血小板输注中抗体检测和配合试验的应用

目的：提高血小板制剂对血小板减少、尤其是血小板输注无效症（PTR）患者的输注疗效,避免宝贵血源的浪费。方法：应用单抗固相微孔板（MASPAT）法检测患者血清中的血小板抗体,进行血小板供者与患者之间的配合试验。结果：2005年6月-2007年11月对109例患者进行了血小板抗体的检测,其中42例患者检出血小板抗体（阳性率38．5％）,对含有血小板抗体的患者经适合性血小板输注后,血小板计数有明显上升。结论：MASPAT法在特异性、敏感性、重复性方面良好,操作快速、简便,判断可靠;易做到规范化,程序化,标准化;据此建立的“适合性血小板输注”对含有血小板抗体的患者是有效的,可用于临床血小板抗体的检测和配合试验。     

Mean platelet volume and mean platelet volume/platelet count ratio in infective endocarditis

Abstract

Infective endocarditis (IE), an infection of the endocardial surface, frequently leads to life-threatening complications, such as thromboembolism due to platelet activation. We investigated the mean platelet volume (MPV) in Korean patients with IE and the serial changes thereof, in comparison with other laboratory parameters. We analyzed 248?MPV results from 22 patients diagnosed with IE in our hospital between January 2011 and April 2012. MPV was measured with an Advia 2120 (Siemens Healthcare Diagnostics, Tarrytown, NY) using EDTA-containing tubes. The mean MPV differed significantly between the patient and control groups, 8.74 vs. 7.96?fl, respectively. In addition, the platelet count and MPV/platelet count ratio were significantly decreased in the patient group. The total platelet mass and platelet size in IE might be increased. Further studies should examine more patients to verify the changes in the MPV and MPV/platelet count ratio in IE and assess in greater detail the relationship between MPV and thrombotic complications caused by platelet activation.     

Phosphorylation of platelet actin-binding protein during platelet activation

In this study we have followed the 32P-labeling of actin-binding protein as a function of platelet activation. Utilizing polyacrylamide- sodium dodecyl sulfate gel electrophoresis to resolve total platelet protein samples, we found 2--3-fold labeling increases in actin-binding protein 30--60 sec after thrombin stimulation. Somewhat larger increases were observed for 40,000 and 20,000 apparent molecular weight peptides. The actin-binding protein was identified on the gels by coelectrophoresis with purified actin-binding protein, its presence in cytoskeletal cores prepared by detergent extraction of activated 32P- labeled platelets, and by direct immunoprecipitation with antibodies against guinea pig vas deferens filamin (actin-binding protein). In addition, these cytoskeletal cores indicated that the 32P-labeled actin- binding protein was closely associated with the activated platelet's cytoskeleton. Following the 32P-labeling of actin-binding protein over an 8-min time course revealed that in aggregating platelet samples rapid dephosphorylation to almost initial levels occurred between 3 and 5 min. A similar curve was obtained for the 20,000 apparent molecular weight peptide. However, rapid dephosphorylation was not observed if platelet aggregation was prevented by chelating external calcium or by using thrombasthenic platelets lacking the aggregation response. Thus, cell-cell contact would seem to be crucial in initiating the rapid dephosphorylation response.     

Inhibition of platelet phospholipid methylation during platelet secretion

A pathway for the synthesis of membrane phosphatidylcholine involving the N-methylation of phosphatidylethanolamine has been detected in several types of mammalian cells. Furthermore, it has been implicated in the coupling of agonist binding to cell response. We examined whether human platelets exhibit this synthetic pathway and whether platelet agonists influence its activity. When washed platelets were incubated with 0.15 microM L-[methyl-3H]methionine at 37 degrees C, they incorporated methyl-3H into their phospholipids linearly at the rate of 1 pmole/10(9) platelets/hr. When incubated with 20 microM radiolabeled methionine, they incorporated about 15 pmole/10(9) platelets/hr. The radioactivity was found predominantly in phosphatidyl- N-monomethylethanolamine, phosphatidyl-N,N-dimethylethanolamine, and phosphatidylcholine. Thrombin caused an immediate (within 15 sec) and sustained (up to 30 min) decrease in the rate and extent of N- methylation of platelet phospholipids. This was accounted for by a decrease in synthesis of methylated phospholipids rather than an increase in their degradation. This thrombin effect correlated with serotonin release and could be dissociated from platelet aggregation and prostaglandin synthesis. Thrombin also decreased the synthesis of phosphatidylcholine when choline was used as the radiolabeled substrate. Other agonists such as epinephrine, adenosine diphosphate (ADP), or A23187 also decreased phospholipid methylation under conditions in which they stimulated serotonin release. These data demonstrate that platelets are capable of synthesizing phosphatidylcholine from phosphatidylethanolamine by N-methylation and that agonists perturb this pathway as they induce platelet secretion. The precise role of phospholipid methylation in either resting or stimulated platelets remains to be established.     

Interruption of tumor-associated platelet consumption with platelet enzyme inhibitors

Twenty dogs with naturally occurring metastatic tumors were treated with anticoagulants (Warfarin) or platelet enzyme inhibitor drugs (dipyridamole, dipyridamole plus aspirin, RA233, sulfinpyrazone, or a combination of RA233 and sulfinpyrazone) to determine if tumor-related reductions in platelet survival and concentration could be reversed. Anticoagulation was ineffective, while platelet enzyme inhibitors were able to produce improvements in platelet survival. Of the 18 dogs with metastatic tumor treated with platelet enzyme inhibitors, only 5 (28%) showed a reduction in platelet survival during the first week of observation on therapy compared to their baseline survivals. This is significantly different than the decreases in platelet survivals observed in 8 of 10 untreated dogs (80%) with metastatic tumor observed for the same interval. Furthermore, 8 of the 18 treated dogs (44%) had platelet survivals within 2 standard deviations of normal, compared to only 1 of 10 untreated dogs. Of the 8 dogs with normal platelet survivals, 6 were treated with a combination of a phosphodiesterase inhibitor (RA233 or dipyridamole) and a cyclooxygenase inhibitor (sulfinpyrazone or aspirin). The combination of RA233 and sulfinpyrazone was the best drug program tested and resulted in normal platelet survivals in 63% and improved platelet counts in 75% of the animals treated. Thus, platelet enzyme inhibitors with different mechanisms of action may have a synergistic effect in reversing the abnormal platelet hemostasis found in a variety of spontaneously occurring canine neoplasms.     

Effect of heparin on platelet count and platelet aggregation

The in vitro effect of heparin on platelet aggregation was studied in three groups: in 26 subjects recently treated with heparin, in 18 subjects on maintenance hemodialysis, and in 20 normal controls. With the aid of Technicon H6000, platelet counts and platelet aggregations were compared in whole blood samples collected in ethylenediaminetetraacetic acid (EDTA) and in heparinized tubes. Although there was no significant difference between platelet count of heparinized and EDTA blood in the control group, the dialysis group and the group recently treated with heparin showed significantly lower platelet counts and more platelet aggregation in heparinized tubes than in EDTA tubes. We speculate that the majority of subjects exposed to heparin develop an antibody or a proaggregator which can aggregate or agglutinate platelets in the presence of heparin and causes destruction of platelets; but only in a small percentage of subjects receiving heparin is this reaction severe enough to cause thrombocytopenia.     

光学法计数在低值血小板检测中的研究应用

目的：评价光学法计数在低值血小板检测中的研究应用。方法：200例低值血小板患者血标本,在血液分析仪上分别进行电阻抗法和光学法血小板计数,同时进行镜检法计数。结果：以镜检法为参考方法,光学法的相关系数为0．994。电阻抗法的相关系数0．921。在小红细胞组、红细胞碎片组和大血小板组,电阻抗法和镜检法计数结果差异有统计学意义（P〈0．05）,光学法和镜检法计数结果差异无统计学意义（P〉0．05）。结论：光学法计数可以满足低值血小板检测准确性的要求,值得临床实验室推广应用。     

Thrombopoietin therapy increases platelet yields in healthy platelet donors

The recombinant thrombopoietins have been shown to be effective stimulators of platelet production in cancer patients. It was therefore of interest to determine if one of these, pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), could be used to increase platelet counts and consequently platelet yields from apheresis in healthy platelet donors. In a blinded, 2-cycle, crossover study, 59 platelet donors were randomized to receive a single subcutaneous injection of PEG-rHuMGDF (1 microg/kg or 3 microg/kg) or placebo and 15 days later undergo platelet apheresis. Donors treated with placebo had a median peak platelet count after PEG-rHuMGDF injection of 248 x 10(9)/L compared with 366 x 10(9)/L in donors treated with 1 microg/kg PEG-rHuMGDF and 602 x 10(9)/L in donors treated with 3 microg/kg PEG-rHuMGDF. The median maximum percentage that platelet counts increased from baseline was 10% in donors who received placebo compared with 70% in donors who received 1 microg/kg and 167% in donors who received 3 microg/kg PEG-rHuMGDF. There was a direct relationship between the platelet yield and the preapheresis platelet count: Placebo-treated donors provided 3.8 x 10(11) (range 1.3 x 10(11)-7.9 x 10(11)) platelets compared with 5.6 x 10(11) (range 2.6 x 10(11)-12.5 x 10(11)) or 11.0 x 10(11) (range 7.1 x 10(11)-18.3 x 10(11)) in donors treated with 1 microg/kg or 3 microg/kg PEG-rHuMGDF, respectively. Substandard collections (<3 x 10(11) platelets) were obtained from 26%, 4%, and 0% of the placebo, 1 microg/kg, and 3 microg/kg donors, respectively. No serious adverse events were reported; nor were there events that met the criteria for dose-limiting toxicity. Thrombopoietin therapy can increase platelet counts in healthy donors to provide a median 3-fold more apheresis platelets compared with untreated donors.     

Factors affecting posttransfusion platelet increments, platelet refractoriness, and platelet transfusion intervals in thrombocytopenic patients

A variety of patient and product-related factors influenced the outcome of 6379 transfusions given to 533 patients in the Trial to Reduce Alloimmunization to Platelets (TRAP). Responses measured were platelet increments, interval between platelet transfusions, and platelet refractoriness. Patient factors that improved platelet responses were splenectomy and increasing patient age. In contrast, at least 2 prior pregnancies, male gender, splenomegaly, bleeding, fever, infection, disseminated intravascular coagulation, increasing height and weight, lymphocytotoxic antibody positivity, an increasing number of platelet transfusions, or receiving heparin or amphotericin were associated with decreased posttransfusion platelet responses. Platelet factors that were associated with improved platelet responses were giving ABO-compatible platelets, platelets stored for 48 hours or less, and giving large doses of platelets while ultraviolet B (UV-B) or gamma irradiation decreased platelet responses. However, in alloimmunized lymphocytoxic antibody-positive patients, the immediate increment to UV-B-irradiated platelets was well maintained, whereas all other products showed substantial reductions. Refractoriness to platelet transfusions developed in 27% of the patients. Platelet refractoriness was associated with lymphocytotoxic antibody positivity, heparin administration, fever, bleeding, increasing number of platelet transfusions, increasing weight, at least 2 pregnancies, and male gender. The only factors that reduced platelet refractoriness rates were increasing the dose of platelets transfused or transfusing filtered apheresis platelets.     

Absence of platelet activating factor (PAF) mediated platelet aggregation: A new platelet defect

The failure of normal human platelets to aggregate in response to platelet activating factor (PAF) has not been previously observed. We report here the first case of a patient whose platelets did not aggregate to PAF on multiple occasions.     

Thrombocytopenia with a circulating platelet agglutinin,platelet agglutinin,platelet lysin and clot retraction inhibitor

A case is reported of thrombocytopenia developing suddenly in a patientwith metastatic carcinoma. Studies of this patient’s serum revealed a plateletagglutinin presumed to be related to the development of the thrombocytopenia in spite of the failure to demonstrate autoagglutination. In addition, atechnique is described for the detection of a platelet lysin and clot retractioninhibitor, both of which could be demonstrated in this patient’s serum. Theidentity of the lysin and retraction inhibitor is suggested.

Studies on the serums of other patients showed a dissociation between theoccurrence of platelet agglutinin on the one hand and of platelet lysin andretraction inhibitor on the other.

Submitted on May 17, 1958 Accepted on July 8, 1958     

The relationship of human platelet density to platelet age: platelet population labeling by monoamine oxidase inhibition

The relationship between platelet density and platelet age appears to vary between species with relatively few labeling studies in humans reported. In this study, irreversible monoamine oxidase (MAO) inhibitors were used to biochemically label the circulating platelet population in 15 humans. Platelet samples were then isolated during the 15 days after drug ingestion. The platelets were separated by density on continuous linear Percoll gradients and the density distributions were divided into five fractions containing approximately equal numbers of platelets. Baseline MAO activity was strongly correlated with platelet density. Twenty-four hours after a single dose of tranylcypromine, platelet MAO activities in the density subpopulations were reduced to 14% to 17% of the baseline values. During the first five days after inhibition, the rates of recovery of MAO activity (percentage per day) were inversely proportional to platelet density. The recovery rates in the two most dense fractions were initially slow but increased after five days. Percentage recovery of MAO activity in the least dense fraction was significantly greater than the percentage recovery in the most dense fraction on days 2, 3, 5, and 8 (P less than .01, sign test). These results support the hypothesis that normal human platelets show a small increase in density with age, but they do not exclude the additional possibility that human platelet lifespan is positively correlated with platelet density.     

Pretransfusion trigger platelet counts and dose for prophylactic platelet transfusions

PURPOSE OF REVIEW: To assess critically both the blood platelet counts that prompt a platelet transfusion (i.e. trigger) in various clinical settings in patients with thrombocytopenia caused by marrow failure and the dose of platelets infused (i.e. number per each transfusion) for optimal hemostasis, feasibility, and safety. RECENT FINDINGS: Definitive studies (e.g. well-designed, prospective, randomized clinical trials) are not available either historically or at present to support evidence-based decisions. Instead, retrospective reviews and anecdotal reports provide observational data to assist in best guess clinical practices. SUMMARY: Reasonable clinical practice, until more definitive data become available, is to transfuse enough platelets per each transfusion to maintain the blood platelet count >10 x 10/L in stable nonbleeding patients, >20 x 10(9)/L in unstable nonbleeding patients, and >50 x 10(9)/L in bleeding patients or in those undergoing invasive procedures.     

Investigation of platelet function and platelet disorders using flow cytometry

Patients with thrombocytopenia or platelet disorders are at risk of severe bleeding. We report the development and validation of flow cytometry assays to diagnose platelet disorders and to assess platelet function independently of platelet count. The assays were developed to measure glycoprotein levels (panel 1) and platelet function (panel 2) in sodium citrated blood. Twenty healthy volunteers and five patients diagnosed with different platelet disorders were included. Glycoprotein expression levels of the receptors Ia, Ib, IIb, IIIa and IX were measured and normalised with forward scatter (FS) as a measurement of platelet size. Platelet function was assessed by CD63, P-selectin and bound fibrinogen in response to arachidonic acid, adenosine diphosphate (ADP), collagen-related peptide, ristocetin and thrombin receptor-activation peptide-6. All patients except one with suspected δ-granule defect showed aberrant levels of glycoproteins in panel 1. Glanzmann's thrombasthenia and genetically verified Bernard–Soulier syndrome could be diagnosed using panel 1. All patients showed reduced platelet function according to at least one agonist. Using panel 2 it was possible to diagnose Bernard–Soulier syndrome, δ-granule defect and GPVI disorder. By combining the two assays, we were able to diagnose different platelet disorders and investigate platelet function independent of platelet count.