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PubMed2000

Plasminogen activator activity in tears after excimer laser photorefractive keratectomy.

PURPOSE: To quantify changes of plasminogen activator activity in tear fluid during corneal re-epithelialization after excimer laser photorefractive keratectomy (PRK). METHODS: Tear samples were collected with glass capillaries from 77 eyes of 42 patients immediately before and immediately after PRK treatment and on postoperative days 3 and 5. In 20 patients, the contralateral eye was similarly sampled to serve as control. Plasminogen activator activity in the tear samples was measured by a spectrophotometric method using human plasminogen and chromogenic peptide substrate, D-valyl-L-leucyl-L-lysine-p-nitroanilide (S-2251). RESULTS: In tears of all eyes that underwent PRK, the plasminogen activator activities were lower immediately after PRK than were the preoperative values. For patient eyes with normal wound healing, tear plasminogen activator activities were significantly elevated above the preoperative level on the third postoperative day and then returned to the preoperative level by the fifth postoperative day. In contrast, tear plasminogen activator activities remained low through the third postoperative day in all (six) eyes in which haze developed after 3 to 6 months. The contralateral control eyes showed no appreciable change in plasminogen activator activity over the 5-day period. CONCLUSIONS: Plasminogen activator activity levels measured in tears of excimer laser PRK-treated eyes may serve as a predictor of wound healing. Extended low levels of plasminogen activator activity through the third postoperative day correlate with the development of corneal healing abnormalities (haze). The low plasminogen activator activity could be not only an accompanying sign but also a cause of defective corneal wound healing.

Adolescent↗
PubMedDate not supplied

Inhibition of tumor growth by plasminogen-related protein-B.

BACKGROUND: Various fragments of the fibrinolytic protein plasminogen can act as antiangiogenic factors and inhibit the growth of primary and metastatic tumors in mice. Plasminogen-related gene-B encodes a putative 9 kDa protein virtually identical to the plasminogen N-terminal activation peptide, a 77-amino acid motif that is liberated from the parent plasminogen molecule during conversion to the serine proteinase plasmin. Previous data have documented enhanced transcription of plasminogen-related gene-B in neoplastic tissues. MATERIALS AND METHODS: We have tested the effects of recombinant versions of plasminogen-related protein-B and the plasminogen N-terminal activation peptide on the growth of tumors in mice, employing murine tumor cell lines implanted subcutaneously. RESULTS: The recombinant plasminogen-related protein-B significantly inhibited the growth of primary tumors in mice, while recombinant plasminogen N-terminal activation peptide elicited only a slight inhibition of tumor growth. CONCLUSION: These data suggest that plasminogen-related protein-B may have utility as a novel cancer therapeutic.

Angiogenesis Inhibitors↗
PubMed1991

Abnormal plasminogen Maywood I.

Maywood I is a dysfunctional plasminogen. It is described in a patient (W.Y.) with a reduced plasma functional activity and with a low normal antigen level. Plasminogen was isolated from the patients plasma by affinity chromatography with L-lysine-substituted Sepharose. The protein yield was 86 mg/l, which was 88% of the plasma Plg antigen level; the specific activity was 24.4 IU/mg protein compared to 28.5 IU/mg protein for the native molecule. The protein was the Glu-form determined by SDS-PAGE and by isoelectric focusing. Six major isoelectric forms were found with isoelectric points between pH's 6.40 and 5.45. Titration of the equimolar plasminogen.streptokinase complex with p-nitrophenyl-p-guanidinobenzoate gave 85% active-sites indicating a homogenous population of molecules; therefore, the propositus is a homozygote. Four different plasminogen activators: a) streptokinase, b) urokinase c) the plasmin-derived light (B) chain-streptokinase complex, and d) tissue plasminogen activator (with soluble fibrin/CNBr-fibrinogen fragments) generated little plasmin from the variant plasminogen (4.5 to 45 nM), 5% or less than that generated from normal plasminogen. At 45 nM plasminogen, the molar ratio of plasminogen:activator was 3.0 for streptokinase, 3.9 for urokinase, 7.1 for the light (B) chain-streptokinase complex, and 155 for tissue plasminogen activator. In the equimolar variant plasminogen.streptokinase complex, the active-site was slowly developed, to a maximum of 85% in 40 min; in the normal complex, 100% active-sites were developed in 15 min. The variant plasminogen forms two equimolar complexes with streptokinase (I and II), with different mobilities in PAGE, in about equal amounts.(ABSTRACT TRUNCATED AT 250 WORDS)

Electrophoresis, Polyacrylamide Gel↗
PubMed1991

Fibronectin decreases the stimulatory effect of fibrin and fibrinogen fragment FCB-2 on plasmin formation by tissue plasminogen activator.

Fibronectin is a dimeric glycoprotein (Mr 440,000) involved in many adhesive processes. During blood coagulation it is bound and cross-linked to fibrin. Fibrin binding is achieved by structures (type I repeats) which are homologous to the "finger" domain of tissue plasminogen activator. Tissue plasminogen activator also binds to fibrin via the finger domain and additionally via the "kringle 2" domain. Fibrin binding of tissue plasminogen activator results in stimulation of its activity and plays a crucial role in fibrinolysis. Since fibronectin might interfere with this binding, we studied the effect of fibronectin on plasmin formation by tissue plasminogen activator. In the absence of fibrin, fibronectin had no effect on plasminogen activation. In the presence of stimulating fibrinogen fragment FCB-2, fibronectin increased the duration of the initial lag phase (= time period until maximally stimulated plasmin formation occurs) and decreased the rate of maximal plasmin formation which occurs after that lag phase mainly by increasing the Michaelis constant (Km). These effects of fibronectin were dose-dependent and were similar with single- and two-chain tissue plasminogen activator. They were also observed with plasmin-pretreated FCB-2. An apparent Ki of 43 micrograms/ml was calculated for the inhibitory effect of fibronectin when plasminogen activation by recombinant single-chain tissue plasminogen activator was studied in the presence of 91 micrograms/ml FCB-2. When a recombinant tissue plasminogen activator mutant lacking the finger domain was used in a system containing FCB-2, no effect of fibronectin was seen, indicating that the inhibitory effect of fibronectin might in fact be due to competition of fibronectin and tissue plasminogen activator for binding to fibrin(ogen) via the finger domain.

Fibrin↗
PubMed1991

Actin accelerates plasmin generation by tissue plasminogen activator.

Actin has been found to bind to plasmin's kringle regions, thereby inhibiting its enzymatic activity in a noncompetitive manner. We, therefore, examined its effect upon the conversion of plasminogen to plasmin by tissue plasminogen activator. Actin stimulated plasmin generation from both Glu- and Lys-plasminogen, lowering the Km for activation of Glu-plasminogen into the low micromolar range. Accelerated plasmin generation did not occur in the presence of epsilon-amino caproic acid or if actin was exposed to acetic anhydride, an agent known to acetylate lysine residues. Actin binds to tissue plasminogen activator (t-Pa) (Kd = 0.55 microM), at least partially via lysine-binding sites. Actin's stimulation of plasmin generation from Glu-plasminogen was inhibited by the addition of aprotinin and was restored by the substitution of plasmin-treated actin, indicating the operation of a plasmin-dependent positive feedback mechanism. Native actin binds to Lys-plasminogen, and promotes its conversion to plasmin even in the presence of aprotinin, indicating that plasmin's cleavage of either actin or plasminogen leads to further plasmin generation. Plasmin-treated actin binds Glu-plasminogen and t-PA simultaneously, thereby raising the local concentration of t-PA and plasminogen. Together, but not separately, actin and t-PA prolong the thrombin time of plasma through the generation of plasmin and fibrinogen degradation products. Actin-stimulated plasmin generation may be responsible for some of the changes found in peripheral blood following tissue injury and sepsis.

Actins↗
PubMed1991

On the mechanism of fibrin-specific plasminogen activation by staphylokinase.

The mechanism of plasminogen activation by recombinant staphylokinase was studied both in the absence and in the presence of fibrin, in purified systems, and in human plasma. Staphylokinase, like streptokinase, forms a stoichiometric complex with plasminogen that activates plasminogen following Michaelis-Menten kinetics with Km = 7.0 microM and k2 = 1.5 s-1. In purified systems, alpha 2-antiplasmin inhibits the plasminogen-staphylokinase complex with k1(app) = 2.7 +/- 0.30 x 10(6) M-1 s-1 (mean +/- S.D., n = 12), but not the plasminogen-streptokinase complex. Addition of 6-aminohexanoic acid induces a concentration-dependent reduction of k1(app) to 2.0 +/- 0.17 x 10(4) M-1 s-1 (mean +/- S.D., n = 5) at concentrations greater than or equal to 30 mM, with a 50% reduction at a 6-aminohexanoic acid concentration of 60 microM. Staphylokinase does not bind to fibrin, and fibrin stimulates the initial rate of plasminogen activation by staphylokinase only 4-fold. Staphylokinase induces a dose-dependent lysis of a 0.12-ml 125I-fibrin-labeled human plasma clot submersed in 0.5 ml of citrated human plasma; 50% lysis in 2 h is obtained with 17 nM staphylokinase and is associated with only 5% plasma fibrinogen degradation. Corresponding values for streptokinase are 68 nM and more than 90% fibrinogen degradation. In the absence of a fibrin clot, 50% fibrinogen degradation in human plasma in 2 h requires 790 nM staphylokinase, but only 4.4 nM streptokinase. These results suggest the following mechanism for relatively fibrin-specific clot lysis with staphylokinase in a plasma milieu. In plasma in the absence of fibrin, the plasminogen-staphylokinase complex is rapidly neutralized by alpha 2-antiplasmin, thus preventing systemic plasminogen activation. In the presence of fibrin, the lysine-binding sites of the plasminogen-staphylokinase complex are occupied and inhibition by alpha 2-antiplasmin is retarded, thus allowing preferential plasminogen activation at the fibrin surface.

Antifibrinolytic Agents↗
PubMed1988

Modified crossed immunoelectrophoresis to study with whole plasma the reversible complex formation of histidine-rich glycoprotein with plasminogen.

To study the reversible complex formation between the plasma protein histidine-rich glycoprotein (HRG) and plasminogen, crossed immunoelectrophoresis of HRG was modified. In the modification, purified plasminogen was introduced into the gel of the first dimension electrophoresis. Two molecular forms of plasminogen, Glu- and Lys-plasminogen, induced a dose-dependent reduction of the electrophoretic mobility of HRG, with a half maximal retardation for both plasminogens at 0.50-0.55 microM of added plasminogen to the agarose gel. HRG in plasma behaved as a uniform fraction with respect to plasminogen binding. In contrast, with the same modified technique another plasma protein, alpha 2-antiplasmin, separated into a retarded plasminogen-binding form and a non-retarded non-plasminogen-binding form. The method can be used to assess several aspects of reversible complex formation between plasma proteins, as demonstrated for plasminogen binding of HRG and alpha 2-antiplasmin in whole plasma.

Binding, Competitive↗
PubMedDate not supplied

[Lysine- and arginyl-binding sites of plasminogen domains].

Hydrolysis of plasminogen permits obtaining its nine fragments. The method of differential scanning microcalorimetry reveals seven domains in plasminogen, and the affinity chromatography--three lysin- and three arginyl-binding sites. The lysin-binding sites of domains (Kringles) K1 and K4 differ in ligand specificity. Benzamidine-binding sites of domain K5 and of plasmin light chain are simultaneously arginine-binding ones. The third arginyl-binding site differing from the benzamidine-binding one is found in fragment K1-3. In the plasminogen-fibrin interaction only lysin-binding sites of plasminogen take part; in the plasminogen fragments-fibrinogen fragments interaction both types of plasminogen sites participate. The heavy chain of plasmin interacts with the E-fragment of fibrinogen by the lysin-binding sites, and the light chain of plasmin interacts with D-fragment of fibrinogen by arginyl-binding sites. Sites complementary to arginyl binding sites of plasminogen are located on the DH-fragment and sites of interaction with lysin- and arginyl-binding sites--on the DL-fragment. The plasmin-fibrin interaction mediated by sites of the first four cringles is not associated with changes in the catalytic function of the active centre. Interaction of Lys-plasminogen with fibrin accelerates polymerization of the latter. The effect of Lys-plasminogen is conditioned by the lysin-binding sites. Glu-plasminogen has no effect on the polymerization process.

Arginine↗
PubMed1988

Tissue-type plasminogen activator binding to human endothelial cells. Evidence for two distinct binding sites.

The endothelium may contribute to fibrinolysis through the binding of plasminogen activators or plasminogen activator inhibitors to the cell surface. Using a solid-phase radioimmunoassay, we observed that antibodies to recombinant tissue-type plasminogen activator (rt-PA) and plasminogen activator inhibitor type 1 (PAI-1) bound to the surface of cultured human umbilical vein endothelial cells (HUVEC). HUVEC also specifically bound added radiolabeled rt-PA with apparent steady-state binding being reached by 1 h at 4 degrees C. When added at low concentrations (less than 5 nM), rt-PA bound with high affinity mainly via the catalytic site, forming a sodium dodecyl sulfate-stable 105-kDa complex which dissociates from the cell surface over time and which could be immunoprecipitated by a monoclonal antibody to PAI-1. rt-PA bound to this high affinity site retained less than 5% of its expected plasminogen activator activity. At higher concentrations, binding did not require the catalytic site and was rapidly reversible. rt-PA initially bound to this site retained plasminogen activator activity. These studies suggest that tissue-type plasminogen activator and PAI-1 are expressed on the surface of cultured HUVEC. HUVEC also express unoccupied binding sites for exogenous tissue-type plasminogen activator. The balance between the expression of plasminogen activator inhibitors and these unoccupied binding sites for plasminogen activators on the endothelial surface may contribute to the regulation of fibrinolysis.

Binding Sites↗
PubMed1996

Functional significance of NH2- and COOH-terminal regions of staphylokinase in plasminogen activation.

Structure/function relationships in the activation of plasminogen with staphylokinase were studied using mutants of recombinant staphylokinase (Sak42D). Deletion of up to 10 NH2-terminal amino acids (Sak42D delta N10) did not affect plasminogen activation, but removal of 11 amino acids completely abolished the ability to activate plasminogen. Elimination of potential plasmin cleavage sites in the NH2-terminal region yielding mutants Sak42D(K8H,K10H,K11H) and Sak42D(K6H,K8H,K11H) did not alter the rate of the exposure of a proteolytically active site (amidolytic activity) in equimolar mixtures with plasminogen, but destroyed the plasminogen activator properties of these muteins. Deleting two residues following the preferred processing site at position 10 (Sak42 delta (K11,G12)) resulted in a mutein also inactive in plasminogen activation. Removal of the COOH-terminal Lys136, yielding Sak42D delta C1, or of Lys135 and Lys136 in Sak42D delta C2 resulted in proteins with strongly reduced plasminogen activation capacity. In contrast, substitution of Lys135 and Lys136 with Ala in Sak42D(K135A,K136A) did not affect activation. Cyanogen bromide cleavage of Sak42D(M26L,E61M,D82E) produced a 61 amino acid NH2-terminal and a 65 amino acid COOH-terminal fragment which did not activate plasminogen, but bound to plasminogen with affinity constants Ka of 4.0 x 10(5) M-1 and 1.4 x 10(7) M-1, respectively (as compared to a Ka of 1.1 x 10(8) M-1 for Sak42D). These results indicate that Lys11 and the COOH-terminal region of staphylokinase play a key role in the activation of plasminogen.

Enzyme Activation↗
PubMed1997

Enhancement of plasminogen binding to U937 cells and fibrin by complestatin.

Plasminogen binds to endothelial and blood cells as well as to fibrin, where the zymogen is efficiently activated and protected from inhibition by alpha 2-antiplasmin. In the present study we have found that complestatin, a peptide-like metabolite of a streptomyces, enhances binding of plasminogen to cells and fibrin. Complestatin, at concentrations ranging from 1 to 5 microM, doubled 125I-plasminogen binding to U937 cells both in the absence and presence of lipoprotein(a), a putative physiological competitor of plasminogen. The binding of 125I-plasminogen in the presence of complestatin was abolished by epsilon-aminocaproic acid, suggesting that the lysine binding site(s) of the plasminogen molecule are involved in the binding. Equilibrium binding analyses indicated that complestatin increased the maximum binding of 125I-plasminogen to U937 cells without affecting the binding affinity. Complestatin was also effective in increasing 125I-plasminogen binding to fibrin, causing 2-fold elevation of the binding at approximately 1 microM. Along with the potentiation of plasminogen binding, complestatin enhanced plasmin formation, and thereby increased fibrinolysis. These results would provide a biochemical basis for a pharmacological stimulation of endogenous fibrinolysis through a promotion of plasminogen binding to cells and fibrin.

Cell Line↗
PubMed1976

Interaction of streptokinase with plasminogen. Isolation and characterization of a streptokinase degradation product.

When streptokinase is incubated with human or rabbit plasminogen, one event which occurs is a specific fragmentation of streptokinase. At least five major identifiable streptokinase fragments appear with time, and they possess molecular weights of approximately 40,000 (SK 1), 36,000 (SK 2), 31,000 (SK 3), 26,000 (SK 4), and 10,000 (SK 5) under denaturing conditions, as observed on calibrated sodium dodecyl sulfate-polyacrylamide gels, compared to native streptokinase of molecular weight 45,000. The amount of each of the fragments generated at given times of incubation of plasminogen and streptokinase depends upon the species of plasminogen employed. Utilizing rabbit plasminogen and streptokinase, the SK 4 fragment was purified. This fragment arises by proteolysis at both the NH2 and COOH regions of native streptokinase. However, when isolated utilizing dilute aqueous buffers, the SK 4 fragment contained a portion of the original NH2 terminus of native streptokinase noncovalently bound to the molecule (SK 4'). SK 4' is capable of activating human plasminogen to plasmin, albeit more slowly than native streptokinase. However, the SK 4'-human plasmin complex possess only very weak plasminogen-activating activity toward sheep plasminogen. Upon removal of the noncovalently bound small NH2-terminal peptide of native streptokinase from SK 4', SK 4 is formed. This particular fragment possesses practically no human plasminogen-activating activity and cannot be used as an activator of sheep plasminogen, even with added human plasminogen.

Amino Acid Sequence↗
PubMed1997

The human ENO1 gene product (recombinant human alpha-enolase) displays characteristics required for a plasminogen binding protein.

Plasminogen binds with low affinity in a lysine-dependent manner to many cell types. Previously, a 54 kDa plasminogen receptor found on the surface of U-937 cells was identified as an alpha-enolase-like molecule. The aims of this study were to determine whether recombinant alpha-enolase (r-alpha-enolase), encoded by ENO1, was a plasminogen binding protein and to generate polyclonal antibodies against this antigen. Plasminogen specifically bound r-alpha-enolase with a Kd 1.9 microM and approached saturation at 10 microM. Lysine-dependent plasminogen binding to r-alpha-enolase was demonstrated by a greater than 80% inhibition of binding by the lysine analogues epsilon-amino caproic acid and tranexamic acid, whilst only 14% inhibition occurred with the arginine analogue benzamidine. Removal of the C-terminal lysine residue of r-alpha-enolase with carboxy-peptidase B significantly reduced its plasminogen binding capacity, suggesting that binding required C-terminal lysine residue of r-alpha-enolase. Binding to r-alpha-enolase enhanced the activation rate of plasminogen by urokinase but prevented alpha 2-antiplasmin from binding plasminogen. Taken together, these data suggest that the gene product of human ENO1 encodes an authentic plasminogen binding protein.

Binding, Competitive↗
PubMedDate not supplied

Augmented pulse-spray thrombolysis with tPA by early pulsed intrathrombic plasminogen enrichment.

PURPOSE: This study was designed to evaluate the efficacy of plasminogen enrichment of subacute thrombus in further accelerating pulse-spray pharmacomechanical thrombolysis (PSPMT) with urokinase (UK) or tissue plasminogen activator (tPA) in a rabbit model. MATERIALS AND METHODS: With use of a subacute rabbit inferior vena cava (IVC) thrombosis model, 78 rabbits were divided into eight groups according to the agents used for thrombolysis: (i) controls (IVC thrombosis, no lysis performed), (ii) pulse-spray thrombolysis with saline only, (iii) PSPMT with UK, (iv) PSPMT with UK, plus interim pulse-spray plasminogen enrichment after 14 minutes, (v) pulse-spray plasminogen enrichment, followed at 10 minutes by PSPMT with UK, (vi) PSPMT with tPA, (vii) PSPMT with tPA, plus interim plasminogen enrichment, and (viii) pulse-spray plasminogen enrichment, followed at 10 minutes by PSPMT with tPA. RESULTS: Intrathrombic pulsed injection of glu-plasminogen after 14 minutes of tPA PSPMT demonstrated significant augmentation of lysis (approximately 31% decrease in residual thrombus) compared with tPA alone (P = .006). Lysis was not augmented significantly when plasminogen was sprayed into thrombus before tPA, or before or after UK. CONCLUSION: Plasminogen enrichment of thrombus after onset of PSPMT with tPA significantly accelerated thrombolysis in a subacute in vivo rabbit model. A clinical trial of this method may be warranted.

Animals↗
PubMed1979

Plasminogen activation transforms the morphology of quiescent 3T3 cell monolayers and initiates growth.

Plasminogen activator of cell origin converts the plasma protein plasminogen to the proteolytic enzyme plasmin. Recently, high levels of activator have been observed to be particularly associated with tumours and transformed cells, and a functional relationship between plasminogen activation and malignancy has been proposed. In this paper we have attempted to induce transformation-like morphology and growth in a population of confluent quiescent cells in tissue culture, by inducing plasminogen activation. Untransformed 3T3 cells grown to confluence in plasminogen-free medium were subjected to plasminogen activation by the addition of urokinase and plasminogen or plasminogen-containing acid-treated serum, or plasmin. Under these conditions, the previously well ordered monolayers became disrupted, with multilayering, and discontinuities in the cell sheet, and the cells simultaneously grew to significantly higher densities. Removal of the plasmin-containing medium supplements effected some restoration of normal morphology. Thus, lhen plasmin was present 3T3 cells did not become transformed, but expresses transformation-like features. Well ordered monolayer morphology and quiescence in 3T3 cells at confluence are therefore dependent upon the absence of plasminogen activation.

Animals↗
PubMed1996

Altered expression of gelatinase and surface-associated plasminogen activator activity by trophoblast cells isolated from placentas of preeclamptic patients.

OBJECTIVE: This study compared in vitro degradative properties of trophoblasts isolated from placentas of preeclamptic patients with those of trophoblasts isolated from placentas of patients with uncomplicated pregnancies. Specifically, the expression of gelatinases, plasminogen activators, and plasminogen activator inhibitor type 1 by these cells was examined. STUDY DESIGN: Gelatinase and plasminogen activator secretion were determined by zymography, whereas antigenic levels of urinary-type plasminogen activator and plasminogen activator inhibitor type 1 in culture-conditioned medium were determined by enzyme-linked immunosorbent assay. We also evaluated expression of cell-surface plasminogen activator activity with a sensitive assay that uses the fluorescent plasmin substrate H-D-Val-Leu-Lys-7-amino-4-methylcoumarin. RESULTS: Cells from both normal and preeclamptic placentas secreted variable levels of gelatinase, of which the most predominant was matrix metalloproteinase-9 (gelatinase B). In 60% of the media conditioned by preeclamptic cells the matrix metalloproteinase-9 present was exclusively of a higher molecular weight (> 92 kd) than the predominant enzyme secreted by trophoblasts isolated from normal placentas. Incubation of the preeclamptic cell culture-conditioned media with p-aminophenylmercuric acetate, an activator of metalloproteinases, resulted in a decrease in the apparent molecular weight of the enzyme, indicating that most of the matrix metalloproteinase-9 contained in these samples was in the inactive form. Although trophoblasts from normal and preeclamptic placentas secreted similar amounts of urinary-type plasminogen activator and plasminogen activator inhibitor type 1, the latter cells expressed significantly less (p < 0.001) cell surface plasminogen activator activity. CONCLUSION: These results suggest that abnormal uteroplacental blood flow in preeclampsia (as a result of either shallow invasiveness of the uterine arteries or excessive fibrin deposition within the intervillous spaces) might result from altered expression of active proteinases by trophoblasts.

Adult↗
PubMed2000

The efficacy of plasminogen-urokinase combination in inducing posterior vitreous detachment.

PURPOSE: To investigate the toxicity of intravitreal plasminogen, urokinase, and their combination, and to evaluate their efficacy in the production of posterior vitreous detachment (PVD) in the rabbit eye. METHODS: Fifty-six albino New Zealand rabbits were examined before and after injection using the indirect ophthalmoscope, slit-lamp biomicroscopy, and electroretinography. Various concentrations of urokinase or recombinant plasminogen or a combination were injected intravitreally into the right eyes of four rabbits for each concentration. The left eyes of the animals served as controls and received 0.1 mL balanced salt solution. Group 1 was injected with pure urokinase (1,000, 5,000, or 10,000 IU); Group 2 with recombinant plasminogen (0.1, 0.4, 1.0, 2.0, 4.0, 8.0, or 16.0 caseinolytic units [CU]); and Group 3 with a combination of 1,000 IU urokinase (highest nontoxic dose) and nontoxic concentrations of plasminogen (0.1, 0.4, 1.0, or 2.0 CU). The animals were killed and the eyes enucleated 15 days after injection. Electron and light microscopy were performed. RESULTS: A concentration of 1,000 IU of urokinase was found to be nontoxic to the retina. Plasminogen concentrations of 2.0 CU or less did not produce retinal toxicity, whereas 4.0, 8.0, and 16.0 CU of plasminogen caused minimal-to-severe inflammatory response in the vitreous without histologic or electroretinographic changes. Neither plasminogen nor urokinase alone was successful in producing PVD. The combination of 1,000 IU of urokinase and 1.0 to 2.0 CU of plasminogen was effective without causing retinal toxicity. CONCLUSION: Posterior vitreous detachment can be produced in the rabbit eye using a combination of plasminogen and urokinase.

Animals↗
PubMed1999

Plasminogen activation in human leukemia and in normal hematopoietic cells.

The active process of pericellular proteolysis is central in tumor invasion, and in particular the essential role of the urokinase-type plasminogen activator (uPA) is well established. uPA-mediated plasminogen activation facilitates cell migration and invasion through extracellular matrices by dissolving connective tissue components. uPA, its receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1) are enriched in several types of tumors. The importance of proteolysis and especially plasminogen activation is less clear in hematopoietic malignancies than in solid tumors. However, patients with leukemia have an increased tendency to bleeding, not always attributable to thrombocytopenia, and tissue infiltration by leukemic cells, processes in which plasminogen activation may be involved. Several studies have indicated that plasminogen activators (PAs) are highly expressed by cultured leukemia cells. Furthermore, differing from adherent tumor cells, leukemic cells have an enhanced capacity to activate pro-uPA and mainly the active form of uPA is released to culture medium. Ex vivo studies have shown that uPAR, uPA and its inhibitors can be found on the surface of normal blood cells and on the blast cell surfaces from patients with acute leukemia as well as from plasma samples. Elevated levels of PAs and their inhibitors have been detected in leukemic cell lysates. Few studies have tried to demonstrate a correlation between prognosis of leukemia and levels of plasminogen activators. More in vivo studies are needed to show, if any of the factors of the plasminogen activation process can be used as tools in subclassification or as markers for prognosis in leukemia. This review article will focus on the in vivo studies of plasminogen activation in leukemia and will present several in vitro findings on PAs in normal leukocytes and leukemic cell lines.

Blood Cells↗