血小板功能检测的研究进展

血小板在执行生理性止血的同时,也在病理性血栓形成过程中起重要作用。血小板功能检测对于临床相关疾病的诊断和抗血小板药物的筛选及相关研究有着重要的意义。目前,血小板功能检测的实验与方法日益增多,但所有这些检测方法均有不足之处,因而有必要研究和发展一种操作简便、检测灵敏度高的方法。本文对近年来血小板功能检测方法诸如血小板的一般功能检测、血小板黏附功能测定、血小板聚集功能测定、血小板释放功能测定、血小板凝血活性检测和流式细胞术在血小板功能检测中的应用等研究进展进行了综述,并对研究前景作了简单的展望。     

Platelets do not adsorb HLA class I molecules during storage of pooled platelet concentrates

The origin of HLA class I molecules on platelets is still under discussion. Adsorption of HLA molecules on platelets using specific experimental conditions has been described. The study presented investigates whether there is a significant elution and adsorption of HLA class I molecules on platelets during storage of pooled random platelet concentrates (PRPC) under routine conditions. Platelet concentrates (PCs) from whole blood were prepared from HLA-A2-positive and HLA-A2-negative donors, pooled and stored under routine conditions. In addition, platelets from HLA-A2-negative donors were pelleted and resuspended in cell-free plasma from HLA-A2-positive donors. HLA-A2-positive PCs (positive control), HLA-A2-negative PCs (negative control) and HLA-A2-negative platelets in plasma from HLA-A2-negative donors were stored simultaneously. Binding of FITC-conjugated monoclonal murine antihuman HLA-A2 antibodies (anti-HLA-A2-mab) was measured during 5-day storage by flow cytometry. An increased binding of anti-HLA-A2-mab during storage was found on HLA-A2-negative platelets (P < 0.005) independently whether they were incubated with cell-free plasma or platelets from HLA-A2-positive donors or autologous HLA-A2-negative cell-free plasma. However, non-specific binding of IgG controls increased equally, whereas anti-HLA-A2-mab binding to platelets from HLA-A2-positive donors did not decrease during storage. This study suggests that there is no significant elution and adsorption of HLA class I antigens of platelets in pooled PCs during storage under the usual conditions for platelet storage. Increased anti-HLA-A2-mab signal was due to non-specific binding. Therefore, HLA class I compatible platelets should maintain their compatibility for an immunized patient when stored in a pool with HLA incompatible platelets and shortened survival after transfusion should not be expected.     

PAS or plasma for storage of platelets? A concise review

Platelet additive solutions (PASs) are becoming increasingly popular for storage of platelets, and PAS is steadily replacing plasma as the storage medium of platelets. PASs are electrolyte solutions intended for storage of platelets, and they are used to modulate the quality of the platelets by adding specific ingredients. All currently available PASs contain acetate. Acetate reduces the amount of glucose that is oxidised into lactic acid and thereby prevents the lowering of pH, which decreases platelet quality. Furthermore, the oxidation of acetate leads to the production of bicarbonate, which serves as buffer. The presence of potassium and magnesium in PAS prevents the lowering of pH and reduces the degree of spontaneous activation of the platelets during storage. In the hospital, platelets stored in PAS result in about half of the number of allergic transfusion reactions as compared with platelets in plasma. Recovery and survival after transfusion, as well as corrected count increments, are at least as good for platelets in PAS as for plasma, and recent data suggest they may even be better. Therefore, with the current generation of PASs, PAS should be preferred over the use of plasma for the storage of platelet concentrates.     

肿瘤患者血小板输注无效的临床分析

目的探讨本院患者血小板输注情况及临床疗效观察,了解血小板输注无效的原因,以便做好预防工作。方法 2010年1月-2012年12月对本院312名血小板减少的肿瘤患者输注机采血小板,并于输注前后检测血小板,以校正患者血小板计数增值(CCI)和24 h血小板回升率,判定输注效果并讨论影响因素。采用简易致敏红细胞血小板血清学技术(SEPSA)检测76名血小板输注无效的肿瘤患者血小板相关抗体,统计抗体分布。结果 75.6%(236/312)的患者输注后血小板数有不同程度的增高,血液病患者输注血小板有效率明显低于实体肿瘤患者(χ2=29.87,P0.05)和其他肿瘤患者(χ2=6.06,P0.05)。输注次数2次的患者(无效率为5.1%)输注血小板无效率明显低于输注次数7次患者(无效率为57.1%),也低于输注3-4次(无效率为39.5%)、5-7次患者(无效率为46.4%),血小板输注无效率随输注次数的增加而增加。分析76名肿瘤患者血小板输注无效原因,其中免疫因素占60.5%,非免疫因素占39.5%。结论反复输注产生血小板抗体是PTR发生的主要原因,多次输注者应进行血小板抗体的筛选,避免或减少造成血小板输注无效的原因,提高血小板输注的有效率。     

血小板抗原基因分型与配型在血小板输注中的应用简

本研究建立已知HPA基因型血小板供者库,开展相同HPA基因型血小板交叉配型后输注,防止血小板抗体的产生,解决血小板无效输注.应用PCR-SSP技术对血小板献血者和患者进行HPA基因分型,采集与患者ABO血型及HPA基因型相同血小板制剂,再加以固相凝集法配型相合后输注.结果表明,牡丹江地区HPA血型分布的遗传特征具有本地区人群特点,具有最高杂合度的是HPA-15,其次是HPA-3.采用ABO血型及HPA基因型均相同加固相凝集配型相合血小板输注有效率94.4％.采用ABO血型相同随机血小板固相凝集配型相合输注有效率77.8％.结论：牡丹江地区HPA血型分布的遗传特征具有多态性,采用血小板HPA同基因型供者采集加配型输注可防止血小板免疫性抗体及输注无效的发生,采用ABO同型随机血小板加配型输注是提高血小板输注的效果、防止输注无效发生的既经济又简便的方法.     

The post-platelet 100,000 count: effects of platelet collection and future

Currently, the majority of platelets transfused in the United States are collected by apheresis. The recent Food and Drug Administration guidance document published maintains that a postdonation platelet count for a donor remain >100,000/μL. During apheresis procedures, platelets are released from the splenic pool into circulation. This allows for higher postdonation platelet counts than anticipated. Some current plateletpheresis instruments take this into account when determining a safe product to be collected. On other instruments, the software does not allow for this correction. This may impact collections, but is expected to be resolved with a software update.     

Transfusing platelets 2 h after the completion of amphotericin-B decreases its detrimental effect on transfused platelet recovery and survival

Platelet transfusion support is required during bone marrow aplasia following ablative chemotherapy and bone marrow progenitor cell transplantation (BMT). Amphotericin-B is frequently given to these patients, both therapeutically and prophylactically, and has been described to have a negative impact on the results of platelet transfusions. We conducted a prospective study of the effect of amphotericin-B on transfused platelet recovery and survival in 81 BMT or acute leukaemia patients. One hundred and ninety-five platelet transfusions administered to 81 consecutive patients were analysed. The platelets were transfused 2 h after the completion of amphotericin-B. Using this schedule resulted in no effect of amphotericin-B on platelet recovery or survival, although platelet increments were modestly depressed in patients receiving high- vs. low-dose amphotericin-B. We conclude that the timing of amphotericin-B infusion be evaluated in patients demonstrating poor platelet recovery and survival. Transfusing platelets at least 2 h after the completion of amphotericin-B decreases the detrimental effect of this antifungal agent on transfused platelet recovery and survival.     

2项联合检测作为血小板活化特异标志物的评价

目的探讨平均血小板平均体积(MPV)和血小板分布宽度(PDW)作为血小板活化特异标志物的可行性研究。方法受试者分3组,其中健康对照组100例,心肌梗死组63例,脑梗死组54例,采用Sysmex XE-2100型全自动血细胞分析仪对所有受试者静脉血MPV、PDW进行测定。同时随机抽取广州医学院第二附属医院住院患者100例,在采血后1、2、3和4h分别检测静脉血MPV和PDW。结果与健康对照组比较,心肌梗死组、脑梗死组的MPV、PDW均升高,差异有统计学意义(P<0.01)。采血后进行不同时间检测发现,随着时间的延长,MPV增加,PDW降低。结论在血栓性疾病中MPV、PDW呈增大趋势;MPV联合PDW检测可作为血栓性疾病病情监测与预后判断的重要指标。     

临床727例次单采血小板输注效果分析

为分析临床单采血小板输注效果及其影响因素,回顾性分析湘雅第三医院2010年9月至2011年5月进行单采血小板输注的254名患者的727例次输注资料,计算血小板计数增高指数(CCI)和血小板回收率(PPR)评价血小板输注效果,统计输注有效率,根据患者病症、输血次数、血型和脾肿大与否分组进行统计学比较。结果表明,727例次输注单采血小板有效456例次,占62.72%,以急性失血性贫血和慢性系统性疾病患者输注效果较为明显,尤其是慢性肾疾病(有效率为94.12%);脾脏肿大影响血小板输注效果(有效率仅为40.35%);血小板输注次数与输注效果呈负相关。结论:脾脏肿大和血小板输注次数是影响输注效果的因素。     

Changes in platelet volume,morphology and RNA content in subjects treated with haemodialysis

During haemodialysis treatment, blood flows from the body to the extracorporeal circuit and vice versa. In this study, pathophysiological defects in platelets indicated by alterations in RNA content and aberrations in platelet volume and morphology are detected before and during haemodialysis treatment. In subjects receiving haemodialysis treatment, qualitative interpretation of platelet characteristics with application of light microscopic evaluation reveals only 19±11?% of platelets with appropriate staining density of the granule‐containing cytoplasm. On the contrary, a reference group of apparently healthy subjects shows 70±12?% platelets with appropriate staining density of the granule‐containing cytoplasm. During haemodialysis treatment, mean values for platelet volume, platelet distribution width and platelet large cell ratio demonstrate a tendency to decrease by 10?%, 11?% and 6?%, respectively, from the mean initial value to the value at t = 150?min. Reduction of the platelet volume parameters just mentioned is hypothesized to be due to platelet degranulation as a result of platelet activation.     

Are platelet volume indices of clinical use? A multidisciplinary review

Abstract

Platelet size correlates with platelet activity and can be assessed by platelet volume indices (PVI). The PVI, mean platelet volume (MPV), is universally available with routine blood counts by automated hemograms and therefore is an attractive index to study in clinical scenarios. PVI are useful in assessing the etiology of thrombocytopenia. In addition, a normal platelet distribution width in the setting of thrombocytosis is highly suggestive of a reactive etiology. Higher MPV is also associated with the presence of cardiovascular risk factors, chest pain due to acute coronary syndrome, and adverse outcome after acute coronary syndrome. Results from studies evaluating MPV in patients with peripheral artery disease, unprovoked deep vein thrombosis, and pulmonary embolism further advocate a potential role for MPV in identifying patients at high risk of thrombosis.

Nevertheless, most of these data come from retrospective studies some of which have small study populations and confounding factors influencing platelet volume. Moreover, the cut-off values derived from these retrospective studies have not been validated prospectively. Despite the potential for clinical utility evident from these studies, the above-mentioned flaws together with technical problems in measuring MPV currently limit its clinical usefulness. Our review provides a perspective on PVI's potential clinical use.     

人冻干血小板复水程序的优化

目的探讨不同复水条件对冻干血小板功能的影响。方法经过添加激活抑制剂(PGE1、左旋精氨酸、植酸钠和百维利肽)、DMSO和海藻糖等低温保护剂、冷冻干燥后获得冻干血小板,在一系列不同复水程序下对冻干血小板复水,应用流式仪检测并分析其CD62p和PAC-1的表达,评价血小板活化状态。通过正交设计,研究不同复水程序包括复水溶液、复水温度、复水方式3个影响因素对冻干血小板复水后活性的影响。结果 3个对冻干血小板复水后活化的影响因素排序:复水温度>复水方式>复水液(P<0.05);37℃条件下冻干血小板复水后血小板膜蛋白CD62p和PAC-1活化率最低,分别为(5.96±2.51)%和(4.55±1.97)%;预水化后再行四步添加复水可降低冻干血小板膜蛋白CD62p和PAC-1的表达率至(6.68±2.62)%和(5.45±2.06)%;添加聚乙烯吡咯烷酮(PVP)和血小板抑制剂的2号复水液对冻干血小板复水后膜蛋白CD62p和PAC-1的表达率影响最低,分别为(6.72±3.35)%和(5.28±2.30)%。总体模型评价进一步显示,第5实验组,即在37℃条件下加入PVP和血小板抑制剂的复水液配方,先预水化后再行四步法添加复水,其复水血小板活化率最低,CD62p和PAC-1的表达率分别为(3.06±1.36)%和(2.70±0.84)%。结论 37℃下,饱和蒸汽环境中预水化后,采用加入PVP及血小板抑制剂的复水液四步添加复水,其复水血小板活化率最低。     

Platelets and atherosclerosis

Abstract. Despite the broad array of mechanisms designed to protect the endothelial lining of every blood vessel in the body and maintain the fluid state of blood, injury does occur. Chronic and recurrent damage result in development of lesions characteristic of atherosclerosis. The loss of vascular integrity associated with the pathological process of atherogenesis triggers the haemostatic mechanism. As a result, fibrin and platelets become a part of atherosclerotic lesions and play a role in their progression. Growth of the plaques and destruction of normal endothelium triggers further involvement of platelets leading to occlusion of arteries in the heart and brain, resulting in myocardial infarction and stroke. Understanding the role of platelets in atherosclerosis and limiting its contribution may reduce morbidity and mortality of this dread disease.     

血小板抗体检测及配型对血小板输注无效的临床意义

目的 通过对比血小板配型前后血小板的输注效果,评估血小板抗体检测及配型对血小板输注无效的临床意义.方法 以出血症状改善情况、血小板计数增高指数(CCI)、血小板恢复百分率(PPR)为标准,对比配型前后血小板的输注效果.结果 25例血小板输注无效患者的血小板抗体筛查阳性9例; 9例血小板抗体阳性患者血小板交叉配型前后血小板输注有效率差异有统计学意义(P<0.01),配型后输注的 1 h和24 h CCI、PPR数值明显高于配型前输注的.结论 血小板抗体检测及血小板配型输注可以为患者选择适用的血小板,提高单采血小板的输注有效率,避免滥用血小板.     

比浊法血小板聚集试验的影响因素研究

目的 在不同的实验条件下,观察影响比浊法血小板聚集试验的因素.方法 选择30例健康对照组和204例病例组,观察不同的血小板计数及诱导剂浓度对糖尿病患者餐前与餐后、采血后的检测时间、诱导剂的种类互补等不同试验条件下的血小板聚集率.结果 随着血小板计数的减少或增加,血小板聚集率相应的减少或增加;二磷酸腺苷(ADP)、胶原(COL)、花生四烯酸(AA)浓度增加,血小板最大聚集率增大,相应的阿司匹林抵抗(AR)或氯吡格雷抵抗(CR)的检出率增高;服用不同的抗血小板药物,糖尿病患者对以ADP、COL、AA为诱导剂测定的血小板聚集率,其影响不同;未空腹及检测时间超过3 h重复性较差;服药2周后,COL、AA诱导的血小板聚集率下降不明的患者,随着时间的延长,6个月后易重新出现CR.结论 血小板计数、诱导剂浓度、糖尿病疾病、空腹状态、检测时间与比浊法检测血小板聚集率相关,以ADP、COL、AA作诱导剂检测血小板聚集率较单一的ADP更全面、准确.     

不同血小板激活剂在薄片法聚集实验中的应用比较

本研究的目的是筛选和评价可用于薄片法血小板聚集试验的血小板激活剂。应用下列方法进行实验 :①测定常用血小板激活剂 ,包括ADP、胶原、肾上腺素、花生四烯酸和瑞斯托霉素及阳离子没食子酸丙酯溶液 (c PG)在塑料薄片上诱导 15名健康献血者血小板聚集强度和时间 ;②测定添加血小板抑制剂PGI2 、cAMP或EDTA前后常用血小板激活剂及c PG在诱导 15名健康献血者血小板聚集时间 ;③测定不同浓度肝素对c PG诱导的 15名健康献血者小板聚集时间的影响 ;④检测c PG诱导健康献血者不同浓度血小板的聚集强度和时间 ;⑤检测c PG诱导服用阿司匹林患者血小板聚集时间。结果表明 :①在塑料薄片上c PG诱导血小板的聚集强度最强 ,肉眼清晰可见 ,需要时间较短 ;②瑞斯托霉素、花生四烯酸和c PG均可检测出抑制剂PGI2 和cAMP对血小板的抑制 ,其中c PG检测能力相对最强 ;③ 0 .5 - 3U ml肝素对c PG诱导的血小板聚集时间无明显影响 ;④健康献血者血小板稀释至 30× 10 9 L仍可被c PG诱导出明显的肉眼可见的聚集 ;⑤c PG诱导服用阿司匹林患者血小板聚集时间明显延长。结论 :与常用血小板激活剂比较 ,c PG应用于薄片法血小板聚集试验中具有明显优势。     

Comparison of two platelet additive solutions

The use of an additive solution for substitution of plasma for storage of leukodepleted platelet concentrates can have many advantages. In this study, a comparison was made between two platelet additive solutions: one containing citrate and acetate (PAS-II), the other also supplemented with additional salts such as magnesium, and with gluconate (Composol-PS). Donor-dependent differences were avoided by applying a paired experimental design (n = 10). The platelet concentrates were prepared by pooling five buffy coats and the additive solution, and prestorage filtration was utilized to remove leucocytes to well below 1 x 106. Storage of platelet concentrates up to 9 days after blood collection revealed that platelet concentrates in Composol-PS maintained an almost constant pH of on average 6.93 from day 2 through day 7, and at 6.90 at day 9. This was in contrast to PAS-II, which showed a gradually decreasing pH from on average 6.97 at day 1 to 6.86 at day 9. In all units stored in both solutions the swirling effect was present during 9 days of storage. In conclusion, both additive solutions allow storage of platelets, derived from pooled buffy coats, for up to 9 days after collection of the whole blood, with maintenance of good quality in vitro. Composol-PS has a slightly better buffering capacity, reflected as a more constant pH throughout the storage period.     

Algorithm for managing the platelet refractory patient

Refractoriness to platelet transfusions continues to be a major problem for many thrombocytopenic patients. A proposed algorithm for managing these patients is presented which proceeds from easily instituted changes in platelet transfusions therapy such as provision of ABO-compatible and “fresh” platelet transfusion to the more difficult and costly process of selecting compatible platelets for patients who are documented to be alloimmunized. For non-immunized platelet refractory recipients, multiple clinical and drug factors that may adversely effect transfusion responses have been identified. Identifying which of these factors are causally associated with poor platelet responses in any given remains a substantial challenge. J. Clin. Apheresis 12:4–9, 1997 © 1997 Wiley-Liss, Inc.     

Platelet indices in athletes performing a race in altitude environment

Physical activity could modify platelet count and platelet indices. Previous reports showed modifications after exercise linked to type and duration of sports performances. The shortage of studies in this field stems from the crucial methodological problem of EDTA (ethylenediaminetetraacetic acid)-dependent, mean platelet volume modifications: the published data on platelets count and indices were obtained by using a light-scattering system without standardization of the period elapsed from drawing to measurement. We present a study of platelet indices performed in athletes participating in an extreme' performance, a race of 30 km in altitude (6,700 m of ascents and descents), using standardized measurement by an aperture-impedance instrument transferred in the field. The platelet count and mean platelet volume significantly increased in athletes, whereas platelet distribution width decreased. The mean initial values were 240.6^*10⁹/L for platelet count, 8.79 fL for mean platelet volume, and 15.79% for platelet distribution width. The correspondent mean final values were 288.4^*10⁹/L, 9.14 fL, and 15.48%. The modifications of platelet count and indices were always in the physiological reference range. The entity and the rapidity of platelet count and indices modifications suggest that the more probable source of variation is the recruitment of noncirculating pools of mature platelets. Strenuous exercise does not show abnormal changes of platelet parameters.©1995 wiley-Liss, inc.