中发挥的作用。方法:电喷雾质谱(ESI-MS)检测血红蛋白与氨基氧化苯胂酸(PAPAO)的结合。在2 mg/L胶原或2μmol/L腺苷二磷酸(ADP)刺激下,比较经10μmol/L ATO处理后,富血小板血浆(PRP)和全血血小板聚集的差异。PRP中加入0、40、60、80、100、120 g/L的血红蛋白经10μmol/L的ATO处理后,检测血小板聚集。结果:血红蛋白β链可与一分子有机砷PAPAO结合,并脱去一分子水。ATO处理PRP以及全血,两者血小板聚集均受到显著抑制,但是对PRP血小板聚集的抑制更加明显。在刺激剂为2 mg/L胶原时,血红蛋白≥80 g/L可逆转ATO对血小板聚集的抑制,并且具有浓度依赖性(r=0.956,P=0.003);同样的,在刺激剂为2μmol/L ADP时,血红蛋白≥80 g/L可逆转ATO对血小板聚集的抑制(r=0.940,P=0.005)。结论:在体外血红蛋白通过与砷结合,能够削弱ATO对血小板聚集的抑制,并且具有浓度依赖性。血红蛋白在ATO抑制血小板聚集过程中发挥重要作用。     

不同剂量氯吡格雷抑制血小板聚集功能时效探讨

目的 探讨氯吡格雷常用剂量下，血小板聚集率的变化规律及其临床意义。方法 40例经体检健康的志愿者，随机分为4组，每组10例，第一组一次服用75mg氯吡格雷；第二组服用氯吡格雷75mg／d，连服3天停药；第三组服用氯吡格雷75mg／d连服7天停药；第四组一次服用300mg氯吡格雷。各组分别在不同时间点进行采样，样本用二磷酸腺苷（ADP）进行血小板聚集实验。结果 第一组在停药后6天血小板聚集率恢复到服药前水平。第二组血小板聚集率在服药的3天、停药后的1天、2天与用药前差异有非常显著性（P〈0．01），第10天后恢复到服药前水平。第三组停药后4天内血小板聚集率受非常明显抑制（P〈0．01），6天内受明显抑制（P＜0．05）。第四组服药后2h即可观察到药物对血小板聚集率的非常明显的抑制作用（P〈0．01），停药4天后血小板聚集率恢复到服药前水平。结论 首服剂量越大，维持服药的时间越长，血小板聚集功能恢复所需时间越长。     

野木瓜多糖对家兔血小板聚集的影响

目的研究野木瓜多糖(SCP)对血小板聚集的影响及可能机制。方法选择成年家兔30只,随机分为空白对照组、SCP 17 g/L组、SCP 34 g/L组、SCP 68 g/L组、阿司匹林组,每组6只。制备家兔富血小板及贫血小板血浆,用Born's比浊法检测17、34、68 g/LSCP对胶原、花生四烯酸(AA)、二磷酸腺苷(ADP)及凝血酶诱导的血小板聚集的影响。检测SCP对胶原诱导后血小板中5-羟色胺含量、肝素凝血酶凝固时间(HTCT)及对凝血酶作用后血小板释放丙二醛含量的影响。结果与空白对照组比较,不同浓度SCP组、阿司匹林组的胶原、ADP、凝血酶诱导的血小板聚集明显降低(P<0.05,P<0.01),SCP 1 7 g/L组AA诱导的血小板聚集无明显变化(P>0.05),SCP34 g/L组、SCP 68 g/L组、阿司匹林组AA诱导的血小板聚集明显降低(P<0.05,P<0.01)。与空白对照组比较,不同浓度SCP组丙二醛含量明显降低、HTCT延长,呈浓度依赖性(P<0.01),除SCP 1 7 g/L组外,其余各组5羟色胺明显降低,呈浓度依赖性(P<0.05,P<0.01)。结论 SCP可以抑制由胶原、ADP、AA和凝血酶诱导的血小板聚集,其机制与抑制血小板释放5-羟色胺、血小板因子Ⅳ及减少丙二醛生成有关。     

Evaluation of an automated light transmission aggregometry

Light transmission aggregometry (LTA) is the “gold standard” for platelet function assessment, but it is time-consuming and labor intensive. Recently, an automated platelet aggregation method has been developed on a routine coagulation analyzer (Sysmex CS-2100i). In this study, the performances of CS-2100i including repeatability, correlation with a reference aggregometer (Chrono-log Model 700), and the threshold limitation of platelet counts in platelet-rich plasma (PRP) were evaluated for clinical use. The agonists were adenosine diphosphate (ADP), arachidonic acid, collagen, epinephrine, and ristocetin. The platelet concentration of PRP was adjusted with platelet-poor plasma (PPP) and physiological saline (PS). The CS-2100i showed an excellent repeatability and a strong correlation with the Chrono-log 700 in performing platelet aggregation, and its threshold limitation of platelet counts in PRP was 80 × 10⁹/L. PPP had an inhibitory impact on platelet aggregation induced by ADP, arachidonic acid, collagen or epinephrine; while PS had an inhibitory impact on ristocetin-induced aggregation. PS should be used to adjust PRP for ADP-, arachidonic acid-, collagen-, or epinephrine-induced aggregation; while PPP was recommended for ristocetin-induced aggregation. The CS-2100i showed an excellent repeatability, a strong correlation with Chrono-log 700, a lower platelet count requirement, a shorter turnaround time for samples, the advantage of being a walk-away technology, and the ability to perform a highly standardized platelet function assessment.     

Platelet aggregation impacts thrombin generation assessed by calibrated automated thrombography

A calibrated automated thrombogram (CAT) is performed usually with human platelet-free plasma (PFP) but may be more relevant with platelet-rich plasma (PRP). In this case, platelets are not stimulated by subendothelial molecules like collagen. Our aim was to assess the consequence of strong (collagen) or weak (ADP) induction of platelet release and aggregation on thrombin generation. Platelet aggregation in PRP was triggered with 10 µg/mL collagen or 10 µM ADP using a lumi-aggregometer. Thrombin generation curves were monitored by CAT in different conditions: PRP, PRP with activated platelets (actPRP), aggregated PRP (agPRP), aggregated platelets resuspended in autologous PFP (resPRP), PFP and PFP obtained after aggregation (agPFP). We found a 3-fold shortening of the lag time and time to peak and a marked increase in velocity and thrombin peak without changes in endogenous thrombin potential (ETP) in agPRP with both agonists compared with PRP. The same holds true in agPFP but with a marked increase in ETP compared with PFP. Similar changes in the kinetics of thrombin generation were observed with actPRP-collagen and to a lesser extent in resPRP-collagen compared with PRP. By contrast, there were no modifications of the thrombin generation curves in actPRP-ADP. Alpha-2-macroglobin-thrombin complexes were unchanged in the different PRP conditions but were increased in PFP prepared from agPFP compared to control PFP. Platelet aggregation during activation by agonists other than thrombin did not increase thrombin generation but accelerated its kinetics mainly via platelet content release and platelet-derived extracellular vesicules formation. In diseases characterized by altered platelet granule content or release as well as altered platelet activation, a platelet aggregation step prior to CAT analysis may be clinically relevant to improve laboratory estimation of the bleeding/thrombotic balance.     

Effects of Cephalothin and Penicillin G on Platelet Function in Vitro

High concentrations of cephalothin or penicillin G inhibit a number of the functions of human or rabbit platelets in citrated platelet-rich plasma (PRP) and in suspensions of washed platelets. The reactions shown to be inhibited are: ADP-induced shape change and the primary and secondary phases of aggregation and release induced by ADP or adrenaline in human citrated PRP; release and aggregation of washed human platelets exposed to collagen, thrombin, vasopressin, or the ionophore A23,187; aggregation of washed human platelets exposed to phytohaemagglutinin from Phaseolus vulgaris (PHA) or polylysine; release induced by concanavalin A or PHA in suspensions of washed platelets from rabbits; platelet adherence to a collagen-coated surface or to the damaged intimal surface of the rabbit aorta; platelet factor 3 availability; lysis of rabbit platelets by an antiserum directed against them; and clot retraction. Neither antibiotic affected serotonin-induced aggregation; a high concentration of cephalothin slightly inhibited the initial rate of serotonin uptake. Penicilloic acid showed about half the inhibitory effect of penicillin G on ADP-induced aggregation. In citrated human platelet-rich plasma, ampicillin and oxacillin inhibited ADP-induced aggregation to the same extent as similar concentrations of penicillin G; in suspensions of washed platelets, however, ampicillin was less inhibitory than penicillin G or oxacillin. Platelet ultrastructure, assessed by transmission electron microscopy, was not visibly altered. Evidence that the antibiotics become bound to platelets is the finding that platelets incubated with the antibiotics and resuspended in fresh media showed less response to aggregating agents compared with control platelets. Penicillin G and related antibiotics may be inhibitory because they coat the platelet surface. Their effects on platelet functions are probably responsible for excessive bleeding and increased bleeding times observed in patients and volunteers receiving high doses of these antibiotics.     

Platelet aggregation occurs in congenital afibrinogenaemia despite the absence of fibrinogen or its fragments in plasma and platelets, as demonstrated by immunoenzymology

Platelet aggregation of an afibrinogenaemic patient's platelet rich plasma (PRP) was greatly decreased when ADP was used for stimulation. In the presence of collagen or arachidonic acid the changes in light transmission recorded during platelet aggregation of patient's PRP were similar to those observed with normal PRP but the size of aggregates appeared to be smaller in comparison with those observed with normal platelets. In addition, thrombin-induced aggregation of washed platelets was similar to normal platelets. The interpretation was made possible because the fibrinogen level in plasma and in platelets was found to be almost nil as demonstrated by both an Elisa procedure described here and the determination of fibrinopeptide A (fpA). Furthermore, fibrinogen fragments, which could result from abnormal synthesis and therefore replace fibrinogen in platelet aggregation, were undetectable by immunological analysis using specific antibodies against A alpha, B beta and gamma chains and 10 different monoclonal antibodies against fibrin degradation products.     

Platelet inhibiton by sodium nitroprusside, a smooth muscle inhibitor.

The effects of sodium nitroprusside (N.P.), a pure smooth muscle inhibitor, on platelet function were studied. Platelet-rich plasmas (PRP) from normal controls and from patients receiving N.P. were studied in vitro for aggregation in response to adenosine diphosphate (ADP), epinephrine, and collagen. Platelet ADP release (release reaction) was also investigated. Normal platelets demonstrated marked inhibition of aggregation when incubated with N.P. for 3 min. Prolonging the incubation was without additional effect. ADP and ATP release from platelets in response to collagen was also inhibited. PRP from patients receiving nitroprusside at concentrations between 25 mug/min an 165 mug/min showed inhibition of aggregation when compared to findings prior to the administration of N.P. N.P. acts by inhibiting contractile proteins and thus platelet ADP release and aggregation may depend on contraction of platelet smooth muscle-like protein, thrombosthenin.     

Inhibition of platelet aggregation by idebenone and the mechanism of the inhibition

The inhibitory effect of 6-(10-hydroxydecyl)-2,3-dimethoxy-5-methyl-1,4-benzoquinone (idebenone) on platelet aggregation was studied in rat and human platelets in vitro, and the mechanism of inhibition was examined in rat platelets. Idebenone inhibited the aggregation induced by collagen and thrombin in washed platelets, and by arachidonate and ADP in platelet-rich plasma (PRP). The inhibition was more prominent in collagen- and arachidonate-induced aggregation. In collagen-induced aggregation of human platelets, idebenone was 8-fold more potent than aspirin. In addition, idebenone inhibited prostaglandin synthesis and thromboxane B2 production, and also increased the cyclic AMP content in platelets. However, the concentration of idebenone required to inhibit thromboxane B2 production was much lower than that required to increase cyclic AMP. These results indicate that idebenone inhibits platelet aggregation by inhibiting thromboxane B2 synthesis rather than by increasing cyclic AMP content.     

Platelet factor XIIIa release during platelet aggregation and plasma clot strength measured by thrombelastography in patients with coronary artery disease treated with clopidogrel

Abstract

It has been estimated that up to half of circulating factor XIIIa (FXIIIa) is stored in platelets. The release of FXIIIa from platelets upon stimulation with adenosine diphosphate (ADP) in patients with coronary artery disease treated with dual antiplatelet therapy has not been previously examined. Samples from 96 patients with established coronary artery disease treated with aspirin and clopidogrel were examined. Platelet aggregation was performed by light transmittance aggregometry in platelet-rich plasma (PRP), with platelet-poor plasma (PPP) as reference, and ADP 5?µM as agonist. Kaolin-activated thrombelastography (TEG) was performed in citrate PPP. PRP after aggregation was centrifuged and plasma supernatant (PSN) collected. FXIIIa was measured in PPP and PSN. Platelet aggregation after stimulation with ADP 5?µM resulted in 24% additional FXIIIa release in PSN as compared to PPP (99.3?±?27 vs. 80.3?±?24%, p?<?0.0001). FXIIIa concentration in PSN correlated with maximal plasma clot strength (TEG-G) (r?=?0.48, p?<?0.0001), but not in PPP (r?=?0.15, p?=?0.14). Increasing quartiles of platelet-derived FXIIIa were associated with incrementally higher TEG-G (p?=?0.012). FXIIIa release was similar between clopidogrel responders and non-responders (p?=?0.18). In summary, platelets treated with aspirin and clopidogrel release a significant amount of FXIIIa upon aggregation by ADP. Platelet-derived FXIIIa may contribute to differences in plasma TEG-G, and thus, in part, provide a mechanistic explanation for high clot strength observed as a consequence of platelet activation. Variability in clopidogrel response does not significantly influence FXIIIa release from platelets.     

Reduced aggregability of platelets in rat anaphylaxis.

In the PRP of anaphylactic rats, ADP, collagen and thrombin induced platelet aggregation was considerably reduced. Reduced aggregability could be transferred to normal platelets by suspending them in the PPP of anaphylactic animals and the impaired aggregation of platelets from animals undergoing anaphylaxis could be restored by exchanging their plasma for that of normal controls. Ellagic acid, a known activator of factor XII, produced similar alterations as obtained in anaphylactic shcok. It is suggested that the inhibition of platelet aggregation is due to the anaphylactic activation of factor XII and this mechanism may be of importance in rat anaphylaxis.     

Effect of heparin on platelet aggregation

The effect of heparin on platelet aggregation was systematically examined on platelets in plasma (PRP), as well as on gel-filtered, washed, and formaldehyde-fixed platelets. Results indicate that, although heparin causes a mild potentiation of platelet aggregation in the PRP systems, a significant inhibitory activity is observed when heparin is added to isolated platelets. This inhibitory activity appears to be specific and not related to the impurities in the heparin preparations, as heparinase, as well as protamine, effectively neutralizes the heparin-mediated inhibitory activity on platelet aggregation. Although heparin-mediated inhibitory activity can be demonstrated in the presence of a number of different agonists (ADP, arachidonic acid, thrombin, Ionophore A23187, epinephrine, and ristocetin), the most pronounced inhibition is seen in the presence of ristocetin. Further studies show that heparin enhances thromboxane generation in isolated platelets. Platelets pretreated with heparin, however, fail to respond to preformed thromboxane. These findings suggest that, in addition to the potentiation of thromboxane production in platelets, heparin may also attribute some change(s) to the platelet(s)/platelet membrane, which interferes with their ability to respond to the agonists of platelet aggregation. This antiaggregatory activity of heparin was found to be inhibited by a factor(s) present in plasma but not in serum.     

In vitro platelet aggregation defects in patients with myeloproliferative disorders and high platelet counts: Are they laboratory artefacts?

It has been recently shown that in vitro platelet aggregation is inhibited when platelet concentration in platelet-rich plasma (PRP) is “normalized” by the addition of platelet-poor plasma (PPP). In this study we tested the hypothesis that the large amount of PPP required to “normalize” PRP in patients with thrombocytosis may result in falsely defective platelet function. To this end, we evaluated platelet aggregation in PRP samples “normalized” with either PPP or buffer in 16 patients with high platelet counts induced by myeloproliferative disorders. Comparison with the results obtained in healthy subjects demonstrated that patients had reduced platelet responses to ADP or collagen in PRP/PPP samples, but normal responses in PRP/buffer. By contrast, the majority of patients had severely defective platelet response to epinephrine independently from the methodological approach. We suggest that the reduced in vitro platelet aggregation previously described in patients with myeloproliferative disorders and thrombocytosis partially derived from a laboratory artefact.