内皮细胞损伤标志物在妊高征患者中的变化

目的：探讨孕妇血浆中内皮细胞损伤标志物内皮素（ET－1）、血栓调节蛋白（TM）、血管性假血友病因子（vWF）水平变化与妊高征（PIH）发病的关系。方法：应用放射免疫吸附法和酶联免疫吸附法，分别测定66例妊高征患者（轻度20例，中度24例，重度22例）和24例正常晚期孕妇（对照组）血浆中的ET－1、TM、vWF水平。结果：PIH各组ET－1水平明显高于对照组（P＜0．01），轻度PIH患者TM、vWF水平与对照组比较，差异无显著性（P＞0．05），中度PIH患者TM、vWF水平与对照组比较，差异有显著性（P＜0．05），重度PIH患者TM、vWF水平与对照组比较，差异有非常显著（P＜0．01）。结论：血管内皮细胞损伤是妊高征的重要发病机制。3个分子标志物水平升高越显著妊高征患者病情就越重，检测其水平对妊高征的诊断，观察病情及防止子痫的发生均有意义。     

妊娠高血压综合征患者血浆血管性假血友病因子及血栓调节蛋白水平的变化

目的 探讨孕妇血浆血管性假血友病因子(vWF)及血栓调节蛋白(TM)水平变化与妊娠高血压综合征(PIH)发病的关系.方法 应用酶联免疫吸附试验(ELISA)分别测定38例PIH患者(PIH组)及16例正常妊娠妇女(正常妊娠组)血浆中vWF及TM水平,并以20名健康未孕妇女(正常未孕组)为对照.结果 正常妊娠组vWF(95.4±40.2)%,轻度PIH患者血浆vWF为(98.1±41.1)%.二者比较,无显著性差异(P＞0.05);正常妊娠组TM为(13.7±4.6)ng/mL,轻度PIH患者为(17.4±7.1)n2/mL,二者比较,有显著性差异(P＜0.05).中、重度PIH患者血浆vWF为(195.6±39.8)%,TM为(21.8±7.4)ng/mL,与正常妊娠组比较,有显著性差异(P＜0.01).PIH患者血浆vWF与TM呈显著正相关.结论 vWF和TM可作为判断PIH病情程度的指标;vWF与TM在PIH发病中起相互协同作用.     

妊娠糖尿病孕妇血管性假血友病因子检测的意义

目的探讨妊振糖尿病(GDM)孕妇血浆血管性假血友病因子(vWF)变化及意义。方法采用酶联免疫吸附测定(ELISA)法检测20例GDM、32例正常妊娠及22例非妊娠妇女血浆vWF浓度。结果与非妊娠对照组相比,正常妊娠组空腹血浆vWF浓度明显升高,GDM组血浆vWF浓度又显著高于正常妊娠组及非妊娠对照组[(329.43±83.67)%vs(205.14±49.25)%vs(108.62±37.58)%],其差异均有统计学意义(P<0.001)。结论GDM孕妇存在较正常孕妇更为严重的血管内皮功能受损。     

正常妊娠和妊娠高血压综合征孕妇舌深静脉观测分析

对100例育龄非孕妇女、64例正常晚期妊娠妇女及90例妊娠高血压综合征患者进行舌深静脉观测，以探讨正常妊娠及妊娠高血压综合征（简称妊高征）妇舌深静脉变化。结果：正常晚期妊娠孕妇合深静脉图象与非妊娠妇女比较有显著差异（P＜0．01），妊高征妇女舌深静脉图象与正常妊娠妇女及非孕妇女比较均有显著差异（P＜0．01），重度妊高征妇女舌深静脉图象与轻度妊高征比较，有显著差异（P＜0．01）。表明：正常晚期妊娠及妊高征孕妇舌深静脉有变化。对探讨正常妊娠妇女和妊高征患者血液流变学变化具有意义，亦可辅助中医望诊。     

妊娠高血压综合征患者血浆纤维结合蛋白含量的变化

目的探讨妊娠高血压综合征（PIH）患者血浆纤维结合蛋白（Fn）含量的变化情况。方法用免疫透射比浊法检测43例妊高征患者（轻度10例,中度19例,重度14例）、50名正常妊娠孕妇及60名健康未孕妇女（对照组）Fn含量。结果正常未孕妇女血浆Fn均值为239±61mg/L;正常孕妇在妊娠早期与正常未孕妇女无明显差异,妊娠中、晚期明显升高,有随孕周增加而升高的趋势。与同期正常孕妇相比,妊高征患者血浆Fn含量明显增高;与病情严重程度和孕周增加有较好的相关性。结论Fn可能参与了PIH的血管内皮损伤,观察血浆Fn含量变化有助于判断妊高征的发生、发展及预后。     

STA-R全自动血凝仪检测孕妇vWF方法的改进

用法国STA全自动血凝仪测定vWF（血管性血友病因子）活性时,血浆先进行1∶2稀释,当vWF活性＞105%时仪器自动进行1∶8稀释测定得出测试结果.妊娠时vWF活性增高,尤其是妊娠高血压综合征（妊高征）患者,一般均＞105%,因此本文建立一种新的孕妇vWF检测方法.     

妊娠高血压综合征患者血清胱抑素C测定的临床意义

目的探讨胱抑素C对妊娠高血压综合征（简称妊高征）早期肾损害的诊断价值。方法收集妊高征患者、正常晚期妊娠孕妇和健康体检妇女血清,采用免疫比浊法测定胱抑素C（CysC）和超敏C-反应蛋白（hsCRP）,用酶法测定肌酐（Ser）和尿酸（UA）。结果CysC在妊高征患者和正常晚期妊娠中含量分别为（1．90±0．52）和（1．23±0．24）mg／L,均明显高于健康体检妇女（0．77±0．15）mg／L,P〈0．05,且妊高征组CysC水平高于晚期妊娠组（P〈0．05）。结论 血清CysC水平是反映妊高征患者早期肾损害的一项敏感和可靠的指标。     

慢性肾脏病患者血浆血管性血友病因子裂解酶活性测定

目的检测慢性肾脏病（CKD）患者血浆血管性血友病因子裂解酶（vWF-CP）活性及血浆血管性血友病因子（vWF）水平,探讨其在慢性肾脏病的意义。方法将139例CKD患者根据不同种类肾病及不同肾功能状态分组,其中狼疮肾炎（LN）31例,原发性肾病综合征（NS）25例,糖尿病肾病（DN）45例及慢性肾炎（CGN）38例;CKDⅠ期38例,CKDⅡ期30例,CKDⅢ期39例,CKDⅣ期19例,CKDⅤ期13例。采用残余胶原结合实验法及ELISA法对患者血浆vWF-CP活性及vWF水平进行检测,观察不同组患者血浆vWF-CP活性及vWF水平,并与正常对照组（40名）比较。结果LN、NS、DN及CGN患者血浆vWF-CP活性显著低于对照组,且以LN、NS组下降最明显,vWF水平均显著高于正常对照组,血浆vWF含量均与血浆vWF-CP活性呈负相关。CKDⅡ期、Ⅲ期血浆vWF-CP活性显著低于Ⅰ、Ⅳ、Ⅴ期．而Ⅰ、Ⅳ、Ⅴ期之间无显著区别,vWF含量各期之间无显著差别。结论CKD患者血浆vWF-CP活性明显降低,其低下程度与肾病的病种有关。内皮损伤、自身抗体可能是致CKD患者vWF-CP活性低下的原因之一。它参与了CKD的发生发展,可能是CKD血栓形成的机理之一。     

两种常见妊娠合并症的血小板参数变化分析

目的：探讨妊高征和妊娠合并慢性肾炎两种异常妊娠合并症的血小板（Plt）参数变化意义。方法：用Sysmex-SE9500全自动血细胞分析仪对102例妊高征、26例妊娠合并慢性肾炎妇女及69例正常妊娠妇女的外周静脉血标本进行Plt、血小板平均体积（MVP）、血小板分布宽度（PDW）和大血小板（P－LCR）4项参数测定。结果：妊高征组的Plt比正常妊娠对照组减少（P＜0．01），而MPV、PDW、P－LCR三项参数均高于对照组（其中MPV、PDW的P＜0．01，P－LCR P＜0．05）；妊娠并慢性肾炎组的Plt、MPV、PDW、PLCR均低于正常妊娠对照组（其中Plt、MPV P＜0．01，PDW、P－LCR P＜0．05）。结论：Plt参数可有效反映妊高患者的Plt破坏程度和凝血异常程度。而且在判断妊高症病程方面有重要意义；观测Plt4参数还可为临床鉴别此两种常见病理妊娠提供有效依据。     

妊娠高血压综合征患者血浆纤维蛋白溶解系统变化的初步观察

目的：观察妊高征患者血浆纤维蛋白溶解系统的变化。方法：测定20例正常孕妇及15例妊局征患者血浆中D-二聚体（D－D）、纤维蛋白降解产物（FDP）、纤维蛋白原（Fg）含量、组织型纤溶酶原激活物活性（t－PA：A）、α2－纤溶酶抑制物活性（α2－PI：A）。结果：非孕妇与正常孕妇、正常孕妇与好高征患者之间D—D含量有明显差异（P＜0．01,P＜0．001）;好高征患者FDP阳性率（100％）明显高于正常妊娠组（15％）;非孕妇与正常孕妇及好高征患者间Fg含量有明显差异（P＜0.01）;非孕妇与正常孕妇之间t－PA:A无差异,但与妊高征之间有明显差异（P＜0.001）,α2－PI：A则无组间差异。结论：正常孕妇处于高凝状态,妊高征者有血栓形成倾向。本文所检测的指标,尤其D－D、t－PA：A和α2－PI：A为纤溶系统分子标志物,可作为监测妊高征纤溶及凝血变化的重要指标。     

Von Willebrand factor and cancer; metastasis and coagulopathies

Von Willebrand factor (VWF) is a multimeric procoagulant plasma glycoprotein that mediates platelet adhesion along the endothelium. In addition to its role maintaining normal hemostasis, more recently novel biological functions for VWF have been described, including inflammation, angiogenesis, and metastasis. Significantly increased plasma VWF levels have been reported across a variety of cancer patient cohorts. Given that VWF is established as a risk factor for venous thrombosis, this is of direct clinical importance. Moreover, elevated VWF has also been observed localized within the tumor microenvironment, correlating with advanced disease stage and poorer clinical outcome. Critically, evidence suggests that elevated VWF levels in cancer patients may not only contribute to cancer associated coagulopathies but may also mediate cancer progression and metastasis. Studies have shown that VWF can promote pro‐inflammatory signaling, regulate angiogenesis and vascular permeability, which may facilitate tumor cell growth and extravasation across the vessel wall. Endothelial secreted VWF multimers contribute to the adhesion and transendothelial migration of tumor cells key for tumor dissemination. In support of this, VWF inhibition attenuated metastasis in vivo. Perhaps most intriguingly, specific tumor cells have been reported to acquire de novo VWF expression which increases tumor‐platelet heteroaggregates and confers enhanced metastatic activity. Current knowledge on the roles of VWF in cancer and in particular its contribution to metastasis and cancer associated coagulopathies is summarized in this review.     

Reduced von Willebrand factor survival in von Willebrand disease: pathophysiologic and clinical relevance

Summary.  Von Willebrand disease (VWD) is characterized by a wide heterogeneity of clinical and laboratory phenotypes. The complexity of the phenotype is further increased by a highly variable removal rate of von Willebrand factor (VWF) released by desmopressin, which is independent of post-infusion peak level. After the initial demonstration that a reduced VWF survival is present in patients with R1205H mutation (VWD Vicenza), several other mutations, mostly occurring in the VWF D3 domain, have been shown to be associated with accelerated removal of released VWF. Normal subjects with O blood group show reduced survival after desmopressin, underlining the role of different VWF glycosylation present in ABO blood group. Recent evidence suggests that liver and spleen macrophages are responsible for VWF clearance through uptake and endocellular degradation, but it is still not known why some VWF mutants are more prone to increased clearance.     

人vW因子A2区基因表达及其生物学活性的研究

血管性血友病因子（vWF）在血管性血友病（vWD）以及血栓性血小板减少性紫癜（TTP）的发病中起着重要的作用,并受血浆vWF裂解蛋白酶（ADAMTS-13）的调节.本研究表达vWF中的A2区蛋白,并研究ADAMTS-13对其裂解活性.首先,应用基因重组技术在大肠杆菌中表达人vWF-A2区蛋白,经纯化、复性获得重组蛋白（rvWF-A2）,并与正常人以及TYP患者血浆进行反应,分析rvWF-A2蛋白的生物学活性.结果表明：重组表达载体pQE-30-vWFA2在大肠杆菌M15中得到高效表达,目的蛋白表达量占菌体总蛋白的42%,Ni-NTA agrose柱层析纯化后,其纯度为98%,经复性的rvWF-A2可作为ADAMTS-13良好的酶解底物,可被枸橼酸钠抗凝的正常人血浆切割,其切割程度随时间的延长而增加,而EDTA抗凝的正常人血浆以及TTP患者血浆无法切割此蛋白.结论：在大肠杆菌中高效表达的rvWF-A2具有良好的生物学活性,为建立定量测定ADAMTS-13活性的方法奠定了坚实的基础.     

Laboratory diagnosis of von Willebrand disease

Von Willebrand disease is the most-common inherited bleeding disorder, including both quantitative (types 1 and 3) and qualitative (type 2) defects of von Willebrand factor. Among patients with suspected von Willebrand disease, the laboratory diagnosis requires three levels of testing: screening tests, specific assays for von Willebrand factor to establish the diagnosis, and discriminating tests to allow accurate characterization of the numerous types and subtypes of the disease. Because of their poor sensitivity, normal screening tests do not exclude the diagnosis. In most cases, specific measurements of von Willebrand factor antigen, von Willebrand factor ristocetin cofactor activity, and factor VIII levels in plasma allow differentiation of quantitative (proportionately decreased levels) and qualitative (discrepant levels) deficiencies of von Willebrand factor. Among the latter, a decreased von Willebrand factor ristocetin cofactor activity/von Willebrand factor antigen ratio is in favor of the three subtypes (2A, 2M, and 2B) defined by an abnormal interaction between von Willebrand factor and platelet glycoprotein Ib, whereas a decreased factor VIII/von Willebrand factor antigen ratio suggests subtype 2N, defined by a defective binding of von Willebrand factor to factor VIII. Several discriminating tests are available to definitively characterize each subtype. Moreover, for all variants, the link between phenotype and genotype is established using DNA analysis. In all cases, the precise characterization of type and subtype of von Willebrand disease remains essential for the choice of optimal therapeutic monitoring of each patient. Presented at the Joint Meeting of the World Health Organization and the International Society for Thrombosis and Hemostasis “Impact, Prevention and Control of von Willebrand’s disease” London, October 12–14, 1998     

Treatment of severe von Willebrand disease with a high-purity von Willebrand factor concentrate (Wilfactin®): a prospective study of 50 patients

BACKGROUND AND OBJECTIVES: A plasma-derived von Willebrand factor (VWF) concentrate with low factor VIII (FVIII) content was specifically developed to treat von Willebrand disease (VWD). Efficacy and safety were investigated by merging the results of two comparable protocols conducted prospectively in 5 European and 12 French centers. METHODS AND RESULTS: Fifty patients with clinically severe VWD (72% had VWF ristocetin cofactor activity less than 10 IU dL(-1) and 46% had FVIII < 20 IU dL(-1)) were treated with the concentrate as the only therapy, except for clinical situations requiring a priming dose of FVIII to rapidly correct an intrinsic coagulation defect. A total of 139 spontaneous bleeding episodes were treated; only 53 (38%) needed a concomitant FVIII dose. Outcome was excellent or good in 89% of the episodes. Forty-four patients underwent 108 surgical or invasive procedures. Outcome was excellent or good in 95 scheduled procedures (only VWF was infused) and 13 emergency procedures (a priming FVIII dose was co-administered with the first VWF infusion). There were no thrombotic complications and none of the 18 patients with type 3 VWD developed anti-VWF or anti-FVIII antibodies. CONCLUSIONS: This concentrate safely and effectively provides hemostasis in patients with clinically severe VWD.     

血管性血友病诊断与分型方法的建立和临床应用

目的 建立系列的血管性血友病（vWD）筛查、诊断和分型的方法。方法 以Ⅲ型胶原包被微孔板，以辣根过氧化物酶（HRP）-兔抗人血管性血友病因子（vWF）IgG为检测抗体，建立vWF胶原结合试验（vWF：CB）；通过垂直式琼脂糖电泳、电转印和化学发光法对vWF多聚体进行分析。对可疑vWD患者测定血小板、出血时间、活化部分凝血活酶时间（Am）、血小板聚集试验（RIPA）、FVⅢ：C、vWF：RCo、vWF：Ag、vWF：CB，并进行多聚体分析。结果 所建vWF：CB线性范围广，灵敏度高，重复性好；所建多聚体琼脂糖电泳分析方法安全、敏感、解像度高。10例可疑病例血小板均正常，大部分出血时间延长、APTY延长、RIPA反应低下、FVⅢ：C降低；vWF：RCo、vWF：Ag含量和vWF：CB均不同程度降低；部分患者vWF：Ag／vWF：CB比值〉2，中、高分子量vWF显著缺如。8例患者可明确诊断为vWD，其中1型2例、2A型4例和3型2例。结论 本研究建立的方法和推荐的组合实验适合于绝大多数vWD患者的诊断和分型需要。     

The von Willebrand factor

Summary Von Willebrand factor (vWf) is a multimeric and multivalent adhesive protein which is essential for platelet adhesion to subendothelium and for stabilization of factor VIII procoagulant activity in circulation. The quantitative measurement of vWf involves essentially two different approaches. The first is based on the interaction between vWf and Gp Ib of the platelet membrane in presence of ristocetin (ristocetin cofactor activity, RiCof) and depends not only on the amount of the factor but also on its ability to bring about this interaction, large multimers being more active. The second approach involves the immunological quantitation of vWf (vWf:Ag) by its interaction with specific polyclonal or monoclonal antibodies as measured by several methods, i.e., electroimmunoassay, immunoradiometric assay and immunoenzymatic assay. Although in the majority of type II von Willebrand disease (vWd) with dysfunctional vWf there is a discrepancy between RiCof and vWf:Ag, it should be emphasized that RiCof activity does not entirely reflect the ‘true’ activity of vWf since it does not explore all the functions of this factor; furthermore, the relationship between degree of multimerization and RiCof level is not always tenable, as for example in vWd ‘Vicenza’. For the diagnosis of congenital and acquired vWd RiCof assay together with family investigation is the eligible test, with an estimated ability to detect at least 50% of the carriers of the abnormal gene, including mildly affected patients; vWf:Ag appears less sensitive and, on the basis of studies carried out in our laboratory, a relative sensitivity of 64% is proposed. Both assays require the definition of separate normal ranges for children and adults and for 0 and non-0 blood group subjects; a nonparametric approach in a large sample of normal subjects is advisable. With RiCof assay performed by an aggregometric method using formalin-fixed platelets an interassay variability of 6% and 8.5% respectively for high- and low-control plasma was found in our laboratory. With vWf:Ag assayed by an ELISA method a variability of 7% for low- and 6% for high-control plasma was found. Thus, both methods appear sufficiently precise for clinical use. The use of an internal pool calibrated against an international standard allows to perform comparable interlaboratory measurements. To further improve standardization of these assays, collaborative studies seem urgently required. Presented at the ‘2nd International Symposium on Standardization and Quality Control of Coagulation Tests: Implications for the Clinical Laboratory’, Rome, September 28–29, 1989.     

Evaluation of a microfluidic flow assay to screen for von Willebrand disease and low von Willebrand factor levels

Essentials

• von Willebrand factor (VWF) function is shear stress dependent.
• Platelet accumulation in a microfluidic assay correlates with VWF levels.
• The microfluidic assay discriminates type 1 von Willebrand disease from healthy controls.
• The microfluidic flow assay detects responses to therapeutic intervention (DDAVP).

Summary

Background

von Willebrand disease (VWD) is a mucocutaneous bleeding disorder with a reported prevalence of 1 in 10 000. von Willebrand factor (VWF) function and platelet adhesion are regulated by hemodynamic forces that are not integrated into most current clinical assays.

Objective

We evaluated whether a custom microfluidic flow assay (MFA) can screen for deficiencies in VWF in patients presenting with mucocutaneous bleeding.

Methods

Whole blood from individuals with mucocutaneous bleeding was assayed in a custom MFA.

Results

Thirty‐two patients with type 1 VWD (10/32) or reported mucocutaneous bleeding were enrolled. The platelet adhesion velocity (r = 0.5978 for 750 s^?1 and 0.6895 for 1500 s^?1) and the maximum platelet surface area coverage (r = 0.5719 for 750 s^?1 and 0.6633 for 1500 s^?1) in the MFA correlated with VWF levels. Furthermore, the platelet adhesion velocity at 750 s^?1 (type 1 VWD, mean 0.0009761, 95% confidence interval [CI] 0.0003404–0.001612; control, mean 0.003587, 95% CI 0.002455–0.004719) and at 1500 s^?1 (type 1 VWD, mean 0.0003585, 95% CI 0.00003914–0.0006778; control, mean 0.003132, 95% CI 0.001565–0.004699) differentiated type 1 VWD from controls. Maximum platelet surface area coverage at 750 s^?1 (type 1 VWD, mean 0.1831, 95% CI 0.03816–0.3281; control, mean 0.6755, 95% CI 0.471–0.88) and at 1500 s^?1 (type 1 VWD, mean 0.07873, 95% CI 0.01689–0.1406; control, mean 0.6432, 95% CI 0.3607–0.9257) also differentiated type 1 VWD from controls. We also observed an improvement in platelet accumulation after 1‐desamino‐8‐d ‐arginine vasopressin (DDAVP) treatment at 1500 s^?1 (pre‐DDAVP, mean 0.4784, 95% CI 0.1777–0.7791; post‐DDAVP, mean 0.8444, 95% CI 0.7162–0.9726).

Conclusions

These data suggest that this approach can be used as a screening tool for VWD.