Blood coagulation and fibrinolysis in patients with acromegaly: increased plasminogen activator inhibitor-1 (PAI-1), decreased tissue factor pathway inhibitor (TFPI), and an inverse correlation between growth hormone and TFPI

Objective Growth hormone/insulin-like growth factor-1(GH/IGF-1) hypersecretion may influence risk factors contributing to the increased cardiovascular morbidity and mortality associated with acromegaly However, so far little is known about the impact of GH/IGF-1 on coagulation and fibrinolysis in acromegalic patients as possible risk factors for cardiovascular disease (CVD). To our knowledge, plasma tissue factor pathway inhibitor (TFPI) and thrombin-activatable fibrinolysis inhibitor (TAFI) levels in these patients have not been investigated. Therefore, the main purpose of this study was to evaluate the markers of endogenous coagulation/fibrinolysis, including TFPI and TAFI, and to investigate the relationships between GH/IGF-1 and these hemostatic parameters and serum lipid profile in patients with acromegaly. Research Methods and Procedures A total of 22 patients with active acromegaly and 22 age-matched healthy controls were included in the study. Fibrinogen, factors V, VII, VIII, IX, and X activities, von-Willebrand factor (vWF), antithrombin III (AT III), protein C, protein S, tissue plasminogen activator (t-PA), tissue plasminogen activator inhibitor-1 (PAI-1), TFPI and TAFI, as well as common lipid variables, were measured. The relationships between serum GH/IGF-1 and these hemostatic parameters were evaluated. Results Compared with the control subjects, fibrinogen, AT III, t-PA, and PAI-1 were increased significantly in patients with acromegaly (P < 0.0001, P < 0.05, P < 0.01, and P < 0.0001, respectively), whereas protein S activity and TFPI levels were decreased significantly (P < 0.05 and P < 0.01, respectively). Plasma TAFI Ag levels did not significantly change in patients with acromegaly compared with the controls. In patients with acromegaly, serum GH levels were inversely correlated with TFPI and apo AI levels (r: −0.514, P: 0.029 and r: −0.602, P: 0.014, respectively). There was also a negative correlation between insulin-like growth factor −1 (IGF-1) and PAI-1 (r: −0.455, P:0.045). Discussion We found some important differences in the hemostatic parameters between the patients with acromegaly and healthy controls. Increased fibrinogen, t-PA, PAI-1 and decreased protein S and TFPI in acromegalic patients may represent a potential hypercoagulable and hypofibrinolytic state, which might augment the risk for atherosclerotic and atherothrombotic complications. Thus, disturbances of the hemostatic system and dyslipidemia may contribute to the excess mortality due to CVD seen in patients with acromegaly.     

凝血酶激活的纤溶抑制物与肺部疾病的关系

凝血酶激活的纤溶抑制物是近年发现的血浆羧肽酶原,在调节血浆凝块的纤溶过程中有着重要的作用,而最近研究发现,其生理功能并不限于控制纤溶系统,还有更广泛的抗炎作用.现就凝血酶激活的纤溶抑制物与肺血栓栓塞症、ALI、肺间质纤维化、支气管哮喘等肺部疾病的相关性作一综述.     

冠心病患者血浆组织因子和组织因子途径抑制物的变化

目的 :通过检测不同类型冠心病 (CHD)患者血浆组织因子 (TF)和组织因子途径抑制物 (TFPI)水平变化 ,探讨其在CHD发病过程中的作用。方法 :以酶联免疫吸附测定法测定CHD患者血浆中TF和TFPI抗原水平。结果 :不稳定型心绞痛 (UAP)和急性心肌梗死 (AMI)患者的血浆TF和TFPI水平与正常对照者和稳定型心绞痛 (SAP)患者相比均有显著性增高 (P <0 .0 5 ) ,以AMI患者尤为明显 (P <0 .0 1) ;UAP和AMI患者的TF PI/TF比值显著降低 (P <0 .0 5 ) ,而SAP患者的上述指标与正常对照者相比 ,其差异均无显著性意义 (P >0 .0 5 )。结论 :UAP和AMI患者TFPI/TF系统失衡 ,标志高凝状态的存在 ;TF和TFPI在这两种类型CHD的发病机制中可能起着重要的作用     

Clinical significance of expression of tissue factor and tissue factor pathway inhibitor in ulcerative colitis

AIM: To investigate the clinical significance of expression of tissue factor (TF) and tissue factor pathway inhibitor (TFPI) in ulcerative colitis (UC).METHODS: Thirty UC specimens taken by colonoscopy from patients with active UC treated at the Department of Pathology, Central Hospital Affiliated to Shenyang Medical College from February 2010 to January 2012 were included in an experimental group, and 30 normal colon tissue samples taken by colonoscopy from non-UC patients were included in a control group. Expression of TF and TFPI in UC and normal colon tissue samples was detected by immunohistochemistry.RESULTS: The positive rate of TF in UC was significantly higher than that in normal colon tissue (63% vs 33%, χ² = 5.41, P < 0.05). The positive rate of TFPI in UC was also significantly higher than that in normal colon tissue (43% vs 17%, χ² = 5.08, P < 0.05).CONCLUSION: Positive rates of TF and TFPI expression in UC are significantly higher than those in normal colon tissue. TF and TFPI may play an important role in the pathogenesis of UC.     

维生素E对组织因子及其抑制物在冠心病中的干预作用

目的 :探讨冠心病 (CHD)的不同类型中血浆组织因子 (TF)及其抑制物 (TFPI)含量的差异变化及维生素E对急性心肌梗死 (AMI)干预作用。方法 :用ELISA方法检测CHD患者 [包括稳定型心绞痛 (SAP)、不稳定型心绞痛 (UAP)、AMI]入院时及治疗 2周时血浆TF、TFPI含量。结果 :①SAP组血浆TF、TFPI水平及TF/TF PI比值均高于正常对照组 ,但差异无统计学意义 (P >0 .0 5 ) ,且治疗前后无明显变化 (P >0 .0 5 )。②UAP组、AMI组血浆TF、TFPI水平及TF/TFPI比值明显高于正常对照组 ,差异有统计学意义 (P <0 .0 1) ,但两组常规治疗前后差异无统计学意义 (P >0 .0 5 )。③AMI加维生素E组干预治疗后TF值显著下降 (P <0 .0 1) ,与正常对照组比较差异无统计学意义 (P >0 .0 5 ) ,TFPI干预治疗后无明显变化 (P >0 .0 5 ) ,而TF/TFPI比值明显降低 (P <0 .0 1)。④各组TF与TFPI呈明显正相关 (P<0 .0 5 ,r=0 .4 32 )。结论 :TF及TFPI在CHD尤其是急性冠状动脉综合征的发生中起重要作用 ,维生素E的干预显著降低患者血浆TF水平 ,降低TF/TFPI比值而对TFPI无影响。     

Increased circulating levels of thrombin-activatable fibrinolysis inhibitor in lung cancer patients

Impairment of fibrinolytic function plays an important role in the mechanism of thrombotic disorders in cancer patients. This study assessed the circulating level of thrombin-activatable fibrinolysis inhibitor in patients with lung cancer and its expression by several lung cancer cell lines. The plasma concentrations of thrombin-activatable fibrinolysis inhibitor were significantly increased in lung cancer patients compared to healthy subjects. The concentration of thrombin-activatable fibrinolysis inhibitor was particularly higher in patients with small cell carcinoma compared to those with adenocarcinoma or squamous cell carcinoma, and in cancer patients that responded to chemotherapy compared to non-responders. In vitro studies showed more expression of thrombin-activatable fibrinolysis inhibitor in small cell carcinoma than in adenocarcinoma cell lines and more expression in lung cancer cell lines sensitive to anti-cancer agents than in resistant cell lines. This study suggests that thrombin-activatable fibrinolysis inhibitor, in part secreted from lung cancer cells, may play a role in the pathogenesis of thrombotic disorders in lung cancer patients.     

溃疡性结肠炎患者血小板功能状态和组织因子途径抑制物变化的临床研究

目的　探讨溃疡性结肠炎 (UC)患者血小板功能状态和组织因子途径抑制物 (TFPI)的变化。方法　使用TYXN 91智能血液聚集仪测定 47例UC患者和 3 0例对照组血小板聚集率(PAg) ,同时采用ELISA法检测血清P 选择素 (Ps) ,采用发色底物法测定血浆TFPI含量 ,并对其进行相关性分析。结果　活动期UC患者PAg明显高于缓解期和对照组 (P <0 .0 1) ,活动期和缓解期UC患者Ps和TFPI含量均高于对照组 (P <0 .0 1和P <0 .0 5 ) ,PAg与TFPI呈正相关 (γ =0 .62 3 ,P <0 .0 1) ,Ps与TFPI无明显相关性 (γ =0 .2 3 5 ,P >0 .0 5 )。结论　活动期UC患者体内存在血小板激活 ,在缓解期血小板活化程度仍增高 ;UC患者血浆TFPI含量升高 ,且PAg与TFPI在UC患者的血栓前状态中可能发挥着协同作用。     

人组织因子途径抑制物腺病毒表达系统的构建及体内表达

目的构建携载人组织因子途径抑制物(tissuefactorpathwayinhibitor,TFPI)基因的重组腺病毒载体,为基因治疗提供实验基础。方法利用基因重组技术,将人TFPI基因连接到穿梭质粒pDC316中,然后将腺病毒骨架质粒pBHGlox△E1,3Cre以及重组穿梭质粒pDC316-TFPI共转染293细胞,并在其中发生Cre重组酶介导的位点,特异性重组及腺病毒包装,扩增后进行滴度测定。将包装成功的携带人TFPI基因的重组腺病毒(Ad-TFPI)转染兔颈动脉,并用携带LacZ报告基因的重组腺病毒(Ad-LacZ)作为对照,3d后RT-PCR、ELISA法检测人TFPImRNA、蛋白的表达。结果得到了携带人TFPI基因的重组腺病毒,包装的病毒蚀斑形成单位(plaqueformationunit,PFU)滴度为7.6×1012/L。在Ad-TFPI转染兔颈动脉后3d,RT-PCR法和ELISA法均检测出TFPI表达,Ad-LacZ转染后未测到人TFPI的表达。结论成功构建了人TFPI腺病毒表达载体,为下一步的基因治疗提供了基础。     

Total thrombin-activatable fibrinolysis inhibitor (TAFI) antigen and pro-TAFI in patients with haemophilia A

Pro-thrombin-activatable fibrinolysis inhibitor (pro-TAFI), also known as TAFI, procarboxypeptidase U, or procarboxypeptidase B, is a relatively recently described plasma glycoprotein synthesized in the liver. It can be catalysed into its active form, TAFI (TAFIa, carboxypeptidase U or B) by a complex of thrombin/thrombomodulin. TAFI can potentially inhibit fibrinolysis by removing carboxyterminal lysine residues from partially degraded fibrin, decreasing plasminogen binding on the surface of fibrin, which thereby results in a decrease of the fibrinolytic activity. As TAFI represents a connection between coagulation and fibrinolysis, it can be expected that TAFI levels are altered in different thrombotic and haemorrhagic diseases, such as haemophilia A. Total TAFI antigen (including pro-TAFI, TAFI and the inactive form of TAFI [TAFIi]) and pro-TAFI were determined in 17 patients with haemophilia A. Thirteen healthy age-matched volunteers served as controls. No significant difference in levels of total TAFI antigen was observed between controls and patients with haemophilia, although it was slightly decreased in patients with haemophilia. Pro-TAFI was significantly reduced in haemophilia patients compared to controls (P=0.0113). TAFI antigen levels similar to controls have already been described in different clinical conditions, including haemophilia A. Decrease of pro-TAFI in haemophilia A can be an additional factor, together with decrease in thrombin generation, which induces impaired activation of pro-TAFI to TAFI, and could cause accelerated fibrinolysis. This supports the validity of usage of antifibrinolytics in the treatment of haemophilia A. In this paper we use new nomenclature for TAFI, and we believe that this recommended terminology for different forms of TAFI can simplify further standardization in TAFI investigation.     

不稳定型心绞痛患者抗凝治疗前后血浆组织因子途径抑制物含量和活性变化

目的 :探讨不稳定型心绞痛 (UA)患者使用低分子量肝素 (LMWH)抗凝治疗前后血浆组织因子途径抑制物 (TFPI)含量和活性的变化及临床意义。方法 :采用ELISA法测定 5 1例UA患者血浆组织因子 (TF)、TFPI含量 ,采用发色底物法同时测定TF、TFPI活性 ,并与 2 2例正常人 (对照组 )比较。 5 1例UA患者又分为LMWH组 32例 ,用LMWH加常规治疗 5d后复查TFPI含量和活性 ;常规组 19例 ,用常规治疗 5d后复查上述指标。结果 :①UA组血浆TF含量和活性均较对照组明显升高 (P <0 .0 5 ) ;血浆TFPI含量较对照组明显降低(P <0 .0 5 ) ;TFPI活性较对照组明显升高 (P <0 .0 5 )。②LMWH组TFPI含量治疗后较治疗前明显升高 (P<0 .0 5 ) ,TFPI活性治疗前后无明显变化 (P >0 .0 5 ) ,常规组TFPI含量和活性均无明显变化 (P >0 .0 5 )。结论 :UA患者存在异常激活的高凝状态 ,LMWH有独特的TFPI释放特性 ,并通过TFPI途径发挥抗凝作用     

In vitro evaluation of clot quality and stability in a model of severe thrombocytopenia: effect of fibrinogen,factor XIII and thrombin-activatable fibrinolysis inhibitor

Background

The treatment options in severe thrombocytopenia (platelet count ≤20×10⁹/L) are limited. The aim of this study was to investigate ways of improving blood clotting and stability in reconstituted thrombocytopenia.

Materials and methods

Thrombocytopenia (platelets [16±4]×10⁹/L) was created by differential centrifugation of normal blood followed by reconstitution of whole blood which was subjected to clotting in a rotation thromboelastometer by CaCl₂ and tissue factor, and to fibrinolysis by tissue plasminogen activator (tPA). In separate experiments, blood was diluted by 40% with TRIS/saline solution. Blood was treated with fibrinogen (fib), factor XIII (FXIII), and thrombin-activatable fibrinolysis inhibitor (TAFI).

Results

The maximum clot firmness of thrombocytopenic blood was approximately 2-fold less than that of intact blood. Supplementation of blood with fib and FXIII improved clot formation. In the presence of tPA, among fib, FXIII and TAFI, only fib stimulated clot propagation whereas each of these agents increased clot strength. There was a synergistic effect when fib was added together with FXIII or TAFI. Fibrinolysis was inhibited by TAFI and to a greater extent by TAFI + FXIII. Fourty percent dilution of blood reduced clot strength and increased susceptibility to tPA. Clot strength was increased by the treatments in the following order: fib/FXIII/TAFI > fib/TAFI > fib > TAFI > FXIII. In the presence of tPA, TAFI and FXIII lysed the clots significantly more slowly. This effect was stronger when blood was treated with the combination of fib/FXIII/TAFI. Doubling the fib concentration, alone or together with other agents, did not improve clot strength or stability.

Discussion

Augmentation of clot formation and anti-fibrinolysis by combining fib, FXIII and TAFI may be beneficial for the treatment of patients with severe thrombocytopenia especially when complicated by haemodilution following introduction of fluids to compensate for massive blood loss.     

厄贝沙坦对心肌梗死后大鼠心肌组织因子和组织因子途径抑制物的影响

目的观察大鼠心肌梗死后心肌组织因子（TF）和组织因子途径抑制物（TFPI）的mRNA表达及厄贝沙坦对其的影响。方法通过结扎大鼠左冠状动脉前降支建立大鼠急性心肌梗死模型组,另设假手术组（20只）。模型组术后24h存活大鼠随机分为安慰剂组（20只）和厄贝沙坦组（50mg·kg^-1d^-1,20只）。用药4周后处死各组大鼠并分别称其心室重量,计算左心室重量指数及心肌梗死面积,应用反转录聚合酶链反应（RT—PCR）方法检测心肌TFmRNA和TFPImRNA。结果模型组左心室重量指数大于假手术组（F=7．83,P〈0．05）。模型组内药物组左心室重量指数明显低于安慰剂组（F=8．96,P〈0．05）,心肌梗死面积比较差异无统计学意义（F=0．96,P〉0．05）;模型组TFmRNA、TFPImRNA表达均高于假手术组（F：8．24,P〈0．05）,其中药物组＇IFmRNA较安慰剂组明显降低（F=8．57,P〈0．05）,药物组TFPI mRNA较安慰剂组增加（F=1．35,P〉0．05）。结论大鼠心肌梗死后心肌TF mRNA、TFPI mRNA表达均明显增高。厄贝沙坦可显著降低心肌梗死大鼠心肌TF mRNA的表达。     

Decreased tissue factor pathway inhibitor (TFPI)‐dependent anticoagulant capacity in patients with cirrhosis who have decreased protein S but normal TFPI plasma levels

Protein S acts as a cofactor for tissue factor pathway inhibitor (TFPI) in the down regulation of thrombin generation, and acquired and congenital protein S deficiencies are associated with a concomitant TFPI deficiency. In contrast, in patients with liver diseases, decreased protein S, but normal or increased levels of TFPI have been reported. We compared TFPI and protein S plasma levels between 26 patients with cirrhosis and 20 healthy controls and found that TFPI levels were comparable between patients (111 ± 38%) and controls (108 ± 27%), despite reduced protein S levels (74 ± 23% in patients vs. 98 ± 10% in controls). Subsequently, we quantified the activity of the TFPI‐protein S system by measuring thrombin generation in the absence and presence of neutralizing antibodies to protein S or TFPI. Ratios of peak thrombin generation in the absence and presence of these antibodies were calculated. Both the protein S and the TFPI ratios were increased in patients with cirrhosis compared to controls. Protein S ratios were (0·62 [0·08–0·93] in patients vs. 0·32 [0·20–0·54] in controls; TFPI ratios were 0·50 [0·05–0·90] in patients vs. 0·18 [0·11–0·49] in controls). Thus, although the acquired protein S deficiency in patients with cirrhosis is not associated with decreased TFPI levels, the TFPI/protein S anticoagulant system is functionally impaired.     

Tissue factor and tissue factor pathway inhibitor levels in unstable angina patients during short-term low-molecular-weight heparin administration

High tissue factor (TF), tissue factor pathway inhibitor (TFPI) levels and a hypercoagulability state have been documented in unstable angina patients. We evaluated whether short-term enoxaparin administration (100 IU/kg b.i.d. for 3 d) reduces the high TF levels and the hypercoagulability state, and whether it influences the fibrinolytic system in 20 unstable angina patients. On d 3, we observed a significant reduction in TF levels both 1 h and 4 h after the morning injection (-25.6% and -21.7%; P < 0.001 respectively) in comparison with the base-line levels. Both 1 and 4 h after the morning injection on the d 3, TFPI levels significantly (P < 0.001) increased (+96.4%, +96.9% respectively) with respect to the base-line values. After enoxaparin administration, at all observation times, thrombin-antithrombin complexes and prothrombin fragment F1 + 2 levels were significantly (P < 0.001) lower with respect to base-line levels. We observed a slight but significant increase in tissue plasminogen activator antigen levels in preinjection samples, as well as 1 h and 4 h after enoxaparin administration, in comparison with the base-line values. This study provides evidence that low-molecular-weight heparin (LMWH) administration, in addition to a reduction of hypercoagulability and a mild fibrinolytic activation, is associated with decreased TF levels, so indicating a novel mechanism of interference of LMWH with the haemostatic system.     

人组织因子途径抑制物基因在兔颈动脉损伤血栓形成中的作用

Objective To observe the effects of tissue factor pathway inhibitor(TFPI) on thrombosis formation in rabbit carotid arteries after ballon injury. Methods Fouty rabbits with the weight 2.5-3.0 kg were respectively divided into 4 groups, Ad-TFPI, Ad-LacZ, PBS and normal control groups. The normal control group was not given any treatment and other 3 groups were given 0.2 ml Ad-TFPI, Ad-LacZ or PBS reproduced by the Dispatch catheter respectively after the PTCA balloon iniury on the right carotid arteries. Ten days after gene transfer the repeated balloon injury was performed in the 3 groups, and the first balloon injury was performed in the normal control group by the same method. The carotid blood flow was recovered immediately after the injury. Thirty minutes later all the animals were sacrificed. The injured carotid arteries and one part of contralateral normal artery were cut down, scissored along the long axis, flattened and fixed in the 2% glutaral. The platelet aggregation and thrombosis formation on the luminal surfaces was observed under electron microscope. Results The electron microscope results showed that the vascular endothelial cell structure was integrated and lined up in order in the nomal artery which had no any injury. After the balloon injury in the normal control group, the structure of the endothelial cell was disintegrated, and there was some platelet aggregation but no fibrosis formation. A large amount of platelet aggregated but no fibrosis formed in Ad-TFPI group after the repeated balloon injury. A large amount of fibrosis formed and red cells piled up in the Ad-LacZ and PBS group. The positive rate of thrombosis formation among groups had siginificant differences(χ2=14.95, P<0.01). The positive rate in Ad-TFPI group(20%) was lower than that in Ad-LacZ group(80%, χ2=7.20, P<0.01) and PBS group(70%, χ2=5.05, P<0.05), but was higher than that in the normal control group(10%, χ2=0.39, P>0.05). The positive rate in Ad-LacZ group(80%) was higher than in the normal control group(10%, χ2=9.90, P<0.01) and in the PBS group(70%, χ2=0.27, P> 0.05). The positive rate in PBS group(70%) was higher than that in the normal control group(10%, χ2=7.50, P< 0.01). Conclusions The repeated balloon injury method can cause a large amount of fibrosis formation in the rabbit carotid. TFPI gene inhibits thrombosis formation in balloon-injured rabbit carotid arteries.