Carboxypeptidase U (TAFIa) Is Rapidly Activated and Deactivated Following Thrombolysis and Thrombectomy in Stroke Patients.

The antifibrinolytic enzyme carboxypeptidase U (CPU, TAFIa, CPB2) is an appealing target for the treatment of acute ischemic stroke (AIS). Increased insights in CPU activation and inactivation during thrombolysis (rtPA) with or without endovascular thrombectomy (EVT) are required to develop CPU inhibitors as profibrinolytic agents with optimal benefits/risks. Therefore, CPU kinetics during ischemic stroke treatment were evaluated. AIS patients with documented cerebral artery occlusion receiving rtPA (N = 20) or rtPA + EVT (N = 16) were included. CPU activation during thrombolysis was measured by an ultrasensitive HPLC-based CPU activity method and by an ELISA measuring both CPU and inactivated CPU (CPU + CPUi). Intravenous blood samples were collected at admission and throughout the first 24 h. Additional in situ blood samples were collected in the rtPA + EVT cohort proximal from the thrombus. The NIHSS score was determined at baseline and 24 h. CPU activity and CPU + CPUi levels increased upon rtPA administration and reached peak values at the end of thrombolysis (1 h). High inter-individual variability was observed in both groups. CPU activity decreased rapidly within 3 h, while CPU + CPUi levels were still elevated at 7 h. CPU activity or CPU + CPUi levels were similar in in situ and peripheral samples. No correlation between CPU or CPU + CPUi and NIHSS or thrombus localization was found. The CPU system was rapidly activated and deactivated following thrombolysis and thrombectomy in stroke patients, suggesting that a CPU inhibitor would have to be administered during rtPA infusion and over the next few hours. The high CPU generation variability suggests that some patients may not respond to the treatment. EudraCT number 2017-002760-41.

Carboxypeptidase U (CPU, TAFIa, CPB2) in Thromboembolic Disease: What Do We Know Three Decades after Its Discovery?

Procarboxypeptidase U (proCPU, TAFI, proCPB2) is a basic carboxypeptidase zymogen that is converted by thrombin(-thrombomodulin) or plasmin into the active carboxypeptidase U (CPU, TAFIa, CPB2), a potent attenuator of fibrinolysis. As CPU forms a molecular link between coagulation and fibrinolysis, the development of CPU inhibitors as profibrinolytic agents constitutes an attractive new concept to improve endogenous fibrinolysis or to increase the efficacy of thrombolytic therapy in thromboembolic diseases. Furthermore, extensive research has been conducted on the in vivo role of CPU in (the acute phase of) thromboembolic disease, as well as on the hypothesis that high proCPU levels and the Thr/Ile325 polymorphism may cause a thrombotic predisposition. In this paper, an overview is given of the methods available for measuring proCPU, CPU, and inactivated CPU (CPUi), together with a summary of the clinical data generated so far, ranging from the current knowledge on proCPU concentrations and polymorphisms as potential thromboembolic risk factors to the positioning of different CPU forms (proCPU, CPU, and CPUi) as diagnostic markers for thromboembolic disease, and the potential benefit of pharmacological inhibition of the CPU pathway.

Overexpression of TAFI promotes epithelial mesenchymal transition in endometriosis.

OBJECTIVE: Endometriosis is a disease that occurs in women. Thrombin-activated fibrinolytic inhibitor (TAFI) is mainly secreted by stem cells and acts as a regulatory role in the body. Epithelial leaf transition plays a leading role in cell growth and invasion. Our study focuses on the mechanism of TAFI in patients with endometriosis. PATIENTS AND METHODS: The expression of TAFI was determined by immunohistochemistry. Reverse transcriptase-polymerase chain reaction (RT-PCR) served to detect the expression of TAFI and the effect of TAFI on overall survival (OS) and progression-free survival (PFS) levels. The changes of primary cytology in patients with endometriosis were observed under a microscope. The cell source was further determined by immunofluorescence labeling of vimentin and cytokeratin, and the expression of TAFI was detected by Western-blot. 3-4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and cell invasion assay were utilized to detect the viability and aggressiveness of cells after epithelial mesenchymal transition (EMT). RESULTS: TAFI was overexpressed in endometriosis tissues and no expression of TAFI was found in normal tissues, which is consistent with RT-PCR results. TAFI overexpressed endometriosis patients had low levels of overall OS and PFS. There were statistically significant differences. Cell morphology shows that endometriosis primary cells are mainly composed of epithelial cells and fibroblasts. Immunofluorescence assay showed that vimentin and cytokeratin were expressed in cells, and the expression of TAFI was detected by Western-blot. Compared with normal tissues, TAFI was considerably higher in patients with endometriosis. The results of Western-blot and RT-PCR showed that the expression of TAFI was significantly increased in patients with endometriosis and the cell proliferation and cell invasion were significantly accelerated. CONCLUSIONS: Our results show that TAFI is highly expressed in endometriosis and causes EMT, which accelerated the cell proliferation and cell invasion. Snail is an inhibitor of E-cadherin, which may participate in metastasis and invasion of endometriosis by mediating EMT. So, we suspect that Snail controls the occurrence of the EMT and then affects the cell metastasis and invasion, which requires further verification.

Activated thrombin-activatable fibrinolysis inhibitor attenuates the angiogenic potential of endothelial cells: potential relevance to the breast tumour microenvironment.

Thrombin-activatable fibrinolysis inhibitor (TAFI) is a basic carboxypeptidase zymogen present in blood plasma. Proteolytic activation of TAFI by thrombin, thrombin in complex with the endothelial cell cofactor thrombomodulin, or plasmin results in an enzyme (TAFIa) that removes carboxyl-terminal lysine residues from protein and peptide substrates, including cell-surface plasminogen receptors. TAFIa is therefore capable of inhibiting plasminogen activation in the pericellular milieu. Since plasminogen activation has been linked to angiogenesis, TAFIa could therefore have anti-angiogenic properties, and indeed TAFIa has been shown to inhibit endothelial tube formation in a fibrin matrix. In this study, the TAFI pathway was manipulated by providing exogenous TAFI or TAFIa or by adding a potent and specific inhibitor of TAFIa. We found that TAFIa elicited a series of anti-angiogenic responses by endothelial cells, including decreased endothelial cell proliferation, cell invasion, cell migration, tube formation, and collagen degradation. Moreover, TAFIa decreased tube formation and proteolysis in endothelial cell culture grown alone and in co-culture with breast cancer cell lines. In accordance with these findings, inhibition of TAFIa increased secretion of matrix metalloprotease proenzymes by endothelial and breast cancer cells. Finally, treatment of endothelial cells with TAFIa significantly inhibited plasminogen activation. Taken together our results suggest a novel role for TAFI in inhibiting tumour angiogenic behaviors in breast cancer.

Thrombin-activatable fibrinolysis inhibitor influences disease severity in humans and mice with pneumococcal meningitis.

BACKGROUND: Mortality and morbidity in patients with bacterial meningitis result from the proinflammatory response and dysregulation of coagulation and fibrinolysis. Thrombin-activatable fibrinolysis inhibitor (TAFI) is activated by free thrombin or thrombin in complex with thrombomodulin, and plays an antifibrinolytic role during fibrin clot degradation, but also has an anti-inflammatory role by inactivating proinflammatory mediators, such as complement activation products. OBJECTIVE: To assess the role of TAFI in pneumococcal meningitis. METHODS: We performed a prospective nationwide genetic association study in patients with bacterial meningitis, determined TAFI and complement levels in cerebrospinal fluid (CSF), and assessed the function of TAFI in a pneumococcal meningitis mouse model by using Cpb2 (TAFI) knockout mice. RESULTS: Polymorphisms (reference sequences: rs1926447 and rs3742264) in the CPB2 gene, coding for TAFI, were related to the development of systemic complications in patients with pneumococcal meningitis. Higher protein levels of TAFI in CSF were significantly associated with CSF complement levels (C3a, iC3b, and C5b-9) and with more systemic complications in patients with bacterial meningitis. The risk allele of rs1926447 (TT) was associated with higher levels of TAFI in CSF. In the murine model, consistent with the human data, Cpb2-deficient mice had decreased disease severity, as reflected by lower mortality, and attenuated cytokine levels and bacterial outgrowth in the systemic compartment during disease, without differences in the brain compartment, as compared with wild-type mice. CONCLUSIONS: These findings suggest that TAFI plays an important role during pneumococcal meningitis, which is likely to be mediated through inhibition of the complement system, and influences the occurrence of systemic complications and inflammation.

Brief report: carboxypeptidase B serves as a protective mediator in osteoarthritis.

OBJECTIVE: We previously demonstrated that carboxypeptidase B (CPB) protects against joint erosion in rheumatoid arthritis by inactivating complement component C5a. We also found that levels of CPB are abnormally high in the synovial fluid of individuals with another joint disease, osteoarthritis (OA). We undertook this study to investigate whether CPB plays a role in the pathogenesis of OA. METHODS: We compared the development of OA in CPB-deficient (Cpb2(-/-) ) mice and wild-type mice by subjecting them to medial meniscectomy and histologically assessing cartilage damage, osteophyte formation, and synovitis in the stifle joints 4 months later. We measured levels of proCPB, proinflammatory cytokines, and complement components in synovial fluid samples from patients with symptomatic and radiographic knee OA. Finally, we used enzyme-linked immunosorbent assay, flow cytometry, and hemolytic assays to assess the effect of CPB on formation of membrane attack complex (MAC)-a complement effector critical to OA pathogenesis. RESULTS: Cpb2(-/-) mice developed dramatically greater cartilage damage than did wild-type mice (P < 0.01) and had a greater number of osteophytes (P < 0.05) and a greater degree of synovitis (P < 0.05). In synovial fluid samples from OA patients, high levels of proCPB were associated with high levels of proinflammatory cytokines and complement components, and levels of proCPB correlated positively with those of MAC. In in vitro complement activation assays, activated CPB suppressed the formation of MAC as well as MAC-induced hemolysis. CONCLUSION: Our data suggest that CPB protects against inflammatory destruction of the joints in OA, at least in part by inhibiting complement activation.

Insights into thrombin activatable fibrinolysis inhibitor function and regulation.

Fibrinolysis is initiated when the zymogen plasminogen is converted to plasmin via the action of plasminogen activators. Proteolytic cleavage of fibrin by plasmin generates C-terminal lysine residues capable of binding both plasminogen and the plasminogen activator, thereby stimulating plasminogen activator-mediated plasminogen activation and propagating fibrinolysis. This positive feedback mechanism is regulated by activated thrombin activatable fibrinolysis inhibitor (TAFIa), which cleaves C-terminal lysine residues from the fibrin surface, thereby decreasing its cofactor activity. TAFI can be activated by thrombin alone, but the rate of activation is accelerated when in complex with thrombomodulin. Plasmin is also known to activate TAFI. TAFIa has no known physiologic inhibitors and consequently, its primary regulatory mechanism involves its intrinsic thermal instability. The rate of TAFI activation and stability of the active form, TAFIa, function in maintaining its concentration above the threshold value required to down-regulate fibrinolysis. Although all methods to quantify TAFI or TAFIa have their limitations, epidemiologic studies have indicated that elevated TAFI levels are correlated with an increased risk of venous thrombosis. Major efforts have been made to develop TAFI inhibitors that can either directly interfere with TAFIa activity or impair its activation. However, the anti-inflammatory properties of TAFIa might complicate the development and application of a TAFIa inhibitor that aims to increase the efficiency of thrombolytic therapy.

Thrombin activatable fibrinolysis inhibitor: a putative target to enhance fibrinolysis.

Thrombin activatable fibrinolysis inhibitor (TAFI) was discovered two decades ago consequent to the identification of an unstable carboxypeptidase (CPU) formed upon thrombin activation of its proenzyme. The antifibrinolytic effects of the activated form (TAFIa, CPU) are linked with its capacity to remove C-terminal lysines from the surface of the fibrin clot. A distinctive characteristic of TAFIa is its temperature-dependent conformational instability: TAFIa activity spontaneously decays with an apparent half-life of 8 to 15 minutes at 37°C. A variety of studies has demonstrated a role for TAFI/TAFIa in venous and arterial diseases. In addition, a role for TAFI/TAFIa in inflammation and cell migration has also been shown. Because TAFI/TAFIa is a potential risk factor for thrombotic disorders, many inhibitors, both at the level of activation or at the level of activity, have been developed and were proven to exhibit a profibrinolytic effect in animal models. Pharmacologically active inhibitors of the TAFI/TAFIa system may open new ways for the prevention of thrombotic diseases or for the establishment of adjunctive treatments during thrombolytic therapy.

Is there any role of thrombin activatable fibrinolysis inhibitor in the development of a hypercoagulable state in gastric cancer.

BACKGROUND: The purpose of this study was to investigate plasma levels of thrombin activatable fibrinolysis inhibitor (TAFI) and TAFI's relationship with coagulation markers (prothrombin fragment 1 + 2) in gastric cancer patients. METHODS: Thirty-three patients with gastric adenocarcinoma and 29 healthy control subjects were prospectively enrolled in the study. Patients who had a history of secondary malignancy, thrombosis related disease, oral contraceptive use, diabetes mellitus, chronic renal failure or similar chronic metabolic disease were excluded from the study. A fasting blood sample was drawn from patients to determine the plasma levels of TAFI and Prothrombin Fragment 1 + 2 (F 1 + 2). In addition, data on patient age, sex, body mass index (BMI) and stage of disease were recorded. The same parameters, except stage of disease, were also recorded for the control group. Subsequently, we assessed the difference in the levels of TAFI and F 1 + 2 between the patient and control groups. Moreover, we investigated the relation of TAFI and F 1 + 2 levels with age, sex, BMI and stage of disease in the gastric cancer group. RESULTS: There were no statistical differences in any demographic variables (age, gender and BMI) between the groups (Table 1). The mean plasma TAFI levels of the gastric cancer group (69.4 ± 33.1) and control group (73.3 ± 27.5) were statistically similar (P = 0.62). The mean plasma F 1 + 2 level in the gastric cancer group was significantly higher than for those in the control group (549.7 ± 325.3 vs 151.9 ± 67.1, respectively; P < 0.001). In the gastric cancer group, none of the demographic variables (age, gender and BMI) were correlated with either TAFI or F 1 + 2 levels. Also, no significant associations were found between the stage of the cancer and either TAFI or F 1 + 2 levels. CONCLUSION: In our study, TAFI levels of gastric cancer patients were similar to healthy subjects. The results of our study suggest that TAFI does not play a role in pathogenesis of the hypercoagulable state in gastric cancer patients.

Role of thrombin activatable fibrinolysis inhibitor in endocrine and cardiovascular disorders: an update.

Thrombin activatable fibrinolysis inhibitor (TAFI) is a zymogene that potently inhibits fibrinolysis through the removal of the carboxy-terminal lysine and arginine residues from partially degraded fibrin polymers. In addition, TAFI has a suppressor effect on conversion of inactive plasminogen to plasmin. Since impaired fibrinolysis is a very well established risk factor for cardiovascular morbidity and mortality, understanding the role of TAFI in cardiovascular disorders, insulin resistance, diabetes and other endocrine problems may hold promise for improving management of these diseases. This paper includes a review of current evidence on TAFI pathway and its alteration in endocrine and cardiovascular disorders and relevant patents.

Secretion and antifibrinolytic function of thrombin-activatable fibrinolysis inhibitor from human platelets.

BACKGROUND: The thrombin-activatable fibrinolysis inhibitor (TAFI) is a zymogen first characterized in human plasma that is activated through proteolytic cleavage by thrombin, thrombin in complex with thrombomodulin, or plasmin. Active TAFI attenuates fibrinolysis by removing C-terminal lysine residues from partially degraded fibrin, thereby inhibiting a potent positive feedback loop in the fibrinolytic cascade. The existence of a separate pool of TAFI within platelets has been described. OBJECTIVES AND METHODS: We aimed to confirm the presence of TAFI in the medium of washed, thrombin-stimulated platelets and to evaluate the characteristics of platelet TAFI by western blot analysis and with a quantitative assay for activated TAFI. We also assessed the ability of platelet TAFI to inhibit fibrinolysis in vitro, using a platelet-rich thrombus lysis assay. RESULTS: Our data are consistent with the presence of TAFI in the α-granules of resting platelets. In contrast to previous reports, platelet TAFI is very similar in electrophoretic mobility to plasma-derived TAFI. We also show, for the first time, that platelet-derived TAFI is capable of attenuating platelet-rich thrombus lysis in vitro independently of plasma TAFI. Moreover, we demonstrate additive effects on thrombolysis of platelet-derived TAFI and TAFI present in plasma. CONCLUSIONS: Taken together, these observations indicate that the secretion of platelet-derived TAFI can augment the concentrations of TAFI already present in plasma to enhance attenuation of the fibrinolytic cascade. This could be significant in regions of vascular damage or pathologic thrombosis, where activated platelets are known to accumulate.

Thrombin activatable fibrinolysis inhibitor (TAFI): a molecular link between coagulation and fibrinolysis.

Although the maintenance of precise balance between coagulation and fibrinolysis is of utmost importance for normal haemostasis, until recently these two systems were considered as completely separate mechanisms involved in the process of formation and dissolution of blood clot. Thrombin activatable fibrinolysis inhibitor (TAFI) is a recently described attenuator of the fibrinolytic rate and is considered to be the molecular link between coagulation and fibrinolysis. TAFI circulates in plasma as an inactive precursor and its conversion in active enzyme (TAFIa) occurs by the action of thrombin or plasmin, but most efficiently by thrombin in the presence of its cofactor thrombomodulin. Once generated, TAFI down-regulates fibrinolysis by removing C-terminal lysine residues from partially degraded fibrin; thereby preventing the upregulation of plasminogen binding and activation. Because TAFI is activated by thrombin on one side, and acts as the attenuator of fibrinolysis on another side, it enables fine synchronization between these two systems. The antifibrinolytic function of TAFI mostly depends on TAFI concentration, the rate of its activation and the half-life of TAFIa in plasma. Changes in thrombin generation can have a profound effect on the rate of TAFI activation, and consequently on the rate of fibrinolysis. Therefore, it has been hypothesized that increased thrombin generation seen in thrombophilia patients may enhance TAFI activation, leading to a hypofibrinolytic state, which may further contribute to the thrombotic tendency. However, the results of several studies, in which relation between TAFI level and the occurrence of thromboembolic complications in carriers of hereditary thrombophilia have been investigated, were not consistent.

Insights into the molecular inactivation mechanism of human activated thrombin-activatable fibrinolysis inhibitor.

SUMMARY BACKGROUND: Thrombin-activatable fibrinolysis inhibitor (TAFI) is a validated target for thrombotic diseases. TAFI is converted in vivo to activated TAFI (TAFIa) by removal of its pro-domain. Whereas TAFI is stable and persists in the circulation, possibly in complex with plasminogen, TAFIa is unstable and poorly soluble, with a half-life of minutes. OBJECTIVES: In order to study the molecular determinants of this instability, we studied the influence of protein inhibitors on human TAFIa. RESULTS: We found that protein inhibitors significantly reduced the instability and insolubility of TAFIa. In addition, we solved the 2.5-A resolution crystal structure of human TAFIa in complex with a potent protein inhibitor, tick-derived carboxypeptidase inhibitor, which gives rise to a stable and soluble TAFIa species. The structure revealed a significant reduction in the flexibility of dynamic segments when compared with the structures of bovine and human TAFI. We also identified two latent hotspots, loop Lbeta2beta3 and segment alpha5-Lalpha5beta7-beta7, where conformational destabilization may begin. These hotspots are also present in TAFI, but the pro-domain may provide sufficient stabilization and solubility to guarantee protein persistence in vivo. When the pro-domain is removed, the free TAFIa moiety becomes unstable, its activity is suppressed, and the molecule becomes insoluble. CONCLUSIONS: The present study corroborates the function of protein inhibitors in stabilizing human TAFIa and it provides a rigid and high-resolution mold for the design of small molecule inhibitors of this enzyme, thus paving the way for novel therapy for thrombotic disorders.

Factor XI deficiency--resolving the enigma?

The management of factor XI deficiency is not straightforward for three reasons: firstly, the role of this factor in the coagulation pathway is not clearly understood; secondly, the bleeding tendency, although mild, is unpredictable and does not clearly relate to the factor XI level; and thirdly, all treatment products, although available, have some potentially serious side effects. These factors (or enigmas) contribute to the variable management of patients with this coagulation factor deficiency, but recent research is helping to clarify some of these areas.

Carboxypeptidase U (TAFIa): a new drug target for fibrinolytic therapy?

Procarboxypeptidase U (TAFI) is a recently discovered plasma procarboxypeptidase that upon activation by thrombin or thrombin-thrombomodulin turns into a potent antifibrinolytic enzyme. Its prominent bridging function between coagulation and fibrinolysis raised the interest of many research groups and of the pharmaceutical industry. The development of carboxypeptidase U (CPU) inhibitors as profibrinolytic agents is an attractive concept and possibilities for rational drug design will become more readily available in the near future as a result of the recently published crystal structure. Numerous studies have been performed and many of them show beneficial effects of CPU inhibitors for the improvement of endogenous fibrinolysis in different animal sepsis and thrombosis models. CPU inhibitors combined with tissue-type plasminogen activator (t-PA) seem to increase the efficiency of pharmacological thrombolysis allowing lower dosing of t-PA and subsequently fewer bleeding complications. This review will focus on recently obtained in vivo data and the benefits/risks of targeting CPU for the treatment of thrombotic disorders.

The role of thrombin activatable fibrinolysis inhibitor in arterial thrombosis at a young age: the ATTAC study.

BACKGROUND AND OBJECTIVES: Thrombin activatable fibrinolysis inhibitor (TAFI) attenuates fibrinolysis and may therefore contribute to the pathophysiology of arterial thrombosis. The aim of the present study was to elucidate the pathogenetic role of TAFI levels and genotypes in young patients with arterial thrombosis. PATIENTS AND METHODS: In a case-control study, 327 young patients with a recent first-ever event of coronary heart disease (CHD subgroup) or cerebrovascular disease (ischemic stroke subgroup) and 332 healthy young controls were included. TAFI levels [intact TAFI, activation peptide (TAFI-AP) and (in)activated TAFI (TAFIa(i)] and TAFI activity were measured and genetic variations in the TAFI gene (-438G/A, 505G/A and 1040C/T) were determined. RESULTS: In the total group of patients, TAFIa(i) levels were higher (145.1 +/- 37.5%) than in controls (137.5 +/- 31.3%, P = 0.02). Plasma levels of intact TAFI, TAFI-AP and TAFI activity were similar in patients and controls. In the CHD subgroup (n = 218), intact TAFI levels were higher (109.4 +/- 23.0%) than in controls (102.8 +/- 20.7%, P = 0.02). In 325Ile/Ile homozygotes, lower TAFI levels and a decreased risk of arterial thrombosis were observed (OR 0.58, 95% CI 0.34-0.99) compared with patients with the common 325Thr/Thr genotype. This association was most evident in CHD patients (OR 0.48, 95% CI 0.26-0.90). Haplotype analyses supported a role for the Thr325Ile polymorphism. CONCLUSIONS: TAFIa(i) levels were higher in patients with cardiovascular disease. Furthermore, the TAFI 325Thr/Ile polymorphism was associated with lower TAFI levels and with the risk of cardiovascular disease in young patients, especially in CHD.

Regulation of tissue inflammation by thrombin-activatable carboxypeptidase B (or TAFI).

Thrombin-activatable procarboxypeptidase B (proCPB or thrombin-activatable fibrinolysis inhibitor or TAFI) is a plasma procarboxypeptidase that is activated by the thrombin-thrombomodulin complex on the vascular endothelial surface. The activated CPB removes the newly exposed carboxyl terminal lysines in the partially digested fibrin clot, diminishes tissue plasminogen activator and plasminogen binding, and protects the clot from premature lysis. We have recently shown that CPB is catalytically more efficient than plasma CPN, the major plasma anaphylatoxin inhibitor, in inhibiting bradykinin, activated complement C3a, C5a, and thrombin-cleaved osteopontin in vitro. Using a thrombin mutant (E229K) that has minimal procoagulant properties but retains the ability to activate protein C and proCPB in vivo, we showed that infusion of E229K thrombin into wild type mice reduced bradykinin-induced hypotension but it had no effect in proCPB-deficient mice, indicating that the beneficial effect of E229K thrombin is mediated through its activation of proCPB and not protein C. Similarly proCPB-deficient mice displayed enhanced pulmonary inflammation in a C5a-induced alveolitis model and E229K thrombin ameliorated the magnitude of alveolitis in wild type but not proCPB-deficient mice. Thus, our in vitro and in vivo data support the thesis that thrombin-activatable CPB has broad anti-inflammatory properties. By specific cleavage of the carboxyl terminal arginines from C3a, C5a, bradykinin and thrombin-cleaved osteopontin, it inactivates these active inflammatory mediators. Along with the activation of protein C, the activation of proCPB by the endothelial thrombin-thrombomodulin complex represents a homeostatic feedback mechanism in regulating thrombin's pro-inflammatory functions in vivo.