高血压病患者淋巴细胞肿瘤坏死因子-β诱导的变化

肿瘤坏死因子(Tumor NecrosisFactor,TNF)不仅对肿瘤细胞有细胞毒性作用,它还通过使内皮细胞ELAM及ICAM的表达水平升高、刺激趋化性因子及集落刺激因子的产生从而促进粒细胞粘附、血管和血细胞生长、提高前列环素的水平和促血小板凝集活性并同时减低凝血调素蛋白C的抗凝活性促进凝血及促进内皮细胞sis癌基因的表达等参与心血管系统细胞功能的调     

纤维蛋白溶解系统的动力学模型与数值模拟

为了探讨蛋白C对凝血酶激活的纤维蛋白抑制剂(thrombin activatable fribrinolysis inhibitor,TAFI)的作用的影响,近而更深一步弄清楚血凝块形成初期TAFI对纤维蛋白溶解的调节机理,根据生物试验建立了一个关于蛋白C和TAFI的凝血动力学模型.通过对模型的动力学分析和数值模拟得到:凝血酶激活的纤维蛋白抑制剂刺激因素必须大于一个阈值纤溶系统才能发挥作用;当TAFI少量缺乏时,对血液凝固的影响不是太大;只有当TAFI的缺乏达到一定程度时,才会导致疾病的发生;但当TAFI过量时对系统的影响很大;系统对TAFI的需求随着蛋白C的增加而增加.     

胆汁中凝血状态与胆囊结石形成关系的研究

目的:通过研究胆固醇结石(简称胆石)病人胆囊胆汁中的凝血状态及其与交联纤维蛋白生成的关系来探讨胆石的形成机制。方法:收集胆石病人胆囊胆汁26份和非胆石病人胆囊胆汁17份, 测定胆汁中的凝血酶原片段F₁₊₂抗原、凝血因子Ⅷ抗原、纤维蛋白原抗原、抗凝血酶Ⅲ抗原和活性、组织因子抗原、组织因子途径抑制物抗原、蛋白C抗原、蛋白C活性、总蛋白S和游离蛋白S。结果:胆石组胆汁中纤维蛋白原抗原、凝血因子Ⅷ抗原、抗凝血酶Ⅲ抗原和组织因子抗原均高于非胆石组(P＜0.01);胆石组凝血酶原片段F₁₊₂抗原、抗凝血酶Ⅲ抗原与活性之比高于非胆石组(P＜0.05), 抗凝血酶Ⅲ活性低于非胆石组(P＜0.05);两组间组织因子途径抑制物抗原和蛋白C抗原无显著差异。结论:组织因子是胆石病人胆囊胆汁中凝血活性亢进的启动因子;胆石病人胆囊胆汁中凝血活性增高, 但抗凝活性并没有相应增强。     

组织因子的分子结构,转录调控及临床意义

组织因子(tissue factor,TF)又称凝血因子Ⅲ,是外源性凝血系统的启动因子,机体内活性最强的促凝物质之一。它在钙离子、磷脂存在的条件下,能与第Ⅶ因子(FⅦ)结合成复合体,同时使FⅦ裂解为有活性的FⅦa。TF-FⅦa复合体迅速活化X和Ⅸ因子,从而启动凝血系统。 九十年代以来,特别是发现组织因子途径抑制物(tissue factor pathway inhibitor,     

遗传性凝血因子ⅩⅢ缺乏症的研究进展

凝血因子ⅩⅢ(FⅩⅢ)是催化凝血过程最后一步反应的凝血因子,在止血过程中起重要作用。FⅩⅢ缺乏将导致凝血块不稳定,引起出血。近年来的研究发现FⅩⅢA链和B链基因突变影响FⅩⅢmRNA、蛋白质的的表达及稳定性,引起FⅩⅢ缺乏。本文就FⅩⅢ基因突变与遗传性FⅩⅢ缺乏症关系作一综述。     

内原性凝血及抑制的动力学模型与模拟

目的 研究正反馈和负反馈反应在凝血放大中的动力学作用。方法 运用模型化方法就凝血的中心反应建立动力学模型。结果 当β≥β0(存在但不易由模拟结果得到)时系统才有正平衡解；逐渐增加κ2的值，发现因子Xa的浓度从轻微的振荡到一个正平衡解改变为逐渐增加到一个正平衡解；当κ3超出正常生理意义的8倍，系统开始出现极限环解。结论 很显然Xase必须大于一个阈值凝血反应才能放大，此阈值随APC浓度的增加而增大。通过进一步分析系统的震荡及周期性与APC的关系，发现震荡有可能发生在病理情况下，另外根据数学动力学原理，尝试着给出了血友病治疗的理论依据。     

对接触系统的再认识

目前人们已认识到接触系统在启动体内凝血方面所起的作用微乎其微，而接触系统蛋白的缺乏却可能是血栓形成的危险因素。研究表明活化的因子Ⅻ(FⅫa)和活化的因子Ⅺ(FⅪa)可裂解纤溶酶原；激肽释放酶能将前尿激酶活化为双链结构的尿激酶；高分子量激肽原和低分子量激肽原能选择性抑制凝血酶诱导的血小板活化；高分子量激肽原可通过置换粘附蛋白抑制细胞粘附，可通过抑制内皮细胞的迁移和增殖抑制血管新生；内毒素可激活接触系统，高分子量激肽原的下降和α2巨球蛋白．激肽释放酶复合物的上升与不可逆低血压的发生显著相关。总之，接触系统在促进纤维蛋白溶解、抗血栓、抗粘附、抑制新生血管形成、介导炎症反应等病理生理方面起着重要作用。     

遗传性凝血因子XⅢ缺乏症的研究进展

凝血因子XⅢ（FXⅢ）是催化凝血过程最后一步反应的凝血因子，在止血过程中起重要作用。FXⅢ缺乏将导致凝血块不稳定，引起出血。近年来的研究发现FXⅢA链和B链基因突变影响FXⅢmRNA、蛋白质的的表达及稳定性，引起FXⅢ缺乏。本文就FXⅢ基因突变与遗传性FXⅢ缺乏症关系作一综述。     

内毒素致弥散性血管内凝血的研究进展

内皮细胞损伤在内毒素致DIC发病中起关键作用 ,内毒素致DIC中凝血系统的激活主要是由组织因子途径介导 ,凝血系统和炎症反应相互作用 ,纤溶系统出现先激活后抑制的双相反应。抗凝血酶与凝血酶调节蛋白等特异性分子标志物的检测可用于早期DIC的诊断 ,双相APTT波形是一敏感指标。活化蛋白C可能是一种新的治疗内毒素DIC的有效药物     

蛋白质C表位附加系统

1　蛋白质 C表位附加系统可以做到以下几点 :1 .1　用有效钙螯合剂或蛋白质 C多肽来洗脱标记蛋白。1 .2　在 Western印记中保证单一带的纯度。1 .3　用单一抗体 C末端或 N末端识别蛋白质C标记。1 .4　无论任何表达系统都能保证较高的特异性。1 .5　不必将钙结合位点导入蛋白质即可保证钙依赖性抗体的结合。蛋白质 C表位附加系统 ( Roche)是唯一允许快速纯化标记融合蛋白并可保证高度特异性和纯度的系统。这种表位标记抗原由蛋白质 C衍生而来 ,蛋白质 C是一种 Vitamin K依赖的原生质酶原 ,它由凝血调节蛋白复合物裂解激活为一种抗促凝酶…     

The role of complex formation and epidermal growth factor-like domains in the regulation of blood coagulation by the thrombomodulin-protein C system

The explosive nature of the coagulation cascade led many scientists to investigate how it is regulated. Proteinase inhibitors such as antithrombin III inhibit active proteases of the coagulation cascade. Cofactors such as factor Va and factor VIIIa are proteolytically inactivated by activated protein C. Protein C is activated by the thrombin-thrombomodulin complex on the endothelial cell surface. Thus, the independent actions of the proteinase inhibitor system and the thrombomodulin-protein C system complement each other to maintain regulation of blood coagulation. The thrombin binding site of thrombomodulin was identified to be the fifth and sixth repeats of the epidermal growth factor-like domain. The same binding template contains sufficient information to block the functions of thrombin as a procoagulant. However, additional repeats are required for the activation of protein C. Thrombomodulin is the first example which illustrates that the epidermal growth factor-like domain functions as a binding template for thrombin and as a switch to turn off the procoagulant activity of thrombin as well as to trigger the protein C anticoagulant pathway. Epidermal growth factor-like structures are found in many of the coagulation factors. Complex formation is a repeated theme not only in the blood coagulation cascade but also in the thrombomodulin-protein C anticoagulant pathway.     

Dynamic model of the role of platelets in the blood coagulation system

In order to confirm which process is the most important in the blood coagulation cascade, a dynamic model of the function of platelets in blood coagulation is provided based on biochemical experiments. A series of conclusions based on qualitative analysis and mathematical simulation are drawn about the influence of the activation rate of factor VIII and factor IX on the generation of thrombin (IIa). It is evident that the pro-coagulation stimulus must exceed a threshold value to initiate the coagulation cascade. The value is related to the rate of platelet activation, the binding constant d2. The stability of the fixed value is also related to the pro-coagulation stimulus. This article also evaluates the influence of the stimulus strength and the activated rate parameter of platelets on thrombin. The proportion of platelets activated at any given time is designated c. To each c, we obtain a maximum concentration of thrombin. It is evident that when the level of factor IX is below 1% of normal levels, the rate of thrombin generation reduces dramatically resulting in severe bleeding tendency.     

Whole blood coagulation on protein adsorption-resistant PEG and peptide functionalised PEG-coated titanium surfaces

The aim of this study was to investigate whole blood coagulation on low blood plasma protein adsorbing surfaces. For this purpose, the polycationic graft copolymer poly(L-lysine)-g-poly(ethylene glycol) (PLL-g-PEG), PLL-g-PEG grafted with a cell adhesive peptide containing the amino acid sequence -Arg-Gly-Asp- (RGD), and PLL-g-PEG with a control peptide -Arg-Asp-Gly- (RDG) were adsorbed onto titanium (oxide), forming stable monomolecular adlayers through electrostatic attraction. Free oscillation rheometry and complementary techniques were used to measure the coagulation time (CT) and other interactions of the surfaces with native whole blood, recalcified platelet-rich plasma (PRP), and recalcified citrated platelet-free plasma (PFP). The results show that the uncoated titanium surfaces (reference) activated platelets and quickly triggered the coagulation cascade via the intrinsic pathway, whereas the PLL-g-PEG surfaces displayed a prolonged CT, approximately 2-3 times longer compared to uncoated titanium. We hypothesise that blood coagulates outside the vascular system independent of low protein adsorption to or activation by surfaces, due to the absence of an active down-regulation of procoagulative processes by the vascular endothelium.     

Stochastic modeling of blood coagulation initiation

A kinetic Monte Carlo simulation was developed using the deterministic reaction network developed by the Mann laboratory for tissue-factor (TF)-initiated blood coagulation. The model predicted thrombin dynamics in recalcified whole blood (3-fold diluted) pretreated with convulxin (platelet GPVI activator) and picomolar levels of TF (0-14 pM). The model did not accurately predict coagulation times at low TF (0-0.7 pM). The simulation revealed that approximately 0.2 pM TF was the critical concentration to cause 50% of reactions containing 3-fold diluted whole blood to reach a clotting threshold of 0.05 U/ml thrombin by 1 h. Simulations of 1 nl of blood (5 pM TF) revealed small stochastic variations in thrombin initiation time, while 16.6 pl simulations were highly stochastic at this level of TF (50 molecules/16.6 pl). Further experiment and simulation will require evaluation of mechanisms of coagulation kinetics at subpicomolar levels of TF.     

Adsorption of coagulation proteins from whole blood on to polymer materials: relation to platelet activation.

A combination of methods, immunoassays of plasma proteins and platelet release of beta-thromboglobulin and chromogenic substrates for enzymatically active coagulation factors, was used to measure the reactions of coagulation proteins upon contact between whole blood and artificial surfaces as a function of time and surface material. Four types of well-known polymer surfaces, polyvinylchloride, polytetrafluoroethylene, polyurethane and silicone rubber, were investigated to elucidate if a simple and fast in vitro experimental set-up can be of guidance in the selection of materials for use in vivo. Platelets were activated at the polymer surfaces whereas the coagulation enzymes showed little activity on the polymer surfaces tested. There was a correlation between the adsorption of adhesins (fibrinogen, fibronectin and factor VIII-related antigen) at the surfaces and the release of beta-thromboglobulin from platelets, suggesting that adsorption of adhesins is a major determinant of blood compatibility of polymer materials. Significant differences between the surfaces were seen--polyurethane being the surface with the least protein adsorbed and least platelet activation initiated. This study shows that it is possible to make a first in vitro choice of possible blood compatible artificial surfaces before expensive and cumbersome in vivo experiments.     

Thrombin: a multifunctional enzyme

Thrombin is the final enzyme of blood coagulation cascade. It belongs to the trypsin family of serine proteases. Its two primary actions are to cleave fibrinogen to release fibrin and to activate platelets through a limited proteolysis of a specific receptor. In addition, thrombin is the major regulator of blood coagulation. It is both a procoagulant enzyme in the activation of factors V and VIII, and an anticoagulant enzyme through the activation of protein C and TAFI. This multi-functionality of thrombin depends upon the conformation of its active site: depth for high specificity and shape for a finely tuned selection of substrates. Since new anticoagulant molecules, some with anti-thrombin activity, are emerging, it is important to understand the mechanisms allowing thrombin to be so specifically multifunctional.     

In vitro blood compatibility of polymeric biomaterials through covalent immobilization of an amidine derivative

We present a surface coating with anticoagulant characteristics showing significantly reduced coagulation activation. The synthesis of a monomeric conjugate containing a benzamidine moiety was carried out and its inhibitory activity against human thrombin, the key enzyme of the blood coagulation cascade, was determined using a chromogenic assay. Based on that, low-thrombogenic interfaces were prepared by covalent attachment of this low-molecular weight thrombin inhibitor on poly(octadecene-alt-maleic anhydride) copolymer thin films and characterized using ellipsometry, XPS and dynamic contact angle measurements. The in vitro hemocompatibility tests using freshly drawn human whole blood showed, in agreement with the SEM images, that a PO-MA film modified with a benzamidine moiety using a PEG spacer decreased the activation of coagulation, platelets and the complement system. The decreased protein adsorption, in addition to the specific inhibition of thrombin, effectively enhanced the short-term hemocompatibility characteristics.     

Lipopolysaccharide-binding proteins of Limulus amebocyte lysate.

Limulus amebocyte lysate, obtained from horseshoe crab (Limulus polyphemus) blood cells, contains a coagulation system which is activated by bacterial lipopolysaccharide (LPS). A chromatographic fraction of Limulus lysate, containing the endotoxin-sensitive factor(s) which initiates the coagulation cascade, was studied. We utilized a photoreactive, cleavable, radiolabeled derivative of Salmonella minnesota LPS, LPS-(p-azidosalicylamido)-1,3'-dithiopropionamide (LPS-ASD), to identify LPS-binding proteins. The lysate fraction was incubated with LPS-ASD, and LPS-binding proteins were identified by autoradiography of sodium dodecyl sulfate-polyacrylamide gels. An 82-kDa protein, a major protein component of this fraction from Limulus lysate, was identified as a LPS-binding protein in a majority of lysates. Incubation of whole Limulus lysate with antiserum to this protein resulted in enhanced sensitivity of the lysate to LPS, suggesting that this 82-kDa protein is a negative regulator of coagulation. A minor 50-kDa protein component of lysate also was identified as a LPS-binding protein and is a candidate for the LPS-sensitive coagulation protein in L. polyphemus.     

Thrombin generation measurement in factor VII-depleted plasmas compared to inherited factor VII-deficient plasmas

Activated factor VII (FVIIa)/tissue factor enzyme complex is the initiator of the coagulation cascade in vivo. FVIIa is of particular interest because it has been found to induce haemostasis in various bleeding disorders. In order to evaluate the FVII threshold that is required to initiate the clotting cascade, we measured thrombin generation in FVII-depleted plasmas spiked with increasing amounts of normal pooled plasma and in inherited FVII-deficient plasmas. According to the literature, only trace amounts of FVII are sufficient to initiate blood coagulation in vitro. By contrast, results on inherited FVII-deficient plasmas showed a wide variety of the amounts of thrombin generated in plasmas with the same FVII coagulant activity levels. This suggests that the threshold of FVII required to initiate haemostasis in vivo depends on one or more, hitherto unknown, plasmatic or cellular factors.     

Mathematical modeling and computer simulation in blood coagulation

Over the last two decades, mathematical modeling has become a popular tool in study of blood coagulation. The in silico methods were able to yield interesting and significant results in the understanding of both individual reaction mechanisms and regulation of large sections of the coagulation cascade. The objective of this paper is to review the development of theoretical research in hemostasis and thrombosis, to summarize the main findings, and outline problems and possible prospects in the use of mathematical modeling and computer simulation approaches. This review is primarily focused on the studies dealing with: (1) the membrane-dependent reactions of coagulation; (2) regulation of the coagulation cascade, including effects of positive and negative feedback loops, diffusion of coagulation factors, and blood flow.