Platelets do not express the oxidized or reduced forms of tissue factor

Background

Expression of tissue factor (TF) antigen and activity in platelets is controversial and dependent upon the laboratory and reagents used. Two forms of TF were described: an oxidized functional form and a reduced nonfunctional form that is converted to the active form through the formation of an allosteric disulfide. This study tests the hypothesis that the discrepancies regarding platelet TF expression are due to differential expression of the two forms.

Methods

Specific reagents that recognize both oxidized and reduced TF were used in flow cytometry of unactivated and activated platelets and western blotting of whole platelet lysates. TF-dependent activity measurements were used to confirm the results.

Results

Western blotting analyses of placental TF demonstrated that, in contrast to anti-TF#5, which is directed against the oxidized form of TF, a sheep anti-human TF polyclonal antibody recognizes both the reduced and oxidized forms. Flow cytometric analyses demonstrated that the sheep antibody did not react with the surface of unactivated platelets or platelets activated with thrombin receptor agonist peptide, PAR-1. This observation was confirmed using biotinylated active site-blocked factor (F)VIIa: no binding was observed. Likewise, neither form of TF was detected by western blotting of whole platelet lysates with sheep anti-hTF. Consistent with these observations, no FXa or FIXa generation by FVIIa was detected at the surface of these platelets. Similarly, no TF-related activity was observed in whole blood using thromboelastography.

Conclusion and significance

Platelets from healthy donors do not express either oxidized (functional) or reduced (nonfunctional) forms of TF.     

Natural deep eutectic solvents for lignocellulosic biomass pretreatment: Recent developments,challenges and novel opportunities

Conversion of lignocellulosic biomass to fuels and chemicals has attracted immense research and development around the world. Lowering recalcitrance of biomass in a cost-effective manner is a challenge to commercialize biomass-based technologies. Deep eutectic solvents (DESs) are new ‘green' solvents that have a high potential for biomass processing because of their low cost, low toxicity, biodegradability, easy recycling and reuse. This article discusses the properties of DESs and recent advances in their application for lignocellulosic biomass processing. The effectiveness of DESs in hydrolyzing lignin-carbohydrate complexes, removing lignin/hemicellulose from biomass as well as their effect on biomass deconstruction, crystallinity and enzymatic digestibility have been discussed. Moreover, this review presents recent findings on the compatibility of natural DESs with enzymes and microorganisms.     

CRUSADE评分联合血栓弹力图对急性冠脉综合征患者出血风险的评估

目的:运用CRUSADE评分系统联合血栓弹力图对急性冠脉综合征(acute coronary syndrome,ACS)患者抗栓治疗中的出血风险进行评估。方法:回顾性分析2013年1月至2013年12月在上海交通大学医学院附属新华医院住院的ACS患者病历249例,用CRUSADE评分联合血栓弹力图评估ACS患者30天出血事件的发生。结果:随访的249例ACS患者,共有46例(18.5%)患者发生了出血事件;按照CRUSADE评分进行危险分层,极低危组、低危组、中危组、高危组、极高危组的出血率分别为:15%、7.5%、21.2%、32.5%26.7%;各组间出血率的比较:中危组、高危组及极高危组各组的出血率均高于低危组,差异有统计学意义;而高危组出血率高于极低危组,差异有统计学意义,而极低危组与其他各组比较,差异无统计学意义;低危组以上患者出血率高于低危组以下,差异有统计学意义。低危组以上中危组、高危组及极高危组各组间出血率比较,差异无统计学意义;按照血小板抑制率中位数分组,大于中位数组的出血率高于小于中位数组,差异有统计学意义。经多因素Logistic分析:PAg T抑制率(ADP)是ACS患者抗栓治疗中出血事件的独立影响因素。利用ROC曲线分析CRUSADE评分、血栓弹力图以及两者联合对患者出血事件发生的评估,两者联合的曲线下面积大于单独利用CRUSADE评分。结论:随着CRUSADE评分危险分层的增加出血的发生率亦呈增加趋势;危险分层低危以上的患者,不论中危、高危、极高危发生出血事件风险较低危险以下有明显增加;血栓弹力图监测血小板抑制率可作为CRUSADE评分的补充,提高对ACS患者出血风险的预测。     

A monovalent agonist of TrkA tyrosine kinase receptors can be converted into a bivalent antagonist

Background

Receptor tyrosine kinases (RTK) act through dimerization. Previously it was thought that only bivalent ligands could be agonistic, whereas monovalent ligands should be antagonistic. This notion changed after the demonstration that monovalent ligands can be agonistic, including our report of a small molecule monovalent ligand “D3” that is a partial agonist of the NGF receptor TrkA. A bivalent “D3-linker-D3” was expected to increase agonism.

Methods

Dimeric analogs were synthesized and tested in binding, biochemical, and biological assays.

Results

One analog, 1-ss, binds TrkA with higher affinity than D3 and induces or stabilizes receptor dimers. However, 1-ss exhibited antagonistic activity, through two mechanisms. One mechanism is that 1-ss blocks NGF binding, unlike D3 which is non-competitive. Inhibition of NGF binding may be due to the linker of 1-ss filling the inter-receptor space that NGF traverses before docking. In a second mechanism, 1-ss acts as a pure antagonist, inhibiting NGF-independent TrkA activity in cells over-expressing receptors. Inhibition is likely due to 1-ss “freezing” the TrkA dimer in the inactive state.

Conclusions

Dimerization of an RTK can result in antagonism, through two independent mechanisms.

General significance

we report a small molecule monovalent agonist being converted to a bivalent antagonist.     

Anticoagulant activity of M-LAO, l-amino acid oxidase purified from Agkistrodon halys blomhoffii, through selective inhibition of factor IX

One of haemorrhagic toxins present in snake venoms is l-amino acid oxidase (LAO), which catalyzes the oxidative deamination of l-amino acids with the generation of hydrogen peroxide. Although it is widely accepted that LAO alters platelet function, the effects of LAO on human blood coagulation remain largely unknown. The present study demonstrated, for the first time, that M-LAO, LAO purified from the venom of Agkistrodon halys blomhoffii (Japanese mamushi), possesses an anticoagulant activity. Thrombelastography (TEG) showed that M-LAO significantly delayed the onset and the progress of the coagulation process. In addition, the enzyme prolonged the activated partial thromboplastin time (aPTT) dose-dependently, but had little effect on the prothrombin time (PT), suggesting that its principal activity was mediated in the intrinsic coagulation pathway. Furthermore, M-LAO reduced factor IX procoagulant activity in a dose-dependent manner and did not affect other coagulation factors. These results indicate that M-LAO has an anticoagulant activity that impairs the intrinsic clotting by inhibiting factor IX.     

Role of thromboelastography in clinical emergency trauma patients

This study aimed to investigate dynamic changes in thromboelastography (TEG) and evaluate its value for emergency trauma patients. TEG values (R, K, Angle, MA and CI) were detected l, 3, and 7 days post trauma. Followup prognosis data were recorded among all patients and correlations between TEG changes and prognosis were assessed. The results revealed that the K and R volumes of the poor prognosis group were obviously higher than those of the good prognosis group at each time point; while the Angle, MA, and CI volumes of the poor prognosis group were relatively lower than those of the good prognosis group, and showed a little rising tendency. This study suggests that variables measured by TEG were closely associated with prognosis evaluation, and highlighted the dangers of coagulation disorder at early stage. TEG is of benefit in evaluating the function of coagulation in clinical emergency trauma patients.     

血栓弹力图测定结果的比对研究

目的:探讨裕泽毅血栓弹力图仪与美国血计公司TEG血栓弹力图仪检测结果相关性,可信度与优劣。方法:根据美国NCCLS-EP9-A,从我院随机抽取60例病人,分别取静脉血2ml、枸橼酸抗凝处理充分混匀后进行测定。用裕泽毅血栓弹力图仪与TEG血栓弹力图仪及两家公司生产的试剂进行平行交叉测定,观察其相关性。结果:美国血技公司TEG血栓弹力图仪对于两家公司的试剂检测结果高度相关(r0.8);裕泽毅公司血栓弹力图仪和试剂对于TEG血栓弹力图仪和试剂检测结果高度相关(r0.8)。结论:裕泽毅公司机器和试剂可以替代TEG机器和试剂进行血栓弹力图检测,能为临床医生提供可靠、准确、有价值的诊断指导依据。