Plasminogen: an enigmatic zymogen.

Plasminogen is an abundant plasma protein that exists in various zymogenic forms. Plasmin, the proteolytically active form of plasminogen, is known for its essential role in fibrinolysis. To date, therapeutic targeting of the fibrinolytic system has been for 2 purposes: to promote plasmin generation for thromboembolic conditions or to stop plasmin to reduce bleeding. However, plasmin and plasminogen serve other important functions, some of which are unrelated to fibrin removal. Indeed, for >40 years, the antifibrinolytic agent tranexamic acid has been administered for its serendipitously discovered skin-whitening properties. Plasmin also plays an important role in the removal of misfolded/aggregated proteins and can trigger other enzymatic cascades, including complement. In addition, plasminogen, via binding to one of its dozen cell surface receptors, can modulate cell behavior and further influence immune and inflammatory processes. Plasminogen administration itself has been reported to improve thrombolysis and to accelerate wound repair. Although many of these more recent findings have been derived from in vitro or animal studies, the use of antifibrinolytic agents to reduce bleeding in humans has revealed additional clinically relevant consequences, particularly in relation to reducing infection risk that is independent of its hemostatic effects. The finding that many viruses harness the host plasminogen to aid infectivity has suggested that antifibrinolytic agents may have antiviral benefits. Here, we review the broadening role of the plasminogen-activating system in physiology and pathophysiology and how manipulation of this system may be harnessed for benefits unrelated to its conventional application in thrombosis and hemostasis.

Molecular characterization of a Trichinella spiralis enolase and its interaction with the host's plasminogen.

The binding and activation of host plasminogen (PLG) by worm surface enolases has been verified to participate in parasite invasion, but the role of this processes during Trichinella spiralis infection has not been clarified. Therefore, the expression and immunolocalization of a T. spiralis enolase (TsENO) and its binding activity with PLG were evaluated in this study. Based on the three-dimensional (3D) molecular model of TsENO, the protein interaction between TsENO and human PLG was analysed by the ZDOCK server. The interacting residues were identified after analysis of the protein-protein interface by bioinformatics techniques. The key interacting residues were confirmed by a series of experiments. The qPCR analysis results demonstrated that Ts-eno was transcribed throughout the whole life cycle of T. spiralis. The immunofluorescence assay (IFA) results confirmed that TsENO was distributed on the T. spiralis surface. The binding assays showed that recombinant TsENO (rTsENO) and native TsENO were able to bind PLG. Four lysine residues (90, 289, 291 and 300) of TsENO were considered to be active residues for PLG interaction. The quadruple mutant (Lys90Ala + Lys289Ala + Lys291Ala + Lys300Ala) TsENO, in which the key lysine residues were substituted with alanine (Ala) residues, exhibited a reduction in PLG binding of nearly 50% (45.37%). These results revealed that TsENO has strong binding activity with human PLG. The four lysine residues (90, 289, 291 and 300) of TsENO play an important role in PLG binding and could accelerate PLG activation and invasion of the host's intestinal wall by T. spiralis.

Platelet-Mediated Modulation of Fibrinolysis.

Platelets are crucial to the hemostatic response. Their role in coagulation is well documented and they have been considered for some time to promote resistance of thrombi to fibrinolysis. Platelets confer resistance to lysis by promoting clot retraction of the immediate fibrin network and through release of plasminogen activator inhibitor-1 from their α-granules. However, recent developments in the field indicate that the role of platelets in fibrinolysis is much more diverse. Indeed, novel studies suggest that platelets form different subpopulations upon activation that play varied roles in regulating hemostasis. Likewise the developments in our understanding of thrombus formation, architecture, and changes in fibrin deposition and composition suggest that these different subpopulations of platelets may populate distinct areas within thrombi and potentially dictate the local hemostatic balance in these areas. This review will discuss the diverse roles of platelets in fibrinolysis and highlight the recent developments in the field and the contribution of both the intracellular pool of modulators as well as the membrane surface in regulating these processes.

Inhibition of plasminogen activation by apo(a): role of carboxyl-terminal lysines and identification of inhibitory domains in apo(a).

Apo(a), the distinguishing protein component of lipoprotein(a) [Lp(a)], exhibits sequence similarity to plasminogen and can inhibit binding of plasminogen to cell surfaces. Plasmin generated on the surface of vascular cells plays a role in cell migration and proliferation, two of the fibroproliferative inflammatory events that underlie atherosclerosis. The ability of apo(a) to inhibit pericellular plasminogen activation on vascular cells was therefore evaluated. Two isoforms of apo(a), 12K and 17K, were found to significantly decrease tissue-type plasminogen activator-mediated plasminogen activation on human umbilical vein endothelial cells (HUVECs) and THP-1 monocytes and macrophages. Lp(a) purified from human plasma decreased plasminogen activation on THP-1 monocytes and HUVECs but not on THP-1 macrophages. Removal of kringle V or the strong lysine binding site in kringle IV10 completely abolished the inhibitory effect of apo(a). Treatment with carboxypeptidase B to assess the roles of carboxyl-terminal lysines in cellular receptors leads in most cases to decreases in plasminogen activation as well as plasminogen and apo(a) binding; however, inhibition of plasminogen activation by apo(a) was unaffected. Our findings directly demonstrate that apo(a) inhibits pericellular plasminogen activation in all three cell types, although binding of apo(a) to cell-surface receptors containing carboxyl-terminal lysines does not appear to play a major role in the inhibition mechanism.

Macrophage gene expression and foam cell formation are regulated by plasminogen.

BACKGROUND: Deciphering the molecular and cellular processes that govern macrophage foam cell formation is critical to understanding the basic mechanisms underlying atherosclerosis and other vascular pathologies. METHODS AND RESULTS: Here, we identify a pivotal role of plasminogen (Plg) in regulating foam cell formation. Deficiency of Plg inhibited macrophage cholesterol accumulation on exposure to hyperlipidemic conditions in vitro, ex vivo, and in vivo. Gene expression analysis identified CD36 as a regulated target of Plg, and macrophages from Plg(-/-) mice had decreased CD36 expression and diminished foam cell formation. The Plg-dependent CD36 expression and foam cell formation depended on conversion of Plg to plasmin, binding to the macrophage surface, and the consequent intracellular signaling that leads to production of leukotriene B4. Leukotriene B4 rescued the suppression of CD36 expression and foam cell formation arising from Plg deficiency. CONCLUSIONS: Our findings demonstrate an unanticipated role of Plg in the regulation of gene expression and cholesterol metabolism by macrophages and identify Plg-mediated regulation of leukotriene B4 as an underlying mechanism.

From fibrinolysis to the plasminogen-plasmin system and beyond: a remarkable growth of knowledge, with personal observations on the history of fibrinolysis.

Great advances have been made in our understanding of the fibrinolytic system from the initial discovery of proteolysis of fibrin by plasmin to the multifaceted and complex role of the plasminogen-plasmin (P-P) system. We now know that the P-P system is composed of several serine proteases and their inhibitors (serpins). This system is involved in many physiological functions, including embryogenesis, cell migration, and wound healing. They also play an important role in the pathogenesis of many diseases, including atherosclerosis, obesity, cancer, and even autoimmune disorders, and neuronal degeneration. Knowledge of their role in cancer enables their use as a prognostic factor. Therapeutic use of various forms of proteases derived from this system has been employed as thrombolytic agents. In addition, small molecules designed to inhibit many of the components of the P-P system are now available for clinical trial, aimed at treatment of these various disorders. The history of such remarkable development of our knowledge on fibrinolysis is reviewed in this article.

Serum-derived plasminogen is activated by apoptotic cells and promotes their phagocytic clearance.

The elimination of apoptotic cells, called efferocytosis, is fundamentally important for tissue homeostasis and prevents the onset of inflammation and autoimmunity. Serum proteins are known to assist in this complex process. In the current study, we performed a multistep chromatographic fractionation of human serum and identified plasminogen, a protein involved in fibrinolysis, wound healing, and tissue remodeling, as a novel serum-derived factor promoting apoptotic cell removal. Even at levels significantly lower than its serum concentration, purified plasminogen strongly enhanced apoptotic prey cell internalization by macrophages. Plasminogen acted mainly on prey cells, whereas on macrophages no enhancement of the engulfment process was observed. We further demonstrate that the efferocytosis-promoting activity essentially required the proteolytic activation of plasminogen and was completely abrogated by the urokinase plasminogen activator inhibitor-1 and serine protease inhibitor aprotinin. Thus, our study assigns a new function to plasminogen and plasmin in apoptotic cell clearance.

Plasmodium ookinetes coopt mammalian plasminogen to invade the mosquito midgut.

Ookinete invasion of the mosquito midgut is an essential step for the development of the malaria parasite in the mosquito. Invasion involves recognition between a presumed mosquito midgut receptor and an ookinete ligand. Here, we show that enolase lines the ookinete surface. An antienolase antibody inhibits oocyst development of both Plasmodium berghei and Plasmodium falciparum, suggesting that enolase may act as an invasion ligand. Importantly, we demonstrate that surface enolase captures plasminogen from the mammalian blood meal via its lysine motif (DKSLVK) and that this interaction is essential for midgut invasion, because plasminogen depletion leads to a strong inhibition of oocyst formation. Although addition of recombinant WT plasminogen to depleted serum rescues oocyst formation, recombinant inactive plasminogen does not, thus emphasizing the importance of plasmin proteolytic activity for ookinete invasion. The results support the hypothesis that enolase on the surface of Plasmodium ookinetes plays a dual role in midgut invasion: by acting as a ligand that interacts with the midgut epithelium and, further, by capturing plasminogen, whose conversion to active plasmin promotes the invasion process.

[Gene-targeted radiation therapy mediated by radiation-sensitive promoter in lung adenocarcinoma and the feasibility of micro-PET/CT in evaluation of therapeutic effectiveness in small animals].

OBJECTIVE: To explore the combined anti-tumor effect of radiation therapy and gene-targeted suppression of tumor neovasculature in lung adenocarcinoma in vivo, and to explore the feasibility of micro-PET/CT in dynamic evaluation of treatment effectiveness. METHODS: Thirty 5-6-week old male BALB/c nude mice were used in this study. The mouse models of xenotransplanted human lung adenocarcinoma were divided into 5 groups at random, six mice in each group: the control group, radiation treatment alone group and three groups of recombinant baculovirus plus radiation treatment (intratumoral injection, tail vein injection, and intramuscular injection). The tumor volume was measured every 2 days. Growth delay time (GD) and growth inhibition rate was calculated. FDG metabolism was evaluated by micro-PET-CT before and after treatment. The expressions of VEGF, CD31 and Ki-67 were detected by immunohistochemistry (IHC). RESULTS: The tumor growth delay was >12 days, and the tumor inhibition rate was >45% in the recombinant baculovirus combined with radiotherapy groups, significantly higher than that of the radiotherapy alone group (P < 0.05). Immunohistochemical analysis showed that the expressions of VEGF, CD31 and Ki-67 were significantly lower than that in other groups (P < 0.05). The micro-PET-CT assessment showed that the FDG-metabolism in the recombinant baculovirus combined with radiotherapy groups was significantly reduced (P < 0.05), and the SUVmax (FDG metabolism) of transplanted tumors after treatment was also markedly decreased in comparison with that of the control group. The tumor volume after treatment was significantly correlated with SUVmax in the recombinant baculovirus intratumoral injection + radiotherapy group(r = 0.976), recombinant baculovirus intravenous injection + radiotherapy group (r = 0.954), recombinant baculovirus intramuscular injection + radiotherapy group (r = 0.929), and radiotherapy alone group (r = 0.871, P < 0.05). CONCLUSIONS: The recombinant baculovirus containing Egr1 promoter and K5 gene combined with radiotherapy enhances the suppressing effect on the growth of lung adenocarcinoma in the tumor-bearing nude mice. The inducibility of Egr1 promoter by radiation allows the targeting and controllability of treatment. Micro-PET-CT results have a good correlation with the treatment effectiveness. Therefore, it can be used in real-time evaluation of tumor metabolic function in vivo.

Plasminogen is a complement inhibitor.

Plasminogen is a 92-kDa single chain glycoprotein that circulates in plasma as a zymogen and when converted to proteolytically active plasmin dissolves preformed fibrin clots and extracellular matrix components. Here, we characterize the role of plasmin(ogen) in the complement cascade. Plasminogen binds the central complement protein C3, the C3 cleavage products C3b and C3d, and C5. Plasminogen binds to C3, C3b, C3d, and C5 via lysine residues, and the interaction is ionic strength-dependent. Plasminogen and Factor H bind C3b; however, the two proteins bind to different sites and do not compete for binding. Plasminogen affects complement action in multiple ways. Plasminogen enhanced Factor I-mediated C3b degradation in the presence of the cofactor Factor H. Plasminogen when activated to plasmin inhibited complement as demonstrated by hemolytic assays using either rabbit or sheep erythrocytes. Similarly, plasmin either in the fluid phase or attached to surfaces inhibited complement that was activated via the alternative and classical pathways and cleaved C3b to fragments of 68, 40, 30, and 17 kDa. The C3b fragments generated by plasmin differ in size from those generated by the complement protease Factor I, suggesting that plasmin-mediated C3b cleavage fragments lack effector function. Plasmin also cleaved C5 to products of 65, 50, 30, and 25 kDa. Thus, plasmin(ogen) regulates both complement and coagulation, the two central cascade systems of a vertebrate organism. This complement-inhibitory activity of plasmin provides a new explanation why pathogenic microbes utilize plasmin(ogen) for immune evasion and tissue penetration.

Involvement of α2-antiplasmin in dendritic growth of hippocampal neurons.

The α2-Antiplasmin (α2AP) protein is known as a principal physiological inhibitor of plasmin, but we previously demonstrated that it acts as a regulatory factor for cellular functions independent of plasmin. α2AP is highly expressed in the hippocampus, suggesting a potential role for α2AP in hippocampal neuronal functions. However, the role for α2AP was unclear. This study is the first to investigate the involvement of α2AP in the dendritic growth of hippocampal neurons. The expression of microtubule-associated protein 2, which contributes to neurite initiation and neuronal growth, was lower in the neurons from α2AP⁻/⁻ mice than in the neurons from α2AP⁺/⁺ mice. Exogenous treatment with α2AP enhanced the microtubule-associated protein 2 expression, dendritic growth and filopodia formation in the neurons. This study also elucidated the mechanism underlying the α2AP-induced dendritic growth. Aprotinin, another plasmin inhibitor, had little effect on the dendritic growth of neurons, and α2AP induced its expression in the neurons from plaminogen⁻/⁻ mice. The activation of p38 MAPK was involved in the α2AP-induced dendritic growth. Therefore, our findings suggest that α2AP induces dendritic growth in hippocampal neurons through p38 MAPK activation, independent of plasmin, providing new insights into the role of α2AP in the CNS.

Plasminogen-plasmin system in the pathogenesis and treatment of lung and pleural injury.

Lung and pleural injuries are characterized by inflammation, fibrinous transitional matrix deposition, and ultimate scarification. The accumulation of extravascular fibrin is due to concurrently increased local coagulation and decreased fibrinolysis, the latter mainly as a result of increased plasminogen activator inhibitor-1 (PAI-1) expression. Therapeutic targeting of disordered fibrin turnover has long been used for the treatment of pleural disease. Intrapleural fibrinolytic therapy has been found to be variably effective in clinical trials, which likely reflects empiric dosing that does not account for the wide variation in pleural fluid PAI-1 levels in individual patients. The incidence of empyema is increasing, providing a strong rationale to identify more effective, nonsurgical treatment to improve pleural drainage and patient outcomes. Therapeutics designed to resist inhibition by PAI-1 are in development for the treatment of pleural loculation and impaired drainage. The efficacy and safety of these strategies remains to be proven in clinical trial testing. Fibrinolytic therapy administered via the airway has also been proposed for the treatment of acute lung injury, but this approach has not been rigorously validated and is not part of routine clinical management at this time. Challenges to airway delivery of fibrinolysins relate to bioavailability, distribution, and dosing of the interventional agents.

Apparent involvement of plasmin in early-stage follicle rupture during ovulation in medaka.

Until recently, the role of the proteolytic system involving serine proteases in follicle rupture during ovulation in mammalian species has been a subject of controversy. We undertook the present study to examine whether proteases play a role in follicle rupture using the teleost medaka (Oryzias latipes) model. Various serine protease inhibitors, including a specific plasmin inhibitor, drastically reduced the rate of ovulation, as assessed by an in vitro ovulation assay, which was established for the fish. Biochemical, molecular biological, and immunological analyses demonstrated that plasminogen/plasmin was present in large follicles destined to ovulate. The active protease, plasmin, was detected in follicles approximately 3-7 h before the expected time of ovulation. Specific antibodies against the medaka plasmin light chain suppressed the ovulation rate of the follicles when antibodies were added to the medium during the period in which active plasmin was generated. This finding was an indication that a plasmin-like protease similar if not identical to plasmin plays a role in follicle rupture during ovulation in the medaka. Our data also indicate that this serine protease participates in the rupture for only a few hours prior to the activation of matrix metalloproteinase (Mmp)-mediated hydrolysis at ovulation. Based on our previous and current data, we propose a follicle rupture model involving two different proteolytic enzyme systems, serine protease and Mmp, in medaka ovulation. The current study is the first to provide evidence of the indispensable role of plasmin or a plasmin-like protease in the ovulation of a nonmammalian vertebrate species.

Plasminogen regulates stromal cell-derived factor-1/CXCR4-mediated hematopoietic stem cell mobilization by activation of matrix metalloproteinase-9.

OBJECTIVE: Granulocyte colony-stimulating factor (G-CSF) is a widespread therapeutic agent for stimulation of hematopoietic progenitor and stem cell (HPSC) mobilization from bone marrow (BM). Plasminogen (Plg) has been shown to be critical for HPSC mobilization. Here, we investigated the role of Plg in G-CSF-induced HPSC mobilization and the underlying mechanism. METHODS AND RESULTS: By using gene-targeted mice, our data show that Plg is required for G-CSF-induced HPSC egress to sinusoidal capillaries in BM and subsequent mobilization to peripheral circulation. G-CSF induced Plg-dependent activation of matrix metalloproteinase-9 (MMP-9) in BM, and MMP-9 neutralization or deficiency suppressed HPSC migration and mobilization. Reconstitution of MMP-9 activity by BM transplantation after lentiviral overexpression rescued HPSC mobilization in Plg-deficient mice, indicating that MMP-9 activation is required for Plg-mediated HPSC mobilization. Interestingly, after G-CSF simulation, Plg downregulated stromal cell-derived factor-1 in BM and spatiotemporally regulated the expression of C-X-C chemokine receptor type 4 (CXCR4) on mobilized HPSCs, and reconstitution of MMP-9 activity in Plg-deficient mice reversed CXCR4 expression on HPSCs in plasma and BM, suggesting that CXCR4 serves as a new downstream signal of Plg/MMP-9 in HPSC mobilization. CONCLUSIONS: Our data elucidated a novel mechanism that Plg regulates MMP-9-dependent CXCR4 expression to facilitate HPSC mobilization in response to G-CSF.

The plasmin-antiplasmin system: structural and functional aspects.

The plasmin-antiplasmin system plays a key role in blood coagulation and fibrinolysis. Plasmin and α(2)-antiplasmin are primarily responsible for a controlled and regulated dissolution of the fibrin polymers into soluble fragments. However, besides plasmin(ogen) and α(2)-antiplasmin the system contains a series of specific activators and inhibitors. The main physiological activators of plasminogen are tissue-type plasminogen activator, which is mainly involved in the dissolution of the fibrin polymers by plasmin, and urokinase-type plasminogen activator, which is primarily responsible for the generation of plasmin activity in the intercellular space. Both activators are multidomain serine proteases. Besides the main physiological inhibitor α(2)-antiplasmin, the plasmin-antiplasmin system is also regulated by the general protease inhibitor α(2)-macroglobulin, a member of the protease inhibitor I39 family. The activity of the plasminogen activators is primarily regulated by the plasminogen activator inhibitors 1 and 2, members of the serine protease inhibitor superfamily.

Plasminogen K5 activates mitochondrial apoptosis pathway in endothelial cells by regulating Bak and Bcl-x(L) subcellular distribution.

Plasminogen Kringle 5(K5) is a proteolytic fragment of plasminogen, which displays potent anti-angiogenic activities. K5 has been shown to induce apoptosis in proliferating endothelial cells; however the exact mechanism has not been well explored. The present study was designed to elucidate the possible molecular mechanism of K5-induced endothelial cell apoptosis. Our results showed that K5 inhibited basic fibroblast growth factors activated in human umbilical vein endothelial cells (HUVECs), indicating proliferation in a dose-dependent manner and induced endothelial cell death via apoptosis. K5 exposure activated caspase 7, 8 and 9. These results suggested that both the intrinsic mitochondrial apoptosis pathway and extrinsic pathway might be involved in K5-induced apoptosis. K5 reduced mitochondrial membrane potential (MMP) of HUVECs, demonstrating mitochondrial depolarization in HUVECs. K5 increased the ratio of Bak to Bcl-x(L) on mitochondria, decreased the ratio in cytosol, and had no effect on the total amounts of these proteins. K5 also did not effect on Bax/Bcl-2 distribution. K5 increased the ratio of Bak to Bcl-x(L) on mitochondrial that resulted in mitochondrial depolarization, cytochrome c release and consequently the cleavage of caspase 9. These results suggested that K5 induces endothelial cell apoptosis at least in part via activating mitochondrial apoptosis pathway. The regulation of K5 on Bak and Bcl-x(L) distribution may play an important role in endothelial cell apoptosis. These results provide further insight into the anti-angiogenesis roles of K5 in angiogenesis-related ocular diseases and solid tumors.

Plasmin induces apoptosis of aortic valvular myofibroblasts.

Previous studies have described remodelling of the extracellular substratum by matrix metalloproteinases (MMPs) in aortic valves. However, involvement of the fibrinolytic system has not yet been analysed. We hypothesized that plasminogen and plasminogen activator(s) are present in aortic valves and that plasminogen activation could induce the degradation of adhesive proteins and apoptosis of the valvular myofibroblasts. We employed ELISA, western blotting, fibrin-agar zymography, and immunochemistry to detect components of the plasminogen activation system, in samples of aortic valves and valvular myofibroblasts in primary culture. Using myofibroblast cultures, real-time measurement of plasminogen activation was performed in the absence and presence of inhibitors (amiloride, epsilon-aminocaproic acid, and an MMP inhibitor); the degradation of fibronectin was visualized on western blots; and the apoptotic process was assessed by detection of phosphatidylserine exposure (binding of FITC-annexin V) and DNA fragmentation (TUNEL and ELISA). We demonstrate that a time- and plasminogen concentration-dependent generation of plasmin occurs on the surface of cultured valvular myofibroblasts expressing both u-PA and t-PA. Only u-PA appears to activate plasminogen as t-PA is essentially found in complex with PAI-1. Plasmin-dependent degradation of pericellular proteins, such as fibronectin, leads to cell detachment and apoptosis. In conclusion, various proteins of the fibrinolytic system are synthesized in vitro by cultured myofibroblasts from aortic valves, leading to plasmin-dependent cell detachment-induced apoptosis, a biological process named anoikis. The presence of plasminogen in aortic valves suggests that this process may be operating in vivo and may participate in valvular tissue remodelling, as also suggested by the finding of apoptotic cells in valvular tissue. This is the first demonstration of the presence and potential role of enzymes of the fibrinolytic system in aortic valves.

Impaired wound healing in mice with a disrupted plasminogen gene.

Activation of plasminogen (Plg) has been proposed to play a role in proteolytic degradation of extracellular matrices in tissue remodeling events, including wound healing. However, there has been no definitive proof of involvement of Plg in such processes. We now report that healing of skin wounds is severely impaired in mice made deficient in Plg by targeted gene disruption. The results demonstrate that Plg is required for normal repair of skin wounds in mice and support the assumption that it also plays a central role in other disease processes involving extracellular matrix degradation, such as cancer invasion.

Plasminogen alleles influence susceptibility to invasive aspergillosis.

Invasive aspergillosis (IA) is a common and life-threatening infection in immunocompromised individuals. A number of environmental and epidemiologic risk factors for developing IA have been identified. However, genetic factors that affect risk for developing IA have not been clearly identified. We report that host genetic differences influence outcome following establishment of pulmonary aspergillosis in an exogenously immune suppressed mouse model. Computational haplotype-based genetic analysis indicated that genetic variation within the biologically plausible positional candidate gene plasminogen (Plg; Gene ID 18855) correlated with murine outcome. There was a single nonsynonymous coding change (Gly110Ser) where the minor allele was found in all of the susceptible strains, but not in the resistant strains. A nonsynonymous single nucleotide polymorphism (Asp472Asn) was also identified in the human homolog (PLG; Gene ID 5340). An association study within a cohort of 236 allogeneic hematopoietic stem cell transplant (HSCT) recipients revealed that alleles at this SNP significantly affected the risk of developing IA after HSCT. Furthermore, we demonstrated that plasminogen directly binds to Aspergillus fumigatus. We propose that genetic variation within the plasminogen pathway influences the pathogenesis of this invasive fungal infection.

[Mechanism of inhibitory effect of angiostatin on plasminogen activation by its physiologic activators].

The influence of angiostatin K1-4.5--a fragment of the heavy chain of plasmin and a powerful inhibitor of angiogenesis--on kinetic parameters (k(Pg) and K(Pg)) of human Glu-plasminogen activation under the action of urokinase (uPA) not having affinity for fibrin and fibrin-specific tissue plasminogen activator (tPA) was investigated. Angiostatin does not affect the k(Pg) value, but increases the value K(Pg) urokinase plasminogen activation. A decrease in the k(Pg) value and an increase in the K(Pg) value were found for fibrin-stimulated plasminogen activation by tPA with increasing concentrations of angiostatin. The obtained results show that angiostatin is competitive inhibitor of the uPA activator activity, while it inhibits the activator activity of tPA by mixed type. Such an influence ofangiostatin on the kinetic constants ofthe urokinase plasminogen activation suggests that angiostatin dose dependent manner replaces plasminogen in the binary enzyme-substrate complex uPA-Pg. In case of fibrin-stimulated plasminogen activation by tPA, both zymogen and tPA are bound to fibrin with formation of the effective triple tPA-Pg-fibrin complex. Angiostatin replaces plasminogen both from the fibrin surface and from the enzyme-substrate tPA-Pg complex that leads to a decrease in k(Pg) and an increase in K(Pg) of plasminogen activation. Inhibition constants by angioststin (Ki) of plasminogen-activator activities of uPA and tPA determined by Dixon method were found to be 0.59 +/- 0.04 and 0.12 +/- 0.05 microM, respectively.