Platelet degranulation and monocyte-platelet complex formation are increased in the acute and convalescent phases after ischaemic stroke or transient ischaemic attack

Flow cytometric studies suggest that platelets are activated in ischaemic stroke or transient ischaemic attack (TIA). However, few studies have measured circulating leucocyte-platelet complexes in this patient population. Whole blood flow cytometry was used to quantify the expression of CD62P-, CD63-, and PAC1-binding, and the percentages of leucocyte-platelet complexes in acute (1-27 d, n = 79) and convalescent (79-725 d, n = 70) ischaemic cerebrovascular disease (CVD) patients compared with controls without CVD (n = 27). We performed a full blood count, and measured plasma levels of soluble P-selectin, soluble E-selectin, and von Willebrand factor antigen (VWF:Ag) as additional markers of platelet and/or endothelial cell activation. The median percentage CD62P expression and the median percentage monocyte-platelet complexes were higher in both acute and convalescent CVD patients than controls (P 相似文献   

There's more to transient ischaemic attack than ABCD

Several important studies have contributed to recent changes in clinical thinking regarding transient ischaemic attack (TIA). The use of risk stratification scores and urgent clinics has become popular and incorporated into recent guidelines, but there may be some pitfalls with this approach. This commentary stresses the need for careful individual patient assessment focussed on prompt determination of the underlying pathogenic mechanism for the TIA. Suggestions on how to incorporate assessment of TIA patients into the current duties of stroke units are made.     

Challenges in secondary stroke prevention: prevalence of multiple metabolic risk factors,including abnormal glycaemia,in ischaemic stroke and transient ischaemic attack

Background: Secondary prevention of ischaemic stroke (IS) and transient ischaemic attack (TIA) mandates identification and treatment of multiple metabolic risk factors. The aim was to determine the prevalence of abnormal glycaemia, hypertension and dyslipidaemia in patients presenting to an Acute Stroke Unit of a tertiary referral teaching hospital with IS or TIA. Methods: We reviewed the clinical characteristics of consecutive patients presenting with symptoms of acute stroke or TIA between 1 February 2006 and 30 June 2007 to determine the prevalence of diabetes, impaired fasting glucose (IFG), post‐stroke dysglycaemia (PSD), hypertension and dyslipidaemia. Results: Mean age ± SD of the 224 patients (84 female) was 71 ± 15 years. Seventy per cent (n= 157) of patients presented with IS and 30% (n= 67) with TIA. Of the cohort, 15% (n= 33) had previously diagnosed diabetes, 10% (n= 22) were diagnosed with diabetes during admission and 19% (n= 42) had IFG diagnosed during admission. A further 4% (n= 9) were classified as having PSD. Sixty‐two per cent (n= 139) of patients had previously diagnosed hypertension; another 7% (n= 15) were diagnosed during admission. Eighty‐eight per cent (n= 197) of patients had dyslipidaemia. Thirty per cent had all three risk factors concurrently. Conclusion: Abnormal glycaemia was present in almost half the patients presenting with IS/TIA, with the majority of cases undiagnosed. One‐third of patients had abnormal glycaemia, hypertension and dyslipidaemia concurrently. Patients presenting with stroke should be routinely screened for abnormal glycaemia in concert with other vascular risk factors.     

Acute stroke and transient ischaemic attack management – time to act fast

Stroke is Australia's second single greatest killer with 53 000 new events each year at a rate of 1 every 10 min. Stroke services should be organized to enable people to access proven therapies, such as stroke unit care and thrombolysis, to reduce the impact of stroke. Timely, efficient and coordinated care from ambulance services, emergency services and stroke services will maximize recovery and prevent costly complications and subsequent strokes. Efficient management of patients with transient ischaemic attack can produce significant reductions in subsequent stroke events and risk stratification using the ABCD2 tool can aid management decisions. Evidence for acute stroke care continues to evolve and it is crucial that health professionals are aware of, and implement, best practice clinical guidelines for stroke care.     

Ischaemic stroke and transient ischaemic attack on anticoagulants: outcomes in the era of direct oral anticoagulants

Clinical and imaging characteristics of patients receiving direct oral anticoagulants presenting with transient ischaemic attack or stroke are lacking. A retrospective review of all patients who presented to a high‐volume primary stroke centre with acute stroke symptoms while prescribed an oral anticoagulant between January 2012 and June 2017. Clinical, radiological characteristics and functional outcomes were examined. Anticoagulated patients diagnosed with stroke or transient ischaemic attack shared similar disease and outcome characteristics irrespective of anticoagulants used. One‐third of warfarin patients with sub‐therapeutic international normalised ratios were treated with thrombolytics but no direct oral anticoagulants level was performed in any of the patients, with only one treated by intravenous thrombolysis.     

Correlation between immature platelet fraction and reticulated platelets. Usefulness in the etiology diagnosis of thrombocytopenia

Background: Reticulated platelets (RP) are a surrogate marker for megakaryocytic activity, but the limitation of this determination is the lack of standardization of methodology. The determination of the immature platelet fraction (IPF) is performed in a simple, automated, and reproducible way between laboratories. We analyzed the correlation between IPF and RP, and usefulness of IPF in patients with thrombocytopenia. Methods: RP were determined by flow cytometry using double staining with thiazole orange and CD61 PerCP^®. IPF was performed with Sysmex XE2100 analyzer. We used a control group with normal platelets, and thrombocytopenic patients were classified into three groups: Group 1. Central thrombocytopenia, Group 2. Thrombocytopenia as a result of enhanced peripheral platelet destruction, and Group 3. Peripheral non‐immune thrombocytopenia by abnormal distribution. Results: Fourteen controls and 66 patients were analyzed. Group 1: 25 patients, they had mean and confidence interval 95% (95% CI) for IPF 8.67% (6.49–10.46%) and RP 4.08% (2.86–5.30%). Group 2: 20 patients, they had mean and 95%CI for IPF 16.80% (12.20–21.39%) and RP 16.14% (9.89–22.40%). Group 3: 21 patients, they had mean and 95% CI for IPF 9.04% (6.95–11.14%) and RP 5.23% (3.41–7.05%). The overall Pearson linear correlation between IPF and RP was r: 0.65. There were statistically significant differences in values of IPF and RP between Group 2 and the other two groups (P < 0.01). Conclusion: There is a good correlation between IPF and RP mainly in thrombocytopenia by peripheral destruction. Determination of IPF is an easy technique in their implementation, standardized and reproducible, so it could be a useful screening technique in patients with thrombocytopenia.     

Low positive predictive value of the ABCD2 score in emergency department transient ischaemic attack diagnoses: the South Western Sydney transient ischaemic attack study

Background: The ABCD² stroke risk score is recommended in national guidelines for stratifying care in transient ischaemic attack (TIA) patients, based on its prediction of early stroke risk. We had become concerned about the score accuracy and its clinical value in modern TIA cohorts. Methods: We identified emergency department‐diagnosed TIA at two hospitals over 3 years (2004–2006). Cases were followed for stroke occurrence and ABCD² scores were determined from expert record review. Sensitivity, specificity and positive predictive values (PPV) of moderate–high ABCD² scores were determined. Results: There were 827 indexed TIA diagnoses and record review was possible in 95.4%. Admitted patients had lower 30‐day stroke risk (n= 0) than discharged patients (n= 7; 3.1%) (P < 0.0001). There was no significant difference in proportion of strokes between those with a low or moderate–high ABCD² score at 30 (1.2 vs 0.8%), 90 (2.0 vs 1.9%) and 365 days (2.4 vs 2.4%) respectively. At 30 days the sensitivity, specificity and PPV of a moderate–high score were 57% (95% confidence interval (CI) 25.0–84.2), 32.2% (95% CI 29.1–35.6) and 0.75% (95% CI 0.29–1.91) respectively. Conclusions: Early stroke risk was low after an emergency diagnosis of TIA and significantly lower in admitted patients. Moderate–high ABCD² scores did not predict early stroke risk. We suggest local validation of ABCD² before its clinical use and a review of its place in national guidelines.     

Relationship between size and thiazole orange fluorescence of platelets in patients undergoing high-dose chemotherapy.

The reticulated platelet count relies upon the assumption that newly formed platelets contain a residual amount of RNA which selectively binds the dye thiazole orange (TO) and greatly enhances its fluorescence signal. It has, however, recently been shown that almost half of the platelet TO-signal is derived from the labelling of dense-granule nucleotides. It is therefore possible that the higher TO fluorescence of young platelets partially derives from the higher granule content due to their larger volume. To investigate the relationship between platelet size and TO fluorescence we studied 13 patients with high-risk breast cancer undergoing high-dose chemotherapy. Mean platelet volume, platelet distribution width, platelet-large cell ratio, membrane content of glycoprotein Ib and IIb-IIIa and platelet aggregation were significantly greater during resolution than during development of thrombocytopenia, suggesting a prevalence of young and old platelets respectively. Mean TO fluorescence per cell was higher in the platelet population enriched in young cells than in that enriched in old cells, but this difference was no longer observed when the ratio TO signal/platelet size was examined. Moreover, RNase treatment and platelet degranulation reduced TO fluorescence to a similar extent in platelet populations enriched in young or old cells. Therefore our data suggest that the higher TO signal of young platelets is derived, to a significant extent, from their larger volume and granule content.     

In vivo biotinylation studies: specificity of labelling of reticulated platelets by thiazole orange and mepacrine

Animal in vivo biotinylation studies have demonstrated that thiazole orange (TO) labels the youngest cells in the circulation. TO has since been widely used for the measurement of reticulated platelets. As recent findings suggest that at high concentrations TO also labels platelet dense granules non-specifically, the value of previous work is unclear. Mepacrine also labels platelet dense granules and can detect storage pool defects. In this study, a mouse in vivo biotinylation model was used to determine the specificity of TO and mepacrine staining on platelets recently released into the circulation. The mean life span of biotin/TO (low), biotin/TO (high) and mepacrine/TO dual-positive platelets was 1.4 d (SD 0.5), 2.2 d (SD 0.2) and 2.3 d (SD 0.3) respectively (n = 6) compared with a life span for biotin-positive platelets of 4.9 d (SD 1.6). TO (low), TO (high) and mepacrine labelled 8.0% (SD 3.1), 43.9% (SD 8.3) and 40.0% (SD 9.9) of the total platelet population respectively (results of 30 samples from six mice), which decreased to 6.8% (SD 3. 9), 26.6% (SD 6.9) and 25.7% (SD 10.6) after thrombin degranulation. The shorter life span and lack of thrombin sensitivity of TO (low)-positive platelets, suggests that TO (low) measures reticulated platelets specifically. The comparative life spans and thrombin sensitivity of TO (high) and mepacrine-positive platelets suggest that TO (high) labels platelet dense granules. These data also suggest that dense granules are lost during platelet ageing.     

Antithrombotic use following transient ischaemic attack or ischaemic stroke among older Australians with atrial fibrillation

Hospital audits may underestimate anticoagulant use among acute ischaemic stroke patients with atrial fibrillation (AF), as treatment may commence after discharge. To account for this, antithrombotic use in the 4 months after hospitalisation for transient ischaemic attack or ischaemic stroke among AF patients was assessed using claims data. Results suggest that treatment may be commenced soon after discharge and should be considered when assessing prevalence of use.     

氯吡格雷在短暂性脑缺血发作后脑卒中二级预防中的应用

目的探讨氯吡格雷在短暂性脑缺血发作(transient ischemic attack,TIA)后预防缺血性脑卒中发作的疗效。方法选择TIA患者279例,随机分为2组:氯吡格雷组158例(氯吡格雷75 mg,1次/d),长效阿司匹林组121例(拜阿司匹林100 mg,1次/d)。患者随访1.5～3.0(2.3±0.3)年,评估2组的安全性。结果氯吡格雷组患者缺血性脑卒中复发率明显低于长效阿司匹林组,差异有统计学意义(5.06% vs 12.40%,P0.05)。全部患者在TIA后缺血性脑卒中复发的危险比为0.4031,氯吡格雷组为0.1284,长效阿司匹林组为0.8129,差异有统计学意义(P0.05)。氯吡格雷组患者发生次要事件概率和胃肠道出血事件明显低于长效阿司匹林组,差异有统计学意义(1.90% vs 8.26%,1.27% vs 6.61%,P0.05)。结论氯吡格雷对TIA患者的缺血性脑卒中的预防,优于长效阿司匹林。     

Patent foramen ovale closure following cryptogenic stroke or transient ischaemic attack: Long‐term follow‐up of 301 cases

Patent foramen ovale has been identified as a conduit for paradoxical embolism resulting in cryptogenic stroke or transient ischemic attack (TIA). We aimed to establish rates of death, recurrent stroke or TIA among patients undergoing PFO closure for stroke or TIA at our unit. A retrospective analysis of all PFO closure patients was performed between May 2004 and January 2013. Follow up was performed by mortality tracing using the Medical Research Information Service of the Office of National Statistics. With regard to stroke or TIA recurrence, written consent forms and questionnaires were mailed with follow up telephone calls. Medical notes and imaging records were consulted where adverse events were noted. 301 patients aged 48.6 ± 11.0 years, 54.4% male, with ≥1 thromboembolic neurovascular event had percutaneous PFO closure with one of eight devices, with successful implantation in 99% of cases. Follow‐up duration was 40.2 ± 26.2 months (range 1.3–105.3); complete in 301 patients for mortality (100%) and 283 patients (94.0%) for neurovascular events. Two patients died during follow‐up (respiratory failure n = 1; road traffic accident n = 1). Recurrent stroke (MRI or CT confirmed) was observed in five patients (0.5%; 0.55 per 100 person‐years) and TIA in 9 (1.1%; 0.98 per 100 person‐years). Atrial fibrillation requiring treatment was documented in 14 patients (1.7%). Percutaneous PFO closure in patients with cryptogenic stroke or TIA is a safe treatment with a low incidence of procedural complications and recurrent neurovascular events. Registry data like these may help to demonstrate the utility of PFO closure in stroke. © 2015 Wiley Periodicals, Inc.     

ABCD~3-I评分法预测短暂性脑缺血发作患者早期脑卒中风险

目的探讨ABCD3-I评分法对短暂性脑缺血发作(TIA)患者早期脑卒中风险的预测价值。方法选取颈内动脉系统TIA患者182例,分别采用ABCD2、ABCD3、ABCD3-I评分标准进行评分,其中ABCD3-I危险分层分为低危40例,中危74例,高危68例,并观察TIA后7d内脑梗死的发生率。结果 ABCD2、ABCD3与ABCD3-I评分法预测TIA后7d内脑梗死风险的ROC曲线下面积分别为0.625、0.713、0.831。TIA患者7d内进展为脑梗死27例(14.8%)。ABCD3-I评分低危、中危和高危患者7d内脑梗死的发生率分别为0、6.8%、32.4%,中危和高危脑梗死发生率明显高于低危患者,高危脑梗死发生率明显高于中危患者,差异有统计学意义(P<0.01);ABCD3-I评分与7d内脑梗死发生率呈正相关(r=0.486,P<0.01)。除年龄因素外,ABCD3-I评分法中各个评分项目对TIA后7d内脑梗死发生率均有明显影响(P<0.05)。结论 ABCD3-I评分法能更有效地预测TIA患者早期发生脑梗死的风险,可作为常规应用于临床,指导TIA危险分层评价和治疗。     

Enhanced ex vivo inhibition of platelet function following addition of dipyridamole to aspirin after transient ischaemic attack or ischaemic stroke: first results from the TRinity AntiPlatelet responsiveness (TrAP) study

Ex vivo dipyridamole ‘non‐responsiveness’ has not been extensively studied in ischaemic cerebrovascular disease. Platelet surface marker expression, leucocyte‐platelet complex formation and inhibition of platelet function at high shear stress as detected by the PFA‐100^® Collagen‐Adenosine‐diphosphate (C‐ADP) and Collagen‐Epinephrine cartridges was assessed in 52 patients within 4 weeks of transient ischaemic attack (TIA) or ischaemic stroke on aspirin, and then 14 d (14 d) and >90 d (90 d) after adding dipyridamole. A novel definition of ‘Dipyridamole non‐responsiveness’ was used. The median C‐ADP closure time increased following addition of dipyridamole, remained elevated at 90 d (P ≤ 0·03), and was unaffected by aspirin dose. 59% at 14 d and 56% at 90 d were ‘dipyridamole non‐responders’ on the PFA‐100. The proportion of non‐responders at 14 and 90 d was similar (P = 0·9). Compared with baseline (4·6%), median monocyte‐platelet complexes increased at 14 d (5·0%, P = 0·03) and 90 d (4·9%, P = 0·04). Low C‐ADP closure times were associated with increased monocyte‐platelet complexes at 14 d (r = ?0·32, P = 0·02) and 90 d (r = ?0·33, P = 0·02). Monocyte‐platelet complexes increased in the subgroup of dipyridamole non‐responders on the PFA‐100 (P ≤ 0·045), but not in responders (P ≥ 0·5), at 14 and 90 d versus baseline. Additional inhibition of platelet function has been detected with the PFA‐100 when dipyridamole is added to aspirin. Elevated monocyte‐platelet complexes may contribute to ex vivo dipyridamole non‐responsiveness.     

Comparison of reticulated platelet count and mean platelet volume determination in the evaluation of bone marrow recovery after aplastic chemotherapy

Reticulated platelet count provides an estimate of thrombopoiesis in the same way as reticulocyte count is a measure of erythropoiesis. We applied thiazole orange (TO) staining, followed by fluorescence-activated flow-cytometric analysis, to platelets in whole-blood samples from normal subjects and 18 aplastic patients after chemotherapy for haematologic malignancies. The percentage of TO-positive platelets in 30 control subjects was 5.7 +/- 2.4% (mean +/- 1 SD), determining the threshold of reticulated platelet positivity as up to 10.5% (mean + 2 SD). In the 18 patients studied, the mean percentage of TO-positive platelets was 4.3 +/- 1.89% during aplasia and 23.3 +/- 9.43% during bone marrow recovery, respectively (P < 0.05). All patients had a percentage of TO-positive platelets of up to 10.5%. In comparison, mean platelet volume during bone marrow recovery increased in 12 cases of the 18 patients studied. We conclude that flow cytometric analysis of reticulated platelets is a sensitive and specific test for evaluating thrombopoiesis recovery during aplastic chemotherapy, and platelet transfusion should be reconsidered in these patients.     

Tracking and characterisation of transfused platelets by two colour, whole blood flow cytometry

We describe a flow cytometric technique to study transfused platelets in patients. By selecting donor/recipient pairs discrepant for HLA-A2, transfused platelets were identified using anti-HLA-A2 with a detection limit of <5%, and accuracy within 4.3% of predicted (r2 > 0.96, P < 0.0001). In two of three episodes, transfused platelets were present in greater numbers than predicted from increment counts. Platelets with bound fibrinogen fell from 20.9 +/- 23.6% of donor platelets pretransfusion to 1.7 +/- 0.3% 1 h post-transfusion, whereas P-selectin-positive platelets fell gradually, from 24.1 +/- 6.7 to 7.3 +/- 3.3% at 6 h. This method avoids radio or chemical labelling, and could be used to assess novel platelet preparations post-transfusion.     

Quality of harvested autologous platelets compared with stored donor platelets for use after cardiopulmonary bypass procedures

Platelet dysfunction has a major contribution in bleeding after cardiopulmonary bypass (CPB) and transfusion of platelets is frequently used to secure haemostasis. Allogeneic platelets prepared for transfusion are functionally impaired. Autologous platelets harvested preoperatively require a shorter storage time before transfusion and their use also avoids the risks associated with transfusion of allogeneic blood products. For the first time, we have compared the functional quality of autologous platelets with allogeneic platelets prepared by two methods, immediately before infusion. Platelet activation was assessed by P-selectin expression and fibrinogen binding using flow cytometry. We also monitored the effects of CPB surgery and re-infusion of autologous platelets on platelet function. Autologous platelet-rich plasma (PRP) contained a significantly lower (P < 0.05) percentage of P-selectin-positive and fibrinogen-positive platelets compared with allogeneic platelet preparations, and also contained a significantly higher (P < 0.05) percentage of responsive platelets. Allogeneic platelets prepared by donor apheresis were more activated and less responsive than those produced by centrifugation of whole blood. In patients' blood, the percentage of platelets expressing P-selectin or binding fibrinogen increased significantly after CPB (P < 0.05), while the percentage of platelets responsive to in vitro agonists was decreased (P < 0.05 in autologous transfusion patients), consistent with platelet activation during the procedure. The percentage of activated platelets decreased (statistically not significant) after re-infusion of autologous PRP. P-selectin expression had returned to pre-CPB levels 24 h post-operatively. Autologous platelet preparations display minimal activation, but remain responsive. Conservation of platelet function may contribute to the potential clinical benefits of autologous transfusion in cardiopulmonary bypass.     

Reticulated platelets in uremic patients: Effect of hemodialysis and continuous ambulatory peritoneal dialysis

Platelet RNA content can be detected by flow cytometry using thiazole orange staining to identify platelets recently released into the circulation. We studied platelet RNA content and platelet function in uremic patients under different treatment regimens. Four groups were studied: (I) 15 end-stage renal disease (ESRD) patients (10M/5F) on maintenance hemodialysis (HD); (II) 11 ESRD patients (6M/5F) on continuous ambulatory peritoneal dialysis (CAPD); (III) 8 patients with chronic renal failure managed conservatively (5M/3F); and (IV) 34 controls (20M/14F). A double color labeling technique using a phycoerythrin-tagged antibody against glycoprotein lb (CD42b) and RNA labeling by thiazole orange was performed and read by flow cytometry. Aggregation studies were made in platelet-rich plasma using ADP, epinephrine, collagen, arachidonic acid, and ristocetin. In group I, samples were also obtained after HD. Platelet counts did not differ among the groups. Aggregation studies showed a lower response to ADP and ristocetin in the HD patients, but not in the CAPD or in the chronic renal failure patients. The percentage of platelets with high RNA content in group I was significantly lower than in controls (3.72 ± 1.72% vs. 9.05 ± 3.53%, P < 0.01), but was also lower than in the remaining groups (I vs. II P < 0.01, and I vs. III P < 0.01). No differences were seen in platelet RNA content among groups II (8.67 ± 2.73%), III (9.14 ± 3.04%) and IV. In group I, the percentage of reticulated platelets decreased further after HD (2.14 ± 1.09%, P < 0.01). Aggregation studies showed a significantly lower response to ADP and ristocetin in group I (P < 0.05), but not in groups II or III in comparison with controls. Aggregation response to ADP and ristocetin decreased after HD (P < 0.05). In conclusion, HD may decrease the percentage of RNA-rich platelets through elimination of the younger and more active platelets and worsen the thrombopathy present in uremic patients.     

Flow cytometric analysis of platelets from childrenwith the Wiskott-Aldrich syndrome reveals defects in platelet development, activation and structure

The pathophysiology of platelet dysfunction in the Wiskott-Aldrich immune deficiency syndrome (WAS) remains unclear. Using flow cytometry, we have characterized the functional properties of platelets from 10 children with WAS. Patients with WAS had thrombocytopenia, small platelets, increased platelet-associated IgG and reduced platelet-dense granule content. Levels of reticulated 'young' platelets were normal in the WAS patients. Although the mean numbers of platelet glycoprotein (GP) Ib, GPIIbIIIa and GPIV molecules per platelet appeared lower in WAS patients than in healthy controls, analysis of similar-sized platelets revealed the mean number of GPIb molecules per platelet to be comparable in patients and normal controls. Surface GPIIbIIIa and GPIV expression was, however, significantly lower on the WAS platelets than on normal platelets. Compared with normal platelets, WAS platelets showed a reduced ability to modulate GPIIbIIIa expression following thrombin stimulation. In addition, thrombin- and ADP-induced expression of CD62P and CD63 was defective in WAS platelets. Phallacidin staining of the WAS platelets revealed less F-actin content than in normal platelets. Together, these data suggest that the reduced platelet number and function in WAS reflects, at least in part, a defect in bone marrow production as well as an intrinsic platelet abnormality.